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Technical Information Center,Office of Public Affairs
ENERGY RESEARCH AND DEVELOPMENT ADMINISTRATION
Gas Bu
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Library of Congress Cataloging in Publication Data
Main encry under title:
Gas bubble disease.
"Conf·7410B."
Includes index.
Supt.of Docs.no.:ER 1.ll:CONF-7410B
1:Gas bubble disease in fish-Congresses.I.Fickeisen.D.H.
II.Schneider,"'ark Joseph,1942·Ill.Battelle Memorial
Instituce.Columbus.Ohio,Pacific Northwest Laboratory,Richland,
Wash.IV.United States,Atomic Energy Commission.Division
of Biomedical and Environmental Research.
(DNLM:1.Gas poisoning-<;ongresses.2.Water pollution,
Chemical-Congresses.QH90.A3 G246 1974)
SH177.G3G37 597',02'4 75·619327
ISBN 0-87079-{1234
Available as CONF·741033 for $6.00 (foreign,$8.50)from
National Technical Information Service
U.S.Department of Commerce
Springfield,Virginia 22161
ERDA Distribution Category UC·12
Composed by Battelle,Pacific Northwest laboratories
Richland.WashingtOfl
Printed in the United States of America
ERDA Technical Information Center.Oak Ridge,Tennessee
February 1976
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CONF-741033
Gas Bubble Disease
Proceedings of-a Workshop held at Richland,Washington,
October 8-9,1974
Cosponsored by Battelle,Pacific Northwest
Laboratories,and U.S.Atomic Energy Commission
Editors
D.H.Fickeisen
and
M.J.Schneider
1976
ARLIS
.Alaska Resources
LIbrary &Infonnation SeTVlces
Anchorage,Alaska
Published by
Technical Information Center,Office of Public Affairs
Energy Research and Development Administration
Foreword
We are happy to host the Nitrogen Task Force and
other interagency participants of this workshop
on gas bubble disease.Six years ago,Battelle first
participated in a tripartite research program of the
Environmental Protection Agency (EPA),the Na-
tional Marine Fisheries Service,and the Atomic
Energy Commission involving many of the people
here today.One needs only to compare·the state of
knowledge on gas bubble disease at the time of the
tripartite study·with this workshop proceedings
in order to fully appreciate the extent to which
refinement and accurate delineation of the super-
saturation problem has taken place."In 1971,the
effects of gas bubble disease seemed to be con-
founded with those of other stressors;in the present
series of papers,we now see it as a quite lethal factor
initiated at a fairly critical level of supersaturation.
We also know a good deal about its pathology.With
the limited resources available to investigators of
gas supersaturation effects,the findings here repre-
sent significant accomplishments.
Several participants have identified areas of
uncertainty requiring continuing research during the
years ahead.I would like to comment on several
points.In listening to the past two days'delibera-
tions,I believe that four areas require early atten-
tion.They require early attention because data
useful in practical applications to minimize gas
bubble mortality will depend in a key way on our
understanding of the underlying processes.First,it
seems to me that determinations are needed of the
vertical distributions of fishes with respect to gas
supersaturation,especially at periods of high runoff
in the river system of the Columbia,for example.To
too large an extent,models in use are approxima-
tions that need fairly systematic validation for each
application.Appropriate field effort,as was de-
'fPA-Co/umbia River Thermal Effects Study
Vol.1 -Biological Effects Studies.January 1971
VoL 2 -Temperature Prediction Studies,January 1971
scribed yesterday,is time-consuming and hence
expensive when applied to validation purposes.It is
also necessary,considering the confoundment
caused by possible habitat preferences of fish.
Second,field effort should be directed to estab-
lishing systematically the population pressures and
habitat preferences of various fish.Fish seem to
have limited ability to distinguish levels or to detect
critical levels of gas supersaturation,per se;thus,
it is not difficult to see how mortality in salmonids
exposed to low levels of gas supersaturation might
be greatly aggravated by flight to avoid predation
by squawfish,for example.
Third,pressure-equilibration relationships in
fish need to be better defined at physiological levels.
We need to keep in mind the experience of hyper-
baric physiologists in other fields;namely that tissue
gas equilibration,while varying inversely with pres-
sure and time,is probably a multi-compartmental
process showing widely differing rate constants.
Thus,a small compartment,slow in equilibrating,
may trigger a neurological incapacitation during
decompression even though the body fluids,gener-
ally,seem to be equilibrated at the lower pressure.
Certain delayed effects,also described in the past
two days,may have a similar explanation.
In the papers and the round table discussions
that follow,a number of related ideas are developed.
If past progress is a guide,I look forward in our
future meetings to definitive explanations of these
problems affecting hydroelectric power development.
Burton E.Vaughan,Manager
Ecosystems Department
Battelle,Pacific Northwest laboratories
iii
-
Preface
Gas bubble disease resulting from exposure of
aquatic organisms to water supersaturated with
dissolved gas was described some 100 years ago,
but only in the past decade has a serious problem
been recognized in a natural river system.Opera-
tion of spillways at hydroelectric generating facili-
ties on the Columbia River and its major tributaries
results in entrainment of air into the·river water,
increasing the dissolved gas content to supersatu-
rated levels.The resulting gas bubble disease is a
factor contributing to observed declines in salmo-
nid fish stocks.Even more recently,a widespread
gas bubble disease problem has been realized with
fish kills in steam generating station discharge
plumes,both at freshwater and marine sites.In
these cases,supersaturation is caused by the de-
crease in gas solubility which accompanies heating
of water used to cool condensers.
In the past several years,intensive research
efforts have been undertaken by several agencies
in the Pacific Northwest.These projects have been
broadly scoped with major goals of prediction of
gas levels and their effects,reduction of impacts
on populations of aquatic biota,and development
of national water quality criteria and standards.
Many of these studies have progressed to a point of
having a sufficient data base to begin drawing
major conclusions and we have been encouraged
by work in other regions.The Nitrogen Task Force
has served as an informal forum for exchange of
ideas and development of plans for future work,
but we felt a need for a Gas Bubble Disease Work-
shop to draw together on-going research within the
Pacific Northwest as well as other regions.The
workshop format included formal presentation of
papers printed herein and a series of informal
round table discussions charged with determining
research needs in specific areas of interest.Notes
of the round table discussions are also included
herein.It is with the hope of stimulating further
research efforts and providing a comprehensive
view of on-going projects that we present these
proceedings.
The success of the Gas Bubble Disease Work-
shop was certainly due to the efforts of participants
for which we thank them.Those who accepted our
invitations to co-chair round table discussions con-
tributed significantly to the workshop.Additionally,
many Battelle-Northwest staff members contributed
significantly to the workshop,and their aid is also
appreciated.In particular,J.C.Montgomery,R.W.
Hanf,Jr.,J.c.Mourich,and J.L.Helbling deserve
special recognition for their hard work.
Duane H.Fickeisen
Mark J.Schneider
Richland,Washington
November 1975
For Battelle-Northwest and the Division of Bio-
medical and Environmental Research of the Energy
Research and Development Administration.
v
Gas Bubble Disease of Salmonids:Variation in Oxygen-Nitrogen Ratio with
Constant Total Gas Pressu re 85
R.R.Rucker
Effect of Gas Bubble Disease on Lateral line Function in Juvenile Steelhead Trout 89
M.H.Schiewe,D.D.Weber
Effects of Stress on Salmonid Blood Clotting Mechanisms 93
E.Casilfas,L.Smith.B.G.D'Aoust
Changes in Blood Chemistry of Juvenile Steelhead,Salmo gairdneri,Following
Sublethal Exposure to Nitrogen Supersaturation 96
T.W.Newcomb
Continuous Monitoring of Total Dissolved Gases,a Feasibility Study 101
T.F.Jenkins
An Electronic Monitor for Total Dissolved Gas Pressure
B.G.D'Aoust,R.White,H.Seibold
106
Round Table Discussions 111
Biological Studies:laboratory Orientation 112
A.V.Nebeker,D.H.Fickeisen
Biological Studies:Field Orientation ,114
W.Ebel,R.McConnell
Analytical Methods.... . . .. .. . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ...116
M.J.Schneider,B.G.D'Aoust
Physics of Dissolved Gases and Engineering Solutions , ,,.118
G.C Richardson,R.Baca
Water Quality Standards.. . . .. . . . . .. . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . ...120
R.L.Rufifson,R.Pine
list of Participants 121
Index
viii
123
-
--
Contents
Foreword iii
Preface v
Effe,&s oflong-Term Exposure to Supersaturation of Dissolved Atmospheric
Gases on Juvenile Chinook Salmon and Steelhead Trout in Deep and Shallow
TestTanks 1
;,,E.M.Dawley,M.Schiewe,B:Monk
Gas Supersaturation Research National Marine Fisheries Service
Prescott Facility-1971 to 1974 11
T.H.Blahm,B.McConnell,G.R.Snyder
Equipment and Techniques for Monitoring the Vertical Distribution of Fish
in Shallow Water 20
W.Marshall
Dissolved Gas Supersaturation:Live Cage Bioassays at Rock Island Dam,Washington 24
D.E.Weitkamp
Supersaturation and Fishery Observations in Selected Alpine Oregon Streams 37
G.R.Bouck
Some Effects of Excess Dissolved Gas on Squawfish,Ptychocheilus oregonensis
(Richardson),......................................................41
W.W.Bentley,E.M.Dawley,T.W.Newcomb
Responses of Coho Salmon (Oncorhynchus kisutch)to Supersaturation at
One Atmosphere 47
D.Beyer,B.G.D'Aoust,L.Smith
Effects of Gas Supersaturated Water on Freshwater Aquatic Invertebrates 51
A.V.Nebeker,D.G.Stevens,J.R.Brett
A Study of the Pathogenesis of Gas Bubble Disease in Steelhead Trout
(Sa/mo gairdneri)...........................................................................•...66
R.K.Stroud,A.V.Nebeker
effect of Temperature on Tolerance to Dissolved Gas Supersaturation of Black
Bullhead,Jetalurus me/as 72
D.H.Fickeisen;J.C Montgomery,R.W.Hanf,Jr.
Gas Bubble Disease Mortality of Atlantic Menhaden,Brevoortia Tyrannus,at a
Coastal Nuclear Power Plant 75
R.A.Marcello,Jr.,R.B.Fairbanks
Observations on the Effects of Gas Embolism in Captured Adult Menhaden 81
A.Clay,A.Barker,S.Testaverde,R.Marcel/o,G.C.McLeod
vii
IE.M.Dawley
M.Schiewe
B.Monk
Effects of Long-
Term Exposure to
Supersaturation of Dissolved
Atmospheric Gases on Juvenile
Chinook Salmon and Steelhead
Trout in Deef)and Shallow
Test Tanks
-
ABSTRACT
Bioassays in shallow (0.25 m)and deep (2.5 m)tanks with dis-
solved atmospheric gas concentrations ranging from 100 to
127%of saturation in water at 10°C were conducted to deter-
mine the lethal and sublethal effects on juvenile fall chinook
Oncorhynchus tschawYlscha and steelhead trout Sa/mo gairdneri.
Juvenile fall chinook (38.7 to 41.3 mm)were much more
resistant to supersaturation than juvenile steelhead (164 to
196 mm).Chinook tested in the shallow tanks at 120%of super-
saturation incurred 500A".mortality after 22 days,whereas steel-
head tested at the same level incurred 500A".mortality in 30 hr.
Gas bubble disease signs were noted on mortalities and on live
subsamples taken every 28 days.Vertical distribution of both
chinook and steel head groups in the deep tanks appeared to
compensate for about 10%and 10 to 15%,respectively,of effec-
tive saturation.Average depths of the fish tested in deep tanks
increased with increase"gas concentration.Significant differ-
ences in growth and condition factor were not found between
stressed and control fish during the test period.
Effects of supersaturation of dissolved atmo-
spheric gases on freshwater fishes have been
studied by many irivestigatorssince the late 1800's.
The current problem of supersaturation in the
Columbia and Snake:Rivers (Ebel,1969;Beiningen
and Ebel,1970;Ebel,1971;Meekin and Allen,1974)
has resulted in renewed interest in effects of super-
saturation on fish,and a great deal of research has
recently been accomplished by fisheries and other
agencies in the Pacific Northwest.Information on
the resistance of indigenous fish species to high
gas concentrations is well-documented for expo-
sure in shallow water for short periods of time
(Rucker and Tuttle,1948;Harvey and Cooper,1962;
Coutant and Genoway,1968;Bouck,et aJ.,1970;Ebel,
Dawley,and Monk,1971;Bouck,1972;Blahm,
McConnell,and Snyder,1973;Fickeisen,et aI.,1973;
Dawley and Ebel,1974)but there are still many un-
Jnswered questions regarding the effects of chronic
low-level exposure on survival.Fish may be subjected
to low levels of supersaturation in two ways.They
may inhabit water areas where they cannot com-
pensate for gas saturation by sounding,or they
may inhabit deep water areas where hydrostatic
pressure offsets the effects of high gas levels.
The National Marine Fisheries Service,funded
in part by the Environmental Protection Agency
(EPA)in 1972,began investigations of chronic
effects of long-term exposure of juvenile fall chi-
nook to various low levels of supersaturation.In
this report we describe those effects observed from
deep and shallow water tanks on juvenile fall chi-
nook salmon Oncorhynchus tschawytscha,and juve-
nile steelhead trout,Sa/mo gairdneri.
MATERIALS AND METHODS
Two bioassays of dissolved gas were conducted
at the Northwest Fisheries Center (Seattle,WA).The
first was completed in 1973 using fall chinook sal-
mon as the test animals,and the second in 1974 using
steel head trout.Assays consisted of 20 simultaneous
tests of chinook and 18 simultaneous tests of steel-
head,in fresh water at 10°C,with various concen-
trations of dissolvee gas in deep and shallow water
tanks.At termination of the tests,surviving fish
were divided into two groups;one group was trans-
ferred directly to salt water to assess the effects of
Dawley,Schiewe,and Monk:National Marine Fisheries Service,
Seattle,Washington.
1
,olIaj;Ift
-
stress from supersaturation on their ability to trans-
fer to salt water;a second group was examined for
signs of gas bubble disease.Groups of chinook and
one group of steel head exhibiting signs were then
placed in equilibrated water (100%T.D.G.)for a
2-week recovery period and subsequently re-
examined for signs of gas bubble disease.
Deep water tanks were 2.44 m (8 ft)deep which
provided a maximum hydrostatic compensation of
0.27 atm or 27%of saturation,and shallow tanks
were 0.24 m (10 in.)deep providing only 0.025 atm
of pressure compensation or 2.5%of saturation.The
shallow water tests on both chinook salmon and
steel head trout consisted of two replicates at 120,
115,110,105,and 100%(control)total dissolved gas
(T.D.G.).Deep water tests with chinook salmon
included tests at 127%(1 tank),124%(1 tank),120%
(2 tanks),115%(2 tanks),110%(2 tanks),105%
(1 tank),and 100%(1 tank).Deep water tests with
steel head trout consisted of two replicates at 127%,
1200,.{"115%and at 110%T.D.G.(Previous work
indicated that tests at 110%of saturation in deep
tanks could serve as a quasi-control).
juvenile fall chinook were acquired from the
Spring Creek National Fish Hatchery in early
February 1973 as buttoned up fry for Use in the first
experiment,and juvenile steelhead were captured
during their seaward migration down the Snake
River on April 30,1974,for use in the second bio-
assay conducted in 1974.Steel head were 1+yr of
age.Chinook and steelhead populations were ac-
climated to our laboratory water system at 10°C,
for 19 and 6 days,respectively,prior to testing.
Before initiation of tests,random samples were
taken from each population to obtain average
weights,lengths,and condition factors (Table 1).
The bioassay with chinook began February 20,
1973,with the introduction of 220 fish per tank
and was terminated on July 8 (127 days of ex-
posure to concentrations of dissolved gas plus
13 days of subsequent tests).Steefhead tests began
May 6,1974,with the introduction of about 80 fish
per tank;these were terminated after 21 days (7 days
of exposure to concentrations of dissolved gases
plus 14 days of subsequent tests).
Once testing began,each tank was examined
four times daily for the first 4 days followed by
three,two,or one times daily throughout the re-
mainder of the test period.During each observa-
tion mortalities were removed,their length and
weight recorded,and signs of gas bubble disease
noted.Vertical distribution of the fish in each deep
tank was also noted in percentage of total popula-
tion at four levels of depth;0-0.6 m,0.6-1 m,1.2-
1.8 m,and 1.8-25 m.Subsampling of each test
group for condition factor and disease signs was
done each 28 days at a rate of 10%(but not less
2 Dawley,Schiewe,Monk
TABLE 1 Meilns ilnd Stilndud Deyiiltlons 01 Weights,
Lengths,ilnd Condition Filctors of Rilndomly Silmpled
filII Chinook ilnd Steelheild Tilken from Test POpuliltions
Before Testing
Filii chinook
n
60
64
n
24
26
29
26
than five individuals)of the surviving population.
Fish from each subsample were weighed,mea-
sured and examined for signs of gas bubble dis-
ease (none were returned to the tests).Fish were
fed an Oregon Moist Pellet@ ration 5 days a week,
at a rate of 4%of body weight/day.Rations for
each test tank were corrected daily for numbers
of surviving fish and corrected each 28 days for
weight change (calculated from size of fish sub-
sampled every 4 weeks).
Dechlorinated water from the Seattle dty
water system which is supplied by the Cedar River
was used in these tests.Temperature was main-
tained at 10°±0.5°C,by mixing hot (27°C)a'nd
cold (7°C)water in a reservoir tank.Water for the
shallow tank system was supersaturated by inject-
ing 0.5 Qlmin air and 0.23 elmin ~into the suction
side oEtwo centrifugal pumps which were plumbed
with a recirculation loop to two closed receivers (52
gal each).Hydraulic pressure within the receivers
was maintained at 2.1 kg/cm 2 (30 psi)where dis-
solved gas content was increased to about 122%
of saturation T.O.G.(Fig.1).Water for the deep
tank system was recirculated by,a pump through
an open reservoir tank 9 m deep x 3 m in diameter
which was tapped at the bottom for distribution
to the test tanks.Air and oxygen were injected into
the recirculating pump at about 2.0 £Imin and 0.2
£/min respectively.This resulted in a stable satura-
tion level of 128%T.D.G.Both sources supplied
individual test tanks through PVC lines which
directed the supersaturated water to a vertical
stack of aluminum trays (28 x 41 cm)placed 5 to
10 cm above one another.One half of each tray
was perforated with 500-3 mm holes and the per-
;;.
~,I
j..•..•....:'..'~.."·'r
BACK PR{SSURt:
VAll'E
FIG.1 Schematic drawing of system used to produce water
supersaturated with dissolved abnospheric gas in shallow
water tanks.
RESULTS
Lethal Effects of Dissolved Gas
Chinook groups held at 120010 and 115%of
saturation in the shallow tanks and at 127%and
124%in the deep tanks sustained substantial mor-
tality (67%-97%)after 60 days of exposure.By the
same time,13%mortality had occurred in groups
held at 110%in shallow tanks and 4%had occurred
in groups held at 120%in deep tanks.Mortality
was insignificant in groups held at lower gas con-
centrations.Curves of accumulative mortality for
all test groups are shown in Fig.2.Average cumula-
tive mortality in the control tanks was minimal
(3%)for the first 60 days,but by day 127 had
sharply increased to 26.3%in the shallow tanks
and 13.6%in the deep tanks.
A change in normal feeding response and swim-
ming behavior of chinook groups (both deep and
shallow)was noticed on day 64 of the test.We be-
lieve these changes resulted from an infedion
caused by Cytophaga psychrophila.All groups (test
and control)were taken from test tanks and bathed
in a 10 ppm solution of terramycin for 1 hr.A supple-
ment to the daily ration of 0.5%oxytetracycline was
administered for the following 10 days,and after a
2-week interval another 0.5%supplement was added
for 10 days.After the second treatment (day 1(0)the
fish in all tanks behaved normally.
Steel head groups held at 120%and 115%of satu-
ration in shallow tanks developed substantial mor-
tality within 7 days,100%and 57%respectively,
"'Trade names referred to in this publication are not an endorse-
ment of commercial products by the National Marine Fisheries
Service.
Dissolved gas analyses were made on each tank
at least once each weekday for the first 2·weeks,
then a minimum of twice each week for the rest of
the test period.Procedures were identical to Dawley
and Ebel (1974).A gas chromatograph was calibrated
for nitrogen and argon using a modified manometric
blood gas analysis apparatus (Van Slyke and Neill,
1924).The modified Winkler procedure (A.P.H.A.,
1971)was used for analysis of oxygen concentrations.
Water samples were collected by use of a siphon
tube from the middle of the water column in the
shallow tanks and from the surface of the deep tanks.
Gas concentrations remained steady throughout the
test periods and mean values for each tank did not
change more than 1%on a weekly basis with stan-
dard deviations for both tests less than 2.6%
T.D.G.overall (Table 3).Samples were taken from
the top,middle and bottom of the water column of
the deep tanks several times and gas concentrations
were found to be uniform throughout the tank.
EQUILIBRATION
lRAYS
AIR EQUILIBRATION
AIR
PR{SSUR{
TANJ:.
forated ends alternated to produce a back and
forth flow of water f~om tray to tray.Water was
then collected in a plexiglass box (18 in.x 18 in.
x 10 in.deep)with a false bottom of porous poly-
ethylene plate through which air was passed.The
level of supersaturation desired for each test tank
was maintained by regulating the number of per-
forated trays and the amount of air supplied to
the collection boxes.Water from each box was
gravity fed to a test tank through a vinyl tube,end-
ing at the water surface.A flow rate of 7.5 £Imin
was maintained which created a circulation at
about 0.2 m/sec within the test tank.
Test tanks were made of green tinted fiber-
glass,1.2 m in diameter and of two heights,0.6 m
and 3.0 m (shallow,deep),holding about 270 and
2700 9.of water,respedively.A plexiglass window
extended from the top to the bottom of the
deep tanks allowing observations to be made over
the entire water column.Curtains covered the
windows and were removed only at times of obser-
vation.Water drained from the bottom of these
tanks through an external standpipe.
lighting was controlled with time clocks to
simulate natural sunrise and sunset and light inten-
sity at the surface of each test tank was from 10
to 20 lumens/ft2 during full intensity periods.
Water quality determinations for parameters
other than dissolved oxygen and nitrogen were
made before testing began and once each week or
once every 4 weeks depending on the parameter
measured (Table 2).Analysis procedures were
those of A.P.H.A.,Standard Methods for the
Examination of Water and Waste Water,1971.
The monthly measurements were made by per-
sonnel of the State of Washington Department of
Ecology using a P &E 303®atomic absorption spec-
trophotometer.All concentrations of heavy metals
or other potentially dangerous compounds fell
below potential danger levels to salmon and steel-
head (McKee and Wolf,1969).
-
.....
Long-Term Exposure of Sa/monids 3
TABLE 2 Range of Concentrations in mgliof Water Quality PUilmeters Measured of Wilter from Testing Filcilities During the
Period February 2O-'une 25,1973 and MilY 6-21,1974
Test tanks
Shallow
.;--
Pilrilmeler 100%105%.110%115%120%100%105%110%115%120%125%128%
6.7-6.9 6.7-7.1 6.7-7.0
.....
Tot.Hard.18-21
Tot.Alk.10-15
pH 7.0-7.3 6.9-7.1
NH,.05 .05
Cll .02
Zn ,n.d.
Cu n.d.
ed n.d.
Pb n.d.
Cd n.d.
6.8-7.1
.05
Weekly measurements
20 19-21
12-13 10-13
6.8-6.9 6.9-7.1 6.9-7.3 7.0-7.3 6.9-7.1 6.8-7.1
.05 .05 .05 .05 .05 .05
.02 .02
Monthly measurements
n.d.
n.d.
n.d.
n.d.
n.d.
.os .05 .os
n.d.=nondetectableon Perkins and Elmer 303~atomic absorption spectrophotometer.Measurements made by Washington State
Department of Ecology,Olympia,Washington.
NOTE:Other ~rameters measured prior to testing:e02(1.2-2.0),chloride (4-6),cyanide (.021,fluoride (,9-1),iron (.1-.6),
nitrate (.03),nitrite (.003),phenol (.011.potassium (.2-11.sulfate (3).
Measurements made by Environmental Protection Agency,Redmond,Washington.
fIG.3 Mortality versus time curves for juvenile steelhead
exposed to various concentrations of dissolved atmospheric
gas in shallow (0.25 m)and deep (2.5 m)water tanks at 10°C.
____0 DEEP
__e SHAUOW
20
100 .----------:..----------------~'(/
j
!e
/
105%
110%
120 140
I 1l5~
I
r--'
./105%
SHAllOW TANKS
- ---DEEP TANKS
100
>->-=-~III
0::
0
~
>-z...60u
Q<'
~
"":>
>-::s 40
:::>~::>u
20
0
0 20 40
,....
-
.-and the two groups at 127%in the deep tanks aver-
aged 25%mortality (Fig.3).Control mortality was
2%and was not used to adjust test mortality curves.
.....
FIG.2 Mortality versus time curves (combined replicates cor-
rected for mortality of control tests)for juvenile fall chinook ex-
posed to various concentrations of dissolved atmospheric gas in
shallow (0.25 m)and deep (2.5 m)water tanks at 10°C.
Progression of Gas Bubble Disease
The frequency of occu rrence of most gas
bubble disease signs increa~ed with increasing
levels of supersaturation.The fall chinook mor-
talities incurred the highest incident rate for
cutaneous blisters on the head and mouth and for
occlusion of gill filaments.Mortalities from 120%
and 115%shallow and from 127"k,deep tanks
showed 40-70%·incidence of these signs.Other
signs which increased with increasing levels of
supersaturation but with lower frequencies of
occurrence were:heart occlusions (14 to 34%).
blisters in the connective tissue surrounding the
eye,and blisters between the fin rays (Fig.4).
In research by Dawley and Ebel (1974),the
appearance of gas emboli in the lateral line was
the first external sign of gas bubble disease to
develop on spring chinook exposed to various
levels of supersaturation.We,however,observed
that gas emboli in the lateral Iine of both fall chi-
nook and steel head trout mortalities were not
prevalent within any group,but did appear in high
percentage (50-100%)on the biological (live)sub-
samples from all test groups.Scattered bubbles (less
than 15%of the lateral line)also appeared on most
mortalities from the control groups.We cannot
account for this observation and for that reason only
•HEAD
e GillS
100 l o MOUTH*FINS
x HEART
+EYE
'V ABDOMEN
80 6 BODY SURFACE
Ul -T LATERAL LINEz
<.:>en .-_.i!:-3 60 I-
.__.........e
:1:
/__-0-...:.
!!l 10
/e/-"'O
LL
1-Zw 110 -I,L'...··...*ua:w / I v-:::~"t<~i.~xa.
201-I!~(/A:::-<_;
~.,~.,oR "~~/~6+/".'.z.""~,~~.,~J:l~T_._T
0
_.---=:;:;::-...~I
100 105 110 115 120 125
TEST UEVEl (%OF SATURATION)
FIG.4 Frequency (%)of dead juvenile chinook bearing gas
bubble disease signs from shallow (0.25 m)test tanks al various
levels of dissolved gas.
ascribe this sign to gas bubble disease when more
than 15%of the lateral line appears occluded.
Disease signs on live fish sampled generally
appeared in rates and patterns similar to those
recorded on dead fish removed from the same test
concentrations.However,emboli in branchial
arteries.gill filaments,and the heart were rarely
observed on live subsamples,but were prevalent
Long-Term Exposure of Salmonids 5
.:~
1601«1
o IJ£EP
•SHAllCM
III 100
DAYS
6DIII
\-
f\\~r\~J j
~.i----
100 0
105
124
127
0.6
Q.&1~
~1.0
~L2
x
lL Ul!l
1.6
1.8
192'1)94'1 S'/69 1'l 899'1 10911912'1
DAYS
AGo 7 Me.Jn depth durins d.llylisbt hours of sroups of juvenile
chinook held in 2.5 m deep t~nlcs ~t concentr.lltions of 100,lOS.
110.11S.120.124..lind 127%of S.lItur.lltion--.ner"sed for periods
of 10 d~ys over 127 d~ys.
FIG.6 Frequency (%)of delod juvenile chinook beuing
selected sn bubble disene signs from sh.llllow (0.25 m)versus
deep (2.5 m)test t~nb ~t nrious levels of dissolved sa.
FIC.5 Exposure times to 25%mortality of juvenile chinook
held .lit nrious levels of S.lIS in deep (2.5 m)versus sh.llllow
w.lIter (0.25 m)t.llnks.
ci O£EPI'JATER TESTS
:-SHALLOW T£STS
~III l>BLISTERS ON HEA.D
~•BLISTER IN MOOTH
~70 0 OCCWS ION OF Gl Ll l>
~:FI~~/NTS:/==-;r=::::.::::::
';;;4J .'1 ,0....
~30 ....,..,a ,.
i:/:/o_~~::::;;~>/
~0 "--,,-
100 lOS 110 115 120 124 lZ7
T£ST LEVEll'!.Of SATURATION Of TOG.
on mortalities examined immediately after death,
indicating these signs are directly associated with
the death of the animal.
The trends in gas bubble disease signs noted
on the steelhead differed from those recorded dur-
ing tests with the fall chinook salmon.Two differ-
ences were:
1.Heart and gill emboli occurred in almost
100%of the dead steelhead,whereas these signs
were rarely noted in dead chinook,suggesting that
decomposition of the fall chinook quickly masked
these signs.
2.Incidence rates of certain signs were
directly associated with duration of the test.live
and dead steel head at test termination showed
light incidence of exophthalmia,cutaneous bubbles
on the head and in the buccal cavity,and no signs
of bubbles on the body surface,whereas the chi-
nook,subjected to supersaturation for a much longer
duration,showed high incidence of these signs.
Also,gas bubbles between the fin rays were not
prevalent on fall chinook,yet showed very high in-
cidence on steelhead mortalities and live subsamples
after 7 days exposure to high supersaturation.
Effects of Water Depth
When mortality rates in the deep tanks are
compared to those in the shallow tanks (Fig.2
and 3),the average depth of the chinook and steel-
head groups in the deep tanks appears to have
compensated for about 10%and 10-15%,respec·
tively,of effective saturation.Fig.5 shows that
the time to 25%mortality of fall chinook at various
levels of dissolved gas concentrations in the deep
tanks were comparable to time in the shallow
tanks at a 9.5 to 10'%lower effective saturation (Le.
25%mortality was reached at 30 days of exposure
in the deep tanks at 124%and in the shatlow tanks
at 115%).Also a comparison of incidence and
degree of G.B.D.signs on dead chinook between
deep versus shallow tanks indicates an effective
decrease in supersaturation of 12 to 15%(Fig.6).
Vertical distributions of chinook groups for
the first 3 days of the test were variable and not
significandy different from one gas level to the
next.After 3 days.however.the fish groups at
higher saturation levels maintained a greater
depth than those at the lower levels and main-
tained this difference the entire test periOd (Fig.
7).Steelhead freshwater tests showed similar
results.i.e,mean depth increased with increasing
gas concentrations.
Night observations showed a depth shift
downward of approximately 0.3 m (O.18-{).42 m)
for each of the test species at each saturation level
(Fig.8 and 9).The increased depth trend with
6 Dawley,Schiewe.Monk
-
,.,..
BOTTOM
0 r--SURFACE
0.5
Vi i
""i 0....•0 0t::i 1.0 -~il ••0x 0kil6il••.....ilc 6
""15 6~(0:00Xl-1l59<lIGHTEO • =1200-l859::
UNLIGHIED il =1<m-0559
2..0
-
2.5
100 lOS 110 US lZO
TEST l£VEL ''10 OF SATURATION)
124 m
These were examined to 1)determine if certain
size portions of the population were more suscepti-
ble to gas bubble disease and,2)detect any
variations in growth caused by chronic exposure
to the various levels of saturation.
,Condition factors of mortalities occurring within
a 16-day range of the monthly subsamples (e.g.,day
of subsample..t.8 days)were compared,by means of
a student's T-test,with the mean condition factor of
these same subsamples.The mortalities in the 110,
115 and 120%saturation shallow tanks and in the
124 and 127%deep tanks had condition factors sig-
nificantly higher than the live subsamples (t =3.78,
32 df,P (0.001,t =3.87,27 df,P (0.001;for deep
and shallow tanks,respectively).Thus,the larger
fish with higher condition factor died at a signifi-
cantly higher rate at these concentrations.
higher dissolved gas concentration noted during
daytime observations also occurred at night.
FIG.9 Mean depth of juvenile steelhead groups in 2.5 m deep
tanb at dissolyed gas concentrations of 110,115,120,and
127%of lliIturation-ayeraged for 2 segments of the day over
15 days.
FIG.8 Mean depths of juvenile chinook groups in 2.5 m deep
tanks at dissolved gas concentrations of 100, 105,110,115,120,
124,and 127%of saturation-averaged for 3 segments of the
day over 30 days.
Effects of Transfer to Salt Water
Subsamples of survivors from combined
replicates of all test groups (chinook and steel-
head)were placed into salt water at 25 ppt salinity
at 10°C,to determine whether prior exposure to
various levels of dissolved gases affected the
Long-Term Exposure of Sa/monids 7
Recovery From Gas Bubble Disease
At completion of the freshwater phase of testing,
chinook and steeJhead groups still surviving re-
tained signs of gas bubble disease similar to those
noted on the monthly subsamples (described earlier).
Subsamples of chinook tested at 110%of saturation
in shallow tanks and at 110,115 and 120%in deep
tanks were placed in fresh water at 100%saturation
for recovery observations.A portion of each of these
test groups had sustained significant mortalities
from gas bubble disease,other groups not included
in the recovery tests had no observable signs of gas
bubble disease.All subsamples sustained mortalities
from 10-16%in the 2-week re'covery period (group
size 27-48 fish).However,these mortalities could
not be attributed to gas bubble disease.After 2
weeks,the survivors no longer exhibited outward
signs with exception of one fish with a hemorrhaged
eye and another with bubbles in the orbit.
Eleven steelhead surviving the deep test tanks
set at 127",.6 saturation were placed in a shallow
tank at 100%saturation.After 3 days,examina-
tion indicated that cutaneous blisters had
decreased both in size and number.(e.g••5 mm
blisters had decreased in size to 2 mm and 20
blisters on the operculum decreased to 4).These
fish were subsequently placed in water at 105%
of saturation and all signs remained the same after
another 4 days,at which time fish were released.
Mortality did not occur in the 7-dayrecovery period.
o
w (weight in grams)x 105
L3 (fork length in millimeters)
K=
,----------SURFACE---------,o
0.5 L1GHIEO 0.0600-1859
UNLlGHTED.c.=lc;ro.0559
Vi
""""1.0.....
""~
x
1.5 ~.....
"-....c 0 0""~a
-<::t.
6 il
2.0
2.5 BOTTOM
100 105 110 115
TEST l£VEL ''10 OF SATURATION)
Effect of Gas Supersaturation on
Condition Factor
Weight and length data obtained from the
live subsamples and the fresh mortalities were
used to calculate a condition factor "K"where:.
-
-
.,...
~~----'-------------:;.'"
7060
NMFS
42mm
30 40 so
NUMBER OF OA YS
70 f-
,,
,t:!,
o 115'-TOG "
j \,£t.(,3mm
I 112%TOG "
ro f-67mm /"1/i
~112%TOG I 0 '
~so ~"li ,tt'
~.<;>"~40 f.I ,t
IX •0 ,~~3Q t.·I'p',4Omm.·
112STDG ~
,Q C,'\'I'
III ~!J c//,.-~=-;{
10 lv'.i ..."NMFS--.""42mm
"
<;>,11".-~-J:]/1l0'r0 TOG.....----o -o 10 20
abi1lity of these fish to make this tranSItion.Chi-
nook groups of 50 fish from each gas level and
from each series (deep and shallow)were trans-
ferred on test day 127.A combined total of 98%
mortality occurred in 3 days.Only 8 fish survived
for a longer time;1 fish from the 105%shallow tank
and 7 fish (14%)from the 1100,{,deep tank;these
lasted the entire 13 days.Results of a statistical
comparison of fork lengths of the survivors (X =
67.3 mm)to those of mortalities (X =52.5 mm)
indicate a definite size correlation with ability to
make the transfer (T =5.73,46 df,P <0.001).This
suggests that the majority of the experimental stock
had not yet reached smolting size and their ability to
transfer to salt water was thus severely lessened.
Steelhead test groups were likewise subsampled
and groups of 10 to 20 fish were placed into salt
wat,er.Mortality varied from 0-16 with no correla-
tion to previous stress experience.However,a size
comparison between mortalities and survivors indi-
cated that mortalities were the smaller of the popu-
lation (T =1.925,51 df,P <0.06)again suggesting that
the dead fish may not have been up to smolting size.
DISCUSSION
Tes:t Results
The mortality curves (Fig.2 and 3)may be
affected by synergistic effect of C.psychrophila
after day 64;however,incidence rate and types of
gas bubble disease signs of dead fish showed no
apparent difference for individual tests between
the first 60 days and ~he last 67 days indicating that
the 'effect was not large.We,therefore,assume the
mortality curves (adjusted for control mortality)
are generally representative of death rates caused
by gas bubble disease at the dissolved gas con-
centrations indicated.The first 60 days have no
qualifications,but the last 67 days may represent a
fish stock with less than normal tolerance to excess
dissolved gas pressure.
As shown in Fig.10,the death rates and curve
shapes correlate well with experiments done by
Meekin and Turner (1974),in which they exposed 67,
53,and 40 mm fall chinook to 122%N1,+Ar plus 74%
02 ('112%T.D.G.).Fish tested by these researchers
showed a definite inverse correlation between
resisf.ance times in supersaturated conditions and
age and growth.larger fish (53 mm,67 mm)suc-
cumbed much more rapidly than 40 mm fish,
tested at the same level of percent saturation (T.D.G.).
This same trend was also shown by Shirahata
(1966),testing rainbow trout from hatching to fry
stage.From this evidence we conclupe that the
times to death at indicated gas concentrations pre-
sented here are typical for these spE'cies at this
size,and that the increase in mortality rates of fall
8 Dawley,Schiewe,Monk
FIG.10 Mortality versus time cUrYes for bio~y 0'dissolyed
gas in shililow tilnlcs (0.25 m or less)ilt 122%N:+At ilnd 74%0'
uturiltion 0:(resulting in 112%T.D.G.)with 'illl chinook at
nnous sizes (Meekin and Turner 1974)and curves at 115 and
110%T.D.G.with fall chinook at 42 mm (NMFS data).
chinook groups as the experiment progressed was
mainly commensurate with aging and growth.
In tests done by Dawley and Ebel (1974)the re-
sistance times of steel head in shallow water tanks at
115%supersaturation was 400"16 longer than our
tests with steelhead at the same saturation level.
Fish tested at that time were hatchery reared and
smaller (130 mm compared to 180 mm)which is
probably the reason for their greater resistance.
The order of magnitude of this difference,how-
ever,is small compared to the 1-to 2-month dif-
ferences in resistance times we observed between
fall chinook and steelhead.This difference correlated
Nell with data by Meekin and Turner (1974t which
also indicates that fall chinook were more tolerant
to exposure to supersaturation than were steelhead
of comparable size and age.This same order of
ranking was noted by Ebel,Dawley,Monk (1971)
and Dawley and Ebel (1974).
Some prominent signs of gas bubble disease
.occurred on dead chinook in association with certain
stages in physical development or stress experi-
ence.Cutaneous blisters in the buccal cavity and on
the body surface and hemorrhages in and around
the eye required more time to develop than other
-
.-
signs,thus did not exist on mortalities from the
higher test levels because of shorter time duration.
Therefore,incidence rate was lower but is entirely
dependent on the time under stress.Exophthalmia
appeared frequently in the 3rd and 4th months,
similar in incidence to blisters on the head.Blisters
at the mid-line of the vertical surface occurred fre-
quently on chinook mortalities taken from the deep
and shallow tanks at the highest levels.This fre-
quency decreased,however,as testing progressed
and by the 4th month there was no evidence of this
sign;it appeared to be related to recent yolk absorp-
tion.Blisters between the fin rays occurred at a very
low incidence (40%)at the highest test levels com-
pared to what had been previously observed by other
investigations in other tests with la,:ger salmon ids
of other races and species.
Comparison of condition factors between live
and dead fish seems to indicate that the larger fish
were more susceptible to gas bubble disease.This
agrees with earlier research by Shirahata (1966)and
Meekin and Turner (1974).The species tested by
these researchers (rainbow trout,salmon,and steel-
head,respectively),became less tolerant with age
and growth,starting as button up fry.
Although a portion of the test groups in the
deep tanks remained at sufficient depth to increase
their resistance time,the depth did not provide
sufficient compensation to prevent mortality,par-
ticularly at levels above 120%
The mortality rates and G.B.D.signs of both
speciE~s also indicated that less hydrostatic compen-
sation was derived due to depth disposition than
expected when the mean depth of the fish groups
is considered.Thus,individual fish must move sub-
stantially from the observed mean depth of the test
lot.If this did not occur t the effect of hydrostatic
compensation would have resulted in a calculated
reduction in effective supersaturation of 12-16%for
chinook and 17-20%for steelhead.Since the actual
mortality rates indicated that only a 10%and 10~15%
(chinook and steelhead,respectively)reduction
occurred,we can assume that the fish were moving
randomly about within the tank.
Application to the River Environment
Certain observations made during these bio-
assays have important implications relative to the
experience of naturally migrating populations of
juvenile salmonids.The Columbia River system is
of major concern in our research efforts,thus the
following discussion is centered on the implica-
tions ~o fish in the Columbia.
Most areas where'salmon and steelhead
incubate and develop in the Columbia River system
are located above dams and,therefore,would be
little affected by supersaturation.However,spring
chinook and steel head on tributaries of the Willam-
ette River,a major tributary of the Columbia,
make heavy use of areas below dams.Also,hatch-
ery water sources in some instances are either
below dams that may produce supersaturation
during the rearing period or are taken from wells
yielding water with high dissolved gas content.In
these instances the early stages of life such as the
period of incubation become quite important.
Data from our bioassays and others-Shirahata
(1969)and Meekin and Turner (1974)-indicate that:
1)yolk sac fry sustain injuries at low levels of dis-
solved gas which become fatal as the yolk is nearly
absorbed;and 2)that after fry have "buttoned up"
tolerance to supersaturation becomes quite high
but decreases gradually thereafter until "time of
seaward migration.Nebeker (1973)indicated that
tolerance of adult salmon ids to supersaturation is
slightly less than that of juvenile migrants.Sig-
nificant changes in tolerance at various life
stages obviously occur and the effect varies de-
pending on the life stage.Equilibration of hatchery
water sources is thus extremely important at cer-
tain stages of fish development.AI!;o,spillway
discharges at certain times will have more'effect
on survival of downstream juvenile migrants than
at others,thus management policies should consider
the changing effects of these discharges.
The spring freshet on the 10wer'Columbia and
Snake Rivers coincides with juvenile salmonid
outmigrations as well as some adult upstream
migrations.Freshet conditions are variable from
year to year,but heavy spillway discharges usually
occur every year for some duration creating super-
saturation from 120%to 1400..1>.During years of high
flow these levels occur throughout long stretches
of the river (650 km and more),resulting in
long-term exposure of some stocks.Rates of juve-
nile migration indicate that at least 26 days is
required for travel from Little Goose Dam to the
Columbia River estuary during the highest flows.
Thus,even if fish are compensating for super-
saturation by sounding a significant portion of the
population is subjected to levels of dissolved gas
supersaturation exceeding 120%.
Data on depth distribution of migrating juve-
nile fish within the Snake River near Lower Monu-
mental Dam (Smith 1974)indicate that 58%of the
chinook and 36%of the steel head were in the upper
3.7 m of the water column.Mean depths for these
portions of the migrating stocks were 1.30 m.and
1.33 m,respectively.This would compensate for
14.5-14.6%effective saturation,'which means
that at higher levels of supersaturation (135%or
greater)both stocks of fish would be exposed to
levels of gas concentration above 120%for at least
28 days during periods of high flow.
Long-Term Exposure of Salmonids 9
These bioassays have shown that although
both the fall chinook and steel head tend to remain
at greater depths with increasing levels of super-
saturation,they are unable to totally compensate
for dissolved gas concentrations above 120%.
Therefore,it is imperative that corrective measures
to reduce supersaturation be implemented as soon
as possible to reduce mortality.
SUMMARY AND CONCLUSIONS
Bioassays in shallow (0.25 m)and deep (2.5 m)
tanks with dissolved nitrogen and argon gas con-
centrations ranging from 100 to 127%of saturation
were conducted to determine lethal and sublethal
effects on juvenile faU chinook salmon and steel-
head trout.Throughout the test,mortalities and
live subsamples were weighed,measured,and
examined for signs of gas bubble disease.After
exposures of 127 days (fall chinook)and 7 days
(steelhead),remaining groups of fish were:1)
put into saltwater tanks to determine the ability
to transfer to salt water;or 2)put into equilibrated
water (100%T.D.G.)to determine the ability to
recover from gas bubble disease.
We concluded from these experiments that:
1)~gnificant mortality of juvenile fall chi-
nook commences at about 115%of supersaturation
(T.D.G.)in shallow tanks where hydrostatic com-
pensation is not possible and at about 124%in deep
tanks where compensation is possible.
2)Significant mortality of juvenile steelhead
commences at about 115%in shallow tanks and at
about 127%in deep tanks where hydrostatic com-
pensation is possible.'.
3)Tolerance to supersatuation of atmo-
spheric gas of both fall chinook and steelhead
decreases with age and growth.
4)Emboli in branchial arteries,gill fila-
ments,and the heart were rarely observed on live
subsamples,but were prevalent on mortalities
indicating these signs are directly associated with
the dei~th of the animal.
5)The average depth maintained by chinook
and steelhead groups when allowed to sound com-
pensated for about 10%and 10 to 15%(respec-
tively)of the saturation value measured and com-
puted on the basis of surface (760 mm)pressure.
6)Both fall chinook and steelhead with
signs of gas bubble disease are able to recover
from e:lC.posure to supersaturation.
.7)Exposure to various levels of supersatura-
tion does not seem to affect the ability of steelhead
to transfer to salt water;data on effect of expo-
sure to supersaturation on ability of fall chinook
to transfer to salt water were inconclusive.
10 Dawley,Schiewe.Monk
REfER~CES
Amencan Public Health Association.1971.Standard Methods
for Ihe ExamillCltion of Water and Wastewater.Thirteenth
Edition.New York,N.Y.674 pp.
Beiningen,K.T.and W.J.Ebel.1970.Effect of John Day Dam
on dissolved nitrogen concentralions and salmon in Ihe Colum-
bia River,1968;,Trans.Am.Fish.Soc.99:664-671.
Blahm,T.H.,R.J.McConnell and G.R.Snyder.1973.Effect of
Gas 6upersaturaled Columbia River Water on the Survival of
Juvenile Salmonids.National Marine Fisheries Service,Prescott
Field Station.(Un pub.Ms.)61 pp.(Processed).
Bouck,G.R.1972.Effects of Gas Supersaturation on Salmon
in the Columbia River.Paper presented at Ecological Society
of America Symposium,August 1972.29 pp.
Bouck,G.R.1970.Gas Bubble Disease in Adult Columbia River
Sockeye Salmon (Oncorhynchus nerka).Pacific Northwest
laboratory,Federal Water Quality Administration,Corvallis,
Oregon,June 1970 (Unpub.Ms.)11 pp.
Coutant,C.C.and R.G.Genoway.1968.An Exploratory Study
of Interaction of Increased Temperature and Nitroge,n Super-
saturation on Mortality of Adult Salmanids.BNWL-1529,
Battelle-Nonhwest,Richland,Washington.
Dawley,E.M.and W.J.Ebel.Lethal and Sublethal Effects of
Various Levels of Nitrogen and Argon Supersaturation on
Juvenile Chinook Salmon and Steelhead Trout.National Marine
Fisheries Service.(Ms;in prepration).
Ebel,W.J.1969,Supersaturation of nitrogen in the Columbia
River and ils effects on salmon and steelhead trout.Fish.Bull.
68:(1):1-11.
Ebel,W.J.19n.Dissolved Nitrogen Concentrations in the
Columbia and Snake Rivers in 1970 and Their Effea.on Chinook
Salmon and Steelhead Trout.NOAA Tech.Report 5SRF-646.
7 pp.
Ebel,W.J.,E.M.Dawley and B.H.Monk.1971.Thermal
tolerance of jwenile salmon in relation to nitrogen super-
saturalion.Fish.8ull.69:833-843.
Fickeisen.D.H.,J.C.Montgomery and M.J.Schneider.1973.
Tolerance of Selecled Fish Species to Atmospheric Gas Super-
saturalion.Unpublished data presented at A.F .5.meeting,
Orlando,Florida.
Harvey,E.N.and A.C.Cooper.1962.Origin and Treatment of
a Supersaturated River Water.Inti.Pacific Salmon Fish.
Comm.Prog.Rpt.No.9,19 pp.
McKee,J.E.and M.W.Wolf.,1963.Water QualityiCriteria,
2nd Edition,Resources Agency of California State Water
Quality Control Board,Pub.No.J-A.550 pp.
Meekin,T.K.and R.L Allen.1974.Nilrogen Saturation Levels
in the Mid-depth Columbia River,1965-19n.Wash.Dept.Fish.,
Tech.Rpt.12,pp.32-77.
Meek!n,T.K.and 8.K.Turner.1973.Tolerance of Salmonid
Eggs,Juveniles and Squawfish 10 Supersalurated Nitrogen.
Wash.Dept.Fish.,Tech.Rpl.12,pp.78-126.
Nebeker,A.V.'.1973.Environmental Protection Agency Progress
Report,Western Fish Toxicology Stalion,EPA,Corvallis,
Oregon.
Rucker,R. R.and E.M.Tuttle.1948.Removal of excess
nitrogen in a hatchery waler supply.Prog.Fish.Cult.10:88-90.
Shirahata,5.1966.Experiments on nitrogen gas disease with
rainbow trout fry.Fresh.Fish.Res.lab.B!JI'.15(2):197-211.
(In Japan,with Eng!.Sun.).
Smith,J.R.197-4.Distribution of seaward-migraling chinook
salmon and sleelhead troul in the Snake River above Lower
Monumental Dam.Marine Fisheries Review,Vol.36,No.8,
August 1974.
Van Slyke,D.D.and I.M.Neill.1924.The determinations of
gases in blood and olher solutions by vacuum extraction and
manometric measurement.I.J.of BioI.Chern.61(2):523-574.
I T.H.Blahm
B.McConnell
G.R.Snyder
In 1969 the National Marine Fisheries Service ini-
tiated "on-site"environmental research on the
lower Columbia River.The research facility
(Snyder,Blahm,McConnell,1970)is housed on two
33.5 x 10 m (110 x 32 ft)barges moored near Pres-
cott,Oregon.(Fig.1).
.Research on the effect of nitrogen supersatura-
tion was begun in 1971.The primary emphasis ~has
been on bioassay of prevailing Columbia River
water at the site;however,several other types of
tests have been conducted e.g.avoidance and
detection of gas supersaturation,vertical depth
distribution of fish,intermittent exposure to super-
saturated and equilibrated N2 levels.In addition
to the biological tests,daily N2 monitoring at
Prescott has been done since 1971;also in relation
to monitoring,a study was initiated (in 1974)to
determine the gas equilibration characteristics in
the Columbia River between The Dalles Dam and
Prescott,Oregon.Exploratory tests have been done,
and will continue,on O 2 consumption and stamina
of fish in relation to N2 saturation.The effects of
hydrostatic pressure on the survival of N2 stressed
fish is also being examined.Results of the explora-
tory tests will not be included in this report.
The objective of this summary report is to out-
line representative tests and results to demonstrate
the types and diversity of studies completed at the
Prescott Facility.
Test results included herein are brief descrip-
tions of general samplings from the total effort;
more detailed information will be made available
on request.A list of published and non-published
data will be included in this report.Following is
the sequence in which the various projects will be
discussed:
Gas Supersatura-
tion Research
National Marine Fisheries Service
Prescott Facility-1971 to 1974
OREGON
1WA"INOm
ABSTRACT
In 1969,the NMFS constructed a field facility for "on-site"en-
vironment,al testing.The facility is housed on two 110 x 32 foot
barges moored on the Columbia River near Prescott,Oregon
(RM 72).Research on the effects of nitrogen supersaturation was
begun in 1971.Survival is better in the 2.5-m deep tanks than in
1-m deep tanks.Results of tests done in "shallow"test tanks are
not representative of what might occur in the river,as fish are
not restricted to "shallow"depths.Intermittent exposure to high
(130,120,ilnd 110%)and equilibrated (110%)levels of N2 satura-
tion generally enhanced test fish survival over that recorded for
fish held in constant high levels.Preliminary tests indicate that
the fish are not able to detect and avoid lethal conditions of
supersaturation.Dissolved gas levels have been monitored at
Prescott since 19n.
~-
,~
-
-
-
FIG.1 Lower Columbia River and locale of Prescott Field
Facility.
Blahm,McConnell,and Snyder:Environmental Conservation
Division,National Marine Fisheries Service,Seattle,Washing-
ton.
11
-
-
-~
'1.Biological
A.Intermittent exposure to supersatu-
rated and equilibrated N z levels
B.Detection and avoidance of N z super-
saturated water
C.Bioassay of prevailing Columbia River
conditions
1.deep versus shallow tanks
2.artificially created N 2 levels
D.Description of the vertical distribu-
tion of fish using depth sounding
gear
2.Physical Monitoring
A.Daily gas saturation levels at Prescott
B.Gas equilibration characteristics in
the Columbia River between The
Dalles Dam and Prescott,Oregon
INTI:RMITTENT EXPOSURE
The intermittent exposure tests were planned
to assess the effect of intermittently exposing fish
to either 130,120,110 or 100%N z saturation.Infor-
mation from these tests would help determine if
manipulation of water flows (spill)at dams could
possibly afford the fish some relief from gas super-
saturated water conditions during critical N z
periods.
Groups of at least 10 fish were held in separate
175 ~tanks in which the N z levels 130,120,110,
and 100%saturated were alternately switched as
per tine following diagram:
Percent N z
130 120 110
t t t
100 100 100
The time cycle,in two test tanks,was based on 24 hr,
e.g.CIne tank,8 hr at 130%and 16 hr at 100%and the
other,16 hr at 130%and 8 hr at 100%Nz;the same
pattern was used for the 120 and 110%N z tests.
The switching was done by valving,which elimi-
nated handling the fish.Test temperatures were
between 10 and 13°C.The alternating cycle was
continued for 192 hr (8 days).For this report the
results of the 130%to 100%Nz tests are used to
present the general results.Table 1 summarizes the
results of intermittent exposure to 130 and 100%N z•
Compiled in Table 1 is the time (hr)to 50 and 100%
mortality when the fish were exposed to 130%N z
for 24,16,and 8 hr.
With the exception of steel head and white-
fish the 16-or 8-hr exposure enhanced survival
time to 50%mortality over that recorded for a
constant (24-hr)exposure to 130%N z•At the 100%
mortality level the 16-or 8-hr exposure enhanced
survival of all species over that recorded.for fish
held constantly at 130%N z (Table 1).
DETECTION AND AVOIDANCE OF N2
SUPERSATURATION
Juvenile salmonids are being tested at the
Prescott Facility to determine if they can detect
and/or avoid Nz supersaturation.Homogenous
groups of fish are introduced into the end of a test
tank (Fig.2)which provides a lateral choice of two
channels,one containing diSSOlved nitrogen at
130%saturation and the other at 102%saturation.
Water depth in the channels is maintained at 0.33 m
to eliminate the effects of hydrostatic pressure.
External influences are minimized by placing cur-
tains around the test tanks and by limiting inspec-
tion and water sampling to twice daily.Test dura-
tion is 192 hr or until 50%mortality occurs.A
·f
TABLE 1 nme in hr to 50 and 100%Mortality for Fish Subjected
to 130%N1 Saturiltion for Either 24.16,or 8 he of EKh 24-hr Cycle
Durins 192-hr Intermittent Exposure Test.The fISh were
ilkematel,.exposed to 130 ilnd 100%Nz,.
,....
--
-largemouth bass
Rainbow
Chinook
Cutthroat
Whitefish
Coho
Steel head
50%MorU6ty 100%Morblity
Number of hr during NCh 24-hr cycle that
fish were subjected to 13O%NJ
24 16 8 24 16 .,8
Tune-hr nme -hr
93.5 173.5
47.0 69.5 117.5 141.5 •
24.0 120.0 •48.0 ••
24.0 72.0 103.5 39.0 ••
23.5 13.5 •13.5 47.5
22.0 ..78.0
16.0 16.0 •31.0
12 BEahm.McConnell.Snyder
COLUMBIA RIVER WATER
130%N2 102%N2
I I
I I
I IIIIII,
I II
130%102%
I 1
I I
I I
I i
I I
I ,
t t
124%108%
DRAIN
o
FIG.2 Plan view of 1.5 x 3.3 m tank used to determine if juve-
nile fish can detect and avoid Nz supersaturated water.
replicate test is conducted (using fish from the
same population)by switching the "high"N2
and "low"N2 channels.A sample of 20 fish is used
for each test.
Preliminary tests have been conducted on two
species of fish;juvenile steelhead trout and fall
chinook salmon.During the first test 5001..,of the
steel head died in 42.5 hr;in the replicate test a
50%mortality was reached in 43 hr.The survivors
from both tests had external gas bubble disease
symptoms.No mortalities occurred in the two
replicate chinook tests (within 192 hr)and only
10%showed external N2 symptoms.These two
preliminary tests indicate that the juvenile steel·
head did not avoid the high gas concentration
while fall chinook salmon did.Results from the
intermittent exposure tests in Table 1 of this report
show juvenile steel head to be less tolerant to con-
centrations of dissolved nitrogen than chinook.
N2 BIOASSAY TESTS
Two types of N2 bioassays have been con-
ducted at the Prescott Facility since 1971:1)
tests using deep (1.8 x 2.5 m)and shallow (1.8 x
1 m)test tanks at prevailing river levels 6f N 2
and 2)tests using small 175 P .tanks (0.33 m water
depths)at constant N2 levels of 130,120, 110,
and 100%saturation.
Mortality in Deep and Shallow Tanks
The primary objective of these tests was to
determine if water depth enhanced fish survival.
Table 2 summarizes the species used,number of
fish in each test,test duration,range and average
nitrogen levels and the total percent mortality that
occurred during the test.The N 2 levels,as indicated
in Table 2,were either created artificially.or were
TABLE 2.Summary of the Numbers of Each Species Held in the Three Tanks (Two Each 1-m
Deep and a 2.5-m Deep Tank)Used for Bioassay Tests at the Prescott Facility.Included in the
table is the number of days the fish were held and the Nz saturations.The percent mortality
that occurred in each tank is shown for each spec:ies.
Test tank water depth Percent Nz
Number Range
Species of fISh 1 m
t 1m 2.5 m Duration during test Average
Percent Mortality
Cutthroat 50 10 60 40 59 119 -136 124
Cutthroat 50 8 40 27 49 112 -130 120
Steelhead 80 10 80 6 55 112 -129 120 R2
Chinook 9S 0 80 11 SS 112 -129 120 R
Smelt 7S 30 100 40 12 119 -122 121
Smelt 50 24 100 23 5 117 -121 119 R
Crappie 50 0 0 0 20 117 -123 120 R
Squawfish 20 0 0 0 35 115·124 120
'Control tank gas equilibrated
2R =natura[ly occurring river levels
Research at Prescott 13
TABLE 3 Species and nme,in hr,to 50%Mortality for
Groups of F'1Sh Held at 130,120,110,and 100%Nz Saturation.
Included is a ranking from most to least tolerant.
to 130%N2 saturation.The ranking changes at the
120%level,but not drastically (Table 3).These
tests reflect general trends.and one could sur-
mise that the synergistic effect of various stresses
(temperature,disease.maturity. etc.)could alter
the pattern derived from these series of tests.
DESCRIPTION OF VERTICAL DISTRI-
BUTION OF FISH USING DEPTH
SOUNDING GEAR
In 1972 a Benmar depth sounder was modified
for use in a 1.8 x 3 m redwood test tank.Pre-
liminary tests in the tank indicated that the sound-
ing gear would work satisfactorily to determine
depth distribution of fish.After examining the
resulting tapes of several 24-hr tests we found
that the fish were generally below 1 m water depth
(this would enhance their survival from that in a
1 m deep tank).Two transducers were used in the
test tank;one at the water surface and one on the
bottom.A printer/counter system provided a fish
count for each 0.6 m interval of water in the 2.5 m
tank.(A description of the technical aspects of the
system is attached to th is report.)
The next step was to test this equipment in
the river.The two-transducer;arrangement was
modified to a 10-transducer array (Fig.3)which
could be placed on the bottom of the river at gently
sloping beaches.This configuration was used at
two locations near Prescott for a total of 75 hr
during day and night.Fig.4 summarizes the
results:of 776 fish approximately 72%were
detected between 0.9 to 2.1 m (3 and 7 ft).Many
more fish were detected during darkness than day-
light.While species could not be differentiated by
the sounder.a minimum seine effort (2 sets)on the
•=No 50%mortality level
1 Number in parenthesis indicates ranking of tolerance
.-~
*
*
*
*
*
*
*
*
*
100
*
*
*
*
*
*
*
*
*
110120
*(1)
*(2)
141.5 (5)
*(3)
119.5 (6)
SO.5 (8)
•(4)
72.0 (9)
30.0 (7)
Percent Nz saturation
nme to 50%mortality -hr
93.5 (1)1
55.0 (2)
47.0 (3)
24.0 (4)
24.0 (5)
23.0 (6)
22.0 (7)
16.0 (8)
5.5 (9)
130
Largemouth bass
Crappie
Rainbow
Chinook
Cutthroat
Whitefish
Coho
Steel head
Smelt
Species
the naturally prevailing Columbia River levels dur-
ing the test.The shallow control tank was supplied
with gas equilibrated Columbia River water.The
following is a list of water quality parameters that
were monitored in each test tank during the N-2
bioassay studies:
"I.Dissolved oxygen -02
2.Nitrogen gas-N 2
3.Carbon dioxide-C02
4.Ammonia-NH]
5.Conductivity
6.Alkalinity
7.Turbidity
8.pH
With the exception of N2 ,all parameters re-
mained within safe biological ranges throughout
test period.
Examining Table 2,we see that survival was
better in the deep tanks.This is what one would
expect knowing that as hydrostatic pressure
increases (with water depth)the percent of nitro-
gen decreases.In these tests the crappie and
squawfish were the most tolerant while the smelt
were the least tolerant.Within the salmonids
tested,the cutthroat were slightly more tolerant
than either the chinook or steelhead,while the
latter two species showed comparable tolerance.
These general conclusions apply only to the N2
level!.as indicated in Table 2.
Small)Tanks-130,120,110,and 100%
N~S;aturation
The tests in the 175 ~tanks were designed to
provide added information on the effect of nitro-
gen on fish survival.Thirteen bioassay tests have
been done with nine species.GrouPs of from 5 to
20 fish were held in separate tanks at 130, 120,
110,cmd 100%N2 saturation.Each test was con-
tinued for 192 hr during which mortality was
recorded.As with the preceding tests (deep versus
shallow tanks)water quality parameters remained
in acceptable biological ranges with the exception
of Ni.Test temperatures were between 10 and
13°C..Table 3 summarizes the species of fish used
and It he time (hr)to 50%mortality at 130 and
120%N~saturation.The 50%death level was not
reached in any of the 110%N2 saturation tests
nor did any mortality occur in the 100%N~sat-
urated control tests.Bass and crappie were·the
most tolerant of the species used in these tests
while smelt were the least tolerant.At the 130%
N~level the rainbow and steelhead trout were the
most and least tolerant,respectively,of the salm-
onid species.The remaining salmon ids,including
whitefish,seemed to be grouped at around 24 hr
survival for 50%of the test animals subjected
-
-
-
14 B.'ahm.McConnell,Snyder
FIG.3 Phl~to of lG-transducer sonic array and recorder.
120
lID
III
120
lWl
5.0
FEB MAR APR MAV JUN JUL AUG
12.5 ---SPILL DISCHARGE
-N2ANDAr l,
10.0 ,\/-_...
I \
1.5 ,\
t \
I \
5.0 I
I
·River mile (RM),rather than river kilometer,is used in this
report because most current references do not include the
metric equivalent.
FIG.5 Weekly averages of Bonneville "spill"and percent N.
saturation at Prescott,Oregon.
GAS SA TURA TlON LEVELS AT PRESCOTT,
OREGON-1971 ..1974
Since January 1971 approximately 1,000
Columbia River water samples have been analyzed
for gas content.During this period samples have
been taken twice weekly from the Oregon side of
the Columbia River (RM 72)·;daily samples were
taken when the N2 level exceeded 110%.The
highest level recorded at Prescott was 136.6%
Ni (132.6%total gas saturation)on June 25,1974.
Nitrogen concentrations in the lower Columbia
River exceeded the provisional standard (110%
N2)each month from November 1973 through
August 1974.Additional information was
obtained from samples taken weekly on a hori-
zontal transect of the river.N2 saturations on the
Washington side of the river are usually higher
than on the Oregon side;highest levels are gen-
erally recorded from the ship channel.
The saturation levels at Prescott,Oregon,are
influenced by the volume of water being spilled at
:Bonneville Dam,for example,when weekly aver-
age spillway discharges at Bonneville reaches
4.247 KCMS (150 KCFS)the N2 saturation levels at
Prescott exceed 115%(Fig.5).Gas saturation levels
during the first 8 months of 1972 and 1974 are
cOl!lparable (Fig.6);both being associated with
4O-yr high-water flows.In 1973 N2 levels were
considerably less than in previous years;outflow
was 69%of a 15-yr average.
The data collected from this sampling pro-
gram is tabulated and stored in the Corps of Engi-
neers,North Pacific Divisions'ADP system.
--...-::::'"~--
""--"~~..
DAYliGHT
40 60 III 100 120 I..,
MJMBER OF FISH
20
beaches netted 37 juvenile chinook,21 crappie,17
perch,16 stickleback,9 flounder,2 peamouth chub
and a whitefish.This indicates that the "sounder"
results can be approximately quantified for species
composition with a minimum beach seining effort.
We feel this method of determining vertical fish
distribution will eliminate some of the problems
associated with gill netting.
FIG.4 Numbers of fish detected (at 0.33 m intervals)by sonic
gear during 75 hr of operation at Columbia River beaches
near Prescott,Oregon.
,~
""""
-
Of-0.3
0.3i-0.6
0.6-0.9
0.11-1.2
1.2-L5
1.5-1.8
1.11-2.1
;:;;
'"~2.11-2.4
2.11-2.7
:z:t:11-0.3~
'"0.]-0.3l':!-<0.<,-0.93:
0.'1-1.2
1.:1-1.5
~~
1.:5-1.8
1.Il-2.1
2.1-2.4
2.4'2.7
-
Research at Prescott 15
-
-
......
130,...-------------------,
- ---1974
125 ---1m
--Icm
FIG.6 Ten-d;ay uerilge Nt lilS level ilt Prescott,Creson.
)anuilry Ithrough ~1972.1973 and 1974.
GAS EQU ILiBRA rlON CHARACT.ERISTICS
IN THE COLUMBIA RIVER BETWEEN THE
DALL£:S DAM AND ASTORIA,OREGON
Tw()surveys were done during 1974 to deter-
mine the mixing characteristics and rate of gas
equilibration in the Columbia River between The
Dalles Dam and Astoria,Oregon.One survey was
done between Bonneville Dam and Prescott on
April 9 and 10 and another between The Dalles
Dam and Astoria during June 11 to 14,1974.
Methods
An 18-ft survey boat floated with the water
mass following a 2.4-m long buoy which lwas.sub-
merged except for the top 0.6 m;the buoy was
equipped with a light for night drifting.Transec-
tional and/or single water samples were taken
at least every 10-12 river miles (Fig.7)and
returned to Prescott for analysis.Time delay on the
samples,between collection and analysis,was
never more than 12 hr.
Mixing and Equilibration Between
Bonneville and Prescott-April 1974
Table 4 is a summary of the results of
the survey.The "f1oae'time between BonnevillE
and Prescott was apprOXimately 26 hr.The-average
total outflow from Bonneville Dam was 8.1 KCMS
(286 KCFS)during this period.Bonneville fore-
bay and power house samples each showed 115%
N2 saturation at the beginning of the survey.
The N2 saturation in the spill race (3 transect sam·
pies)ranged from 123 to 130".-6.Table 4 shows side-
to-side mixing (within error of analysis)at Rooster
Rock (RM 130),approximately 15 miles down river
from Bonneville Dam.Below Vancouver the N2
values were apparently influenced (decreased)
by the inflow from the Willamette River.The N2
level decreased approximately 10%between Bonne-
ville and Prescott.
16 Blahm.McConnell.Snyder
AG.7 s.mpling lontions used during il study to determine giIS equilibriltion ch.nKteristics in the Columbiil River between The Dillies
0.-~"sIoria.Ot'egon.
",
~'.
:;:-
WASHINGTON
STEVENSON
BONNEVI U£DAM t
145 ;.::::;::::::::;:::;;:::::;-~:::::::::~
1«1.2
WILLAMETTE RIVER
OREGON
-
TABLE 4.Percent Nz Saturation of Water Samples Taken Between Bonneville Dam and
Prescott,Oregon;the Survey was Done on April 9-10,1974.The average water flow past
Bonneville Dam during this period wu 8.1 KCMS (286 KCfS).
Mixin,;and Equilibration Between The
Dalles and Astoria-June 1974
This sampling effort was begun on June 11 at
The Dalles (RM 192)and completed on June 14 at
Astoria"Oregon (RM 14).No significant equilibra-
tion occurred in the 46-mile reservoir between
The Dalles and Bonneville Dams (Table 5).Spill-
way discharge at The Dalles was held constant at
6.8 KCMS (239 KCFS)for a 7 hr period prior
to the start of the survey;total outflow during this
period was 10.07 KCMS (379 KCFS).The average
total flow at The Dalles on June 10-11 was 10.1
KCMS (386 KCFS)while the average spill was
4.8 KCMS (170 KCFS).In reference to nitrogen level
the flow seems to be mixed (side-to-side)immedi-
ately below the dam and remains so to the Bonne-
ville forebay.
After CI 22-hr time lapse the survey was con-
tinued from Bonneville to Astoria on June 12.
The Bonneville forebay and power house samples
Research at Prescott 17
115.6
116.0
112.2
115.0
112.7
114.0
118.2
118.5
118.2
112.9
116.0
117.3
115.7
117.0
124.0
118.5
115.0
122.1
127.1
118.2
120.5
120.8
114.7
123.0
127.1
129.5
114.9
18.0
ml/Q %
17.3
17.3
17.7
17.5
18.1
18.3
18.1
17.7
18.4
18.4
18.4
18.4
18.8
18.85
18.4
18.1
18.3
19.3
19.2
19.9
20.2
17.95
17.95
19.1
19.8
17.9
8.1
8.4
8.0
7.6
8.1
8.0
8.2
7.9
7.9
8.0
8.1
8.0
7.9
Temp.
7.8
7.8
8.0
7.9
7.8
7.9
7.8
7.8
7.8
8.0
8.0
7.9
7.9
8.0
06:20
23:05
09:00
02:35
Time of
sample
09:05
08:25
08:35
17:00
08:10
19:40
07:35
11:15
08:20
Center
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Center
Wash.
Location
in river
Center
Ore.
Center
Wash.
Center
Ore.
Center
Wash.
Center
Center
99.0
72.0
88.0
78.5
River
mile
101.0
107.0
118.5
140.2
144.4
130.0
145.0
145.0
146.0
Vancouver
Morgan Turn
April 10
Sauvie
Prescott
Above Willamette
River
Kalama
Rooster Rock
Dodson
Troutdale
Tanner Creek
April 9
Bonneville
Forebay
Bonneville
Power House
Sample site
Bonneville
Spillway
I~
TABLE S.Percent N~SOIturiltion of W.;aler SOImples Tilken Betweea The O.iiJ1es .;and AstOriil,-Oregon.The sUl'Yey w;;as done on June 11-14,1974.The ilVerilse w.;atel'flow past both The Dillies
ilnd BonnewiUe D.ilIIIS w;;as .;approllim.;ately 11.3 KCMS (_KCfS).
River Loc.;ation TIme of N2
Silmplesite mile ...river S.ilI11ple Temp.miil %
June 11
The Dalles 192.0 Wash.06:50 13.5 17.6 127.2
Docks 189.0 Ore.06:55 13.4 17.6 126.9
Center 13.4 17.8 128.3
I~Wash.07:00 13.4 17.7 127.6
Crates Point 187.0 Ore.07:10 13.4 17J 127.6
Center 13.4 17.6 126.9
Wash.07:15 13.4 17J 127.6
Lyle 160'181.0 Ore.07:35 13.5 17.0 122.8
marker "56"Center 13.4 17.5 126.2
Wash.07:45 13.4 17.0 1226-Bingen 171.0 Ore.08:05 13.6 17.5 126.7
Center 13.5 17.5 126.4
Wash.08:10 13.5 17.5 126.4
Cook.161.0 Ore.08:3{)13.5 17.4 125.7
Center 13.5 17.5 126.4
Wash.08:40 13.4 17.5 126.2
Stevenson 150.0 Ore.09:05 13.6 17.1 123.8
Center 09:10 13.5 17.6 127.2
Wash.09:15 13.4 17.6 126.9,-
Bonneville 146.0 Ore.09:35 133 17.6 126.7
Forebay Center 13.2 17.6 126.4
Wash.09:25 13.4 17.6 126.9
June 12
ApprollilNtely 22-hr Ii~Ltpse
Bonneville 146.0 Center 07:40 13.6 16.4 118.8
Forebay-Bonneville 145.0 Center 08:50 13.8 16.5 120.0
Powet"House
Bonneville 145.0 Ore.08:55 13.8 18.6 135.3
Spillway
Tanner Creek 144.4 Ore.09:10 13.8 16.9 122.9
Center 13.8 17.4 126.5
Wash.09:15 13.8 19.7 143.3
18 Blahm.McConnell.Snyder
~...
were approximately the sam'e,119 and 120%Ni.
respectively (Table 5);while the three transect
spiHrace samples ranged from 135 to 142%Nz.
Side-to-side mixing had apparently occurred at
Rooster Rock (RM 130).As with the April survey
there was an approximate 10%decrease in N2
level between Bonneville and Prescott and about
18%between Bonneville Dam and Astoria,Oregon.
ThE~Bonneville average total flow during June 12-
14 was 11.7 KCMS (413 KCFS)while the average
spill was 8:1 KCMS (286 KCFS).
In summary it was noted that:1)during both
surveys side-to-side mixing,in reference to N1
level,occurred at Rooster Rock,15 river miles
below Bonneville Dam,2)both surveys showed
approximately 10%equilibration between Bonne-
ville Dam and Prescott,Oregon,3)18 to 20%
equilibration occurred between Bonneville Dam
and Astoria,Oregon,4)no significant gas equili-
bration occurred between The Dalles and Bonneville
Dams,and 5)side-to-side mixing was charac-
teristic in all transects between The Dalles and
Bonneville forebay.These conclusions should be
considered valid only in relation to flows (total and
spill)which occurred during the surveys.
TABLE S.(Continued)
Sample site
RiYer
mile
location
in riYer
TIme of
umple Temp.mill %
-
Dodson
Rooster Rock
Troutdale
Vancouver
Above Willamette
River
Willamette River
Morgan Turn
Sauvie 1
June 13
Kalama
Prescott
Eagle Cliff
June 14
Astoria
140.2
130.0
118.5
107:(J
101.0
103.0
99.0
68.0
78.0
72.0
51.0
14.0
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Center
Wash.
Center
Center
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Center
Wash.
Ore.
Center
Wash.
09:45
09:55
13:10
13:15
13:20
16:05
16:10
16:15
18:15
18:17
18:20
19:55
.20:10
20:50
20:55
21:00
06:25
06:20
06:15
08:35
08:25
08:20
15:00
14:55
14:50
07:50
08:10
08:20
13.8
13.8
13.8
14.6
14.2
14.2
14.8
14.4
14.4
15.0
14.6
14.7
14.8
15.8
14.6
14.6
14.4
14.4
14.2
14.3
14.5
14.4
14.3
14.9
14.6
14.5
15.0
14.4
14.3
18.0
18.5
19.1
18.6
18.6
16.7
16.5
16.7
16.1
16.4
16.5
18.2
16.9
17.9
18.1
18.3
17.3
17.5
16.9
16.6
17.4
17.5
16.8
16.8
16.6
15.5
15.3
15.7
130.9
134.5
136.9
137.5
136.4
137.4
136.1
137.6
134.8
136.0
136.9
135.0
127.8
132.3
133.8
134.6
127.3
126.3
124.2
126.4
126.0
128.6
124.9
124.2
122.4
115.4
112.6
115.4
-
-~
'Sample omitted by error
REFERENCES
Snyder,G.R.,T.H.Blahm and R.J.McConnell.1970.Floating
Laboratory for Study of Aquatic Organisms and Their Environ-
ment.U.S.Dept.of Comm.fish.Ore.356.
Reports Emanating from Prescott
Blahm,T.H.R.j.McConnell and G.R.Snyder.1974.Effect of
Gas :Supersaturated Columbia River Water on the Survival of
Juvenile Chinook and Coho Salmon.SSR-F (in press).
Blahm,T.H.,R.j.McConnell and G.R.Snyder.1972.Gas
Supersaturation Research.1972 Report to North Pacific Division
Corps of Engineers DACW57·72F:60 pp.
Blahm,T.H.,R.j.McConnell and G.R.Snyder.1973.Gas
Supersaturation Research.1973 Report to North Pacific Division
Corps of Engineers DACW57-73F:19 pp.
Blahm,T.H.,R.j.McConnell and G.R.Snyder.1973.Gas
Supersaturation Research.1974 Report to North Pacific Division
Corps of Engineers DACW57-74F:33 pp.
McConnell,R.I.,T.H.Blahm and l.G.Davis.1974.Daily
Concentrations of Dissolved Gas in the Lower Columbia River
at Prescott,Oregon 1971-1974.National Marine FisheriesSer_
vice.Seattle,Washington.
McConnell,R.J.and T.H.Blahm.1974.Occurrence of Fish
Near the Kalama Nuclear Power Plant Site.Columbia River
October 1970-0ctober 1973.National Marine Fisheries Service;
Completion Report to Clark and Cowlitz County PUD's:22 pp.
Research at Prescott 19
J.......•..._f~..·c'
~-
•W.Marshall
112 IN
":YW':""'":''f ~,'--,'-..,,"~"~:T
5 IN
L--
---'~:1CHARTPAPER-
-100
AGo 1 A typical moving-stylus depth sounder ~hogram when
the lransducet'is used inside a 10-fl deep test tank.
depth display to the full 5 in.,the stylus must move
across the paper at a speed 10 times faster than
before.Construction of a mechanical train to move
the stylus this fast is practically impossible.
The Benmar DR-680®sounder eliminates prob-
lems associated with a moving-stylus·recorder _by
using 320 individual non-moving stylii.A 400kHz
carrier frequency provides high resolution of small
targets;individually detected fish are precisely
recorded (Fig.2).Custom-made wide beam trans-
ducers were obtained from Webster Transducers
and provided a detection cone with a base diameter
of 3 ft in an 8·ft deep by 6-ft diameter test tank
(Fig.3).
PREI.IMINARY TESTS
In May 1973,a series of 24-hr tests was con-
ducted with juvenile coho salmon in the 8-ft deep
/l.1arshaU:Marine Fish and Shellfish Division.National Marine
Fisheries Service,Seattle.W<lshington.
eTra-de names referred to in this publication do not imply en-
dorsement of commercial products by National Marine
Fisheries Service.
ABSTRACT
A rE~ording echo sounder was modified to scan multiple custom
transducers to record the depth distribution of fishes.The sys-
tem has been used in tanks as well as the Columbia River and
eliminates the objections that other sampling methods influence
deplth distribution.
Bioassay tests condutted at the NMFS Prescott
Facility in 1972 revealed that salmonids tested in
8-ft deep wooden tanks survive the stress of gas
SUpE~rsaturated water more readily than fish tested
in shallower tanks (Blahm,1972).The hydrostatic
pressure of the water at depths effectively prevents
gas bubbles from forming in the organs of the ex-
posE~d fish.Therefore,few mortalities occur.
This paper describes an acoustic counting sys-
tem developed to provide information on the vertical
distribution of Columbia River fishes.Because of
the river's turbidity and various practical problems
involved with employing fishing gear,an acoustic
system was considered a desirable alternative to
possible optical or direct capture methods.The
acoustic system was initially developed to record
the vertical distribution of fish in the Prescott lab-
oratory's 8-ft test tanks.later,equipment was
developed to monitor the vertical distribution of
fish occurring along river beaches near Prescott.
ECHOSOUNDER LIMiTATIONS
[)epth sounding equipment has been used suc-
cessfully for locating fish for both sport and com-
merdaJ fishing.However,the use of conventional
recording depth sounders to indicate the depth of
fish in shallow water (0 to 10 ft)is difficult because
of mlechanical limitations.The minimum full chart
depth displayed by a single-stylus sounder is typi-
cally 100 ft.If 5-in.wide chart paper is used,fish at
depths to 10 ft wilt be displayed in a O.5-in.wide
section of the chart (Fig.1).If there are many fish.
reeon:fed in this 0.5-in.space,it is difficult to dif-
ferentiate individual fish.To spread this 0-to 10-ft
Equipment and
T'ech n iq ues for
Monitoring the Vertical Distribution
of Fish in Shallow Water
20
-
"""
fiG.2 An echogrU'l from the Benmar DR~showing indi-
vidual targets of J-in.long cohc.salmon.Water depth is 8 ft.
Individual targets of fish could have been
manually counted to quantify the percentages of
fish in specific depth intervals;however,this proved
to be difficult because there were approximately
300 fish in the test tank..
COUNTER/PRINTER SYSTEM
An electronic target echo counter and tape
printer system were designed and constructed so
that the numbers of fish at specific depth intervals
could be quantified and recorded automatically.
This system was constructed with standard tran-
sistor-transistor-Iogic gates on printed circuit
boards manufactured at the Prescott Facility.The
printer used was the Model B5-102 Moduprinter®
manufactured by Practical Awtomation,Inc.
Two transducers were used with the counter
system so that a maximum volume of water inside
the tank could be sampled.One transducer was
mounted at the bottom of the tank;the other just
beneath the water surface.Four detection areas
were defined in the intersecting detection cones
(Fig.4).Because the entire volume of the tank is not
sampled,all results of the tests are based on the
assumption of homogenous horizontal distribution
of fish in the tank.
Basic operation of the counter/printer system
is illustrated by the timing diagram in Fig.5.The
instrument can be programmed to generate a trigger
pulse once every 5,10,20,or 60 min.Whenever
this pulse occurs,it turns on the sounder,auto-
matically switches to the top transducer,and
initiates the control logic for the counting of a
sample.Forty sec after this pulse occurs,the con-
trol logic activates electronic counters #3 and #4.
During a sampling period of 1 sec,all fish echos
REDWOOD
//TANK
r-----~-+-+_~T..NSOUCER
T
8'
......
fig.3 Transducer in test I:iIInk showing conical detection
yolumes of original trilnsducelr (slNded)and the Webster wide
beilm trilnsduce,.
test tank.The transducer was mounted at the bottom
of the tank so that as much as possible of the water
at the top of the tank could be monitored for fish.
The sounder was a<:tivated for approximately 2.5
min once each hour by an electrical timer.The
resulting echograms revealed two distind depth
behavior patterns:1)during daylight hours the fish
were quite active and homogenously distributed
below 2 ft,2)at night the fish concentrated at mid-
depth (2 to 6 ft deep)and were inadive.
COUNTER 1
COUNTER 2
fiG.4 Detection areas inside the intersecting conical detection
yolumes.
Monitoring Vertical Distribution 21
FIG.S Basic:liming diagnm of the elec:.tronic counter/printer
system.
---.,---,~--..-.
......~
-"-_.~-~~....'~',
AG.6 Photograph of the la-transducer anilY.
AG.7 The eJectronic sequencer used with !he 10 transducers.
These tests demonstrated that the Benmar@
sounder could be used effectively for monitoring
the depth of fish in shallow water.
MONITORING THE VERTICAL DISTRIBU-
TION OF FISH IN THE LOWER COLUMBIA
RIVER
An attempt to use the sounder from a boat in
the river was unsuccessful.The presence of the boat
seemed to "spook"the fish and very few were
detected.Subsequently an array of 10 transducers,
mounted on an aluminum sled (Fig.6),was placed
on the bottom of the river to overcome this problem.
An eledronic sequencer,with transistor-tran-
sistor-Iogic gates and relays (fig.7),was designed
to serially switch the sounder output to each trans-
ducer.Fig.8 is a block diagram of this unit~
III
L
110
I : ,
)0 ~SO
TIME IS£CONOSI
n
10
_____,-fl _
__________----Ifl-
___________-JIL
BOTTOM CCUlll
PRINf
1IfS£T COUll1US
TOPCWIII
TRICCUPlJIS£n~_
J
RESULTS OF TANK TESTS
During 1974,separ,iite tests were conduded
continuously for a week to monitor the diel depth
behavior of squawfish,smelt,crappie,and cutthroat
trout in the test tank.IFor these tests,the trigger
pulse was programmed to count a sample once an
hour,day and night.Fish depth data were recorded
by the automatic countl:!r/printer system;the task
of manually counting fish echos was eliminated.
Data from the tapes were tabulated and standardized
to compensate for the unequal water volumes of the
detection areas (Fig.4)"Although some variation
was observed.in the results of these tests,all species
generally avoided the top 2 ft of the tank and tended
to concentrate in the middle areas. The test tank
environment is obviously different from that of the
river;however,we did not have to contend with the
problem of false echo!.caused by other fish or
debris.
22 Marshall
detected by the sounder in the lower half of the
tank are counted and stored·in the counters
(Fig.4).Sixty transmit pulses occur during this
sample period;thereforl:!,a single fish in detedion
area #3 for the sampling period would result in the
number 60 being stored in counter #3.Two fish in
the area would resu It in the number 120 being stored
in the counter.The sounder transmit pulse width is
50 psec.At a velocity of sound in water of 5000 ftl
sec,this represents a distance of 3 in.Therefore,
two fish closer together than 3 in.could possibly
be deteded as a single fish by the sounder.Fifty
sec after the trigger pulse,the bottom transducer
is automatically switched to the sounder and after
a 1Q-sec delay the second sampling period is used
for counting the number of fish echos in detection
areas #1 and #:2 (Fig.4).Seventy sec after the:trigger
pulse the data stored in the counters is printed
on the chart paper.The counters are then reset
to zero and the sounder and control circuits are
turned off.When the next trigger pulse occurs,
the sequence is repeated!.
I~
....,
,"""
.....
r---------------------------- ---- ---,
I +12 I
I I
I VOLTAGE :
REGULATOR !
I
I
I
I
I
I
I
I
I
I
1
1
I
.J
~
CABLE TO SOONOER
FIG.8 Schematic diagram of the electronic sequencer.
The sled is anchored underwater on a gently
sloping beach with the transducers "looking"up-
wards (Fig.9).Cables from each transducer are
connected to the sequencer on the beach.A single
coaxial cable connects the sequencer to the sounder.
After the gear is in position on the beach,the
sounder is set to record for 5 min once an hour.
Each transducer is gated on for 10 sec.The re-
sulting echogram is a series of "stair step"patterns
because of the different depth of each trans-
ducer (Fig.10).The counter/printer system has
not been adapted for automatically recording data
from the transducer array and the fish echos must
be counted manually.
This system proved to be reliable and relatively
trouble free.Data on the vertical distribution of fish
in a natural environment was collected and reported
on by the staff of the Prescott Facility (Blahm,
McConnell,and Snyder,11974).
FIG.9 Transducer array in position underwater with the elec-
tronics equipment on the beach.
FIG.10 An echogram from the transducer array instaUed
underwater on a sloping beach.
REFERENC-=S
Blahm,T.H.1972.Gas Supersaturation Research.Report to
North Pacific Divisic~>n,U.S.Army Corps of Engineers.
Blahm,T.H.,McConnell,R.).and Snyder,G.R.1974.Gas
Supersaturation Research 1972-1974.National Marine Fisheries
Service,Prescott Field Station.
Monitoring Vertical Distribution 23
,,~
Disso IVE~d Gas ID,E.Weitkamp
Supersaturation:
Live Cage Bioassays at
Rock Is~and Dam,Washington
..-
-
~-
ABSTRACT
Three live cage bioassays using juvenile chinook salmon
(0.rshawytscha)were conducted in supersaturated Columbia
River water at the Rock Island Dam fore bay.The tests of 10 and
20 days'duration utilized volition,specific depth,and inter-
mittent exposure cages whi'ch were suspended between the sur-
face and a depth of 4 m.lhe volition cages extended from the
surface to depths of 2.3,and 4 m.The four specific depth cages
were 1 m deep cages suspended at 1 m intervals between the
surface and a depth of 4 m.Iintermittent expoSures were achieved
by raising and lowering three 1 m deep cages in the water
column daily to change Ithe actual level of supersaturation
experienced by the test fish ..
At total dissolved gas supersaturations of about 120%in the
reservoir water,significant mortalities were encountered only
in fish held within 1 m of Ilhe surface.Fish held within 2 m of
the water's surface for a period of 16 hr per day suffered
significant monalities only when the supersaturation rose to
about 125%and above.The effects of supersaturations above
about 125%appear to be much greater than supersaturations
below this level.Fish allow~~to seek the depth of their choice
in the 4 m deep volition cage did not suffer monalities at satura-
tions of 119%and 128%,all.hough some developed gas bubble
disease lesions.Most fish showing slight to severe gas bubble
disease lesions were able to recover when placed at a depth of
3 to 4 m in the supersaturatE!d water.
One of the obvious means of determining the effect
of a given water quallity parameter on fish is the
bioassay experiment.This technique has been
applied to the investilgation of the effects of dis-
solved gas supersaturation on fish.Bioassays in gas
supersaturated water have been conducted by
many workers under both artificial laboratory and
natural field conditions.Most of the experimental
work dealing with the effects of dissolved gas
supersaturation has been conducted in the labora-
tory using artificially supersaturated water or,in
some cases,supersaturated Columbia River water
in shallow tanks.Several live cage bioassays study-
ing the effects of depth in relation to supersatura-
tion have been conducted in the Columbia and
Snake Rivers by EbeJ (1969 and 1970)and Meekin
and Turner (1974).
24
The live cage bioassays conducted by Ebel at
Priest Rapids and Ice Harbor Dams tested varying
high levels of supersaturation.The nitrogen levels
during the three Priest Rapids tests in 1967 ranged
from a high of 143%at the start of the first test to
118%at the end of the third test.The 1970 Ice
Harbor tests exposed juvenile chinook for 7 days to
nitrogen levels of 127%to 134%.At some time dur-
ing most of these tests,supersaturations of 1300.ki or
greater were experienced.It is therefore difficult
to determine if the mortalities resulted from the
highest levels encountered or from continuous
levels above 120%or 125%.
The following study was therefore designed to
test juvenile chinook under similar field conditions
at lower,but more consistent levels of supersatura-
tion.The snowpack of the 1973-74 winter was
sufficient to ensure at least moderate levels of
supersaturation in the mid region of the Columbia
River during the 1974 runoff period.The study was
conducted at Rock Island Dam on the mid Columbia.
Favorable weather and water conditions allowed
the levels of supersaturation to remain quite con-
stant at the test location during test periods.
The study was also designed to test the effect of
intermittent daily exposures to gas supersaturation.
It is believed that under natural conditions juve-
nile saJmonids and other fish are most likely ex-
posed to high gas levels on an intermittent basis.
In some areas this is due,in part,to the inter-
mittent production of supersaturation conditions,
as the spill may occur intermittently at dams on
smaller rivers.In certain areas,such as the mid and
lower Columbia River,however,supersaturation
can be present continuously for several months at a
time.Fish are intermittently exposed to this con-
tinous supersaturation by their vertical move-
ments in the water column.The live cage bioassay
Weitkamp:Parametrix,Inc.,Environmental Services Section,
Seattle,Washington.
",
L
fiG.1 Schematic of fish Rve cages used in supersaturation
bioassays showing the depths occupied by the various uges.
FIG.2 Live cages being assembled ;at the study site;1 m inter-
mittent exposure and fixed depth cages.and 3 m volition cage
are shown.
a zippered opening at the bottom of one side.When
the cages were raised each day,these openings
permitted removal of dead fish from the bottom of
the cage.These openings also permitted observa-
~ion of the test fish in shallow water when the cages
were raised.The specific depth and intermittent-
exposure cages were provided with additional zip-
pered openings on the top for introducing the fish
into the cages.The assembled frames and nets are
shown in Fig.2.
Two cages were suspended within each float.
This provided a minimum distance of 1-1/2 ft
between cages in the same float and about 6 ft
between cages in adjacent floats.Fig.3 shows the
cage and float assemblies in position adjacent to a
barge in the Rock Island forebay.
Test Periods
The first test was conducted for a period of
10 days while the second and third tests were
extended to 20 days each.Because Ebel's (1969 and
1970)previous experiments were conducted for
AIO-[m
8tH IHrr 16hr
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I I I II I
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160M 12hr 8hrIIATl-4mIlNIERMlmNIEXPOSUR£
D-4m D-lm 0-2m D-lm l-2m 2-lm
WAlER
u::_"'[]DIpm
2m-Utj
3m -
'm-
\VQITlON II FIXEDDEPnl
PROCEDURES
Location of Study
A major objective of the dissolved gas bio-
assays was to test the survival of fish in water con-
taining moderate,but relatively constant,levels
of supersaturation under natural conditions.The
Rock Island Dam forebay on the mid region of the
Columbia River was chosen as a site where these
conditions could be obtained during the""'1974 runoff
period.This dam provided <l location where the live
cages could be placed in water at least 20 ft deep
and with a desirable current (0.1 to 0.5 m/sec).
The moderate current ensured good circulation of
water through the live cages without stressing the
test fish.
Submerged Cages
The cages used to conduct the supersaturation
bioassays were of three basic types,volition,fixed-
depth,and intermittent-exposure.All of the cages
were constructed with horizontal dimensions of
4 ft by 4 ft.The volition cages extended from the
surface to depths of 2,3,Clnd 4 m.The four fixed-
depth cages were each 1 m deep and were placed
at depths of 0 to 1 m,1 to 2 m,2 to 3 m and 3 to 4 m.
Following the first test,the 3 to 4 m cage was used
for testing the recovery of fish showing signs of
gas bubble disease (GBD).The intermittent-expo-
sure cages were also 1 m deep,but were constructed
so that they could be placed at desired depths
between 0 to 1 m and 3 to 4 m.In the first test,the
upper depth of the intermittent-exposure cages
was 1 to 2 m.The upper depth was raised to 0 to 1 m
for Tests "and III.The lower depth was 3 to 4 m
for all three tests.A schematic of the various live
cages is shown in Fig.1.
The cages were constructed with a framework
of 1 in.angle aluminum held together by stainless
and cadmium plated bolts and nuts.Diagonals of
1/2 in.aluminum conduit were used to stabilize
the deeper cages.The frames for the volition and
fixed-depth cages were constructed with 6-ft cross
pieces at the top to suspend the cages within the
floats.Suspension of the intermittent-exposure
cages was accomplished by placing removable
6-ft pieces of aluminum tubing horizontally under
the cross pieces which were located at the appro-
priate depths.
Knotless nylon netting of 1/2 in.stretched
mesh was used to enclose the cages.Each cage had
tests were designed to investigate this type of
intermittent exposure that fish experience as they
undergo diel migrations in continuously super-
saturated reservoirs.Intermjttent exposure due to
daily changes in depth should,at least theoreti-
cally,reduce the effects of supersaturation.
--
.-
Live Cage Bioassays at Rock Island 25
-
-
-
-
FIG.3 Assembled liwe agel;in f~ts.Note extended frame of
raised intermittent-exposure 4:age in background.
periods of 7 to 12 days.it was felt that tests of
10 days'duration would be practical.However,the
mortality rates of the first test were much lower
than originally anticipated.so the second and third
tests were lengthened ito 20 days.
Fish Used
The juvenile chinook salmon used in these
tests were provided by the Washington Department
of Fisheries from its ha.tchery facilities at the Wells
Spawning Channel.Much of the success of these
tests can be attributed to the excellent condition of
these upper Columbia.River summer run chinook
salmon.The fish were free of complicating
diseases and appeared very healthy when placed
in the live cages.The fish showed no signs of
smolting at the time they were placed in the live
cages.The fish had an average fork length of
97 mm in Test I,105 mm in Test II,and 112 mm in
Test 111.One hundred fish were placed in each of
the 1 m deep cages.Between 100 and 200 fish were
placed in the deeper volition cages,depending on
the number of fish provided.
Feeding of Fish
No food was given to the fish during Tests I
and III.The fish used in Test II were fed from the
26 Weitkamp
11th day to the end of the test period.It was decided
to begin feeding the Test II fish with Oregon Moist
Pellets@ when some began to show signs of starva-
tion.They were fed at times when the cages were
brought to the surface to check mortalities.It was
hoped that feeding the fish only when the cages
were raised would prevent an artificial depth dis-
tribution in the volition cages that might occur with
surface feeding when the cages were at their
normal depth.Although feeding with the cages
near the surface was successful,it did appear to
cause at least some of the fish to spend more time
near the surface.For this reason no feeding was
attempted in Test III.
Mortalities
Each live cage was raised and checked for
mortalities between 0800 and 1100 daily"'for each
test.During this period the cage was held with its
bottom several inches below the water's surface
so that the fish cou Id be observed for signs of gas
bubble disease (GBD)or any other problems that
might occur.This procedure permitted close obser-
vation of the fish each day without unduly stressing
the fish.The procedure also permitted cleaning
and inspection of the cage netting.Dead fish float-
ing in the cages that extended to the surface were
also removed at 1600,2000,and 2400 hr when
the depth of the intermittent-exposure cages was
changed.All three of the intermittent-exposure
cages were lowered to a depth of 3 to 4 m at 0800
each day.One of the cages was then raised to the
surface position at 1600.2000,and 2400 hr to pro-
vide the surface exposures of 16,12 and 8 hr,
respectively.
The mortalities were checked immediately
following their removal from the cages for external
signs of GBD.The locations of external emboli and
hemorrhages were recorded as well as the fish
length.The abdominal cavities of all mortalities
were checked for internal signs of GBD and the
presence of food in the digestive tract.
Dissolved Gas Analysis
During the morning inspection of the live
cages,the dissolved gas levels present in the fore-
bay were measured using a Weiss satlJrometer.The
measured saturations given in this report are most
likely lower than the levels actually present in the
forebay water.An on-site comparison of the satu-
rometer used in this study was made with several
other saturometers,indicating readings were 1%
to 3%low.Routine monitoring by the Chelan
County PUD indicated that supersaturation in the
study area ranged from 120%to 131%during the
test periods.The saturometer readings were also
taken at a time of day when river supersaturations
were near their lowest level.A 24-hr monitoring
0-lm
8 10 12 14 16 18 20
DAYS
FIC.4 Test I,total dissolved gas levels and %cumulative
mortalities for the 0 to 1 m cage,May 24 10 June 3,1974.No
mortalities occurred in the other cages.
These temperatures were recorded between 0800
and 1000 hr.Some days surface water tempera-
tures were a degree or more higher later in the day.
Secchi disc readings taken during this test ranged
from 0.91 to 1.22 m.
In Test I no fish were lost due to handling
stress.The first mortaility occurred during the 2nd
day in the 0 to 1 m fixed.;.depth cage.The mortality
rate iflcreased somewhat in the 0 to 1 m cage after
the 4th day,reaching a cumulative mortality of 53%
at the end of 10 days.A single mortality occurred
in the 2 to 3 m fixed-depth cage during this test.
This single mortality was obviously due to trauma,
not to GBD.No mortalities occurred in any of the
other cages during this 10-day test.The percent
cumulative mortality during this test is shown in
Fig.4,and Table 1 lists the daily cumulative mor-
tality for Test I.
The surviving fish were examined for signs of
GBD at the end of the test.Ninety percent of the
survivors in the 0 to 1 m cage showed some signs
of GBD.These varied from a few bubbles in one fin
to bubbles in most fins,the head,and mouth,as
well as hemorrhaging at the base of the fins.In the
1 to 2 m cage 15%of the fish showed signs of GBD.
Only 3%of the fish in the 0 to 2 m cage showed
signs.In both cages the signs of GBD were re-
stricted to bubbles in the caudal fins.
Live Cage Bioassays at Rock Island 27
130
z0 125;::-<""120:::II-<'"115~
no
100
80
~
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20
00 2
RESULTS
Test I
The fish for the first live cage bioassay were
placed into the cages in the Rock Island forebay on
May 24,1974.At this time,the total dissolved
gas saturation in the rivler water was 122%.During
the first 4 days of the study the saturations remained
near 122%.The saturations then dropped to about
120%,and remained near 120%for the remaining
6 days of the test.The total dissolved gas levels
measured at the study location are shown in Fig.4.
Saturometer readings were also taken inside the
cages during all three tests.Saturations inside the
cages were not more than 1%of saturation lower
than the readings for the surrounding water.
The water temperatures which were recorded
daily during Test I ranged from 10AOC to 10.9°C.
study conducted in the Rock Island forebay by the
Chelan County PUD in August 1974,showed that
dissolved gas levels change by about 3%of satura-
tion during a 24-hr period.The lowest gas levels
during this 24-hr period occurred between 0300 and
0900 hr,which includes the time that the gas
content was measured during the live-cage tests.
Some daily variation in Ithe gas levels observed in
the live-cage bioassay is due to differences in the
water temperature which varied according to the
time of day during which measurements were taken.
A number of times during the tests,dissolved
gas supersaturations were measured inside some
of the cages.Th is was done to detect any measur-
able effect of the cages on the gas content of the
water as it passed through the cages.....The gas con-
tent of the water inside the cages was always lower
than that of the water outside of the cage by less
than 1%of saturation.
Dissolved oxygen levels were measured by
the Winkler method.Water temperatures were
measured at the surface using a mercury thermom-
eter accurate to 1:.0.1 0c.Light penetration readings
were taken daily adjacent to the cages using a
Secchidisc.
An attempt was made to record the fish
depths inside the 0 to 4 m volition cage.A Benmar
DR-681@ echo sounder with a minimum range of
o to 50 ft was used to make the recordings.The
DR-681 was the only shallow water echo sounder
,that could be acquired for these tests.No useful
\
ecordings were obtain~~d with the echo sounder
"'""due to apparent interferen~e from the cages.
As the depth distriibution of the fish in the
.olition cages was of considerable importance,an
""'"tempt was also made to observe the fish using
uba gear.Even with a strong underwater light,
he diver was unable to observe any of the fish due
o the turbidity of the river water.
.-
....
-
TABLE 1 Yes'l I,Percent Cumulatiye Morbllity Due to Gas Bubble Disease in Juyenile Chinook
5.1lmon bPOSlI!d to 119%to 123%Supersatur~tion ~t Rock IsI~nd FOle~y,M~y 24 to June 3,1974.
~Holdinl
Cage depth (meters)
time 1-2 m and 3-4 m '
(d~ys)0-4m G-3m G-2m 6-1 m 1-2 m 2-3 m 3-4m t 16 +8 t 12 +12 t B +16,
2 1
3 2
4 4-5 14
6 21
7 23
8 35
"""9 42
10 0 0 0 53 0 0 0 0 0 0
Survivors
with GBD
lesions (%)0 0 3 90 15 0 e 0 0 0
1 tindicates hr/day at 1-2 m depth,}indicates hr/day at 3-4 m depth.
Most of the survlvmg fish with evidence of
GBD were placed in the 3 to 4 m cage to determine
if they could recover.A few of the severely affected
fish died before the cage was submerged.The
remainder were held as part of the second experi-
ment.All surviving fish with no indication of GBD
were released in the Rock Island forebay.
first in the 0 to 1 m cage (Table 2).The daily mor-
ality rate in Test II (Fig.5)was similar,but not as
consistent as that of Test I.The rate at which mor-
talities occurred in the 0 to 1 m cage between
FIG.5 Yest II.toQI dissolved SillS levels iIInd %cumubtive
IIlOIUlities pNter th~n 1%'01'ill CiIIges,June 5-25,1974.
O~---l.._....I-_L..---l.-e:::::..:==:..=I:=:"='=::l....ii=--.Io24 6 8 10 12 14 16 18 20
DAYS
,.
I
20
130
z 125!2-<Ca:120::>-<VI 115III
110
100
G-lm
G-2m
III G-3m
~-.-h6t8~-60 _......+12t12a:
~
III
~;::.to<=l
:E::>
C,,)
Test II
The total dissolved g;as saturation of the river
water at the beginning of the second test (June 5)
was down to 120%(Fig.5)as measured at the test
site.By the 4th day (June 9),the gas.saturation had
risen to 122%,and remained constant until the
10th day when it rose to 123%.During the last
11 days (June 15 to 25)of the test,the saturation
varied between 123%and 126%.
The water temperatuflE~s at the beginning of the
second test remained near 10.7°C for the first
3 days.During the remainder of the test the water
temperatures showed a steady increase of about
0.2°C per day,reaching 14°C by the end of the test.
The light penetration decreased considerably dur-
ing the last half of the ~;econd test.For the first
8 days,the Seechi disc readings ranged from 0.91 m
to 1.22 m.During the la!it half of the test,Secchi
disc readings decreased to 033 to 0.61 m.
Two fish were lost at the beginning of this test,
apparently due to handling stress.The other test
fish showed no signs of GBD at the time of these
mortalities.The mortalities occurring due to GBD
during the second test did not begin until the 6th
day.As in the previous test.mortalities occurred
28 Weitkamp
-~
TABLE 2 Tesll II,Percent Cumulative Mortality Due to Gas Bubble Disease in Juvenile Chinook
Salmon Expo!ied to 120%to 126%Supersaturation at Rock Island Forebay,June 5-25,1974.
Recovery
~m Wm ~m ~m Mm Mm ~m
Cage depth (meters)
0-1 m and 3-4 m'
116 +8 112 ,J..12 18 ,p6
1
4
1
1
1
1
1
1
2
2
2
4
7
12
2
4
7
7
7
10
10
10
10
10
10
10
10
10
1
1
1
1
1
1
1
1
2
5
18
30
31
36
43
54
72
74
77
81
84
85
88
2
2
3
3
8
11
17
1
1
2
2
3
0.5
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
Holding
time
(days)-
Survivors
with GBD
lesions (%)38 64 77 91 o 2 9 69 24 15
1 1indicates hr/day at 0-1 m d.~pth.findicates hr/day at 3-4 m depth
*Approximately 1/2 of the fish lost through small tear in cage.
days 5 and 14 during Test II was very close to that
of the same cage between days 2 and 10 during
Test I.The mortality rate in the 0 to 1 m cage of
Test II appeared essentially the same as that of the
same cage in Test I,e):cept that it began about
3 days later.A cumulative mortality of 53%was
reached on day 10 of Test I,and a similar mortality
of 54%was reached on day 13 of Test II.
.The mortality rate in the 0 to 1 m cage de·
creased during the last 6 days of Test II.From
day 14 to the end of the test,the cumulative mortal-
ity only increased from 7'2%to 88%All but one of
the fish remaining in the 0 to 1 m cage at the end of
the test showed signs of GBD.
Unlike the first 10-day test,mortalities oc-
curred in a number of thl:!other cages during Test II.
These mortalities began to occur prior to 10 days,
but at a very low rate (Table 2).The 0 to 3 m cage
lost one of 200 fish on the 8th day.No other mor-
talities occurred in this cage until the 16th day.No
mortalities occurred in the 0 to 4 m cage during
Test 11.The size of the test group in the 0 to 4 m cage
was reduced around the 18th day when slightly
more than half of the group was lost through a
small tear in the netting.The 0 to 3 m cage fish
experienced a few mortalities during the last
5 days with a final cumulative mortality of 3%.
Mortalities began to occur in the 0 to 2 m cage on
the 14th day and increased to 17%at the end of
Test II.
A portion of the survivors in each of the three
volition cages showed signs of GBD.In the 0 to 4 m
cage 38%of the survivors showed indications of
GBD,as did 64%of those in the 0 to 3 m cage and
n%in the 0 to 2 m cage.
Only a single mortality occurred in each of the
two fixed-depth cages between 1 and 3 m.In the
1 to 2 m cage a fish died on the last day,and in the
2 to 3 m cage a mortality occurred on the 13th day.
No survivors in the 1 to 2 m cage showed signs of
GBD;however,2%of those in the 2 to 3 m cage had
GBD.
Some mortalities occurred in each of the
intermittent-exposure cages which were raised to
a depth of 0 to 1 m in Test II rather than to 1 to 2 m
as in Test I.In the cage spending 16 hr/day at the
Live Cage Bioassays at Rock Island 29
'.~
FIG.6 Test III,total dissolved gas lewels and %cumulative
mortalities for the wolidon and fRed depth cages,rune 28 to
July 18.1974.
j
~l
---O-lm
-----0-2m
•••••••••lr3m
---2-3m
---I-2m
___J--
--,-~,--_t--_-,*_r
I,__I
_J
j"'..._.J-"'--'_J,J-
j-J
I r-PO,~,.
I ••~:""•••
o La-L..E;:::-=i..:4 =..1:6=-=8:"=±10="==1J:=2=--:t14:=-~1:=6 =-=7:18:=-==:ZO=='
DAYS
Mortalities also occurred in the two fixed-
depth cages between 1 and 3 m,but at a much
lower rate than in the 0 to 1 m cage.In the 1 to 2 m
cage mortalities first occurred on the 9th day of,
the test (Table 3).In this cage the mortalities then
remained constant giving a cumulative mortality
of 70%at the end of 20 days.In the 2 to 3 m cage,a
single fish died with signs of GBD on the 2nd day.
No other mortalities occurred in this cage.Thirty-
three percent of the survivors in the 1 to 2 m cage
showed signs of GBD,while none of the survivors
in the 2 to 3 m cage showed GBD lesions.
Fish in the shallowest volition cage began to
suffer mortalities on the 2nd day (Fig.6).After a
high mortality of 17%between the 2nd and 3rdday,
the mortality continued at a relatively constant
rate.The cumulative mortality at the end of the
test was 61%in this cage (Table 3).In the 0 to 3 m
cage,the first mortality did not occur until the 5th
day.and only 7.5%had died at the end of the test.
No mortalities of any kind occurred in the 0 to 4 m
cage during the 20-day test period.Signs of GBD
were present in 85%of the survivors in the 0 to 2 m
cage,35%in the 0 to 3 m cage,and in only 8%of
the survivors in the 0 to 4 m cage.The 8%with GBD
lesions in the 0 to 4 m cage had only slight signs,
usually only a few bubbles in a single location.
80
~
;;i....
""60~
rIt...::-....~O<~
~::>
<.)
20
130
z 1250;::::
<""120::>....<'"115rIt
110
100
30 Weitkamp
surface and 8 hr/day at 3 to 4 m the first mortality
occurred on the 9th day with the cumulative mor-
tality reaching 12%by the end of the 20-day test.
Mortalities did not occur until the 18th day in the
other two intermittent-exposure cages.The cumu-
lative mortalities at the end of the test were 4%in
the cage spending 12 hr/day at the surface and 1%
in the cage spending 8 hr/day at the surface.
The portion of the survivors showing signs of
GBD was related to the length of the surface expo-
sure.Signs of GBD were found in 69%of the sur-
vivors in the cage spending 16 hr/day at the sur-
face,24%in the 12 hr/day exposure cage,and 15%
in the 8 hr/day exposure cage.
In Test II the 3 to 4 m cage was used to test the
recovery of fish from Test I that showed signs of
GBD.These fish began to suffer mortalities on the
7th day,and had a 10%cumulative mortality after
20 days.None of the fish in the recovery cage died
after the 12th day.
These mortalities appeared to be due to severe
fungal infections resulting indirectly from GBD.
All dead fish had the caudal fin completely eroded,
and were covered with fungus as far forward as the
anus and dorsal fin.Most of the fish placed in the
recovery cage lost all sligns of GBD by the end of
Test II.Only 9%of the surviving fish in this cage
showed any signs ofGBD when they were released.
The remaining signs were slight cases of exoph-
thalmia.Bubbles and hemorrhages were not ob-
served in any of these fish at the time of their
release.
Test III
The supersaturation of the water at the test
site in Rock Island forebay at the initiation of the
third test was much higher than at the beginning of
the two previous tests.During the first 9 days the
supersaturation dropped slightly and ranged from
120%to 125%at the end of the test.
During the third test.water temperatures
were also higher than th'ey had been during the two
previous tests.At the beginning of Test III,the sur-
face water was 143°C.lrhe temperature continued
to show a steady incr,ease throughout the test,
reaching 16.1°C at the end.Unlike the previous
tests.light penetration showed a steady increase
through the third test.Secchi disc readings were at
OJ6 m at the beginning and reached 1.89 m near
the end of the test.
The initial mortalities occurring in most of the
cages during the thinj test were considerably
higher than in the two previous tests.In the 0 to
1 m cage 6%of the fj~~h had died with signs of
GBD (Fig.6).Eighty-three percent were dead by the
end of the 2nd day.and aU test fish had died with
GBD lesions by the 3rd day.
~,
-
",.,.
TABLE 3 Tesl lilt,Percenl Cumulative Mortality Due 10 (las Bubbre Disease In Juvenile Chinook
Sillmon Exposed 10 120%10128%Supel'5illuralion OIl Rock Island Forebay,June 2810 July 18,1974.
Holding
Cage depth (meIers'
0-1 m and 3-4 m'time Recovery
(days)0o4m O..3m 0·2 m 6-1m 1-2 m 2-3 m 34m t16 ~8 t12 ,j,.12 t 8 p'
1 6 3
2 3 83 1 21 9
3 17 100 1 45 20 1
4 21 1 48 22 1
5 0.5 23 1 52 23 1
6 0.5 26 1 2 57 27 3
7 0.5 28 1 2 61 27 3
8 1.0 30 1 2 64 28 3
9 3.0 35 3 1 2 68 35 5
10 4.0 42 5 1 2 68 35 7
11 4.5 43 6 1 2 68 38 7
12 4.5 45 7 1 5 68 38 7
13 5.0 48 10 1 5 68 38 7
14 5.5 51 10 1 5 68 38 7
15 6.0 52 11 1 5 68 38 7
16 7.0 55 17 1 6 68 38 7
17 7.0 57 22 1 7 68 38 7
18 7.5 57 24 1 8 68 38 7
19 7.5 60 29 1 10 68 38 7
20 0 7.5 61 30 1 13 70 39 7
Survivors
with GBD
lesions (%)8 35 85 33 0 5
4 0 0
1 tindicates hr/day at 0-1 m depth,~indicates hr/day at 3-4 m depth
The fish in the intermittent-exposure cages
suffered much higher mortalities during the third
test than during the two previous tests.Almost all
of these mortalities occurred during the first half
of the test.In the cage occupying the 0 to 1 m depth
for 16 hr/day,the fish began to die with evidence
of GBD on the 1st day (Fig.7).This mortality
reached 45%by the 3rd day and then leveled off.
At 9 days the mortality in this cage was 68%and
this only increased to 70%at the.end of 20 days.
The mortality in the 12 hr/day cage was similar to
that of the 16 hr/daycage,but at a lower rate.In
the 12 hr/day cage,the mortality was 20%by the
3rd day and 35%by the 9th day.The final cumula-
tive mortality in this cage was 39%.The first death
in the 8 hr/day exposure cage occurred on the 3rd
day.This cage suffered a much lower mortality
than the other intermittent-exposure cages at only
7%in the first 10 days.No additional mortalities
occurred in the 8 hr/day exposure cage.
A number of wild fi!;h residing in the area of
the live cages were examined for signs of GBD.
One or more juvenile chinook salmon about 40 to
80 mm long were trapped on top of the deeper live
cages each day as the cages were raised to check
for mortalities.These small chinooks were residing
around the Jive cages from late May to the end of
the 3rd test on July 18.Three-spined sticklebacks
(Gasterosteus aculeatus)were also common around
the live cages.These fish were occasionally trapped
on top of the cages,but many were captured with a
dip net when they were at the reservoir's surface.
As many as 15 sticklebacks were captured at one
time in the evening when they were normally found
at the surface.None of the juvenile chinooks or the
sticklebacks captured adjacent to the live cages
showed any signs of GBD.It is uncertain how many
fish were checked as the fish were returned to the
reservoir and some were undoubtedly captured
more than once.
DISCUSSION
Objective
The major objective of this study was to test
the effects of naturally occurring moderate levels
of dissolved gas supersaturation (120 to 130%)on
juvenile salmonids.This objective was achieved in
field experiments consisting of one la-day test and
Live Cage Bioassays at Rock Island 31
.t.;-.
Intermittent-Exposure
The occurrence of GBD was also directly re-
lated to the daily duration of exposure to super-
saturation resulting from raising and lowering fish
in the water column in the intermittent-exposure
cages.As with the deeper volition cages,both mor-
talities and the percentage of survivors with signs
of GBD were lower than those occurring in the 0 to
1 m cage.In both of the 2O-day tests,the 16 hr/day
near-surface exposure cage had cumulative mor..!
talities and percentages of survivors with signs of
GBD that were more than twice those of the fish
receiving only 12 hr/day of exposure at the sur-
face.Those fish receiving only 8 hr/day of exposure
experienced monalities and showed evidence of
GBD only when the supersaturation ranged from
123%to 128%,as occurred during the last 10 days of
Test I and during the first 18 days of Test II.
Depth Effect
The distinct mitigating effects of small differ-
ences in depth was demonstrated by these three
live cage tests.In Tests I and II,at supersaturations
of 119%to 123%,only fish restricted within 1 m of
the surface suffered mortalities during the first
10 days.In Test II the fish held within 2 and within
3 m of the surface experienced 17%and 3%mor-
talities,respectively,during the last 10 days of the
20-day test when the supersaturation varied be-
tween 123%and 126%.The number of survivors
with signs of GBD was likewise directly related to
the depth at which the fish were restricted,and
thus their exposure to supersaturation.Those fish
exposed to effectively decreased supersaturation
levels,due to the greater depth of their cage,had
both more survivors and a smaller percentage of
survivors with signs of GBD .
It is highly significant that no fish were killed
by supersaturation in the 0 to 4 m volition cage
during any of the three tests at saturations of 120%
to 128%.The number of fish in this cage with signs
of GBD was low,ranging from 0%at the end of the
first test to 33%at the end of the second test.The
lack of mortalities and low incidence of GBD in this
cage is of panicular importance,as these are the
test conditions most representative of what actually
occurs in the reservoirs with unrestricted fish move-
ment.Yet it should be noted that even in this 0 to
4 m cage,fish are confined in shallower water than
they normally occupy in the reservoirs according to
available depth distribution data.If juvenile chi-
nooks are able to tolerate supersaturations between
120%and 128%in this cage for a period of 20 days,
it is highly likely that they would do at least as well
without the restraints of the cage during their
normal migration.
10 12 14 16 18 20
DAYS
r-------....r-
.------------_:-
_..J".J
I("-__I,_I
j-
l ...----r-----
:r-,.J...~...""-
:J
-----tI6 p
---+12,12
..8.16
o 0 2 4
20
FIG.7 Test III.toU!disso'lfed gOIS Jewels ind %cumubtiYe
morWities for the intermitllent -exposure Citges.June 28 to
July 18,1974.
no 1..-._
100
two 2a-day tests.The presence of nearly constant
levels of supersaturation in the 120%to 130%range,
combined with the,uSle of healthy test fish,per-
mitted the accurate determination of the biological
effects of a narrow ranEre of supersaturation,rather
than those resulting from a wide range of super-
saturation.The gas levels tested werE~.also of par-
ticular significance as they were approximately
equal to the highest levels suggested in any pre-
vious repons as permissible without producing
GBD (Egusa,1959 and Shirahata,1966).
This study concentrated on the mitigating
effect that a fish's depth in the water column has
on the level of supersaturation which it can tolerate.
Since most fish in a reselrvoir occupy depths greater
than 1 m most of the time,they should theoreti-
caUy be able to withstand higher levels of super-
saturation than indicated for depths of less than
1 m.The increase in pressure associated with this
increase in depth of 1 m reduces the in situ super-
saturation value by about 10%(Leman,19n).Thus,
fish deeper than 1 m would be subjected to no more
than 110%saturation when the surface saturation
is 120%.This apparentlly explains why fish con-
tinue to survive in reservoir areas experiencing high
levels of supersaturation well above 130%in
surface waters.
130 .--------.-----------,
~125 ~---~!5120~
~11S
....
'"'"
....
-
"'"
-
32 Weitkamp
Effect Above and Below 125%Saturation
Although quite constant levels of supersatura-
tion were experienced during the three tests,the
slight variations in the supersaturation of the
.....reservoir were significant,as demonstrated by the
resulting mortalities.During the first test and the
first half of the second test,the supersaturation
remained below 123%.Almost all mortalities during
~this time occurred in the 0 to 1 m fixed-depth cage.
For the second half of Test III land the first half of
Test III,the supersaturation rose to between 124%
-and 128%,dropping to 123%on only one day.
During this period of higher gas content,mortali-
ties occurred in most of thecages.Again during the
second half of Test III,few mortalities occurred in
~any cage except the 1 to 2 m fixed-depth "'Cage.The
level of supersaturation during this last period had
again dropped to between 120%and 125%.
"""These results indicate that juvenile chinook
remaining below a depth of1 m can only be ex-
pected to suffer significant mortalities over a 20-
_day period when supersaturation is above 123%.
Fish held within 1 m of the surface for no more than
16 hr/day also suffered significant mortalities only
when the supersaturation rose to about 125%or
.-higher.At supersaturations of about 125%and
higher there was a marked increase in mortalities
of fish which spent 8 hr/day or more within 1 m of
.....the surface,and also in fish which were held con-
tinuously between 1 and 2 m.These mortalities
were reduced in fish which received not more than
16 hr/day of surface exposure when the super--saturation dropped below 125%.Fish held between
1 and 2 m appear to suffer mortalities only after
exposures of 9 or 10 days at supersaturations near
125%.This is indicated by comparing the mortalities
in the 1 to 2 m fixed-depth cage of Tests II and III.
In Test II only a single mortality occurred in the
_1 to 2 m cage on the last day after 10 days of expo-
sure to supersaturations between 124%and 126%.
In Test 1/1 the mortalities in the 1 to 2 m cage began
on the 9th day following exposures to supersatura-
-tions of 125%to 128%.
The percentage of survivors having signs of
GBD in the intermittent-exposure cages at the end
_of Tests II and III is another indication of the effects
of supersaturations below 125%as contrasted to
those above 125%.At the end of Test II,GBD lesions
were present in 15%,24%,.and 69%of the survivors
from the 8 hr/day,12 hr/c1ay and 16 hr/day near-
surface exposure cages,respectively.The super-
saturation was low (120 to 123%)during the first
.-.10 days of Test II,but higher (124 to 126%)during
the last 10 days of this test.At the end of Test III,
signs of GBD were absent in the survivors from the
8 hr/day and 12 hr/day exposure cages,and signs
I"'"were present in only 4%of the survivors from the
16 hr/day exposure cage.The supersaturation was
between 125%and 128%during the first 10 days of
this test,but dropped to between 120%and 125%
during the last 10 days.A comparison of these two
tests indicates that fish spending any appreciable
time in water less than 1 m deep are affected by
supersaturation above 125%,but recover rapidly
when it drops below 125%.
As indicated above,there appears to be a sig-
nificant difference in the effects of supersaturations
above and below about 125%.Above 125%the
effects of supersaturation are much greater pro-
ducing GSO faster in a greater percent of test fish
and in fish held in deeper water.Supersaturations
below about 125%appear to produce GBD only in
fish held within 1 m of the surface at least for expo-
sure periods of about 10 days.
Severe Mortality in 0 to 1 Meter Cage
The severe mortality.that occurred in the 0 to
1 ,m fixed-depth cage of Test III cannot be fully
explained.This 100%mortality within 3 days
occurred at supersaturations (125 to 128%)that were
only slightly higher than the supersaturations
(123 to 126%)present during the last 10 days of
Test II.The final cumulative mortality for the same
cage in Test II was 88%.
There are several factors that may have influ-
enced the sensitivity of the Test III fish.They may
have been stressed more than the fish used in pre-
vious tests due to high atmospheric temperatures
during their transport to the test site.There was,
however,no indication that the fish were stressed
when they were placed in the cages.The fish in
Test II/were slightly larger than those of Test II.It
.is doubtful that this slight size difference was suffi-
cient to alter the sensitivity of the test fish to pro-
duce the great differences between Tests 11 and III
mortalities.
The only real differences observed between the
two tests are the supersaturations at the beginning
of each test and the water temperatures during the
tests.The water temperatures during the first few
days of Test III were less than 1°C higher than dur-
ing the last few days of Test II.It seems unlikely
that this slight increase in temperature would have
been responsible for thf:-much higher mortality
rate during the first few days of Test III.Test II had
lower supersaturation initially (1200.-b),rising to the
higher levels only during the last half of the test;
higher levels of supersaturation (125 to 126%)
were present at the beginning of Test III.The fish
may have been seeking a way out of the cage dur-
ing the first few days,and were therefore spending
more time near the top of the cage at the time of
the high supersaturations in Test III,whereas by the
time the highest supersaturations occurred in
Live Cage Bioassays at Rock Island 33
-
,~
,~
-
Test II,the fish may have adjusted to the cage.The
Test III population was also reduced by about 50%
by the time of the highest supersaturation,enabling
the population to occupy a smaller space,perhaps
near the cage bottom.
Depth Distribution in 0 to 4 Meter Cage
It was hoped that some additional information
on the depth distribution of juvenile salmon ids
could be gained by recording the depths occupied
by the fish in the 0 to 4 m volition cage.However,
the depth distribution of the fish in the 0 to 4 m
cage could not be dletermined by either electronic
means or by direct observation.The depth distri-
bution can be estimated by comparing the cumula-
tive mortalities and the percent of survivors with
signs of GBD in this volition cage to the mortali-
ties and survivors in the fixed-depth and intermit-
tent-exposure cages.The comparisons of mortali-
ties strongly suggest that the fish in the 0 to 4 m
cage remained somewhat below a depth of 1 m
most of the time during all three tests.In the third
test,both the mortalities and the percent of sur-
vivors with GBD in the 1 to 2 m cage were greater
than those in the 0 t.o 4 m cage.Both mortalities
and the percentages of survivors with signs of GBD
were greater in all of the intermittent-exposure
cages than in the 0 to 4 m cage during Tests II and
III.These data indicate that the fish in the 0 to 4 m
volition cage spent something less than 8 con-
tinuous hr/day above a depth of 1 m.
Gas Bubble Disease Lesions
An attempt was made to correlate the obvious
external GBD lesions (symptoms,signs)with the
severity of the disease in each of the three tests.
The appearance of bubbles along the lateral line
was not used due to the difficulty of detecting this
lesion in the field.In most fish the gas bubbles
almost always appeared first between the rays of
the caudal fin (Fig.8).Fish with bubbles in this fin
only were considered to have slight GBD lesions.
Bubbles appeared next most frequently in the dor-
sal fin,followed by the anal (Fig.9)and paired fins.
Bubbles were also,in several instances,observed
in the adipose fin.Fish with bubbles in several fins
and/or a few bubbles on the head,or with a mild
case of exophthalmia were considered to have
moderate GBD lesions.Bubbles on the head
(Fig.10)generally occurred only after several of the
fins were involved.In the area of the head,bubbles
appeared on the opercles,jaws,around the eyes
and inside the mouth.In very severe cases,the
head appeared to be a foamy mass due to the bub-
bles covering all surfaces (Fig.11).
Exophthalmia has often been considered to be
a classic indication of GBD.In these live cage tests,
34 Weitkamp
FIG.8 Bubbles in the aud.J1 fin of a ju¥enile chinook.This is
USWlJly the first indication of ps bubble diseOlSe.
FIG.9 Numerous bubbles in the anal fin of a ju¥enile chinook.
AG.10 The head of a juvenile chinook showing numerous
bubbles .il'Ound the eyes.mouth.and the opercIe.
-
-
-
FtG.11 Severe signs of gas bubble disease with ellophth~lmia
of the Ieh ere,accompanied by hemorrhaging and bubbles
uound the eye.
exophthalmia was present in about 20%of the fish
with GBD lesions.Meekin (1974)found exoph-
thalmia in about 5%of the juvenile chinook suc-
cumbing to GBD in a similar study.It occurred both
as the only sign of GBD,and with other signs of
GBDsuch as hemorrhages and bubbles (Fig.12).
Hemorrhages occurred most frequently at the base
of the paired fins,but also occurred around the
eyes,in the jaws,gills and opercles,and occasion-
ally at the base of the anal and dorsal fins.Hemor-
rhage at the base of tlhe caudal fin was not ob-
served.Those fish possessing hemorrhages,exoph-
thalmia and/or extensHve bubbles on the head
were considered to have severe GBD lesions.
Although many of the mortalities displayed
severe GBD lesions,"it was not uncommon for fish
with only slight or moderate indications of GBD to
"die.It appears that the severity of the external
lesions is not a good indicator of the length of sur-
vival or of probable mortality.Some fish with
extremely severe external signs of GBD survived
the 2O-day test and recovered.
Recovery from GBD
The apparent ability of fish exhibiting GBD
lesions to recover has been reported in several
studies (Gorham,1901:;Rucker and Hodgeboom,
1953;Rukavina and Varenika,1956,and Ebel
et aI.,1971).Pauley and Nakatani (1967),however,
questioned the apparent ability of fish to recover
stating that recovery was probably related to the
level of supersaturation,duration of exposure,and
water temperatures,as well as the size and species.
The destructive tissue changes that Pauley and
Nakatani described as part of the early stages of
GBD are inconsistent with recovery of the fish.
Due to the significance of recovery of fish with
GBD,the 3 to 401 fixed-depth cage was converted
to a recovery cage in Tests"and 1/1.The fish placed
FIG.12 Exophthalmia,gas bubbles,and hemorrh~ging around
the eye of a juvenile chinook salmon with severe signs of gas
bubble disease.
in this cage during each test possessed a variety of
GBD lesions ranging from a few bubbles in one fin
to extensive bubbles in most fins and the head,as
well as hemorrhages and exophthalmia.Most of
the fish placed in the recovery cage displayed at
least moderate GBD lesions (bubbles present in
several fins).
The recovery of fish with evidence of GBD is
also indicated by the survivors of the 0 to 4 m voli-
tion cage in Tests 1/and III.At the end of Test II,
follOWing 10 days of supersaturation between 123%
and 126%,GBD lesions were present in 33%of the
survivors.During Test HI saturations ranged from
125%to 128%during the first 10 days and then
dropped to between 120%and 125%.Only 8%of the
jurvivors of the 0 to 4 m cage showed evidence of
GBD at the end of Test 111.If saturations of 123%
to 126%for 10 days produced GBD lesions in 33%of
the 0 to 4 m cage fish during Test II,it is likely that
a similar portion of the fish were affected during the
first 10 days of Test 1\1 when saturations were even
higher.If a third of the fish were affected during
the first half of Test III,then most of them subse-
quently recovered and lost all external evidence of
GBD during the last 10 days of this test when satura-
tions did not exceed 125%.
A few mortalities did occur in the recovery cage
during both Test II and III (lOOk,and 13%,respec-
tively).The dead fish for the most part did not
appear to have died as a direct result of GBD as did
the majority of mortalities in the other cages.Most
of the mortalities in the recovery cage developed
fungal infections of the caudal fin (Fig.13)which
were apparently secondary infections due to gas
bubble damage.The remaining mortalities in the
recovery cage either showed hemorrhage in the
Live Cage Bioassays at Rock Island 35
-
Re.1)Eroded caudal f..,and funpl growth typical of fish that
did not recoftl'from ps bubble disease.
area of the gills and eyes or no signs of GBD.Some
of the mortalities still shc)wed bubbles in the roof of
the mouth although all other external indications
of GBo had disappeared"
The loss of only about 10%of the fish with GBD
in the recovery cage indicates most of the fish
showing GBD lesions are able to survive and re-
cover within a 2O-<iay period.This recovery appears
to be complete in most cases.Only 9%and 5%of
the fish in the recovery cage of Tests II and III,
respectively,showed any indication of GBo after
20 days.This experiment indicates recovery of the
majority of fish with GBD lesions when they are
subjected to increased water pressure.It is not
known if the same effect 'Would be derived by plac-
ing the sick fish in saturated water 'at the same
depth or pressure as they occupied when they
acquired the disease.No attempt was made to
correlate survival with the severity of the GBD
lesions at the beginning of these tests.The presence
of hemorrhages in some of the survivors from the
recovery cage is an indication that some of the fish
that began the test with severe signs of GBo had
lost some of the signs of GBD and recovered and
survived for 20 days.
Previous Uve Cage Studies
The results of this live cage study are quite
similar to the results of Meekin and Turner (1974).
Meekin and Turner tested juvenile chinook in river
36 Weitkamp
water with nitrogen supersaturation at 124%and
an oxygen supersaturation of 117%(about 120%
total dissolved gas).In their study most fish held
within 0.61 m (2 ft)at the surface died with GBD
lesions within 7 days.Juvenile chinook held at
1.52 to 213 m (5 to 7 ft)for 14 days had few mor-
alities (4 to 16%)and the fish held at 2.44 to 3.05 m
(8 to 10 ft)had no mortalities or survivors with
GBO lesions.
Ebel's Priest Rapids (1967)and Ice Harbor (1971)
live cage studies indicated juvenile chinook must
remain below 2.5 m to be free of GBO symptoms.
The difference between Ebel's results and the re-
sults of this study appear to be due to the higher
levels of supersaturation experienced in Ebel's
studies.In Ebel's studies nitrogen supersaturation
reached 127%to 143%during the various tests.
These higher nitrogen supersaturations would
provide total dissolved gas saturations of about
125%or higher.The higher supersaturations in
Ebel's tests would account for the greater mor-
talities,according to the more severe effects of
supersaturations of 125%and higher indicated by
our 1974 study.
REFERENCES
Ebel,W.J.1969.Supersaturation of nitrogen in the Columbia
River and its effect on salmon and steelhead trout.Fish.Bu//.,
68:1-11.
Ebel,W.].19n.Dissolved Nitrogen Concentrations in the
Columbia and Snake Rivers in 1970 and Their Effect on
Chinook Salmon and Steelhead Trout.NOAA Tech.Report
SSRF~,7 pp.
Egusa.S.1959.The gas disease of fish due to excess of
nitrogen.Hiroshima U.,Jour.Fae.Fish.An.Husb.,2:157-182.
leman,B.1971.A Discussion of Nitrogen-Supersaturation
of Columbia River Water in Relationship 10 Basic Phenomenon
and the Physical laws of Gases.Presented at the Association
of Power Biologists Conference.sept.22,1971,Wenatchee,
Washington,Unpublished.
Meekin.T.K.and 8.K.Turner.1974.Tolerance of Salmonid
Eggs,Juveniles,and Squawfish to Supersaturated Nitrogen.
Wash.Dept.fish .•Tech.Rpt.12,pp.78-126.
Pauley,G_8.and R.E.Nakatani.1967.Histopathology of
"gas bubble"disease in salmon fingerlings.Jour.Fish.Res.
Bd.~Can.,24:867-871.
Rucker.R. R.and K.Hodgeboom.1953.Observations on gas
bubble disease of fish.Prog.Fish.Cult.,15:24-26.
Rukavina,J.and D.Varenika.1956.Air bubble disease of
trout of the source of the river Sosna.Acta Ichthyologica
Bosniae et Hercegovinae,fditum 1,x(7):5-12 (from Renfro,
1973).
Shirahata,S.1966.Experiments on nitrogen gas disease
with rainbow trout fry.Bull.Fresh.Fish.Res.lab.,15(2):
197-211.
'.".
'i'
·G.R.Bouck
Bouck:U.S.Environmental Protection Agency,Western Fish
Toxicology Station,Corvallis.Oregon.
37
Several years experience with the lethality of
supersaturation has led the author to the conclusion
that the typical laboratory testing program is not
sufficiently robust to meet its ultimate purpose.For
example,the laboratory testing circumstance typi-
cally uses the "worst possible"conditions that a fish
might conceivably experience,such as crowded
conditions,shallow water,continuous exposure and
.frequent disturbances.Moreover,the nutritional,
immunological,and acclimation state of the test
fish is typically unknown.All of these reasons
emphasize the need for evaluating the problem
in situ.
Attempts to determine the impact of supersatu-
ration in the Columbia and Snake Rivers have been
frustrated,if not stym!ed,by the magnitude of the
required resources and efforts needed for its accom-
plishment.Detection Iimits for supersaturation
mortality therein are frightful,because ocean mor-
tality kills upwards of 98%of the salmon;hence a
slight change in marine survival can have major
effects on the adult salmon run.Mortality detection
limits are stiJl bad in the Columbia River where a
flow of 250 x 103 efs discharges about 1.35 trillion
Ib of water per day;this would require a daily kill
of 13,500 lb of fish to achieve a 1 ppm detection
limit.Adult fish passage records at dams help reduce
the detection limit,but the accuracy of these records
has been disputed and in any event they fail to
indicate the cause of death.Thus,it would seem
that the circumstances in a small supersaturated
stream might present the required blend of natural
circumstances and appropriate size which might
overcome most of the previously listed objections.
A fortuitous fish kill and subsequent investiga-
tion in the Klamath Basin revealed that several
Alpine Oregon streams were naturally supersatu-
rated.This in turn generated further studies which
are still in progress.Therefore,this is a preliminary
report.
SlJpersatlJration
and FishE~ry Observations in
Selected Alpine Oregon Streams
ABSTRACT
Several Alpine Oregon streams were sampled and found to con-
tain excess levels of dissolved g,as tension.Effects of this natu-
rally supersaturated water on hatchery operations-and results of
sampling of natural populations of aquatic organisms are
presented.
Hyperbaric dissolved gas pressures may occur natu-
rally in streams for one or more of several reasons.
These reasons include geothermal heating of
groundwater (Egusa,1959),solar or seasonal heat-
ing of lakes and reservoirs (Harvey,1967),water
falls (Harvey and Cooper,1962),photosynthesis
(Renfro,1963)and high stream velocity (Lindroth,
1957).While these sources of supersaturation have
existed since time began and have been studied to
some extent by researchers,there has been rela-
tively little study of the biological impact of natu-
rally occu rring supersatu ration on wild aquatic
organisms.Conversely most of the supersaturation
bioassay research has been done in hatchery,Iive-
box,or laboratory circtJlmstances.Therefore,it
seemed prudent to investigate the limnology and
fishery biology of naturailly occurring supersatura-
tion,as is described in this report.
Although supersaturation may be a new water
quality parameter to many people,it is not a new
phenomenon to fish or aquatic invertebrates in
general.During eutrophic periods of our geological
history,photosynthetically produced high oxygen
supersaturations must have occurred countless
times and impacted the evolving aquatic fauna.
In this regard,it is interesting to note that Rucker's
(1974)study provides strong evidence that fishes
tolerate oxygen supersaturation much better than
they tolerate nitrogen supersaturation (given iso-
baric total dissolved gas pressures in either case).
Likewise,many trout and salmon streams are
derived completely from springs that discharge
geothermally heated,hen ,ce usually supersaturated
groundwater.Such streams present a naturally
existing opportunity for studying the impact of
supersaturation on aquatic life,and as such,provide
an opportunity for a much needed field verification
of laboratory bioassay data.
-
-
SPRING
-i
t
I
J---------------
able between the springs and in one instance ranges
from 6°C t011°C(42-51°F)'within 100 m of each~other.
There are many spring-fed rivers in this basin,
but only five are being monitored at present.These
are Spring Creek,Williamson River,Crooked Creek,
Fort Cr.eek and Wood River.Additionally,the springs
supplying water to the Klamath Hatchery are being
monitored.Each of these is discussed below,and
the data are presented in Table 1.
Klamath Hatchery Spring and Crooked Creek
Circumstances at the Klamath Hatchery have
been of concern for some time because several fish
kills have occurred previously and were attributed
to gas bubble disease.Another kill began in early
August of 1974;Eagle lake rainbow trout of fry size
were dying and emboli were evident in the afferent
arteries of the gills.Few emphysema were evident.
Dying fish were cultured for common infectious
disease agents but the results were all negative.The
possibility of involvement with chemical toxicity
was not ruled out,but its likelihood was remote.
Therefore,it was concluded that the fish were dying
of gas bubble disease from supersaturation.
The suspect water supply is drawn from the
North settling basin which is uncovered and con-
tains several springs.Total dissolved gas pressure
was about 50 mm Hg above air pressure (aP)or
about 1.076 atm.In the raceways where fish were
dying,the 6P was on Iy 35 mm Hg above air pressure
or about 1.053 atm.Possibly as many as 25%of these
fry died at these relatively low hyperbaric levels.
Remedial action was taken to degas the water
by cascading it into buckets and almost immediately
thereafter the fish kill abated.
Many springs contribute to Crooked Creek in
addition to the hyperbaric hatchery effluent.Dis-
solved gas pressures were measured about 1 mile
below the hatchery and showed a.:lP of 46 mrn Hg
or about 1.070 atm.As noted above,a slightly lower
level of supersaturation was found in the hatchery
where fish were dying of gas bubble disease.
Gas pressures were measured again at about
monthly intervals.The August and September data
indicate that total dissolved gas pressure in the
spring is dropping gradual/y,i.e.,6P=50-48-36.
Possibly this trend will prove to be a cyclic pattern
because fish kills here have occurred both earlier
and later than August.Whatever the case,these
resu Its ind icate tha t the d isso Ived gas pressu re in
this spring water can vary significantly without an
equally evident change in temperature.
Damage to fish or invertebrates in this stream
was not-readily apparent by qualitative observations.
Trout were feeding in the stream at the surface and
their light color did not indicate stress.Examination
of the benthic fauna revealed sculpins,larval
I
TOO£MJLT
OREGON
PROJECT
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Hill
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J8 Rouck
DESCRIPTION OF STUDY AREA
The upper Klamath Basin of Oregon is the
general study area (Fig.1)with surrounding peaks
ranging to about 10,000 ft above sea level.The basin
floor is situated at about 4100 above sea level.This
area is located between Crater lake and Klamath
lake at about 42°40'north latitude and 121°56'
west longitude.The terrain was formed mainly by
volcanic activity which resulted in various deposi-
tions of ash,lava,and pumice and was modified by
glaciation and errosion.Most of the watershed does
not appear to have noteworthy amounts of organic
material in the soil and th is is covered primarily by
lodgepole pine at upper elevations and ponderosa
pine at lower elevations.Altitude increases rapidly
from the valley (basin)floor to the top of steep-
sided foothills.Human habitation is very scarce
except on the valley floor where a few cattle ranches
or summer homes dot the area.The nearest settle-
ments are Chiloquin and Ft.Klamath.
Many springs can be found at the base of these
steep-sided hills.Typically the springs have a basin
area 10 m or more across,and collectively discharge
several hundred gpm of crystal clear water.These
discharges form rivers which flow swiftly across the
valley floor into Agency lake or Klamath lake.The
water is alkaline ranging from pH 7.4-8.0,but is
poorly buffered with only about 3()"40 ppm total
alkalinity (as CaC03).Dissolved oxygen in the spring
wate~is typically above 90%of the saturation value
for that altitude.Water temperatures are rather vari~
-
.~
-
-
lamprey,and abundance of invertebrates including
stonefly and mayfly nymphs,caddis fly larvae,
midge larvae,snails,and sponges.
Fort Creek
This stream originates from springs similar to
Crooked Creek.The August samplings showed dis-
solved gas levels (LW)of 36 mm Hg (1.055 atm}and
this was 27 mm Hg (1.043)in September.rhese
levels may indicate a pattern of declining gas levels.
Fort Creek has a reputation for being a good
trout stream,but neither the fish nor the inverte-
brate fauna were sampled.
Wood River
This isa stream which is both too swift and deep
to wade safely.Its total gas saturation I~vel has
been about 1.050 atm in both August and September.
The fish and invertebrates in it were not sampled.
Spring Creek
This stream arises from springs in a basin about
2 km long and perhaps 100 m wide.At its outlet,the
discharge is at most a few hund red cubic feet per
second and like Wood River,it is generally too deep
and swift to wade safely.It flows less than 1 km and
joins the Williamson River.
Supersaturation in A/pine Streams 39
-
-
-
-
-
-
The situation in this large but shallow spring
basin is somewhat similar to that of a lake and it
has a diurnal change both in temperature and in
dissolved gas pressure.During August and Septem-
ber,early morning water temperatures were about
GOC,and the saturation level was hardly above air
pressure (1.012 atm).As the sun rose,water tem-
peratures climbed rapid Iy to goC and the dissolved
gas level reached 78-80 mm Hg (1.108 atm or higher).
Shortly after the sun set,the water cooled and gas
levels returned to normal.
Much of the spring basin is rather shallow and
trout of a/l sizes can be seen.Dead fish have been
seen on the basin bottom,but fishermen are the
suspected cause because it is a popular place to fish.
Trout fry in shallow water were observed to be light
colored in the morning and darkened by late after-
noon,indicating stress.Some of these fry were
"pinheaded"indicating possible fasting.However,
human residents report that trout have been seen
spawning in this basin each fall (brook trout?)and
spring (rainbow trout?)..
The invertebrate fauna was extremely abundant
in this apparently very productive stream.Bottom
samples contained mayfly nymphs,stonefly
nymphs,caddis larvae,midge larvae,limpets,
snails,leeches,and oligocheates.Apparently,this
potentially lethal supersaturation is not having a
devastating effect in its diurnal form.
Williamson River
This stream was sampled only once in August
and it was slightly hyposaturated at that time.It is
famous for its good trout fishing and it has a prodi-
gious benthic invertebrate population.
DISCUSSION
Supersaturation in the Klamath Basin streams
begins with cold rain and melting snow percolating
into the ground thus recharging aquifers,but in so
doing,the water is geothermally warmed enough to
supersaturate most of its dissolved gases.Dissolved
oxygen levels were near saturation values which
according to Rucker (1974)may diminish the like-
lihood of gas bubble disease.Even so,the result of
supersaturation has been significant mortality from
gas bubble disease in the hatchery (at 105%).Even
higher levels of supersaturation (107%)occur con-
tinuously below the hatchery in Crooked Creek and
still higher levels ("0%)occur diurnally in Spring
Creek.The impact to the wild fish is still undeter-
mined but wild trout are present and feed during
40 Bouck
supersaturation apparently reproducing (judging
from the presence of trout fry).Also,freshwater
invertebrate populations were well represented with
desirable types being present in abundance.
It is possible that the low saturation levels
which killed fish in the hatchery are being aggra-
vated by solar heating of their bodies as well as by
heating of the water.These waters are crystal clear
and since sunshine can be very intense,sunburning
of the fish has been a significant problem in previous
years.Since it is theoretically possible to raise the
gas pressure within the fish by solar heating,sun-
shine may convert an otherwise tolerable gas pres-
sure into an internally lethal gas pressure.If so,it
might account for the fish mortality in the hatchery
raceway.
This leaves the unanswp.red question as to why
the wild fish (and invertebrates)seem to be thriving
at higher gas levels than killed their hatchery
counterparts.One possibility is that present bioassay
methods result in hypersensitivity among the test
fish.For example,it seems rather unlikely that a
wild fish could expose itself continuously for 10 days
to a given uncompensated lethal hyperbaric dis-
solved gas level.One can th ink of other possible
factors,but whatever they may be,one sees a sig-
nificantly different problem when they compare
hatchery and laboratory bioassays to instream
conditions.After these phenomena are given further
study the results may shed some much needed light
both on "backgrou nd"levels of supersaturation and
on how much supersaturation is too much in nature.
REFERENCES
Bouck,G.R.,D.W.Bridges,J.P.Clugston,P.Cufpin,R.Eisler,
D.Hansen,J.S.Hughes,H.E.Johnson,D.Narver and N.Rath-
burn.1974.A Survey of Manpower,Funding,and Biological
Research in Water Pollution Abatemel'lt Among Natural Re-
source Agencies of Canada and the United States.Unpublished
Report of the Water Quality Committee,American Fisheries
Society.
Egusa,S.1959.The gas disease of fish due to excess nitrogen.
Hiroshima Univ.j.Fac.Fish.An.Hush.2:157-1~2
Harvey,H.H.1967.Supersaturation of lake water with a pre-
caution to hatchery usage.Trans.Am.Fish.Soc.96:194-201.
Harvey,H.H.and A.C.Cooper.1962.Origin and Treatment
of a Supersaturated River Water.Inti.Pacific Salmon Fish.
Comm.Prog.Rpt.No.9,19 pp.
Lindroth,A.1957.Abiogenic gas supersaturation of a river
water.Archiv.:Fur Hydrobiologic.53:589-597.
Renfro,W.C.1963.Gas-bubble mortality of fishes in Galves-
ton Bay,Texas.Trans.Am.Fish.Soc.92:320-322.
Rucker,R. R.1974.Gas-bubble Disease of Sa/monids:Varia-
tion in Oxygen-Nitrogen Ratio with Constant Total Gas Pres-
sure.National Marine Fish.Ser.Project Completion Report.
12 pp.(Mimeo).
IDAHO
<---J....-lo12
UTILE GOOSE"
ICE HARBOR ~
SNAKE RIVER
~
WASH I NGTON
Bentley,Dawley,and Newcomb:National Marine Fisheries
Service,Seattle,Washington.
fiG.1 Vicinity map showing Iociltion of Lower Monumental
and Little Goose Dams.
prey influences their selectivity of daily food intake
(Thompson,1959).Thompson found that approxi-
mately 63%had empty stomachs and only 7.5%
showed any evidence of eating juvenile salmon.
Hamilton et al.(1970),in the Lake Merwin investi-
gation,found that 70%of the stomachs examined
were empty,but concluded at the end of his study,
that predation precluded the use of the Lake Merwin
Reservoir as a rearing area for coho salmon.Brett
and McConnell (1950)used an estimated consump-
tion rate of 140 salmon fingerlings per squawfish
per year,a figure which was accepted as reasonable,
to account for calculated losses of sockeye juveniles
from lakelse lake,British Columbia.If we use these
figures for the Snake River during a 45-day juvenile
outmigration,each adult squawfish might consume
I.W.W.Bentley
E.M.Dawley
T.W.Newcomb
ABSTRACT
In the spring of 1974,large numbers of squawfish were en-
countered in the Snake River between Lower Monumental and
Little Goose Dams ..Squawfish exh ibited gas bubble disease
symptoms within 1 week after the onset of 125 to 135%satu-
ration.A 12·day bioassay in shallow tanks to determine tolerance
levels and resistance times at various gas concentrations was
conducted.We found squawfish to be similar to juvenile salmon
and steelhead trout in their resistance to supersaturated con-
centrations of dissolved gas.Feeding response changed after
stress to high concentrations of dissolved gas.Average daily
food consumption of test groups decreased with increased
supersaturation.Squawfish captured in the field during periods
of high supersaturation were less abundant and only a small por-
tion of them had been feeding compared with survey results
taken during lower supersaturation.Nitrogen supersaturation
could be an important factor in assessing the effects of predation
on juvenile salmonid migrants in the Columbia River system.
Some Effects of
Excess Dissolved
Gas on Squawfish)Ptychocheilus
oregonensis (Richardson)
,~
,
Squawfish have attracted the interest of many
investigators in the past,primarily because they
have been regarded as an efficient predator.Our
attention was focused on them as being one of the
possible causes of mortality of seaward migrating
juvenile salmon and steelhead trout in the Snake
River.
In the spring of 1974 large numbers of squaw-
fish were encountered in the Snake River between
Lower Monumental and Little Goose Dams (Fig.1).
At this time,an abnormally high runoff occurred,
resulting in high nitrogen gas concentrations of
prolonged duration (Table 1).The squawfish ex-
hibited gas bubble disease symptoms within 1 week
after the onset of high gas saturation.laboratory
experiments and field observation were conducted
to determine the tolerance of squawfish to super-
saturation of dissolved gas and how it affected their
food intake or predation rate.We wish to report on
;here'aspects and discuss problems that may require
....../further clarification.
/Predation and eating habits have been examined
by several investigators.Squawfish in the Columbia
River were determined to be opportunists in their
eating habits and,by and large,the availability of
41
"'-1
moving upstream to potential spawning areas of
rip-rap dikes adjacent to the dam.I n addition to
learning something of their movements,we wished
to gain some knowledge regarding the numbers of
squawfish in that section of the river.Also,purse
seining at the dam during and after spill might tell
us whether there would be any change in behavior
or response to high nitrogen levels.In this area,
seining began on April 24 and concluded on August 8.
Since we wished to tag and recover the fish in this
area of the dam,we did not sacrifice the fish for
stomach analysis but examined the stomachs by
firmly pressing the lower ventral area and working
forward to the pectoral area.
Water samples were taken biweekly·by air-
craft;dissolved gas values were determined in our
Seattle laboratory by techniques described by Ebel
(1969)and Beiningen and Ebel (1970).
Six laboratory tests were conducted where dis-
solved gas concentrations averaged 126.1,120.4,·
117.2 and 99.8%of saturation of total dissolved gas
(T.D.G.).Average variation from the desired test
concentration was ±1.1%of T.D.G.Test duration
was 12 days.Simultaneous replicates were made of
tests at 117,110 and 100%saturation.
All test tanks were 12 m in diameter with
water depths of 25 em (hydrostatic compensation
0.025 atm of pressure,or about 2.5%of saturation
decrease).Water flow was maintained at 7.5 IZ/min
at 10°±1°C.Tests were conducted with about
10 fish per tank over 12 days starting April 17,1974.
Mean size of the test fish at introduction was 364 mm
and 534 g.Test fish were starved for 16 days prior
to testing.A sample population was fed to deter-
mine the maximum weight of food an unstressed
fish might consume in a 2-week period.A mixed
diet of live steel head (average size -21 g,80 mm)
and dead smelt (average size -30 g,170 mm)was
used.On the basis of food intake of the sample
TABLE 1 Temperature,Oisso/Yed Gases.Spill and Toyl Wate!'Flow (in Thousands of c.f.s.)at ~
Lower Monumental and Uttle Goose DiUIIS from April to August.1974.DaY obtained by biweekly
airplane ffights and analyzed by staff in SeaRle.
Little Goose Dam forebay Lower Monument~1 Dam forebay
Date Temp°C(F)%O~%N z ToO.Co SpiU ToL flow'Temp°C(F)%0'2 %N~T.O.C.Spill Tot.flow
4/19 8.3(46.9)99.6 101.4 101.0 86.0 131.0 8.5(47.3)117.8 122.1 120.9 83.0 150.0
4/23 9.6(49.1)102.5 106.8 105.8 61.0 130.0 9.9(49.8)130.8 U4.6 133.3 59.0 126.0
5/7 11.6(52.9)105.9 110.3 109.2 96.0 162.0 12.1(53.8)130.8 135.8 134.2 95.0 164.0
5/21 11.4(52.5):98.5 104.1 102.8 17.0 83.0 11.3(52.3)114.2 121.5 119.7 15.0 80.0
6/4 12.2(54.0)99.2 102.9 102.0 92.0 161.0 11.9(53.4)127.9 141.1 137.7 90.0 159.0
6/19 12.8(55.0)102.4 111.5 109.4 228.0 300.0 12.5(54.5)128.7 143.8 140.0 220.0 294.0
7/2 16.9(62.4)99.5 106.8 105.1 0.0 126.0 16.7(62.1)124.6 131.8 129.8 66.0 133.0
7/16 17.4(63.3)103.4 106.7 105.9 19.0 63.0 19.0(66.2)102.4 106.4 105.5 3.0 68.0
7/30 24.5(76.1)117.7 110.3 111.5 0.0 47.0 23.2(73.6)110.3 106.7 107.2 0.0 44.0
8/13 21.7{71.1)99.4 104.9 103.7 0.0 44.0 21.7(71.1)101.8 105.8 104.9 0.0 43.0
17.4 salmonids causing a loss of 5.2 million finger-
lings if a population of 300,000 squawfish exists for
the stretch of the river between Ice Harbor to little
Goose Dam.
The effects of dissolved gas on squawfish had
been examined by Meekin and Turner (1974)and
Blahm (1974).Blahm's results indicate that squaw-
fish were more resistant than juvenile salmonids
when stressed with supersaturation.Meekin and
Turner indicated slig'ltly less tolerance than sal-
monids and that predation ability was substantially
reduced when they were in supersaturated ~ondi
tions.We attempted to place specific values On the
dissolved gas tolerance of the squawfish,to substan-
tiate and enumerate changes in the predation rate,
and to discern if any correlations exist between
laboratory experiments and field observations.
TECHNIQUES AND MATERIALS
Squawfish for our laboratory experiment in
Seattle were captured in a Lake Merwin trap in-
stalled in the Palouse River arm at Lyons Ferry,
Washington.This unit was identical to those
described by Hamilton et al.(1970)in the lake
Merwin study in the lewis River.Purse seining at
little Goose Dam was accomplished with a seine
15-ft deep and 525-ft long operated from a power-
driven barge similar to that described by Durkin
and Park (1969).This shallow net was employed
because of the depth limitations near the navigation
locks and spill gates where we concentrated much
of our effort.
All squawfish were tagged with a Floy anchor
tag,FD 67@,and released.Besides the tag,some
were branded using the liquid nitrogen technique
,-described by Mighell (1969);none were fin-clipped
or operculum-perforated.
We purse seined at little Goose Dam tailrace
,-under the assumption that squawfish would be
42 Bentley,Dawley,Newcomb
-
-
-
Juvenile
F.chinook
42mm
0.4 g
10°C
7 day 22 day
57 day 106 day
20 day 47 day
Effects on Squawfish 43
Juvenile
S.chinook
120 mm
15 8
15°C
LE"LE~
19 hr 27 hr
11 hr 14 hr
>35 day >35 day
100
FIG.2 Gas bubble disease symptoms in northern squawfish
with increasing T.D.G.saturation.
VI
~00
Ii:
:E>-
VI
E 60a::
:I:
Vl
u..
u..
o 40
~.....
~
It
100
2J
120%saturation test and many survIVing the 117%
T.D.G.tests.Gill emboli were not detected in squaw-
fish exposed to lower saturation.
Active feeding decreased from an average of
14.3 g food per fish per day for squawfish exposed
to 100%saturation down to 2.3 g food per fish per
day for squawfish exposed to 120%T.D.G.satura-
tion (Fig.3).These data indicate that feeding would
be reduced by 50%at about 115%saturation.Below
test saturations of 120%,squawfish showed a pref-
erence for live steelhead over the dead smelt (Fig.4).
The number of test days the ration of live steelhead
was consumed was reduced about 40%when test
fish were exposed to 117%T.D.G.saturation.Squaw-
Juvenile
steelhead
135mm
208
15°C
tEll
26 hr 33 hr
10 hr 13 hr
>35 day >35 day
LE II LE~
Squawfish
364mm
534 8·
10°C
4.8 day >12 day
41 hr 9.7 day
19 hr 20 hr
>12 day >12 day
TABLE 2 A Comparison of Lethal Exposure Times (or Squawfish and Potential Salmonid Prey
with 2.5%Hydrostatic Compensation
110
115
117
120
126
Percent
saturation
(T.D.G.)
population we established a daily ration for test
-groups of four dead smelt and one live steelhead
fingerling.One-half of the smelt introduced as
food were cut in half to accommodate the smaller
_squawfish.
RESULTS AND DISCUSSION
_Laboratory Bioassay
Substantial squawfish mortality occurred in
tests at 126,120 and 117%saturation.One hundred
percent mortality occurred in 20 hours at the 126%
I""'"level;60%loss occurred within the 12-day test period
at the 120%level;and 32%at 117%saturation.
Mortality from gas bubble disease did not occur at
,_110,107 or 100%T.D.G.saturation.Lethal exposure
times for 10 and 50%mortality (LE 10 and-'lE so )for
squawfish are compared in Table 2 with LE,o and
LE 50 values established (Dawley and Ebel,1974 and
-Dawley et aI.,1975)for potential salmonid prey.It
is evident that squawfish are somewhat more toler-
ant than juvenile steelhead and spring chinook,
"""but have much less resistance than fall chinook fry.
Gas bubble disease signs were found in all fish
exposed to 126,120 and 117%T.D.G.saturations.
Eighty-nine percent of the squawfish exposed to
110%saturation also had gross signs of gas bubble
disease;one of ten fish exposed to 107%T.D.G.satu-
ration exhibited signs.No signs were noted at 100%
""'"T.D.G.saturation (Fig.2).Gross gas bubble disease
signs included hemorrhage and subcutaneous
blisters present over large areas of their bodies.All
fish showing signs of gas bubble disease exhibited
grossly distended blisters between the fin rays.
Exophthalmia ("pop-eye")occurred in only two
experimental fish.Gaseous emboli were noted in
the blood vessels of at least one gill arch in all gas
bubble disease mortalities.Emboli were also ob-
served in the gills of all squawfish surviving the
-
-
1,.0
-
>=
C3
I 10.0
V>
~
E
S'
z
0
~
:E t!A::::l
V>\:2
0 5.0u
0
00"-
.::.
OL-__l..--__l-__.L-__--'-_-----l
100 110 120
PERCENT SATURATl0!'iJIT.O.G.I
FIG.J Feeding inhibition of northern sqUilwllSh on a weight
of food basis.
in the reservoir of lower Monumental Dam on the
Snake River the dissolved gas level remained near
140%for an extended period of time this year and if
the predatory squawfish had maintained an average
daily depth of 276 m (9 ft),the effective saturation
would have been 110%-low enough to allow the
squawfish safe sojourn with no effective curtailing
of predatory capacities.during experiments done
earlier at the Prescott Field Station (Blahm,1974),
as well as those reported here,the squawfish
tended to reside on the bottom of the tanks.How-
ever,this behavior may be an artifact due to the
unnatural conditions of the laboratory test tanks.
Nevertheless,the behavior of remaining on the
bottom of the tanks changed the effective satura-
tion value of the Prescott 1 m depth test from 119.7
to 109.1%T.G.P.and the 25 m test from 119.8 to
95.8%T.G.P.saturation.As a result,no mortality
was reported in tests at Prescott,but gas bubble
disease signs were apparent on all fish from the
1 m test after 35 days.
Seattle and Prescott data suggest that knowl-
edge of the depth distribution is needed on the
squawfish before its role as a juvenile salmonid
predator can be correctly defined.
Merwin Trap a Purse Seine
Fish taken in our Merwin trap located in the
Palouse River arm from January 1 to August 12,
1974,numbered 80,060 (Table 3).Squawfish totaled
16,626,with the majority taken in April,May and
June,when dissolved gas saturation,temperatures
and water flows were increasing (Table 1).Large
numbers of squawfish may have concentrated in
the Palouse River arm to escape the high dissolved
gases in the Snake River.The Palouse River arm
was sampled on July 2,1974,and showed 106.8%
nitrogen saturation at 21.7°C (71.1 Q F).Recaptures
of marked fish released in the vicinity of the trap
were 1,590,indicating that a high percentage
remained in the area.Purse seining the navigation
locks and spill area at Uttle Goose Dam captured
2,101 squawfish which were tagged and released.
Fifty-five of those marked at the purse seine were
subsequently recaptured in the seine at Little Goose
Dam;and 97 that had been marked at the M~rwin
trap,over 11 miles downriver,were also taken in
the seine at Little Goose Dam.Thirteen that were
tagged at the dam appeared in the trap.One 370 mm
squawfish marked at the trap made a round trip to
the dam and back to the trap.Approximately 300 of
the 1,590 recaptures at the trap were multiple
recaptures.One 359 mm squawfish appeared in the
trap 10 times.
Tagged recoveries showing movements between
lyons Ferry and Little Goose Dam indicate that high
nitrogen values in surface waters appear to be
OL..-.l..-__..l..-__..J-__....L._-...J
100 110 lZl
PERCOO SATURATION H.O.G.\
100 ~-__
fish held at 120%saturation ate live steelhead only
on the first test day.
Lethargy exhibited by the highly stressed fish
may be of significance in regard to squawfish preda-
tion on juvenile salmonids migrating down the
Columbia and Snake Rivers.However,the average
water depth inhabited by squaw fish may compen-
sate for the effects of supersaturation.For example,
44 Bentley.Dawley,Newcomb
AGo 4 SeIedWe feeding inhibition of .-them sq~wfish-%oi
test cL1ys when ration is compietely consumed.
,-
-
VI aJ>-
~
l-
F"'"
V>•I=!60 .::."-0
I-Z.....
Ua:
~«l
TABLE 3 Summary of Merwin Trap Catch at Lyons Ferry in the Snake River,
~T~January to August,1914 .
'an.30 Apr.5 May 29 'une 21 July 30
""'"Species to to to to to Totals
Apr.4 May 28 'une 26 July 24 Aug.12
Chinook,Oncorhynchus lschawytscha 1 1 4 0 0 6
~Coho.Oncorhynchus kisUlch 0 0 0 0 0 0
Sockeye,Oncorhynchus nerka 0 1 1 0 0 2
Steelhead,Salmo gairdneri 21 14 10 0 1 46
Shad,A/osa sapidissima 0 0 0 0 0 0
~Squawfish,Ptychocheilus oregonensis 1475 6685 7944 240 282 16626
Whitefish,Prosopium wil/iamsoni 6 16 0 0 0 22
Yellow bullhead,Ictalurus natalis 692 488 320 380 92 1972
Sucker,Catostomus macrochei!us 340 5288 14343 2617 4192 26780
Crappie,Pomoxis annu/aris 6273 11655 3764 3136 1347 26175
Yellow perch,Perca flavescens 125 123 21 138 20 427
Sunfish,Lepomis sp.5 6 5 25 15 56
Shiner,Notropis sp.51 58 73 15 6 203
Madtom,Noturus gyrinus 11 1 0 8 0 20
Bluegill,Lepomis sp.18 6 42 17 9 92
Chiselmouth,Acrocheilus a/ulaceus 195 950 1211 64 182 2602
Carp,Cyprinus sp.209 201 816 293 272 1791
Channel catfish,Ictalurus punctatus 59 504 884 1212 462 3121
Dolly varden,Sa/velinus malrna 4 '1 1 0 0 6
Chub,Hybopsis sp.20 0 1 0 0 21
Bass l.M.,Microplerus salmon ides 1 0 0 0 0 1
Bass S.M.,Microplerus do/omieui 0 4 5 0 2 11
Lamprey,Entosphenus lriden/atus 1 0 0 0 0 1
Sturgeon,Acipenser sp.0 2 0 2 0 4
Peamouth,My/ocheUus caurinus 0 1 48 11 8 68
Sucker,Unident.,Caloslomus sp.0 5 7 1 0 13
Total fish 80060
seine set were '11.7,indicating that there were few
available.When dissolved gas saturation returned
FIG.5 Bimodal length frequency of squawfish taken by purse
seine in the tailrace at Little Goose Dam from 'uly 31st to
August 8,1914.
-
-
avoided during transience,but whether the squaw-
fish were able to detect and avoid this area is
unknown.The depth they normally inhabit may be
sufficient to adequately compensate for the levels
of supersaturation which occurred.
Length frequencies of those taken in the trap
were different from those captured in the purse
seine.In Fig.5 a bimodal frequency is shown for
fish sampled at little Goose Dam in July and August
with the lesser mode which is similar to the length
frequency of fish sampled at the trap (Fig.6).In the
upper mode a group of squawfish,between 300 and
460 mm,had not been encountered before,suggest-
ing that they may represent an entirely different
population.One population may be reservoir
spawners and the other tributary spawners.We
have had tags turned in from.the Palouse and
Tucannon Rivers,the mouth of the Snake River and
the Columbia River at Kennewick,Washington.
Purse Seine -Feeding
Purse seine catch at Little Goose Dam is shown
in Table 4.When dissolved gas saturation was high,
176 squawfish were taken;five showed evidence of
feeding.The numbers of squawfish taken per purse
90
ro
70
::t:60
'"u:
u..500
~....
<10a:>
:E
::::l
Z
30
20
10 ~~rHlf
200 240 200 320 360
LENGTH (mm)
N =519
X=266.6 mm
N =973
X=345.8 mm
400
Effects on Squawfish 45
TABLE 4 Pune Seine Results from Ciltches in the Tililrilce
of UttIe Goose Dilm,1974
FIG.6 Length frequency of sqiRwfish taken in Merwin lrilp i1t
Lyons Ferry in June,1974.
:,••;!(.••.,
-;":-
j
.iff
ration returned to normal they moved in to feed on
whatever was available,which was mainly lamprey
ammocetes.On the other hand,it is possible that
large numbers of squawfish may have been present,
but below the depth of our net where they would
also be at sufficient depth to compensate for gas
supersaturation and thus be safe from the disease.
We donot know if they are at this depth.Thus,what
appeared to be a response to high nitrogen levels
could be the result of normal behavior patterns in
the vicinity of spill gates and turbine discharge
draft tubes at dams.It is evident,however,that
those squawfish taken during high dissolved gas
saturation were not effective predators.
REFERENCES
Beiningen,K.T.,and W.J.Ebel.1970.Effect of John Day
Dam on dissolved nitrogen concentrations and salmon in the
Columbia River,1968.Trans.Am.Fish.Soc.99:664-6n.'
Blahm,T.1974.Squawfish Bioas.say.Bonneville Power Ad-
ministration Quarterly Report,National Macine Fisheries
Service,Prescott Field Station,unprocessed.
Brett,J.R.and J.A.McConnell 1950.Lakelse Lake sockeye
survival,Jour.Fish.Res.Bd.,Can.8(2):83-110.
Dawley,E.M.and W.J.Ebel.1973.Lethal and Sublethal
Effects of Various Levels of Nitrogen and Argon Supersatura-
tiOfl Ofl Juvenile Chinook Salmon and Steelhead Trout.National
Marine Fisheries Service Bulletin.In press.
Dawley.E.M.,M.Schiewe.B.Monk and F.Ossiander.1975.
Effect of Long-term Exposure to Supersaturation of Dissolved
A.tmospheric Gases on Juvenile Chinook Salmon and St.eelhead
Trout in Deep and Shallow Tanks.National Marine Fisheries
Service Bulletin,Seattle,Washington.(Ms.in preparation).
Ebel,W.J.1969.Supersaturation of nitrogen in the Columbia
River and irs effect on salmon and steelhead trout.Fish.Bull.
68:1-11.
Hamilton,J.A.R••L.O.Rothfus,N.W.Erho,and J.D.Remington.
1970 Use of Hydroelectric Reservoir for the Rearing of Coho
Salmon (Oncorhynchus kisutch).Wash.Dept.Fish .•Research
Bull.No.9,6S pp.
Meekin,T.k.and B.k.Turner.1974.Tolerance of Sa/monid
Eggs,Juveniles and Squawfish to Supersaturated Nitrogen.
Wash.Dept.Fish.,Tech.RpL No.12,pp.78-126 (9 figures,
15 tables,9 references).
MigheU,J.l.1969.Rapid cold-branding of salmon and trout
with liquid nitrogen.Jour.Fish.Res.Bd.,Can.26:2765-2769.
Thompson,R.B.1959.Food fo the squaw fish.Pr.ychocheilus
oregonensis (Richardson)of the lower Columbia River.Fish.
Bull.60(159):43-58.
CONCLUSION
Laboratory studies indicated that adult squaw-
fish are susceptible to supersaturation of atmospheric
gas at or exceeding 117%and exposure to these
levels significantly reduced their food intake.
Field studies indicated that exposure of squaw-
fish to supersaturation could be an important factor
in assessing the effects of predation on juvenile
salmonid migrants in the Columbia River but more
information is needed on their movement,behavior
and depth distribution before an accurate assess-
ment can be made.
260 300 340
LENGTH (mm)
220
I
1111
Sets ~mple size Food
2 3 none
4 26 1 unident.
1 22 none
2 12 none
3 17 2 unident.
2 18 none
1 78 2 unident.
15 176 5
llJ per set
2 26 lamprey
3 398 lamprey
4 125 lamprey
1 542 lamprey and
1 699 unident.
1 145 fish
12 1935
161.2 per set
N =897
X:264.1 mm
to normal,catches increased to 1,935 in 5 days of
seining with an increase to 161.2 fish per set.All
fish checked in the latter sample showed evidence
of feeding heavily on lamprey ammocetes
Entosphenus tridentatus,(Gairdner).
We might interpret the presence or absence of
squawfish in the tailrace at Little Goose Dam as
being influenced by high dissolved gases.Satura-
tion in the spill and adjacent areas around the dam
tended to keep squawfish away;and after gas satu-
46 Bentley,Dawley,Newcomb
July 17
IfI!"ltiIi,18
31
August 2
6
8
Dille
-April 24
25
May 3
16
30
31
July 12
~
160 c-
140 ~
:I:120 f-
Vl.......
......100 -
0
~00 -UJ
to
:2:
::l
Z 60 c-
40 ~
20 ~
0
I"'"
MATERIALS AND METHODS
Pressure Chamber
A 4-Q stainless steel pressure chamber,capable
of withstanding 10 atm test levels,was used for
our tests (Fig.1).The chamber has viewing end-
plates,sampling ports,and attached pressure
Beyer,Smith:University of Washington,and D'Aoust:Virginia
Mason Research Center,Seattle,Washington.
Contribution Number 409,College of Fisheries,Seattle,Washing-
ton.
°In these studies,the terms depth,atmosphere (atm),and pres-
sure are interrelated as follows:1 atm =34 It of fresh water
=14.7 Ib/in 2•
In our studies,we have been investigating the
responses of salmon ids to acute supersaturations
(greater than 150%saturation)to determine the
relationships among supersaturation,gas uptake,
and death due to bubble formation.
To study these relationships,we have exposed
fish to three separate types of supersaturation
conditions (internal,external,and a combination
of both).The direction of net gas movement (air in
these tests)in and out of the fish and the initial
site of supersaturation imposed on the fish were
varied in each condition.Internal supersaturations
were produced by decompression from saturation
where a net outward movement of gas occurred as
the fish was desaturated.In the external tests,fish
were placed in supersaturated water,and thus the
net movement of gas was inward.When the inter-
nal and external conditions were combined,the
supersaturation occurred both inside and outside
the fish and there was initially no significant net
.movement of gas except that allowed by bubble
formation in the pressure chamber.Our pre-
liminary studies have centered on the differences
in the occurrence of bubble formation among these
three types of supersaturations ("treatments").
10.Beyer
B.G.D'Aoust
L.Smith
-Responses of
Coho Salmon
~
(Oncorhynchus kisutch)to
~Supersaturation at One Atmosphere
Since Marsh and Gorham (1905)implicated excess
dissolved gas as the causative agent of gas bubble
disease in fish,most studies of the problem have
centered around:1)defining the critical levels at
which problems due to supersaturation arise,and
2)describing occurrences of and solutions to
specific outbreaks of the disease.It seems,how-
ever,that some very interesting and important
studies may have been overlooked.These other
studies involve the actual dynamics of gas bubble
formation within fish tissues and the relationship
of bubbles to gas uptake and elimination rates.One
only needs to look at the example of fish dying due
to supersaturations of around 110 to 120%total gas
pressure.Such saturations in human divers are
readily tolerated and according to current practice
(U.S.Navy,1973),staged decompression from satu-
rations of less than 200%of 1 atm (dives of less than
30 ft)*is not needed.Therefore,the sensitivity of
fish to gas bubble disease appears to be somewhat
anomalous to the response of air breathing animals
(including man)to gas supersaturation imposed
by decompression,primarily because the fish are
affected at such low levels.
ABSTRACT
While saturation limits of 110-120%have been established as
minimum lethal levels,mortalities may not be entirely related
"""to gas saturation or desaturation rates.To compare maximum
saturation times and.capacities for inert gas with bubble forma-
tion,groups of small coho salmon (60·100 mm)were exposed
to supersaturations that were induced either internally (by
.....decompression)or externally (by placing fish in supersaturated
water).A minimum of 1 hr at depth was required to obtain maxi-
mum lethality from decompression.Assuming maximum lethality
was associated with maximum gas absorbed for anyone decom-
--pression,it was concluded that for any given gas pressure,satu-
ration was completed within 60 to 90 min.Thus,the well-
documented lethal times of 24 hr or more for an over-saturation
of 122%of 1 atm(Meekin and Turner,1974)indicate a time lag
.....in achieving maximum effect which cannot be related to gas satu-
ration or desaturation rates.The external supersaturations
showed that 2.50 to 500%total gas pressure would result in com-
plete mortality in 10 to 30 min.
47
.-,
.....
fiG.1.four-i pressure ch~mber used to test the responses
of s.almonids to Yilrious supersaturiloon conditions.Chamber
is mounted on lilting Uble whkh facilitates handling of fish.
gauges and f10wmeters for mon itoring and regulating
the conditions to which the fish are exposed.The
chamber is mounted on a tilting table which facili-
tates the transfer of fish in and out of the chamber.
Test Evaluation
The procedure to estimate time to complete
saturation of critical tissues and organ systems
(tissues and organs that are of vital importance to
the fish,e.g.heart)has been an acute bioassay tech-
niqu~where coho salmon (60 to 100 mm)were
exposed to a certain supersaturation condition and
a dive score taken of the response.The dive score
was calculated as follows:2 points for a dead fish,
1 point for loss of equilibrium and 0 for neither of
these.Signs of distress such as increased irritability,
excitement,and rapid ventilation were not counted.
Most of the fish responded to the given stress either
within 30 min.or at some time much later (hrs).
Therefore,dive scores were recorded at 15 and 30
min after any particular exposure,and an average
of the two scores was used for data.
A gross autopsy of mortalities was conducted
to locate the bubbles within the fish tissues to deter-
mine if there were differences in the pathologies
caused by the different procedures used to impose
supersaturations in the fish.
Internal Supersaturation
To create internal saturation,the fish were
placed in the chamber and saturated at various
depths (by bubbling gases under pressure through
the dlamber at 2 \Umin).*After various lengths of
time (exposure)at depth,the fish were rapidly de-
compressed to the surface at 100 ft/min,removed
48 Beyer,D'Aoust,Smith
from the chamber,placed in water containing gas
at 1 atm,and a dive score recorded.
External Supersaturation
In this series of tests,coho (at surface satura-
tion)were placed directly tnto supersaturated water
and dive scores recorded.Initial saturation levels
ranged from 200 to 700%of one surface value,but
due to mechanical manipulation of the chamber
and oxygen consumption by the fish,these levels
decreased during the test.In these preliminary tests,
no additional gas was added to compensate for the
decrease.
Combined External and Internal
Supersaturation
During the internal supersaturation tests,the
gas in the fish was allowed to diffuse outward into
fresh water when the fish was removed from the
chamber.If this outward diffusion was eliminated
for short periods of time,more gas should remain
in the fish and the severity of bubble formation and
its effects should increase.To minimize this out-
ward movement of gases,fish were saturated at
depth (100 ft only in these tests)as in the internal
supersaturation tests,but instead of placing the fish
in water containing gases at 1 atm,the fish were
held in the chamber (in the supersaturated water)
for various lengths of time (up to 15 min)and a dive
score recorded.
RESULTS AND DISCUSSION
Internal Supersaturation
As the pressure of saturation and resulting
supersaturation after decompression were increased,
the response increased as reflected by the dive scores
(Fig.2).No signsof bubble disease were noted after
decompressions from 66 ft (300%saturation).At
100 ft no scores were recorded;however,these fish
were on the threshold of distress because any
additional stress such as an induced fright
response applied to the fish after decompression
resulted in signs of bubbles.At pressures greater
than 100 ft,dive scores increased at each level until
100%mortality was reached at decompressions
from 200 ft (700%saturation).From 60 to 90 min
exposure was required to obtain maximum
lethality from decompression from anyone depth
(e.g.,no significantly greater increase in percent
"As pressure is increa~ed,water and tissues can keep greater
amounts of gas in solution (according to C (concentration)
=P (pressure)x c (solubility),if gas is bubbled through fresh
water at 34 ft.the water will take up approximately twice
the concentration of gas found in water at the surface).
.".;
',I""!
-.
~
100%
700%
l00"'{'
400%
66%
250%
{)
200%
%Dive Score
External and Internal Supersaturation
When the supersaturation gradient between
the fish and the water was eliminated by simul-
taneously internally and externally supersaturating
the fish,·the response was increased (Fig.3).These
results indicate that the longer the fish are held in
supersaturated water after decompression,the
greater the dive score.The occurrence of bubbles in
this series of tests was similar to the internal super-
saturation with bubbles found throughout the tisssues .
Response of Coho to Supersaturation 49
COMMENTS AND SlJMMARY
From the results of these acute tests,it is clear
that bubble formation can be induced by two
Initial level of supersaturation
TABLE 1 Responses of Coho Salmon to Acute External
Supersaturations.N =20 fish (10 fish x 2 tests at each
saturation).
Bubbles were found in relation to the degree
of perfusion of a particular tissue,indicating the
importance of total gas transport.The frequency of
occurrence was much higher in the blood than in
the white muscle and fat.As the depth of exposure
was increased,there was obvious increase in fre-
quency of bubble formation in all tissues.Bubbles
in the blood were probably the most critical to the
fish because these bubbles cut off circulation to
other organs and systems,occluded the gills,and
frequently filled the chambers of the heart.
External Supersaturation
Fish were more susceptible to external than
internal supersaturation (Table 1)'with fish dying
at 250%saturation (a concentration approximately
equal to water saturated at 50 ft)in 30 min.Most
of our results with this method were preliminary,
but it appears that a slightly different mechanism
related to the direction of the supersaturation
gradient and the total volume of gas available
accounts for this increased response.Also,in con-
trast to the internal supersaturations,bubbles
were primarily found in the dorsal aorta,coronary
artery,and in the heart.Very few bubbles were
found within the organs and tissues,mainly because
the bubbles in the circulatory system were effective
in causing death long before any bubbles were able
to form in the other tissues.
200 FTl100%SAT,I
i .----.----.t i_~__,
l~.-1";FTlcro%SAT.l
T
I
I ~
T I 133FT I
I .----~::15OJ~,SA Ll 1r------r \OOFn4OO'1o SAT.I
~60 00 m
EXPOSURE Wtf (MINUTES)
dive score was observed after 60 to 90 min at 133 ft;
a stress producing a 35%dive score).Theoreti-
cally,it takes the same time period for a tissue
to reach equilibrium at 1 atm (total gas pressure)
as it does at 5 atm.This arises from the fact that
although a greater amount of gas must go into
solution in the fish's tissues,the rate increases
proportionally to the gradient and thus the total
saturation time remains the same.Therefore,
assuming maximum lethality.is associated with
maximum gas absorbed for any given gas pres-
su re,it was concluded that saturation of critical
tissues was completed within 60 to 90 min for
this size fish.Since this time of equilibration,as
indicated above,should apply to any saturation
level,the well-documented lethal times of 1 day
or more for a saturation of 122%(Meekin and
Turner,1974)indicates a time lag in achieving
maximum effect which cannot be related to gas
saturation or desaturation rates per se,but also
involves the amount of gas transfer.Inother words,
the critical tissues of fish that die from long-term
c~ronic bioassays (greater than 10 hr)are satu-
rated after only 60 to 90 min of exposure.There-
fore,other factors related to gas bubble formation,
growth,and effects within the tissues are causing
the mortalities.Other potential mechanisms are
unclear at this time but additional studies by our
group (Casillas,Smith,and D'Aoust,1975)are
investigating possibilities such as alterations in
the blood clotting mechanism and/or dissemi-
nated intravascular coagulation which has been
recently implicated in the sequelae of decompres-
sion sickness in man (DCIEM Conference,1973).
~~
...e:;'"0u
'"...
==60c....
Z...u 40'"~
20
0
0
FIG.2 Responses of coho salmon to decompressions (internal
supersaturations)from various depths.Exposure time indicates
the length of time the fish were maintained at any particular
depth before decompression.The initial saturation is indicated
in parentheses.N =15 fish (3 tests x 5 fish per test),ranges
indicated by yertical broken lines.
--
.-
".."
-
-
-
f"""""
100
o lSM1NUTES
8l I 5 MINUTESI•
~-;-I II oMINUTES
0 I
u T t ...
'"60 ,I I....:6--9 0:::I
Q I.....
'/'I
J.'
~~if/~~u
.--•.-r'"Q I .,lo.,_~
20 j '~~•
/•J.
0
0 15 30 45 60 ~1<!3
EXPOSURE Tip,<[lMlNUlESI
FIG.3 Preliminary results of intem~and exterru.11y c0m-
bined supergturatiom.Exposure time indiates the length of
time the fISh were INintained ill deptb(100 ft only in dIese
tests)before decompression.Zero,50 oand 15 min indicoate the
length of time the fISh were hekl in supergluroated wOlter nter
decompression-Most doaL1 points represent 30 fISh (10 fISh l[3
tests)oand ranges olIre indinted by verticoal broken lines.Where
no vemul lines olIppeollr,only one test WH conducted and N
=10 fish.
separate procedures·and the combination of the
two is somewhat more effective than either taken
alone.Use of these different procedures explains
the anomaly between human diving physiology
and the problems fish encounter naturally.It is
the internal saturation that parallels human diving
decompressions.The interesting and significant
difference between man and fish is that fish of
this size are actually more tolerant of supersatura-
tion (400 versus 200%)than are humans.As long as
a fish is allowed the opportunity and sufficient time
to eliminate the gas,it can survive high internal
supersaturation.
The fish has no opportunity to eliminate gas
when either exposed to external supersaturation
or exposed simultaneously to internal and exter-
nal supersaturation.Thus,the effects of the latter
treatment are synergistic.Furthermore,the bubbles
do not necessarily have to be found throughout the
fish,nor does the fish have to be anywhere near
fully saturated;bubbles in the coronary arteries
are sufficient to cause death.
To summarize our studies and put the results
into perspeaive,several statements can be made:
1.Equilibration of highly perfused tissues of
coho salmon (60-100 mm)occurs in 60 to 90 min
at any saturation.Therefore,below a certain satu-
"Temperature-induced bubbles will be examined in future
studies.
50 Beyer,D'Aoust,Smith
ration level (less than 150%)mortalities cannot be
directly related to equilibration time only because
death in these chronic tests occurs hours or even
days after equilibration is reached.The total
amount of gas necessary to cause mortality is a
critical factor but it is unclear at this time what
other factors are involved in the chronic tests.
2.Fish can withstand high supersaturations
for short periods as long as they are able to elimi-
nate the excess gas.
3.Depth is critical in keeping gas in solution.
If a fish in a river sounds he can redissolve bubbles.
Water often has the same gas concentration
throughout the water column (due to turbulent
mixing).Therefore,a sounding fish may redis-
solve the bubble,but will not be able to eliminate
the gas and will still be supersaturated on surfac-
ing again.If,however,the fish (at the surface)
swims into an area of non~upersaturated water
(e.g.,another stream)he can desaturate quite
easily although some sublethal effects may be
incurred due to the previous supersaturation (e.g.,
fish that we have exposed to sublethal supersatura-
tions are extremely susceptible to disease).If the
fish stays at depth all of the time,no bubbles
should appear.
FUTURE STUDIES
The main emphasis in our future studies will
be to investigate further the relationship of gas
saturation rates and bubble formation to different
sizes,temperatures,activity rates,and other gases;
resolye the differences between our acute test
results and ch ronic tests (the time lag):and explore
the additional mechanisms (blood clotting,etc.)
that could be related to this time lag.
ACKNOWLEDGMENTS
Supported by National Institute of Health
Grant #Hl 16254-02.
REFERENCES
Ackles,K.N.,ed.1973.Blood Bubble Interaction in Decom-
pression Sickness.Def.and Civillnst.Envir.Med.Proc.,302 pp.
Marsh.M.C.and F.P.Gorham.1905.The gas disease in
fishes.:In:R.eport 01 the Bureau 01 fisheries.1904.pp.343-376.
Meekin,T.K.and 8.K.Turner.1974.Tolerance of salmonid
eggs,juveniles.and squawfish to supersaturated nitrogen.In:
Nitrogen Supersaturation Investigations in the Mid-Columbia
River.Wash.Dept.Fish.,Tech.Rpt.12.pp.87-a8.
U.S.Navy.1973.U.S.Navy Diving·MailUal (Vol.1),
NAVSHIPS 0994-001-9010.Washington,D.C.
t .
and fish:Daphnia magna;western crayfish,
Pacifastacus leniusculus;three stoneflies,Acro-
neuria californica,Acroneuria pacifica,and Ptero-
narcys californica;and juvenile steel head,Salmo
gairdneri.
Nebeker,Stevens,and Brett:U.5.Environmental Protection
Agency,Western Fish Toxicology Station,Corvallis,Oregon.
FIG.1 View of interior of test tank showing open tank and net
cages for isolating craytish.
MATERIALS AND METHODS
Physical Cond itions
Five 6000-~fiberglass tanks,60 cm deep,were
used for maintaining four different levels of super-
saturated water and one saturated control (Fig.1).
A supersaturation generator system (Fig.2)was
used to control the gas level in each tank.Test
water was obtained·from two wells located about
30 m from the WiJlamette River.Water was chilled,
aerated to saturation,heated to a given tempera-
ture with immersion hea,ters,pumped under
IA.V.Nebeker
D.G.Stevens
J.R.Brett
The effects of supersaturated water on fish,the
so-called gas bubble disease (GBo),have been
familiar to aquarium and hatchery workers for many
years (Marsh and Gorham,1905;Embody,1934;
Rucker and Hodgeboom,1953).Supersaturated
water (up to 150%total dissolved gas)created by
dams on the Columbia River,and its effect on fish,
have recently been described by Westgard (1964),
Pauley and Nakatani (1967),Ebel (1969),and others.
Three recent literature reviews (Rucker,1972;Weit-
kamp and Katz,1973;and Bouck,1974)adequately
summarize previous work on gas bubble disease
and supersatu rated water problems.laboratory
studies have been conducted recently by Blahm et
al.(1973)and Bouck et al.(1973)to determine
comparative sensitivity of va rious fish species to
supersaturated water,but little work has been
completed with fish-food organisms,especially
freshwater invetebrates.
This study was conducted to determine the
effects of various levels of supersaturated water on
the following freshwater aquatic insects,crustacea,
Effects of Gas
Supersaturated
Water on Freshwater Aquatic
Invertebrates
ABSTRACT
Tests with the stoneflies Acroneuria californica,Acroneuria
pacifica,and Pteronarcys californica;Daphnia magna;crayfish
(Pacifastacus /eniuscu/us);and young steelhead trout (Sa/mo
gairdneri)were conducted to determine the sensitivity of fresh-
water insects,crustacea,and fish to gas supersaturated water.
5toneflies and crayfish were tolerant of supersaturation levels
(125%)that killed trout;however,survival was similar in Daphnia
and trout.Crayfish died at 150"k and 140%;some deaths and
sublethal signs occurred at 130%.5toneflies were immobilized
at 135%and exhibited buoyancy problems at 125%,but were
unaffected at 115%.Daphnia were killed at 120%and exhibited
partial mortality at 115%;air in the gut caused food blockage
and subsequent starvation.Bubbles were observed in body fluid
and tissues,and general body distention occurred before death
in Daphnia,crayfish and stoneflies.The open circulatory system
of invertelbrates,relatively simple compared to fish,appeared
to be the main reason for the greater tolerance of insects and
crustacea to gas bubble disease.They do not have the complex
capillary blood vessel system of fish which is rapidly blocked Iby
bubbles,or emboli,that form in the blood.
-
-
.....
51
EXPOSURE
TANK
-
~--=..:_-------
AIR ESCAPE VENT
/
Gas,Chemical,and Data Analyses
A Weiss saturometer (fig.4),modified and
adapted as a routine analytical laboratory instru-
ment,was used for measuring total dissolved gas
pressures in \the exposure tanks (Table 1).It was
calibrated and checked periodically with the Van
Slyke Gas Analyzer and the Winkler method for dis-
solved oxygen.Several saturometer sensors,gauges,
and a mercury manometer were compared periodi-
cally to ensure accuracy.The formula:BP +~P
-VP/BP x 100 =%saturation,where BP =Barometric
Pressure,~P =Saturometer reading,and VP =Water
Vapor Pressure,was used to calculate total percent
satu ration.
back to normal atmospheric pressure.The degree
of supersaturation was controlled by the amount
of compressed air added to the water.
Test animals were either allowed to move
about freely in the open tank or were restricted
to four net cages (Fig.1)or to stainless steel wire
cages (Nebeker and Lemke,1968)suspended in the
water.Two sizes of wire cages,7.5 cm in diameter
by 12.5 cm high,and 20 an in diameter by 25 cm
high,were used.They were shaped like a standard
cylindrical tin can with a screen bottom on the
large cage and one set of small cages and a petri
dish bottom on the other set of small cages.
The water in Tank 1 was supersaturated at
125%total gas saturation and was used for testing
directly and also as a water source for siphoning
into other smaller tanks and aquaria holding addi-
tional test animals.Water flowing at a rate of
9 Qlmin (Tank la -500 Q),and 6.3Qlmin (Tanks lb,
1c,ld -19 Q)through the siphon lines from Tank 1
maintained the smaller tanks at 125%.
~:.
RETENTION
1
TANK
.\
PUMP TURBUlENCE lOOP
IA I R+WA TEID
HEATER
WELL -e:=I:=::::;"l
WATER ~+-__~
AGo 3 Dii1sr-of supet'Siltura.tion generiltor system.
AIR LINE
AIR COMPR£SSED
COMPR£SSOR GAS AERATION
TANK
pressure,and mixed with compressed air (Fig.3).
Water and any remaining air not in solution flowed
through the retention tank where excess air was
vented off.Test water then flowed into the exposure
tank,where it became supersaturated when released
FIG.2 Su~f'5ilturiU:ion &eneriltor system where ilir ilnd wiltet'
ilre mixed under pressure to produce required percentilSe of
loul gas SItI~f'5ilturill:ion.
-
-
~,
-
52 Nebeker,Stevens,Brett
"'""
5 148.8 1.1 147.3 to 150.4
4 135.5 0.4 135.0 to 135.9
3 119.2 1.1 118.0 to 120.0
3 115.4 0.6 114.8 to 115.9
3 114.8 0.4 114.4 to 115.3
3 108.7 0.6 108.1 to 109.3
5 140.6 1.4 139.0 to 142.0
5 130.5 1.1 129.0 to 131.8
5 119.4 0.5 118.6 to 120.0
N Mean.±.SO Range
45 124.7 0.7 123.2 to 126.3
21 141.4 1.5 139.0 to 143.6
25 130.2 1.2 128.0 to 131.8
29 119.4 0.8 117.6 to 120.9
3 150.4 1.2 149.1 to 151.2
45 124.7 0.7 123.2 to 126.3
10 139.9 0.9 138.0 to 141.3
12 119.3 0.8 117.6 to 120.9
11 150.9 1.1 148.8 10 152.6
7 139.8 1.1 138.0 to 141.3
10 128.6 1.4 126.6 to 130.7
10 119.1 0.8 117.6 to 120.0
5 150.2 0.9 148.8 to 151.2
Measured total gas saturation (%)Z
125
135
150
140
130
120
150
125
140
130
120
150
120
:115
115
110
1
3
4
1
1
2
3
4
1
2
3
4
1 1403
2 130
3 120
1
2
3
4
9
7
8
5
4
3
6
2
Nominal
total gas
Test Test saturation
No.'chamber (%)
Daphnia Test Methods
Known-age Daphnia magna from cultures main-
tained at the Western Fish Toxicology Station
(WFTS)were used for testing.Rearing cultures were
maintained at 15 ±2°C on a 12-hr photoperiod with
one white-light and one Grow-Lux fluorescent bulb.
They were held in 3.8-~glass jars and fed a com-
bination of Oregon Moist Pellets®,pulverized in
water,and a mixed algae culture.Water was
changed biweekly.All Daphnia were transferred
with a large dropper from their rearing jars into a
transfer beaker and then gently poured into the test
cages.They were fed twice daily during testing and
had food in their guts when placed in the super-
saturated water.Lack of a heart beat or movement
when disturbed was used as the criterion for death.
TABLE 1 Nominal and Measured Test Levels
of Total Gas Saturation
'Test No.=WFTS Test No.:1=31,1 =34,1 =38,~=8,2'"29,
6 =30 7 =32 8 =8 9 =39.
iAnaly;is with \Veiss'Sc;turometer.
3Control tanks remained near 100%at all times.
'BOOk animals Were accidentally killed.
PRESSURE
GAUGE
..GAS P!:.RMEA BLE MEM3RAN£
"~..(TUBING)
SA TU ROMETE R
The well water was relatively soft with the
following mean chemical characteristics:hard-
ness,34 mg/2,alkalinity (as CaCOJ ),31 mg/2;
pH,6.7.The pH increased to 7.2 to 7.4 after water
aeration.Complete analyses of test water were
summarized by Samuelson (1975).flow rates into
the test tanks were set at 38 Vmin,and water
velocity varied from less than 3.0 cm/sec inside the
cages to up to 21 cm/sec in the open tank.
Deaths and signs of GBD were observed and
recorded daily,or more often during Daphnia expo-
sures,and data were analyzed according to methods
modified from Sprague (1969).Time to death was
plotted on graph paper.For Daphnia each point
represented the average of duplicate tests at the
given concentration.For crayfish each point repre-
sented an individual crayfish.Time to 50%death
was determined by using straight-line graphical
interpolation on log-probit paper,where each death
in each test replicate was plotted,a line fitted
through the points,and SOCk.death determined
where the line crossed 50%mortality.Lethal thresh-
old mortality concentrations were determined by
plotting time to 50%and 20%death at each test con-
centration for Daphnia and time to 20%death for
crayfish.
fiG.4 D~gram of modified Weiss saturometer•
""'"
-
-
-
....
-
,.....
Effects on Freshwater Invertebrates 53
TABLE 2 MeMi TIme to SO%and 20%Death for
Daphnia,Steelhead,Crayfish,and Insects
quate.The screen served as a suitable substrate so
no additional material was placed in the cages.
"Not ochieved-Insufficienl number of deoths.
'Determined from straight-line plots on log-probit graph paper.
'130'\,onimals were accidentally killed.
35
•
•
•
•
•
38
45
48
•
22
137
49
122
66
131
92
72
82
454
130
40
Mean time'
to 20%
deadt (hr}
•
50
•
•
•
•
91
65
71
93
165
123
330
94
210
130
123
101
Mean time'
toSO%
death (hr}
150
125
120
130
140
120
130
140
150
110
115
120
125
120
130
140
150
1202
140
150
125
115
120
135
Nominal
percentage
541 lul'a tion
9
(Inse<IS)
8
(Insects)
5
(Crayfish)
4
(Crayfish)
6
(Crayfish)
7
(Crayfish)
3
(Daphnia)
2
(Daphnia)
1
(Daphnia)
4
(Sleelhead)
Test number
(test animal)
RESULTS
Daphnia Tests
Test 1 Young non-egg-carrying adults were tested
for 96 hr at 140%,HOOk"120"k"and 100%total dis-
solved gas at 12°C (Table 1)and were held in the
20 by 25 em wire cages.Ten Daphnia were placed
in each cage,with one cage immersed in each of
the four exposure tanks.
Time to 50%death (Table 2)determined at 140%
total dissolved gas was 71 hr.When the test was
54 Nebeker,Stevens,Brett
Crayfish Test Methods
Crayfish used in Test 4 were collected from
Beaver Creek in Benton County,Oregon;those in
Tests 5,6,and 7 were collected from the Alsea River
in Benton County,Oregon.Before testing,crayfish
were held in aquaria with aeration and continual
flow-through of water and were fed young salmon
that had died in other supenaturation tests.The
test gas levels were set at the desired percentage,
and the crayfish were taken directly from the hold-
ing tanks (11 to 12°C)and placed in the open tanks,
in the net cages,in the large screen cages,or in
Tank la,depending on test needs.
Observations on the Branchiobdellid Annelid
worms living on the crayfish exoskeleton were
recorded but no detailed tests were conducted with
them.
Fish Test Methods
Twenty young steelhead trout (SaJmo gairdneri)
were tested for 2 weeks with crayfish and aquatic
insects at 125%.Time to death was determined,and
weight,length,and sublethal signs of GBD were
recorded.Five young sockeye salmon were also
tested with crayfish at 140%.
Insect Test Methods
Thestoneflies used were collected from the
Calapooia River,5 miles upstream from Holly,Unn
County,Oregon,and were placed in holding tanks
with flowing water and adequate water movement.
They were acclimated to test water and temperature
(12°C)for at least 24 hr before testing.Three 19-~
aquaria,17.5 em deep,were used as test chambers
for Test 8,two containing water supersaturated at
125%and one with saturated (100%)water as a con-
trol.Siphon lines delivered water to each aquarium,
maintaining adequate water movement to simulate
stream flow required by stoneflies.Rocks and stkks
were placed on the aquaria bottoms for substrate.
Test 9 was conducted in the large tanks.Two
cylindrical stainless steel wire cages (20 em in diam-
eter by 25 em high)were suspended near the water
surface in the open current (21 em/sec)in each tank
so water movement through the cage would be ade-
The small screen cages with petri dish bottoms were
ideal for Daphnia as they provided minimal current,
and food retention was not a problem.Water flow
through the cages was sufficient to circulate the
test water and maintain gas levels but not disturb
the Daphnia.The cages were hung on the outer lip'
.of the tank in the open current (Test 1),sheltered
from the current (Test 2),and on the net cages in
the center of the tank (Test 3)where the water
velocity was less than 5 em/sec.
-
-
-
-
Effects on Freshwater Invertebrates 55
TABLE J Daphnia Mortality After 96 hr at 140%,130%, 120%,and 100%Total Gas
Saturation at 12°C,and Observed Signs of Gas Bubble Disease'(Test 1).
2 dead,but with live young,no bubbles
Signs of gas bubble disease (GBD)
(8 =Bubbles)
Test 3 Daphnia were tested at 120".-b,115%,110%,
and 100%saturation at 12°C for 1 week.Two 7.5
by 12.5 cm screen cages with petri dish bottoms
At 150%all were alive after 24 hr but many
had air in the gut and brood pouch and were swim-
ming at the surface.The first deaths occurred at
52 to 53 hr at 150%and 140%,indicating a delayed
lethal effect not directly attributable to blockage of
vital fluids,as in fish (Table 4).Mean time for 50%
death at 150%was 101 hr.Mean time to 50%death
was 123 hr at 140".-b saturation,and all had died at
198 hr.Fifty percent were dead after 130 hr at 130%,
and many had air in the gut and were held at the
water surface (Table 4).Mean time to 50%death at
120%was 210 hr (Table 3),and 70%were dead when
the test was terminated after 264 hr (Fig.7).
No bubbles apparent
4 small B clinging externally
Many B-in gut,brood pouch and under carapace
2 B under carapace,some fungus
No B-carrying 2 small eggs
Gut full of air
No B,carrying embryos
Gut full of air,B in brood pouch with 2 young
Gut 4/5 full of air,carrying several young
Gut 1/2 full of air,carrying several young
No internal B,3-4 clinging externally
One B inside carapace -body badly fungused
One B inside carapace -fungused
Fungused
Fungused
Fungused .
No bubbles or fungus
Gut full of air -fungused
Gut full of air,no heart beat,slight movement of appendages
Gut full of air,heart beating well,carrying ephyppia (2 eggs)
One B im.ide carapace -body fungused
No B -lying on cage bottom
B in brood pouch.fungused
B in gut and carapace
3-4 B under carapace
No bubbles
One B under carapace
Body badly fungused
No heart beat -no B,slight movement,half out of molt
Food in fore-and midgut,B filling hindgut,B in brood pouch crowding 3 young
Test 2 Young non-egg-carrying adults were tested
at 150%,140%,130%,120".-b,and 100%total dissolved
gas at 12°C for 11 days and were held in the 7.5 by
12.5 cm screen cages sheltered from direct current
by a larger screen cage (Fig.6).Ten Daphnia were
placed in each cage,and two cages were immersed
in each of the five exposure tanks.
terminated 80".-b were dead after 96 hr (Table 3),and
50%had observable bubbles in the gut or brood
pouch (Fig.5).Time to 50".-b death at 130%was 65 hr,
and 80%were dead after 96 hr.Ten percent at 140%
and 130%were carrying eggs.The time at which
50%died at 120%saturation was 91 hr.Sixty per-
cent were dead after 96 hr at 120%and all were
carrying young,indicating much better reproduction
at the lower gas level (Fig.6).
-
-
.-Percentage Daphnia TIme to
saturation No.death (hr)
~140%1 40
2 40
3 64
4 64
5 64
6 70
7 70
8 88
9 96*
10 96**,
130%1 40
2 40
~3 40
4 64
5 64
6 64
~7 70
8 88
9 96*
10 96**--120%1 40
2 40
3 64-4 88
5 88
6 91
7 96**
8 96**r-9 96**
10 96**
100%1-10 Controls-(Control)
'WFTS Test No.31.
·Barely Alive -96 hr
,.-··Alive and Active -96 hr.
-
100
10
8)/()1'In OA PHN IA
....60 ~4~1~'"~40 £/1~/-END OF TEST
0
>-100 140'>1lJ"I....!L---.:....._0.....l5O'.lo"...?0_0-0-°i;:8)
0:N ;(0/l2O'..........ja....60
::E .....L -:f//.-~/0 .-.I....40 4>./.-z ()_...;,c::0 /END Of lEST .1....20u
0:
.../I....00.
160 200 240
100 I
8)",,12OS
..,
60
.-.--.I
....•I_END Of lEST
'".-~40 ./____f 1l5~
20
0 .~v llll'ro
0 40 8)120 160
Tlh£IHOIJRSI
.....
-
FIG.5 Dapbn~with ilir bubbles in the gut (upper and lower
ilnimills);center i1nitnill without ilir bubbles is control
FIG.6 TIme 10 deilth for Daphnia magna exposed 10 110%"
115%,120%,130%,140%.and 150%tolill dissolved gas saturiltion
(Test 1,2 and 3).
TABLE 4.Daphnia Mortality After 11 DilYS ilt 150%,140%,130%.120%,and 100%Total GiiS
Saturiltion of 12°C,and Observed Signs of Gas Bubble Disease'(Test 2).
56 Nebeker,Stevens,Brett
-
,,....
Percenlilge
saturation
150%
Cage A
Tank 4
1SO"A.
Cage B
Tank 4
140%
Cage A
Tank 1
Dilpbniil TIme to Signs of gilS bubble diseilse (GBD)
No.death (hr)(B =Bubbles)
1 72 No B,gut with food
2 102 Gut full of air,B under carapace
3 102 B in brood pouch and under carapace
4 102 B in gut and brood pouch
5 102 3 B under carapace
6 102 B under carapace
7 102 B uncer carapace
8 102 B under carapace
9 119 Small B under carapace
10 150 No apparent GBD
1 52 No obviousB.some fungus1
2 72 B in gut and brood pouch
3 102 B under carapace.fungus
4 102 B in gut and brood pouch
5 102 B in gut
6 102 B under carapace
7 119 Gut 3/4 full of air.B in brood pouch and under carapace
8 119 4-5 small B under carapace
9 150 5-6 small B under carapace
10 150 No apparent GBD
1 53 large B filling brood pouch,some fungus
2 53 large B in brood pouch
3 102 Gut full of air,large B in brood pouch,B under carapace
4 102 B in brood pouch and under carapace
5 102 2 small B under carapace
6 150 No apparenl GBD
7 150 Gut and brood pouch full of air
8 150 1 B under carapace,some fungus
9 150 No apparent GBD
10 198 2 small B under carapace
TABLE 4 (continued)
,~
Percentage Daphnia Time to Signs of gas bubble disease (GBD)
saturation No.death (hr)(B =Bubbles)
140%1 53 No apparent GBD,fungus
Cage B 2 77 B in gut and brood pouch.fungus
Tank 1 3 102 B under carapace
4 119 Brood pouch full of air
.~5 119 Gut full of air
6 119 No apparent GBD,fungus
7 150 No apparent GBD
8 150 No apparent GBD,tissues fragmented
,-9 150 No apparent GBD,tissues fragmented
10 150 No apparent G8D,tissues fragmented
130%1 77 No apparent GBD,fungus
Cage A 2 77 No apparent GBD,fungus
~Tank 2 3 .102 Air in gut,1 B under carapace,fungus
4 150 No apparent G8D
5 150 No apparent GBD,tissues fragmented
6 169 No apparent GBD,tissues fragmented
7 169 No apparent GBD,tissues fragmented
8 217 No apparent GBD,tissues fragmented
9 243 No apparent GBD,tissues fragmented
10 lost Apparently disintegrated2
~130%1 78 No apparent G8D,fungus
Cage 8 2 78 No apparent G8D
Tank 2 3 102 Gut full of air
4 119 No apparent GBD
5 119 No apparent G8D
6 119 No apparent GBD
7 150 Big B in brood pouch separating carapace halves,gut 1/2 full of air
8 150 Large B separating carapace,gut full of air
9 169 No apparent GBD
10 198 No apparent G8D
120%1 150 No B,fragmented,fungus
Cage A 2 150 No B,fragmented,fungus
Tank 3 3 150 No B,fragmented,fungus
4 150 1 small 8 under carapace
5 243 B in gut and brood pouch
6 264 No apparent G8D
7-10 264*No apparent GBD
120%1 102 No 8,fungus
Cage 8 2 119 No .8
Tank 3 3 150 No 8,carapace torn
4 150 No B,fragmented
5 169 3 small B under carapace
6 217 No apparent GBo
7 243 2 small B under carapace~8 264 No apparent GBD
9-10 264*No apparent G8D
100%1 150 Carapace torn
I""'"Control 2-10 264*No apparent GBD
Cage A
lank 5
100%1-3 150 3 lost or fragmented
Control 4-10 264*No apparent GBo
Cage B
Tank 8
F'~
'WFTS Test No.34
'Fungus begins growing on Daphnia soon after death...,..#Alive after 11 days.
Effects on Freshwater Invertebrates 57-
,-
-
~
00 DAPHNIA d
>-1Il j .'l2$
P-I I
.....70<o /.
P-60""liS H',-;t.0 50:E ----0--\---
P-«I o •185 HR.mzf.t<IORTAlITY....30u
""oj if....20a.
10 0 •'I I
I I I 1
60 1Il1lXl ISO em 300
Tl ....1E (HOURS)
FIG.7 Tune to SO%de~th of Daphnia magna ~t 130%~nd 120%
total dissolYed g.1S wtur~tion.
were suspended in each exposure tank,and five
Daphnia,10 days old,were placed in each cage.
About half the Daphnia were beginning to develop
eggs in the brood pouch.
The first death occurred after 48 hr at 120%
(Table 5),and air was observed in the gut.One
Daphnia,the only one observed during these tests,
had a small bubble in the body fluid of the head
next to the eye.fifty percent were dead after 93 hr
at 120%;bubbles were observed in the gut and on
the carapace of several survivors.The food eaten by
Daphnia was unable to pass through the gut when
air bubbles were present,and many Daphnia
apparently died of starvation.At 115%saturation
30%had died when the test was terminated after
170 hr of exposure.Eighty-five percent were carry-
ing eggs or young after 96 hr.Mean time to 20%
death was determined to be 137 hr.
At 110%the first and only death occurred after
48 hr,but there was no evidence of GBD and it had
young in the brood pouch.Sixty percent were carry-
ing young after 96 hr of exposure.Only 10%were
dead when the test was terminated after 170 hr,
but one was at the surface with a large bubble in
the brood pouch.
lethal threshold concentrations for Daphnia in
gas supersaturated water were determined by using
the time to 50'%and 20%death.The saturation per-
centage at which 50%were no longer killed was
near 128%.The level at which less than 20%of the
Daphnia were killed,a possible safe level,was near
111%total dissolved gas (Fig.8).
Crayfish Tests
Test 4 The water in Tank 1 was supersaturated
at 125%total gas saturation and used both for test-
ing and as a water source for siphoning into Tank 1a
58 Nebeker.5te...ens.Brett
where crayfish were tested with young steel head.
Tank 2 was used as a control and contained satli-
rated water.All tanks were maintained at 12°C.
Crayfish were placed in the net cages in Tank 1
(fig.1)and in Tank1a;water flowing at a rate of.
9 Vmin maintained Tank 1a at 125%.Four large;
crayfish (30 to 40 g)were placed in net cage C of
Tank 1 and 10 medium-sized crayfish (20-30 g)
were placed in net cage D.Three medium-sized and
11 small crayfish (10 to 15 g)were placed in Tank 1a
(with steelhead).They were exposed for 12 days.
Test 4 was a survey test at 125%total gas
saturation to compare the susceptibility of crayfish,
"....insects,and fish.No signs of GBD were observed in
trayfish,but the fish died rapidly at 125%(Table 6)
and all but one were dead after 12 days.
r-Test 5 Crayfish were tested at 150%total gas
saturation to determine their tolerance and to
expose 10 animals at a high gas level and observe
......signs of GBD in individual animals.The test expo-
sure·lasted 6 days,at which time all animals had
died.
One male died after 23 hr and was necropsied
to determine signs of GBD.Gills were"slide-
mounted along with those of a control animal that
was sacrificed.Bubbles were readily apparent in
-the gills of crayfish from 150%,but no bubbles were
observed in gills of the control animal.Three more
male crayfish (No.2,3,4)were moribund after 40 hr.
__They were removed from supersaturated water and
placed in saturated water to observe them for pos-
sible recovery.The others remaining in the tank
were sluggish and unresponsive.Bubbles were in
-the body fluids of the moribund crayfish and they
could be seen easily through membranous joints
(Fig.9)and the ventral abdom inal sternites.There
.....were no bubbles in the control animals.The three
moribund crayfish (No.2,3,4)showed no recovery
Tank 1 Tank 1a
Fish chamber B with crayfish3
1 10 10
2 14 14
3 18 18
4 38 18
5 48 25
6 48 27
7 50 27
6 54 30
9 54 30
10 75 32
11 75 36
12 95 38
13 116 48
14 121 54
15 125 119
16 142 120
17 142 121
16 166 142
19 190 195
20 268 224
(alive)
TABLE 6 Time to Death of Young Steelhead Trout Tested
Concurrently with Crayfish and Insects to Verify Lethality of
Test Water at 125%Total Gu Saturation (Test 8)1
'WFTS Test No.6.
2No crayfish or insect deaths occurred during
the time period of this test at 125%.
Time to deatb
in saturated water after 24 h r;one was dead,but
there were stil/a few slight movements of abdomi-
nal appendages in the other two.
Crayfish No.5 was moribund after 47 hr and
was sacrificed for necropsy.There were bubbles
under the membrane between the carapace and
abdominal tergites (Fig.9).The body cavity was
opened,and many bubbles were found in the body
fluids,and lodged between the internal membranes,
organs,and muscles;there were also bubbles in the
heart.Bubbles were very conspicuous in the gills
(slide mount)and the main gill rib (Fig.10).Bubbles
were also obvious in the body fluid of the plates
protecting and lying with the gills.
Effects on Freshwater Invertebrates 59
DAPHNIA
110 120 IXl 140 -150
PERCl:NT SUPERSAnJRATlON
_-A--1 Ef!D Q.F~S!._
I
I
I
I
\2O'fo,,
\
\
~,° •
'0,.~
"'''''--8 ~----8----0
275
250
Vi"""llB=~:::;
~150
0
:lE
~
0 100...
~;:::
50
fiG.8 Threshold concentration detennined for Daphnia magna
by using times to 50%imd 20%mortality to plot the cunes.
.....
~.
-
.-
FIG.!I Dors.J1 view of crilyflSh.leh:with bubbles under the
distended membrilne between the CHapilCe i1nd the mdomiRilI
segments.Right:control
FIG.10 Crilyfish giRs showing bubbles inside the gil mid-rib
i1nd filaments.
Crayfish No.6 was alive after 47 hr,but was
unable to control chelapods or to pinch.Crayfish
No.7 was alive and,with difficulty,was able to
manipulate chelapods.Crayfish No.8,9,and 10
were alive and able to swim with good coordination.
Another large male,necropsied after 64 hr,was
turgid and swollen from internal pressures of gas
and from water taken up because of osmotic
imbalance (generalized stress reaction in inverte-
brates).Many bubbles were present in the body
fluid,gills.and associated structures;bubbles could
also be seen through the abdominalsternites.One
crayfish died after 88 hr;three remained alive.Bub-
bles were conspicuous in the body fluid of the dead
crayfish and could be seen through the body wall.
60 Nebeker,Stevens,Brett
They could be observed through the thin portions
of the exoskeleton,legs,ven tral abdominal ster-
nites,and membranous joints.The last three cray-
fish were dead after 6 days.
Water supersaturated at 150%total dissolved
gas,acutely lethal to trout in less than an hour
(Nebeker,unpublished data),was acutely debilitat-
ing to crayfish,though·not immediately lethal,as
they remained in an advanced state of immobility
with no coordinated movements for several days
after a possible "ecological death point"was
reached.
Test 6 Crayfish were tested at 150%,140%,130%,
120%,and 100%total dissolved gas saturation at
12°C for 30 days (Table 7).Ten crayfish were
exposed to each gas level,five each in two of the
four net cages in each tank.An additional 10 cray-
fish were placed in the open tank at 150%saturation.
Two stainless steel wire cages were also suspended
in the test water in each tank,and one crayfish was
placed in each cage.
Thirty percent were dead after 40 hr (Fig.11)
at 150%,and many bubbles were readily apparent
in the body fluid,gills and other tissues.There was
no correlation between crayfish size and time to
death in any of the tests.All were dead after 96 hr
at 150%.Fifty percent were dead after 330 hr
(Table 2)at 1~,and most exhibited some degree
of bubbles in body fluids and tissues (Fig.9 and
10).There was an obvious difference in feeding
behavior between crayfish at 1~and controls,
(100%)after 96 hr.Crayfish at 140%moved slowly
or hardly at all,whereas the controls jumped and
swam when startled and moved about freely when
disturbed.All crayfish tested at 140%were dead
after 595 hr (24 days).
Two crayfish died at 130%,one after 215 hr and
one after 453 hr (Table 7);both had bubbles in their
body fluid.No crayfish died at 120%during the
30-day test,and no changes were apparent in feed-
ing behavior when compared to control animals.
Water supersaturated at 120%was apparently safe
for crayfish over a 30-day period.
Bubbles were abundant in the small branchioh-
dellid worms Jiving on the carapace of the test cray.;.
fish from 140%,but were not present in those from
control water.The five young sockeye salmon
placed in 140%were immediately affected;one had
died,one had lost equilibrium,and one had hemor-
rhage popeye after 4 hr of exposure.
Test 7 Crayfish were tested at 150%,140%,130%,
120%,and 100%total dissolved gas saturation at
12°C for 2 weeks.Males and females were tested
separately with five males in one net cage and five
females in another,giving a total of 10 crayfish
per gas level.
500200300400
TIME (HOU RSl
100
1~~I~~
-e----------..---------...----\-f?
ell
CRAYfiSH 0/I
o~1
~:
/0 END OF TEST-I
..___0 1
e °130'1.Io~I
/IoIo~C::::r::::~_L.....I..---L.--"-_L___.L___.l_L.......1JJ
o
>-100...
...z
~40
0::
w
c..20
fiG.11 Time to death for the crayfish Pacifastacus teniu5Cu/us
exposed to 120%,130%,140%,and 150%total dissolved gas
(Test 6).
The first death occurred after 49 hr of exposure
at 150%,and many bubbles were observed in the
""'"body fluids.Fifty percent were dead at 150%after
123 hr (Table 2),and all had bubbles in the body
fluids (Table 8).There was no difference in the sus-
ceptibility of males and females and all were dead
-after 262 hr of exposure.Fifty percent had died at
140%after 165 hr and all had bubbles in the tissues
and body flUids.All had died after 329 hr (2 weeks)
_when the test was terminated.Crayfish exposed to
130%were accidentally killed and no data were
obtained.No deaths occurred at 120%,and no
apparent signs of GSD were observed during the
2':week exposure period.
A lethal threshold concentration for crayfish in
supersaturated water was determined by using time
to 20%death as the criterion (Fig.12).The thresh-
old concentration where crayfish were apparently
safe,at least for 30 days,was near 127%total dis-
solved gas.
_700;:-------~--~-\--QlO_OF.1E~--
,VI
<>:is 6lXJ L
:5
>-I-
::;.!~~\
~CRAYFISH
o
I-200~~
I-•
I 1'-----'O,I-._--J'--_---l__---l__---I.__~
o 110 120 130 140 150
PERCENT SUPERSAl\IRATlON
fiG.12 Threshold conc:entriltion determined for the crayfISh
Pacifastacus leniu5Cu/us.
Insect Tests
Test 8
The stonefly species Pteronarcys californica
and Acroneuria pacifica were tested at 125%total
dissolved gas saturation for 12 days at 12°C to
determine their comparative sensitivity with cray-
fish and young steelhead trout.Ten stoneflies,five
Effects on Freshwater Invertebrates 61
-
.-
late instar nymphs of each species,were placed in
each test chamber.All stoneflies appeared well
adjusted to the test system.Bubbles were seen
adhering to stoneflies during the first few hours
after supersaturated water was introduced into the
aquaria.Three hours after the test began two
Acroneuria were floating and covered with bubbles;
the other animals also had bubbles adhering to their
bodies.They were checked every 4 hr during the
next 2.days and were apparently unaffected.How-
ever.Acroneuria had a few bubbles clinging exter-
nally.One Acroneuria successfully molted after
24 hr at 125%and appeared normal.Most Acro-
neuria had some bubbles clinging to them.and they
would periodically rise to the surface when they lost
hold of the substrate,but would manage to get back
to the bottom as many bubbles would burst when
they sHuck the water surface.Pteronarcys had few
bubbles on the external body wall,but did have
many small bubbles adhering to the ventral tracheal
gill masses (Fig.13);however,bubbles did not
appear to greatly affect behavior or movements.
62 Nebeker,Stevens,Brett
FIG.13 Venttill view of stonefty with bubbles on the gills.
All stoneflies were on the rock substrates and
were alive after 96 hr with no abnormal behavior
patterns,abdominal distention,or signs of stress;
only one Acroneuria was observed with a few bub-
bles on it.All were apparently unaffected after
8 days,but on day 11 one Acroneuria was at the
surface and was unable to get to the bottom because
of gas bubbles.Its body was partially distended and
had external bubbles among the gills.On the 12th
day one Acroneuria in Tank lb and one in Tank 1c
were at the water surface and their bodies were
partially distended.Bubbles buoyed them to the sur-
face and they became trapped in the aquaria cor-
ners.This would not have occurred in streams
where substrate would be more suitable than the
slick glass surface.However,they would.be more
vulnerable to predators in nature as they would
float to the surface more frequently because of their
additional buoyancy.All stoneflies were alive and
apparently unharmed after 12 days'exposure at
125%saturation.
Test 9 The stonefly species Acroneuria californica
was tested at 135%.120%,115%,and 100%total dis-
solved gas saturation.Ten late instar nymphs were
tested for 11 days at 12°C at each gas level.
No effects on stoneflies were observed after
96 hr at 120%,115%,and 100%saturation.A few
bubbles were observed on the stoneflies,but caused
no difficulties.At 96 hr the gas in Tank 1,120%,
was reset to 135%and left at that level for the
remaining 7 days of the test.One stonefly molted
successfully at 100 hr when the gas (130%)was in
transition between 120%and 135%.
The stoneflies appeared unaffected after 48 hr
at 135%.However,after 120 hr of exposure,two
Acroneuria in cage A had distended bodies but were
active when disturbed.Two stoneflies in cage B
also had distended bodies and one was having dif-
ficulty moving normally.Controls were normal and
showed no signs of GBD.Stoneflies were held at
115%for 10 days without any observable effect.
Those at 1135%were more sluggish than the controls
and thosE~at 115%and had bubbles adhering to
their external body wall.Two stonefJies in cage A,
135%,were full of air bubbles;the bubbles were
concentrated in the body fluids of the thorax at the
base of the legs and gills.Their bodies were fully
distended,appearing expanded like balloons.A
small midge living on one stonefly was filled with
air,also looking like a tiny balloon.Two stoneflies
in cage B,135%,also had bubbles in the body fluids,
and their bodies were distended to the capacity of
the body wall.Bubbles were visible through the
body wall at the base of the legs,the gills,and also
scattered throughout the body fluids,such as in the
mandibles,etc.
The stoneflies in 135%were removed from the
supersaturated water after 7 days'exposure and
placed in control water.After 4 hr no bubbles could
be found in the body fluids and the insects were no
longer distended,although they were still somewhat
sluggish.They were able to recover from short
periods of relatively high levels of supersaturated
water.Thus stoneflies are much less susceptible to
gas-supersaturated water than are fish,especially
salmon and trout,but the floating and unnatural
buoyancy may be important in increasing drift and.
predation.
DISCUSSION
Insects and crayfish were more tolerant of gas
supersaturated water than any of several fish spe-
cies tested (Nebeker,unpublished data).Daphnia,
although able to withstand short exposures,were
unable to avoid the problem of food blockage by air
in the gut and died at levels similar to those that
affected young salmon.Only one instance was
observed where an air bubble was seen in the body
fluid of Daphnia,and the heart was never observed
to be impaired by emboli.The circulatory system of
Daphnia is so simple that the problem of capillary
blood-vessel blockage by air,crucial to fish,was
nonexistent.The open circulatory systems of the
insects and crayfish,though more complex than
that of Daphnia,are relatively simple compared to
fish and appeared to be the main reason for the
greater tolerance of invertebrates to supersatura-
tion.The external exoskeleton apparently prevents
much of the surface injury and secondary infections
that are common to fish exposed to supersaturated
water (Nebeker and Brett,1975).
The few reports of GBD in invertebrates that
have been found are generally incidental observa-
tions of bubbles on or in the tissues,and little or no
quantitative gas data are given.Gorham (1901)
reported that scallops,hydroids,and squids showed
signs of GBD.Evans and Walder (1969)used the
shrimp Crangon crangon to study bubble formation
under decompression because its transparent exo-
skeleton allowed any bubbles formed to be immedi-
ately seen.The shrimp,when subjected to 400
kglcm2 for 10 min and then removed,were not as
active as before exposu re but rapidly recovered
without apparent harm.The stoneflies in the pres-
ent study responded similarly,although exposure
conditions and purposes were quite different.
Hughes (1968)reported gas bubble disease in lob-
sters when exposed to water supersaturated by air
leaking into the hatchery water supply,but no gas
levels were given.The occurrence of GBD in three
species of bivalve molluscs was described ~y
Malouf et al.(1972),but dissolved gas levels again
were not given.Massive blisters were formed on the
Effects on Freshwater {nvertebrates 63
I~
valves of oysters,and bubbles were observed in
gill filaments.The cause,increased water tempera-
ture with subsequent supersaturation of dissolved
gas,is a common occurrence and surely is respon-
sible for much more damage to aquatic animals
than is documented in the literature.Gas bubble
disease in larval and juvenile brown shrimp
(Penaeus aztecus)was recently described by
Lightner et aJ.(1974).Stage II protozoeal,larval
shrimp developed the disease after being placed
in water warmed in a closed heater that did not
allow excess gas to escape.Ten percent of the
shrimp were affected (5%died)and had gas bub-
bles under the carapace and either in the gut or
the hemocael surrounding the gut.No saturation
levels were given.Most freshwater invertebrates
are probably less sensitive to GBD than fish,
although there are exceptions,like Daphnia.In
general,if fish are protected by reasonable water
quality standards,the invertebrates will probably
be protected also.However,further work should be
done with important freshwater and marine inverte-
brate species to determine their comparative toler-
ance and to define their most sensitive life stages.
CONCLUSIONS
Daphnia magna was affected by gas-saturated
water ~115%.Signs of gas bubble disease ranged
from no obvious effects to massive air bubbles in
the gut,brood pouch,and under the carapace.Bub-
bles were rarely observ~d in the body fluid.Death
in most cases was due to physical blockage of the
gut by air emboli,with subsequent starvation.The
most obvious sign of gas bubble disease was the
buoyancy created by bubbles,which caused the
Daphnia to float at the water surface.The heart
was apparently unaffected.
Crayfish were tolerant of supersaturated water
and were alive with no apparent effects at 120%
and 125%,levels that were lethal to the young
steel head trout tested with them.Bubbles were
found in body fluids,gills and other tissues of cray-
fish that died at 140%and 150%total gas pressure.
They were more resistant to supersaturated water
than any of the 12 fish species tested at the Western
Fish Toxicology Station (Nebeker,unpublished
data).
Aquatic insects.represented by the three stone-
fly species.were also comparatively tolerant of gas-
supersaturated water.No deaths occurred at 125%,
but some insects were immobilized and had air bub-
bles in the body fluids.They recovered rapidly
(4 hr)when transferred from supersaturated to
saturated water.Increased buoyancy was observed
and coukl increase mortality of insects from
predation.
The open circulatory system of invertebrates,
64 Nebeker,Stevens,Brett
relatively simple compared to fish,appeared to be
the main reason for the greater tolerance of insects
and crustacea to gas bubble disease.They do not
have the complex capillary-blood-vessel system of
fish,which is rapidly blocked by the bubbles or
emboli that form in the blood.
The tough exoskeleton of insects and crustacea
prevents much of the surface injury and secondary
infections that are common to fish exposed to
supersatu rated water.
ACKNOWLEDGMENTS
We wish to thank Mike McCarthy,jim Fendrick,
and Lynn Jones,Oregon State university student
aids,for help with the numerous gas and water-
chemistry analyses;Robert Rulifson for the initial
saturometer and aid during construction and use of
the present models;and Earl Dawley and Larry
Davis for assistance with saturometer calibration.
We also thank Dee Boczkiewicz,Don Samuelson,
Gerald Bouck,Gary Chapman,and Ronald Garton
for assistance and Robert Trippel for help with
construction and operation of test equipment.
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Blahm,T.H.,R.J.McConnell and G.R.Snyder.1973.Effect
of Gas Supersaturated Columbia River Water on Survival of
Juvenile Sa/monids.Final Report,Part 1.National Marine
Fisheries Service,Prescott Field Station,60 pp.
Bouck,G.R.1974'.Total dissolved gas pressure.In:Water
Quality Criteria.National Academy of Sciences.(in press)
pp.13S-139.
Bouck,G.-R.,A.V.Nebeker and D.G.Stevens.1973.Acute
Lethality of Supersaturation to Pacific Salmonids and Other
Species.Task 04 Completion Report.EPA,Western Fish Toxi-
cology Station,Corvallis,Oregon (Unpublished).
Ebel,W.J.1969.Supersaturation of nitrogen in the Columbia
River and its effect on salmon and steelhead trout.Fish.Bull.
68:1-11.
Embody,G.C.1934.Relation of temperature to the incubation
period of eggs of four species of trout.Trans.Am.Fish.Soc.
64:281-292.
Evans,A.and D.N.Walder.1969.Significance of gas micro-
nuclei in the aetiology of decompression sickness.NalUre 222:
251-252.
Gorham,F.P.1901.The gas bubble disease of fish and its
cause.In:Bull.U.s.Fish.Comm.1899.19:33-37.
Hughes J.T.1968.Grow your own lobsters commerdally.
Ocean Industry 3(2):46-49.
Lightner.D.V.,B.R.Salser and R.S.Wheeler.1974.Gas-
bubble disease in the brown shrimp (Peaaeus aztecus).Aqua-
cuhure 4:81-34.
Malouf,R.,R.Keck,D.Maurer and C.Epifanio.19n.Occur-
rence of gas bubble disease in three species of bivalve mol·
luscs.J.Fish.Res.Bd.,Can.29:588-589.
Marsh,M.C.and F.P.Gorham.1905.The gas disease in
fishes.In:Repcw-t of the Bureau of Fisheries 1904,pp.343-376.
Nebeker,A.V.and J.R.Brett.1975.Effects of Gas Super-
saturated Water on Survival of Pacific Salmon and Steelhead
Smolts (in press).
Nebeker,A.V.and A.E.Lemke.1968.Preliminary studies on
the tolerance of aquatic insects to heated waters.J.Kansas
Entomo/.Soc.41(3):413-418.
Pauley,G.B.and R.E.Nakatani.1967.Histopathology of
"gas bubble"disease in salmon fingerlings.J.Fish.Res.Bd.,
Can.24:867-871.
Rucker,R.R.1972.Gas Bubble Disease of Sa/monids:A
Critical Review.Bur.Sport Fish.Wild/.,Tech.Paper.11 pp.
Rucker,R.JR.and K.Hodgeboom.1953.Observations on gas
bubble disease of fish.Prog.Fish.Cult.15:24-26.
Samuelson,D.1975.Seasonal Comparison of Basic Water
Quality for the Western Fish Toxicology Station,the Willamette
River and Major Western Oregon Rivers (1972-1974).Western
Fish Toxicology Station,Corvallis,Oregon (in press).
Sprague,J.B.1969.Measurement of pollutant toxicity to fish.
I.Bioassay methods for acute toxicity.Water Res.3:793-821.
Weitkamp,D.E.and M.Katz.1973.Resource and literature
review.Dissolved Gas Supersaturation and Gas Bubble Disease.
Seattle Marine laboratory.Seattle,Washington.60 pp.
Westgard,R.L.1964.Physical and biological aspects of gas
bubble disease in impounded adult chinook salmon at McNary
spawning channel.Trans.Am.Fish.Soc.93:306-309.
Effects on Freshwater Invertebrates 65
I R.K.Stroud
A.V.Nebeker-A Study of the
Pathogenesis
of Gas Bubble
Disease in Steel head Trout
(Sa/rna gairdneri)
.-.
-
-
ABSTRACT
Steelhead trout were exposed to supersaturated water and
randomly removed for necropsy at predetermined intervals.
Lesions included emphysema of fins and opercular,gas emboli,
exophthalmia,and hemorrhaging.The hypothesis that death
from gas bubble disease is due to hemostasis due to blockage of
blood flow by emboli was supported by the necropsy results.A
"cascading bubble effect"is described to explain bubble
formation.
Although gas bubble disease (GBD)in salmonid
fish has been studied extensively in connedion with
supersaturation of the Columbia River by hydro-
electric projects,few studies of the pathogenesis of
GBD have been reported.Observations of lesions
such as emphysema of the fins and body,exoph-
thalmia,gas emboli within the circulatory,system,
gill hemorrhage,and others have been made on
fish exposed to supersaturated water in hatcheries,
under experimental conditions,and in natural situ-
ations (Bouck et al.,1970;Ebel,1969;Harvey and
Smith,1961;Marsh and Gorham,1905;Rucker and
Hodgeboom,1953;Rucker,1972;Shirahata,1966;
Wyatt and Beiningen,1971).Observations in these
studies were primarily made on fish that had died
or were near death.
This study attempts to document the events
leading to the death of fish and describe the
sequence of developmen t and incidence of certain
gross lesions in juvenile steelhead trout (Sa/mo
gairdneri)continuously exposed to three levels of
supersaturated water in shallow tanks.
MATERIALS AND METHODS
Steelhead trout obtained as eyed-eggs from
the Alsea Hatchery of the Oregon Wildlife Commis-
sion were raised for experimental purposes at the
Western Fish Toxicology Station,Corvallis,Oregon.
These fish ranged from 17 to 24 em fork length,with
the majority of fish in the 18 to 20 cm range at the
time of use.
66
The experimental apparatus consisted of four
12-£1 (3.75 m)diameter circular tanks containing
well water supersaturated to various levels by con-
trolled air injection.Water in the tanks was main-
tained at a depth of 24 in.(60 cm)and lO°e.A more
detailed description of the experimental tanks and
method of supersaturation has been reported else-
where (Nebeker and Stevens,1975).Supersaturation
levels were monitored daily with a Weiss saturom-
eter.Supersaturation levels were maintained within
~1%of 120%,115%and 110%which were the three
exposure levels used in this experiment.A 100%
saturated tank was used as a control.
A four-compartment submersible cage was
placed in the center of each ofthe four tanks.Thirty-
eight randomly selected fish were divided into four
lots and placed in the four compartments of the
cage in the 120%supersaturated water.Twenty-
nine fish were similarly caged in 115%water,29 in
110%and 12 in 100%.Although caging fish in this
manner prevented access to peripheral areas of the
tank where the water current was greatest,it did
facilitate sampling of fish.
Random samples consisting of five fish were
taken from each tank at predetermined intervals to
check the development of lesions associated with
GBD.The fish were immediately placed in a high
concentration of MS222.Complete anesthesia was
achieved in less than 2 min.The water used as a
vehicle for the MS222 was dipped from the same
tanks from which the fish were taken.This proce-
dure kept the fish continually exposed to the same
level of supersaturation until they were necropsied.
Sampling intervals varied according to previously
Stroud:~partment oi Veterinary MediCine,Oregon State
University.Corvallis.Oregon;and Nebeker:United States
Environmental PrOlec1,on Agency.Western Fish Toxicology
Laboratory.Corvalli,.Oregon
"'"'
acquired lC50 data (that level at which 50%died)
for the various saturation levels so that specimens
could be obtained which would be in progressive
stages of the syndrome.
Each fish was measured and examined for
external gross lesions.A gill arch was cut to observe
if gas emboli were present within the conus arteri-
osis and the ventral aorta.An entire gill arch was
removed and examined under a dissecting micro-
scope for evidence of gas emboli in the afferent
lamellar gill vessels.The body cavity was opened
and the heart,kidneys,liver,and internal vessels
were examined for the presence of gas emboli and
other pathology.Samples of the fins,gills,various
internal organs,and the head were preserved in
Bouin's solution for histological examination.These
observations are the subject of another report and
are not included here.
At 120%supersaturation,19 fish died during
the 54-hI"experiment.Those fish which died during
the daywere examined in the afternoon of that day.
Fish that died during the night were examined the
following morning.There was no more than a 12-hr
maximum time between death and necropsy.All
dead fish remained in the supersaturated water
until necropsied.Necropsy procedures were the
same as for the sampled fish.
RESULTS
Occurrence of major gross lesions at selected
time intervals is presented in Tables'.l,2,and 3.The
lesions in fish still alive when sampled are com-
pared to lesions in fish that died.Two fish sampled
while still alive after 20-hr exposure were in terminal
convulsions.These fish contained gas emboli within
the afferent gill vessels,heart and major vessels.
Only one other fish sampled while alive in 120%
supersaturated water contained gas emboli.This
fish did not appear to be in terminal convulsions.
Gas emboli were in the gills,heart and major ves-
sels in all fish dying of GBD,but were not observed
in fish sampled alive except for the three mentioned
above.Because mortality did not occur in the groups
.exposed to 115%and 110%supersaturation (after
TABLE 1 Occurrence of Lesions in Live Steelhead (Sa/rna gairdneri)Smolts Exposed
to 120%Supersaturation Level and Sampled After B,20,30,and 42 hr Compared with Fish Dead
from the Effects of 120%Supersaturation at lB-,JO-,40-,and 54-hr Exposure
Fish alive Fish dead
Time sampled,hr 8 20 30 42 Total 18 30 40 54 Total
--Number of fish in sample 5 6 3 4 18 7 3 4 5 19 '".
Number with emphysema of:
"'""tail 2 3 6 2 3 4 4 13
anal fin 1 3 5 1 3 1 5
pelvic fill 3 1 5 4 2 2 1 9
pectoral fin 2 1 3 4 2 1 2 9
dorsal fin.....operculum 2 1 4
Number having gas emboli in:
afferent gill vessels 2*3 7 3 3 5 18
conus awteriosis 2*2 6 3 3
4 16
atrium 3 1 1 2 7
other vessels or tissues 2 1 1 4-Number with:
exophthalmia 1 1
hemorrhage from gil!2 1 2 5
fin hyperemia 1 1 2
stomach filled with food 4 4 3 1 12 4 3 1 3 11
intestine filled with food 2 2 4 1 1 2 2 6
~excess mucus in intestine 1 1 1 1 2 3 7
other eye pathology
~"fish in terminal convulsions when sampled.
Pathogenesis of Gas Bubble Disease 67
TABUl .Occurrence of Lesions in Steelhe~d (Sa/mo gairdnenl Smolts Exposed 10 115%
Supel'Sillurated Water ~nd S~mpled After 45,93.143,216.255,~nd 336 hr
Number of hr of exposure
45 93 143 216 255 336 Tol~1
Number of fish in s.Jmple 5 5 5 5 5 4 29
Number with emphysema of:
tail 3 4 2 5 1 16-anal fin 1 2 1 3 7
pelvic fin 1 1 1 1 4
pectoral fin 1 2 2 1 7
dorsal fin 1 1
.-operculum 1 1 1 3 6
Number having gas emboli in:
afferent gill vessels
"""conus arteriosis
alrium
olher vessels or tissues 1·1
Number with:
exophthalmia 2 1 1 2 6
hemorrhage from gills 1 1
fin hyperemia 2 1 2 5
stomach filled with food 2 1 3
inlestine fil·led with food 2 3 5
excess mucus in intestine 1 1 3 5
other eye pathology
·Under peritoneum along kidney.
TABU 3 Occurrence of Lesions in Steelhead (Sa/mo gairdneri)Smolts Exposed to 110%
Supenalur~ted W~ter ~nd ~mpled After 69,142,213.310,360,and 408 hr
Number of hr of exposure
69 142 213 310 360 408 To~1
Number of fish in umple 5 5 5 5 5 4 2'9
Number with emphysema of:
tail
anal fin
pelvic fin
pectoral fin
donal fin
operculum 2 1 3 6
Number having gas emboli in:
afferent gill 'Je5Se1s
conus arteriosis
atrium
other vessels or tissues
Number with:
exophthalmia
hemorrhage from gi lis 2 3 4 5 3 17
fin hyperemia
stomach filled with food
intestine filled with food 3 3
excess mucus in intestine 3 4 3 10.-other eye pathology 1·3··\.'.,,..7
·Corneal opacity.
··Hemorrhage in anterior chamber of at least one eye.
68 Stroud,Nebeker-
DISCUSSION
The cause of death of fish by acute GBD has.....been reported by many investigators as hemostasis.
This is caused by blockage of blood flow tllrough the
heart and gills from the accumulation of gas emboli
-within the capillaries of the gills and results in
anoxia and death.Th is study supports this conclu-
sion.All fish that died at 120%supersaturation level
,_had large accumulations of gas emboli in the heart,
ventral aorta,and gills.However,with the exception
-
--
336 and 408 hr,respectively),direct comparisons
could not be made with those that died after expo-
sure to 120%.However,visible gas emboli were not
found in the heart,gills,or major vessels of any
fish exposed to 115%and 110%supersaturated
water.Emboli were absent in the controls.
At 120%and 115%supersaturation,gas accumu-
lations in the interray membranous tissue (emphy-
sema)and within the venules adjacent to the carti-
lagenous rays appeared more often in the tail than
in similar locations in the anal,dorsal,and paired
fins.Gross emphysema did not develop in the fins
of fish exposed for over 408 hr {17 days}to 110%
supersaturation.Subcutaneous emphysema along
the opercula was found in a significant number of
fish from all three levels and apparently is associ-
ated wit\h longer periods of exposure than that
necessary to produce emphysema of t\he fins.Fin
and tail emphysema is apparently a more acute
lesion and is associated with high levels of
supersaturation.
Exophthalmia,a lesion frequently associated
with GBD,occurred in six fish after 93 hr exposure
to 115%supersatu ration.Exophthalmia associated
with GBD is caused by the accumulation of gas
within the fatty tissues of the periorbital space,
resulting in abnormal protrusion of the eye.Other
ocular lesions including blood in the anterior cham-
ber were seen on Iy at 110%supersaturation after
long periods of exposure.
The entire group of fish were fed immediately
prior to the experiment,but not during the experi-
ment.This afforded an opportunity to observe the
effects of supersaturation stress on intestinal motil-
ity.According to Klontz,·food should pass through
the stomach and intestinal tract of trout in 8 to
10 h'r depending on the water temperature.Food·
consisting of Oregon Moist Pellets@ was retained in
the stomach for up to 54 hr at 120016 and 93 hr at
115%.Food observed in the stomach appeared to
have undergone little change due to the digestive
process.A simi lar delay in the emptying of the
intestine was also observed.Controls and fish held
at 110%did not have undigested food in the stomach
when necropsied.'The intestines of many fish also
contained an excess of thick bile-stained mucus.
of two fish in terminal convu Isions and one fish that
appeared norma I,fish exposed to 120%and sampled
prior to death did not have visible accumulations of
gas within major vessels,heart or gills.Visible
emboli were not seen in any fish at 115%or 110%
levels.Based on these observations,the formation
and/or migration of macroscopic emboli within the
blood vascular system appears to be an acute
terminal or near terminal phenomenon.Otherwise,
gradual accumulation of gas emboli would be visible
in the gill filaments of fish still alive under super-
saturated conditions.No such lesions were found in
this experiment.
The data indicate that initiation of bubble
growth from pre-existing nuclei and/or bubble dis-
lodgement from -the periphery of the body may
occur under certain physiological conditions that
can trigger a "cascading bubble effect"of emboli
into the gill capillaries with resultant \hemostasis.
The physiological conditions involved in the initi-
.ation of this apparently irreversible cascading effect
are unknown for fish.Many studies have been con-
ducted on mammalian systems in connection with
decompression sickness or the "bends."Similar
studies should be done to define the physiological
processes occurring in fish dying of GBD and find-
ings shou Id be considered in the determination of
acceptable levels of supersaturation.
For example,supersaturation levels as high as
120%may be tolerated by some salmonid fish under
certain circumstances,whereas levels as low as
115%may be acutely lethal under a different set of
conditions.Circumstances that force fish to swim
rapidly while in sublethal supersaturated water such
as excessive water flow,flight from predators,etc.,
may initiate the "cascading bubble effect"through
increased muscu Jar activity.Muscular activity is
known to contribute greatly to the development
and release of visible gas emboli in cats during
decompression experiments (Harvey,1944b).The
.effect of muscular activity was especially important
at the lower levels of pressure change.The explana-
tion given by Harvey is that muscular contraction
favors bubble formation by further reducing the
hydrostatic pressure causing the formation of large
vapor cavities into which gas can diffuse.If the
vapor cavity persists long enough,a visible gas
bubble can be formed in liquids with low gas ten-
sion.The bubble may gradually dissolve,but before
this happens,it might move into the general circu-
lation.Gas accumulations are known to form in
muscle even at relatively low levels of supersatura-
tion or decompression (D'Aoust,1974;Stroud et aI.,
1975).
·Personal communrcatlon,George W,Klontz.Professor of
Fisheries.University of Idaho,Moscow,Idaho.
Pathogenesis of Gas Bubble Disease 69
......
..,..
I
,.,.,.
Another explanation of bubble formation
related to muscular activity is that an increase in
blood temperature as it passes from the gills to the
systemic circulation causes dissolved gas to come
out of solution (Marsh and Gorham,1905).Increased
muscular activity causes a rise in body heat even in
poikilothermic animals.
This experiment and ·others have shown that
fish in supersaturation experiments frequently die
of gas embolism shortly after disturbances such as
netting live fish,removing dead fish,taking super-
saturation measurements with the Weiss saturom-
eter,etc.This is probably due to the "cascading
bubble effect"initiated by muscular activity or
excitement as a response by the fish to the distur-
bance of the environment.
If muscular activity or stress precipitates the
formation andlor release of gas nuclei from
peripheral vessels,it can be assumed that gas
emboli may be present in the blood vascular system
prior to the activity.Gas-filled venules were seen in
the fins of fish exposed to 120%and 115%levels of
supelrsaturation prior to the appearance of emboli
in the gills.At 110%supersaturation,similar obser-
vations have been made in previous experiments,
but were not seen in this experiment.Undoubtedly,
gas emboli exist in venu les th rough out the rest of
the body,but are most easily demonstrated histo-
logically in the fin and ta il.
Supersaturation levels causing tissue emphy-
sema and the development of gas emboli in
peripheral locations such as the fins may vary from
130%to 110'X,or below.However,the severity and
rate of developmen t of such external lesions as bub-
bles in the fins and tail and subcutaneous emphy-
sema on the body surface seem to have a direct
relationship to time of exposure as well as the level
of supersaturation (Le.,lesions develop more
rapidly at 120%than at 115%).Fish dying of gas
embolism at 120%supersaturation had a greater
incidence of fully developed lesions than surviving
fish even though exposure times were approxi-
mately equal.This indicates that fish which were
more susceptible to extemal lesion development
may also be more prone to the "cascading bubble
effect."
The incidence of extemal bubbles in steel head
trout is greater in the tail than in the fins.It has
been reported from physical models that bubbles
tend to form along lines of material stress (Harvey
et aI.,1944a).Material stress may be related to
swimming movement.The tail has a greater fre-
quency of movement than the fins when fish are in
moving water.Therefore,it is possible that increased
evidence of gas blisters in the tail is due to increased
movement.Similarly,the high incidence of emphy-
sema on the opercula may be related to mobility.
70 Stroud.Nebeker
If muscular activity is as important a factor in
emboli formation and release from peripheralloca-
tions in fish as Harvey indicates it is for mammalian
species,then the results of static water supersatura-
tion bioassays should be carefully examined and
compared to bioassays done in systems requiring
fish to swim against a current.Direct comparison
of supersaturation tolerances of salmonids in static
water versus situations where fish are forced to
swim have not been reported.Fig.1 shows the
results of such a study in largemouth bass (Microp-
terns sa Imo ides)where a significant difference in
tolerance to supersaturated water developed (Bouck
et aI.,1975).Such experiments on salmonids would
provide data more representative of GBD in wild
fish and would be useful to determine permissible
supersaturation levels.
Another interesting aspect of the pathogenesis
of GBD indicated by this data is that exposure to
sublethal levels of supersatu rated water may cause
decreased peristaltic movement of the gastro-
intestinal tract.Food was held in the stomach of
test fish (up to 54 hr at 120%and 93 hr at 115%)
much longer than could normally be expected.
In mammals,stimulation of the sympathetic ner-
vous system by stress factors can slow passage of
food in the gastrointestinal tract (Guyton,1966).
High protein food may tend to "ferment"during
this period of delayed passage through the gut.
This process may permit the abnormal buildup of
gas,pathogenic bacteria,toxic products,heat,or
other causes of irritation which could damage the
intestinal lining.Excess dark bile-stained mucus
with some hyperemia of the intestinal surface
which was seen at necropsy in this experiment
indicates intestinal irritation.Such damage may
100
15°C o~
/0---
>-lIJ 0 ......0 SWIMMING.-
.....0 ......
-<;::/"I-60""0 .--0 p::;:
"iI-0
0'z «l 0--....•u _0'.....-.""....•Q..20 •__-RESTI NG.--
a
0
FIG.1 Effects of swimming activity on the mortality of lilrgt!!:c~
mouth b.ss (Micropterus sa/moidesj in wilter supersa.turated.to c'
1~";
....
....
....
result in an increased chance of systemic bacterial
infection from foci of infection in devitalized areas
of the gut.
Although the sample size is small,data from
this experiment indicate that several aspects of the
pathogenesis of GBD need further investigation.A
study of factors contributing to the acute death of
fish through blockage of the circulatory system by
gas emboli is needed to determine if proposed per-
missible supersaturation levels will assure survival
of fish concurrently stressed by multiple factors.
A second study designed to investigate disease
susceptibility in fish surviving sublethal supersatu-
ration levels is needed.Vibriosis,a septisemic
bacterial disease of salmon,is present within the
saltwater estuaries of the Oregon Coast and may
cause high mortalities in salmonid fish (Cisar and
Fryer,)969).Present knowledge indicates that this
disease invades the fish through the gastrointestinal
tract.If the integrity of the gastrointestinal tract
is signifkantly disturbed by sublethal supersatura-
tion levels,vibriosis or other fish pathogens may
cause increased losses.
REfERENCES
Bouck,G.R.,G.A.Chapman,P.W.Schneider,Jr.and D.G.
Stevens.1970.Observations on Gas Bubble Disease in Adult
Columbia River Sockeye Salmon (Oncorhynchus nerka).Pacific
Northwest laboratory,Federal Water Quality Administration,
Corvallis,Oregon,June 1970 (unpublished manuscript)19 pp.
Bouck,G.R.,A.V.Nebeker,and D.G.Stevens.1975.Acute
Lethality of Supersaturation to Pacific Salmonids and Other
Species.Western Fish Toxicology Station,EPA,Corvallis,
Oregon (unpublished report).
Cisar,J.O.and J.l.Fryer.1969.An epizootic of vibriosis in
chinook salmon.Bull.Wildlife Disease Assoc.,5:73-76.
D'Aoust,B.G.and l.Smith.1974.Bends in fish.Camp.
Biochem.Physio/.,49A:311-321.
Ebel,W.J.1969.Supef5aturation of nitrogen in the Columbia
River and its effect on salmon and steel head trout.Fish.Bull.,
68:1-11.
Guyton,A.C.1966.Text book of Medical Physiology,3rd
edition,W B.Saunders Co.,Philadelphia,PA,/(p.876),11182 pp.
Harvey,E.N.,D.K.Barnes,W.D.McElroy,A.H.Whiteley,
D.C.Pease and K.W.Cooper..1944a.Bubble formation in
animals.I.Physical factors.}.Cell and Compo Physio/.,24:
1-n.
Harvey,E.N.,W.D.McElroy,A.H.Whiteley,G.H.Warren and
D.C.Pease.1944b.Bubble formation in animals.III.An
analysis of gas tension and hydrostatic pressure in cats.J.Cell
and Compo Physiol.24:117-132.
Harvey,H.H.and S.B.Smith.1961.Supersaturation of the
water supply and occurrence of gas bubble disease at Cultus
lake Trout Hatchery.Can.Fish.Culturist,30:39-46.
Marsh,M.C.and F.P.Gorham.1905.The gas disease in
fishes.In:Report of the Bureau of Fisheries,1904,p.343·376.
Nebeker,A.V.and D.G.Stevens.1975.lethal and sublethal
effects of gas supersaturated water on fresh water aquatic
invertebrates.Proceedings of the Gas Bubble Disease Workshop,
October 8-9,1974,Battelle-Northwest,Richland,Washington.
Rucker,R.R.1972.Gas Bubble Disease of Sa/monids:A
Critical Review.Bur.Sport Fish.Wild!.,Tech.Paper,11 pp.
Rucker,R. R.and K.Hodgeboom.1953.Observations on
gas bubble disease of fish.Prog.Fish.Cult.,15:24-26.
Shirahata,S.1966.Experiments on nitrogen gas disease with
rainbow trout fry.Bull.Fresh.Fish.Res.lab.,15(2):197-211.
Stroud,R.K.,G.R.Bouck and A.V.Nebeker.1975.Pathology
of acute and chronic exposure of salmonid fishes to super-
saturated water.In:Chemistry and Physics of Aqueous Gas
Solution Symposium,Toronto,May 11-16,1975 (submitted).
Wyatt,E.J.and K.T.Beiningen.1971.Nitrogen gas bubble
disease related to a hatchery water supply from the forebay of
a high head re-regulating dam.Fish Commission of Oregon
Research Report,];3-12.
Pathogenesis of Gas Bubble Disease 77
]
1
1
Fickeisen.Montgomery.and Hanf:Battelle-Northwest,
systems Department.Richland.Washington.
of the Columbia River in the McNary Wildlife Refuge
at Burbank,Washington.The common stock was
held in circular tanks receiving flowing Columbia
River water and were fed trout pellets.As required
for prophylaxis,the stocks were treated with mala-
chite and with Diquot as well as fed pellets contain-
ing antibiotics.Prior to being tested,the required
number of fish were brought to the test temperature
at a rate of 1°C per day.They were held at the test
temperature for a period of at least 10 days.Tem-
perature fluctuation during this acclimation period
was Jess than 1°C.Throughout the holding period
all of the stock tanks were heavily aerated and
experience has shown that under these conditions
gas tensions range generally between 95 and 105%
of equilibrium value.
,Water supersaturated with atmospheric gases
was generated in a pressure vessel which received
an air-water mixture at about 40 psig.Control of
the dissolved gas tension was provided by an adjust-
able standpipe with a float valve to release excess
gas,thereby permitting an adjustable head space of
air over the water by adjusting the backpressure
and turnover time,and by adjusting the rate of air
inflow.
The testing facility consisted of four rectangular
tanks,each 1.17 by 0.46 m with the water level
maintained at a depth of 35 cm to minimize hydro-
static pressure compensation for excess gas tension..
One of the tanks was supplied with water from the
supersaturation system.Two of them recelved-
supersaturated water mixed with normally saturated;
water to control gas tension,and the founh tank;
was a control and received normally saturated'
water.Turnover time in each tank was calculated to:
be less than 20 min based on the flow rate.Tem-'
perature in all tanks was maintained within O.SoC
10.H.Fickeisen
J.C.Montgomery
R.W.Hant Jr.
ABSTRACT
Black bullhead.Icta/urus me/as,were acclimated to 8,12,16.and
20°C in Columbia River water,and were tested at each of the
acclimation temperatures to determine acute tolerance to dis-
solved atmospheric gas tensions in excess of equilibrium satura-
tion.The data were subjected to probit analysis and mean 96-hr
TL 50 values were 126.7%of equilibrium saturation at 8°C,125.1%
at 12"C,123.8%at 16°C,and 124.4%at 20°C,indicating a slightly
elevated tolerance at the lowest test temperature.These values,
while indicating a statistically significant difference in tolerance,
do not indicate an ecologically significant effect of temperature
on acute tolerance of black bullhead in the range of tem-
peratures tested.
Preliminary bioassays of tolerances of several
seleded freshwater teleost species to dissolved
atmospheric gas supersaturation which were con-
dUded at 20°C (Fickeisen,Schneider,and Mont-
gomery,1973)demonstrated an apparently lower
tolerance for the species having "cold water"
preferenda than for the "warm water"species
tested.In addition,for several years the literature
has held that increased temperature decreases
tolerance to supersaturated dissolved gases,but
little experimental evidence has been available,
especially for non-salmon ids (Anonymous,1971;
Weitkamp and Katz,1973).We therefore began an
extensive series of acute bioassays of the effect of
excess atmospheric gas tension at different tem-
peratures on black bullhead (Ieta/uTus me/as),
pumpkinseed sunfish (Lepomis gibbosus),and rain-
bow trout (Sa/me gairdneri).Tests were to be con-
dUdedi under non-shock conditions,that is,the fish
were to be acclimated to the test temperature prior
to exposure to supersaturation_This paper is a
progress repon of studies that are continuing,and
describes the effects of temperature on tolerance
of black bullhead.
MATERIALS AND METHODS
Adult black bullhead were collected by beach
seine and by hook and line from a backwater pond
72
Effect of
Temperature on
Tolerance to
Dissolved Gas Supersaturation
of Black Bullhead,Icta/urus me/as
-
.....
,j'
!,
135
135
135
105 110 115 120 125 130
GAS SATURATION (MEAN TOTAL 01SSCtv£0 GASt
105 110 115 120·125 130
GAS SATURATION (MEAN TOTAl DISSQVED GASI
lOS 110 115 120 125 130
GAS SATURATION IMEAN TOTAL DISSQV£D GASI
EACH POINT REPRES£NTS 10 INDIVIDUAlS
nOTAl'&II
EACH POINt REPRESENTS 10 INDIVIDUALS
nOTAL'&II
l.Or-----~--------~,.."...---..,
9b-tlOURS •.16°C
EACH POINT REPRESENTS 10 INDIVIDUAlS
(TOTAL'1601
0.0 L-_....J...."--_..L-_---'I.-_-'-""-........L-_---'__....
100
0.0 L..-_-J.-........_.J.-_---l__-'-......._lI.o.."---......__...
~oo
0.0 1-_-.l...__..l-_---l__-l.__...Jl.-__......_-1
100
~0.8
~
'"~O.b
~
2
li 0.4
~oa:
"-
1.0 .----------------"7""--..,
9b-HOURS.ItC
tension may,however,dictate that each potentially
affected area be independently assessed prior to
instigating mitigation plans.
FIG.1 Acute bioass.JIYs of dissolved gas tolerances of Ictalurus
me/as (Black Bullhead)at B°C.
0.2
0.2
FIG.3 Acute bioass.JIys of dissolved gas tolerances of Icta/urus
melas (Black Bullhead)Oil 16°e.
Effect of Temperature on Tolerance 73
1.0..---------------------,
~0.8
-';!::
~O.b
~
2
~0.4
o
'""-
FIG.2 Acule bioassays of dissolved gas tolerances of Ictalurus
me/as (Black Bullhead)at l20 e.
RESULTS AND DISCUSSION
Gross necropsies demonstrated that in nearly
_aI/cases examined the cause of death was clearly
gas bubble disease,manifest by massive cardiac
blockage by emboli.Emphysema and petechial
hemorrhaging were commonly observed as was the
-presence of emboli in blood vessels,gills,and
organs.Cases of not clearly gas bubble disease
mortalities were due to indeterminant causes,but
-.were likely due to unobserved gas embolism.In no
case were there any mortalities in the control tank
and at the termination of the test control fish showed
no external signs of gas bubble disease while they
-were common among survivors which were exposed
,to supersaturated water.
Plots of proportional mortality against total gas
,-tension for each of the test temperatures are shown
in Fig.1 through 4.It is important to note that the
curves are quite steep,indicating a narrow range
.-between non-lethal and lethal levels of gas tension.
This factor could be significant in planning mitiga-
tion activities and in defining water quality standards
as even a relatively small decrease in dissolved gas
.-content might have a large effect on resulting mor-
tality,provided that the new,lower gas tension was
below the lethal threshold.Species differences in
,-tolerance and depth distribution as it relates to
hydrostatic pressure compensation for excess gas
of the desired test temperature and was continuously
recorded.
During a test 10 fish were placed in each of the
four troughs receiving artificially supersaturated
water generated by injecting compressed air into
r-water under pressure in a receiver.The tanks
were covered with black plastic to minimize dis-
turbing effects of wet lab activity.Troughs were
checked at least daily for mortalities which were
I"""removed.About one-third of the mortalities were
randomly selected for gross necropsy to determine
cause of death.At least th ree times during each
"""4 day test dissolved gas tension was measured using
the Weiss saturometer with a mechanical shaker.
Water temperature was also recorded as was baro-
___metric pressure using a mercurial barom~Jer.From
these data the percentage of equilibrium saturation
was computed for total dissolved gases.In addition,
dissolved oxygen was measured by Winkler
,-.titration,and pH was measured once during each
test.At each temperature it was necessary to run
replicate tests varying the gas tensions somewhat
in order to obtain a sufficient data base for probit
analysis to determine TL so values.In most cases,
three tests were required at each temperature.
Mean gas tensions and mortalities at the end of
~96-hr were analyzed by computer probit anaJys,js to
determine fiducial limits about the fitted curve of
proportional mortality versus total gas saturation.
128
Vi
~...
5 126....
z
i 124
'"""~
%
is!122y:....
tion temperature not equal to test temperature).
and supersaturation,it is probable that a different
effect would be observed.
120 '--_-L--..l._L-..J--..L---l._L-...L-....J...--'_.l..--'----!::"--'
12 16
TEMPERATURE !TEST AND ACCLllMlION,°0
ACKNOWLEDGMENTS
Research on which this paper is based was
reported under Atomic Energy Commission Con-
tract Number AT(45-1)-1830.
REFERENCES
Anonymous.1971.Columbia River Thermal Effecls Study.
Volume 1:Biological Studies.Environmental Protedion
Agency.In Cooperation with the Atomic Energy Commission
and the National Marine Fisheries Service.102 pp.
Fickeisen,D.H.,M.J.Schneider and J.C.Montgomery.1973.
Tolerance of Selected Fish Species to Almospheric Gas Super-
saturation.American Fisheries Society 103rd Annual Meeting,
Orlando,Florida,Seplember 10-12,1973.BNWL-SA-4794.
Weitkamp.D.E.and ,\-1.Katz.1973.Resource and Litera-
ture Review of Dissolved Gas Supersaturation in Relation to
the Columbia and Snake River Fishery Resources.Report to
the Northwest Utilities Cooperative by Seattle Marine
Labor atories.
fiG.5 Effect of temperature on tolerance of Ictalurus mefas
(Slack Bullhead)to excess dissolved g<1S tension.Bars represent
liduciallimits <1bout the R;e nlue.
Q
lOS 110 Il5 120 125 130
CAS SA TURA liON jlllA.!11 TO TAL IllS SOt YEO CA SI
0.0 '--__....J.__...L.__.l-_.....,J__...u.__....J..._---'
100
0.2
'l6-HOUR S.2Ifc
=:0.8 EACH POINT REPRESENTS 10 INDIVIDUAlS
~(TOTAl.•1201
!0.6
<Czo -------------
Q;:0.4
~oeo:
C1.
F:ig.5 is a plot of the 96-:hr Tl so values with their
fiducial limits at each of the test temperatures.The
tolerance at the lowest test temperature is statis-
tically significantly greater than that at the higher
temperatures.It is important to note that the dif-
ferences are small and not of ecological significance
as greater perturbations in gas tension take place
over short periods of time in nature.An active fish
moving vertically in the water column will also
experience greater changes in the compensatory
effect of hydrostatic pressure.
Tests with rainbow trout are continuing,but we
are able to report that at 20°C the TL5()is about 118%
total gas supersaturation,a value significantly lower
than that for black bullhead,both from a statistical
and an ecological basis.
To summarize our findings,the effect of tem-
perature within the range tested has little effect on
tolerance of black bullhead acclimated to the test
temperature.Future tests are planned at tempera-
tures ranging up to 30°C.However,if exposed to
the combined effects of a thermal shock (acclima-
to .--------------"..----....".---,
74 Fickeisen,Montgomery,Hanf
FIG...Acute b~5QYS of dissolved g~.oIerillnces of IClafurus
melas (Sbd Sullhe<1d)ill'20 D C.
-
-
"""
.-
i
I
I,I
~I
I..I
1
:I,i
,I
i
1·1
1
j
water is subjected to a substantial temperature
increase upon passage through a power plant con-
denser system without allowing equilibration with
the atmosphere,the water will become gas super-
saturated.The return to normal saturation levels
under such conditions by gaseous diffusion across
the air-water interface appears to be a slow process
and usually does not take place before the discharge
water is again cooled to ambient by entrainment
mixing and heat loss to the atmosphere.Fishes
attracted to heated effluents supersaturated with
dissolved gases may develop gas bubble disease
and are subject to possible mortality.
Mortality of fish due to gas bubble disease in
the heated effluent of power plants has been
observed (DeMont and Miller,1971;Marcello and
Strawn,1973).However,with only a few reported
cases of gas bubble disease mortality of fish in the
thermal effluents of power generating stations,
documentation of all such incidents is needed to
enhance our understanding of what environmental
conditions and power plant design features and
modes of operation may lead to gas bubble mortal-
ity of fish.This paper documents a substantial mor-
tality of Atlantic menhaden,Brevoortia tyrannus,
that occurred in the discharge channel and thermal
plume of Boston Edison Company's Pilgrim Nuclear
Power Station Unit 1 during April 8 through 24,1973.
Marcello:Boston Edison Company,Nuclear Engineering
Department,Environmental Sciences Group,Boston,Massa-
chusetts;Fairbanks:Massachusetts Department of Natural
Resources,Division of Marine Fisheries,Sandwich,Massa-
chusetts.
PILGRIM NUCLEAR POWER ST AnON
Pilgrim Nuclear Power Station is situated on the
western shore of Cape Cod Bay in the town of
I R.A.M.arcello,Jr.
R.B.FairbanksGasBubble
Disease Mortality
of Atlantic
Menhaden,BrevoortiaTyrannus"at
a Coastal Nuclear Power Plant
ABSTRACT
A substantial mortality of Atlantic menhaden,Brevoortia tyran-
nus,occurred in the discharge channel and discharge plume
area of the Boston Edison Company's Pilgrim Nuclear Power
Station Unit 1 during the period April 8 through April 24,1973.
Gas bubble disease was implicated as the cause of their death.
¥easurements of dissolved gas concentration of the station's
intake and discharge water during this fish mortality are pre-
sented.Observations on the behavior and results of the patho-
logical elCamination of menhaden afflicted with gas embolism
are discussed.
Water supersaturated with dissolved gases can have
detrimental effects on fish and other aquatic organ-
isms.The manifestation of these effects in fish is
generaHy referred to as gas bubble disease.While
fish appear to tolerate mild cases of gas bubble
disease,extreme cases have resulted in mortalities
(Marsh and Gorham,1905;Woodbury,1942;Renfro,
1963;Ebel,1969;Beiningen and Ebel,1970;DeMont
and Miller,1971;and others).
Gaseous supersaturation of natural waters has
been attributed to increased photosynthetic activity
(Woodbury,1942;Renfro,1963),the falling of water
into plunge basins below dam spillways (Harvey and
Cooper,1962;Ebel,1969;Beiningen and Ebel,1970),
and the drawing in of air at water pump intakes,
leaky pipelines,and similar situations (Marsh and
Gorham,1905;Harvey and Smith,1961).More
recently the passage of cooling water through the
circulating water systems of electric power gener-
ating stations has also been identified as having the
potential for creating gas-supersaturated conditions
in the cooling water discharge (DeMont and Miller,
1971 ).
Dissolved gas supersaturation in the thermal
effluent of a power plant can occur due to the
inverse relationship between temperature and gas
solubility in water.When natural waters are at or
near saturation levels with dissolved gases and the
I~
75
RG.1 PiJrrim Nuclear Power Sutioa Unit 1 site plan.Arilbic numerals indicilte wiltet'samplins mtions.
76 Marcello,Fairbanks
channel by Pilgrim Station personnel on April 6,
1973.It was not until April 8,1973,however,that
Pilgrim personnel began to observe several mori-
bund menhaden in the discharge effluent.By
April 9,1973,the number of menhaden observed in
the discharge channel dying and exhibiting signs of
stress characterized primarily by erratic and unco-
ordinated movements had increased substantially,
and as a result the Massachuse"tts Division of Marine
Fisheries was notified that a fish kill was in progress
at Pilgrim Station.With the exception of two adult
menhaden collected on the station's intake screens
on March 15,1973,no menhaden had been recorded
prior to April 6,1973,in the vicinity of the station
during routine biological sampling conducted by
the Massachusetts Division of Marine Fisheries
throughout the winter months.Menhaden do not
usually appear in the Cape Cod Bay region until
May and generally depart by November.
Based on visual observations it was estimated
that several thousand menhaden were in the dis-
charge channel and an additional 75,000 to 100,000
were schooling immediately off the discharge chan-
nel in the thermal plume where water temperatures
ranged between 50°F and 62°F.The fish were
MENBADEN MORTALITY
Following a several-day station outage,the Pil-
grim reactor was restarted and the generator phased
to gridl on April 6,1973.During startup operation
and until full power level was achieved,the tem-
perature of the discharge water increased an aver-
age 0.5°F/hr above that at the intake.Small num-
bersof live menhaden were seen in the discharge
Plymouth,Massachusetts (Fig.1).The Pilgrim nuc-
lear unit is a direct cycle boiling water reactor with
a design power rating of 1998 MWt and a net power
output of 655 MWe.The station uses once-through
cooling with an open channel surface jet discharge
system.Water used for condenser cooling is drawn
from Cape Cod'Bay at a normal rate of about
310,000 gpm which removes about 4.5 x 109 Btu/hr
of heat from the condenser resulting tn a water
temperature increase of about 29°F above the water
temperature at the intake.The cooling water is
returned to Cape Cod Bay via a 900-ft!discharge
channel at velocities varying from about 2 fps
(MHW)to 8 fps (MlW).The general configuration
of the intake,discharge channel and breakwater
jetties os shown in Fig.1.
.-
-
-
approximately 30 to 40 cm in total length and about
500 g in weight.
Menhaden from the discharge effluent dis-
played two apparent external lesions,i.e.,hemor-
rhage and subcutaneous emphysema.Subcutaneous
emphysema were evident between the fin rays of
all body fins especially the dorsal and pectorals
(Fig.2)and within the oral cavity.Subcutaneous
ecchymoses (0.5 to 2 cm in size)were evident in
various body locations but particularly frequent
about the head.In addition,some evidence of pro-
nounced exophthalmos was also observed.Fre-
quently menhaden in the thermal effluent were ob-
served jumping clear of the water and propelling
themselves rapidly along the water surface on their
sides.Just prior to death,the fish became dis-
oriented and gyrated below or on the water surface.
On the basis of the external symptoms displayed
and the aberrant behavior observed,gas bubble
disease was diagnosed as the cause of the menhaden
mortalities.At least 90%of the menhaden in the dis-
charge channel and thermal plume displayed
external symptoms of gas bubble disease.
On April 11,1973,several moribund menhaden
were transported to the University of Rhode Island,
Sea Grant Marine Pathology laboratory for confir-
mation of the gas bubble disease diagnosis and to
ensure that no other infectious disease processes
were involved in the menhaden mortality.Cultures
of kidney,liver,and skin lesions from moribund
and recently dead menhaden were made in Zobells
Marine Broth and cultured at room temperature;no
growth was observed after 48 hr.Sections of gill,
heart,liver,spleen,kidney,pancreas,stomach and
intestine were made and were within normal ranges.
Decalcified cross sections of the cephalic area were
examined for olfactory and lateral line changes and
no significant lesions were noted.Sections of the
brain and meninges revealed congestion and foci of
hemorrhage compatible with vascular damage.
Blood cells were also found free within the ven-
tricles.Based on these findings,Wolke (1973)con-
curred with the gas bubble disease diagnosis and
concluded that the menhaden died from gas emboli
resulting in cerebral anoxia and hemorrhage.
An attempt was made to obtain measurements
of dissolved oxygen and nitrogen of Pilgrim Station
intake and discharge water by gas chromatography.
Dissolved gas levels in replicate samples varied
substantially,however.In addition,comparison of
dissolved oxygen measurements determined by gas
.chromatography with those determined by Winkler
titration (which showed good agreement between
replicate samples)revealed such large discrepancies
that the validity of analyses by gas chromatography
in this instance are questionable and are not
presented.
Dissolved oxygen concentrations of intake and
discharge waters were measured throughout the
period menhaden mortality was occurring.Sampling
1:"
i
,....,
-
-
FIG.2 Subcutaneous emphysema between rays of d01'Q1 fin of adult menhaden (photo courtesy
of R.E.Wolke,University of Rhode Island,Kingston,Rhode Island).
Menhaden Mortality at a Power Plant 77
'Saturations calculated from solubility data reported by Weiss
(1970)for a salinity of 30 ppt.
Wolter DissoI"ed oXygen
Time temperature concentration Soltura6on
Date Stiltion (Iu)(F)(mg/l)(%)
TABLE 1 Dissol"ed Oxysen Concentration and Percent
Soituration of Pilgrn Nuclear Power Stiltion Intake and Discharse
Wolter DurinS the April 1973 Menhaden Mortality
96
97
110
105
109
108
95
127
122
129
142
136
136'
95
125
101
132
134
112
108
129
126
118
103
123
122
136
122
112
109
143
138
137
141
104
104
123
124
120
10.2
7.2
10.4
10.6
10.0
10.2
10.2
11.0
9.8
9.6
10.5
10.6
10.0
10.2
10.2
9.8
10.6
10.0
10.3
9.8
9.9
10.1
10.4
10.9
10.5
10.6
11.2
9.7
9.9
9.7
9.9
9.7
10.2
9.5
9.9
9.5
11.7
10.6
10.1
9.2
8.8
8.8
9.4
9.0
9.1
9.8
9.1
9.2
42.5
62.0
59.5
69.0
63.0
48.5
50.9
73.0
74.7
73.0
73.4
44.4
43.9
57.2
57.2
SO.O
43.9
42.8
55.0
49.5
54.1
49.1
44.5
65.5
65.5
52.5
70.0
70.0
<46.0
74.0
73.0
4<1.7
72.0
n.5
47.0
74.0
73.0
April 18 1 1230
3 1233
3 1400
4 1220
4 1250
April 20 1 1140
1 1530
3 1145
3 1500
4 1123
4 1450
April 21 1 1132
2 1140
3 "10
4 1055
6 1150
April22 1 1420
2 1430
5 1435
7 1445
8 1450
9 1445
April 23 1 1351
3 1345
4 1335
April 24 1 1308
3 1300
4 1253
April 26 1 1323
3 1330
4 1315
April 27 1 0845
3 08S0
4 0840
April 30 1 1230
3 1225
4 1213
April12 1 No sample
3 1340 69.0
4 1350 68.0
5 1530 60.0
April13 1 1227 41.0
3 1250 68.0
4 1300 68.0
5 1400 52.0
April 17 1 1130 43.0
3 1110 68.0
4 1115 68.0
5 1215 54.0
stations are shown in Fig.1.Water samples were
collected just beneath the surface with a Van Dorn
bottle,chemically fixed and analyzed for dissolved
oxygen by the Winkler method (APHA,1965).Water
temperatures at the time of sample collection were
obtained using an ARA electronic thermometer.
Saturation levels were calculated using a solubility
curve from the data of Weiss (1970).
In general,all dissolved oxygen measurements
showed that Pilgrim Station intake waters during
the menhaden mortality were near saturation
(Table 1).Dissolved oxygen saturation levels in the
discharge channel and thermal plume usually ranged
from 120%to 140%.
Mortality of menhaden continued until
April 24,1973.However,after April 19,1973,an
apparent decline in mortality occurred.Estimates
of menhaden mortality rates were made on several
occasions during the observed fish kill.Severely
stressed and dead menhaden were collected on the
surface of the thermal plume during 15-min periods.
Time collections of menhaden averaged 10.7 fish
per period (range of 5 to 17 fish per period)or about
0.7 fish per min.For each fish collected at the
water surface at least two additional menhaden
were observed in distress beneath the water surface
that were not collected.Applying an approximate
minimum mortality rate of about 2.7 fish per min,
an estimated 43,000 menhaden died due to gas
bubble disease during the period April 9 through
April 19,1973.
An attempt was made to decrease gas satura-
tion levels in the discharge water to those believed
to be non-lethal to menhaden (i.e.,less than about
120%and 130%saturation for nitrogen and oxygen,
respectively)based on the tolerance observations
of a related species,Atlantic herring (Clupea
harengus harengus),reported by Stickney (1968).
The Massachusetts Water Resources Commission
(Division of Water Pollution Control)requested that
beginning April 20,1973,Boston Edison Company
reduce Pilgrim Station power level by approximately
50%so that the temperature differential between
intake and discharge water would not exceed 15°F.
During the load reduction,studies of the dissolved
gas concentration and saturation of the station
intake and discharge water would be conducted.In
compliance with the Water Resources Commission's
request,pjlgrim Station power level reduction began
at 2400 hr EST on April 20.1973,and by 0600 hr
EST on April 21.1973,the station output was 300 MW
gross (.1t condenser =14.85°F)and remained
approximately the same until 2145 ...hr on April 22,
1973,when the Water Resources Commission per-
mitted Boston Edison Company to begin increasing
the station power level back to full capacity.During
the station power reduction,discharge dissolved
78 Marcello,Fairbanks
.-
-
--
-
oxygen saturation levels were reduced.Menhaden
in the thermal plume still displayed external symp-
toms of gas bubble disease,but very little mortality
was observed.
Field observations of the menhaden continued
on April 23 and 24,1973;and although the majority
of fish still displayed symptoms of gas bubble dis-
ease,they appeared to be more responsive.ind in
somewhat better condition.
The area around and underthe surface thermal
plume was examined on April 23,1973,utili;zing
SCUBA.A large school of pollock (Pollachius virens)
was observed along the interface between the
ambient water and the thermal effluent.Tempera-
tures where the pollock concentrated ranged
between 44°and 48°F.The pollock appeared in
good condition,were actively feeding,and showed
no external symptoms of gas bubble disease.Other
fishes observed and showing no sign of gas bubble
disease were striped bass (Morone saxatilis),short-
horn sculpin (Myoxocepha/us scorpius),and sea
raven (Hemitripterus americanus).No lobster
(Homarus americanus)were sighted but cancer
crabs (Cancer spp.)and green crabs (Carcinus
maenas)were common and feeding upon decom-
posing menhaden on the ocean bottom.
Attempts to remove the school of menhaden
from the discharge area by commercial fisherman
using purse seines were unsuccessful due to the
presence of several large boulders in the discharge
area and also the high velocity of the cooling water
discharge which interferred with normal methods
of purse seining.The school of menhaden was last
observed in the thermal plume area on April 27,
1973.
Another large school of asiult menhaden (esti-
mated 200,000 to 300,000 fish(was observed in the
Pilgrim Station thermal plume in early July 1973.
The menhaden remained in the vicinity of the sta·
,-tion discharge for about 1 day.During the 1-day
observation,no menhaden mortality was noted,
although a few menhaden were observed propelling
themselves on their sides on the water surface and
at times jumping clear of the water.None of the
several menhaden examined showed external signs
of gas bubble disease.The menhaden were con-
I"""centrated in the thermal plume where water tern·
peratures ranged from 68°F to 82°F.Dissolved oxy-
gen concentrations and percent saturation at the
.-periphery of the school of fish ranged from 9.6 to
9.9 mg/Q and 124%to 146%,respectively.
As of the end of September 1974,no large
schools of adult menhaden had been sighted in the
l~immediate vicinity of the station discharge.It
should also be noted,however,that from October
1973 until August 1974,Pilgrim Station has been at
.-.reduced power or not operating,due to refueling,
maintenance and contested licensing hearings
regarding a change in the fuel design.
DISCUSSION
Since the Pilgrim Station menhaden mortality,
concern over the potential problem of gas bubble
disease at coastal electric generating stations has
increased.Recognizing that fishes are attracted to
power plant thermal discharges which may be
supersaturated with dissolved gases,it has become
obvious that the tolerance of important species to
gas supersaturation needs to be determined.
Such studies on the tolerance of menhaden to
supersaturation at varying water temperatures are
currently being conducted (Clay et aI.,1974).Sub·
sequent experiments will attempt to determine
the recovery rates of menhaden after exposure to
supersaturation as well as determining the impor-
tance of other factors (e.g.physiological stress due
to extreme activity levels)in modifying the toler-
ance of menhaden to gas supersaturation.
Other considerations include the ability of some
fishes to detect and avoid water supersaturated
with dissolved gases.Meldrim,Gift and Petrosky
(1973a and 1973b)found that the behavioral
responses of some freshwater fish to supersaturated
conditions varied with the species.They noted that
the yellow perch (Perca flavescens)showed no
definitive response to supersaturated water while
the silvery minnow (Hybognathus nuchalis),golden
shiner (Notemigonus cryso{eucas),and the satinfin
shiner (Notropis ana/ostanus)were very responsive
to supersaturated conditions.Stickney (1968)
observed that the Atlantic herring (a species related
to menhaden)definitely tended to avoid gas-super-
saturated water,but only when saturation levels
were high enough to produce gas bubble disease.
It is possible that menhaden are also capable
of detecting and avoid ing supersaturated
environments.
The ability of fish to detect and avoid super·
saturated environments may be altered by other
factors,however.Meldrim et al.(1973b)have
reported that the behavioral response of the golden
shiner to supersaturated conditions may change
depending on the water temperature associated
with the exposure.They noted that the golden
shiner usually avoided gas supersaturations that
exceeded 110%.However,when temperature
increases of soc and 10°C are associated with the
supersaturated conditions,temperature preference
of golden shiner overrides avoidance of the super-
saturation.It seems likely that similar behavior
occurred during the menhaden incident described
in this paper,i.e.,the preference for above ambi-
ent temperatures within the discharge channel and
Menhaden Mortality at a Power Plant 79
..~
!
\
I
1
....
--
.-
thermal plume may have negated a menhaden
avoidance of the gas-supersaturated cooling water
dis<:harge.
REfERENCES
APHA.1965.Standard Methods for the Examination of Water
and Waste Water Including Bottom Sediments and Sludges.
Amer.Pub!.Health Assoc.12 ed.
Beiningen,K.T.and W.J.Ebel.1970.Effect of John Day Dam
on dissolved nitrogen concentrations and salmon in the Colum-
bia River.1968.Trans.Amer.Fish.Soc.99:664-671.
Clay,A.,A.Barker,S.Testaverde,R.Marcello and G.C.Mcleod.
1974.Observations on the effects of gas embolism in captured
adult menhaden.In:Proceedings of the Gas Bubble Disease
Workshop,October 8-9,1974.Richland,Washington.
DeMont,D.J.and R.W.Miller.1971.First reported incidence
of gas bubble disease in the heated effluent of a steam gener-
ating station.In:Proc.25th Ann.Conf.S.E.Assoc.Game and
Fish Comm.
Ebel.W.J.1969.Supersaturation of nitrogen in the Columbia
River and its effect on salmon and steelhead trout.Fish.Bull.
68:1-11.
Harvey.H.H.and A.C.Cooper.1962.Origin and Treatment
of a Supersaturated River Water.Inti.Pacific Salmon Fish.
Comm.Prog.Rpt.9,19 pp.
Harvey.H.H.and S.B.Smith.1961.Supersaturation of the
water supply and occurrence of gas bubble disease at Cultus
Lake trout hatchery.Can.Fish.Cult.30:39-47 .
80 Marcello,Fairbanks
Marcello.R.A.and K.Strawn.1973.The Cage Culture of Some
Marine Fishes in the Intake and Discharge Canals of a Steam-
Electric Generating Station,Galveston Bay,Texas.Sea Grant
Report TAMU-SG-72-206.267 pp.
Marsh,M.C.and F.P.Gorham.1905.The gas disease in fishes,
In:Report of the Bureau of Fisheries,1904,p.345-376.
Meldrim.J.W.,J.J.Gift,and B.R.Petrosky.1973a.Responses
of Several Freshwater Fishes to Waters Containing Various
Levels of Gas Supersaturation.Ichthyological Associates,
mimeo report,15 pp.
Meldrim,I.W.,J.J.Gift,and B.R.Petrosky,1973b.Responses
of Several Freshwater Fishes to Waters Containing Various
Levels of Gas Supersaturation.I.A Comparison of Two Differ-
ent Methods for Achieving Supersaturation.II.The Effect of
Increased Temperature Associated with Gas Supersaturation
on the Response.Ichthyological Associates,mimeo report.15 pp.
Renfro.W.C.1963.Gas-bubble mortality of fishes in Galveston
Bay,Texas.Trans.Amer.Fish.Soc.92:320-322.
Stickney,A.P.1968.Supersaturation of atmospheric gases in
the coastal waters of the Gulf of Main.Fish.Bull.67:117-123.
Weiss.R.F.1970.The solubility of nitrogen,oxygen,and
argon in water and sea water.Deep Sea Res.,17:721-735.
Wolke,R.E.1973.Necropsy Report,Accession #8174,Uni-
versity of Rhode Island.Dept.Animal Pathology,Kingston,
Rhode Island.1 p.
Woodbury.L.A.1942.A sudden mortality of fishes accom-
panying a supersaturation of oxygen in Lake Waubesa.Wis-
consin.Trans.Amer.Fish.Soc.71 :112-117.
A.Clay
A.Barker
S.Testaverde
R.Marcello
G.C.McLeod
.rDbservations on
r-the Effects of
Gas Embolism
~
in Captured Adult Menhaden
I'I.
I
";!I
~!
f
'I-,
r
,I-
"I
,1
ABSTRACT
The problems of entrapped fish in effluent water of power plants
prompted a study of the parameters that induce gas embolism in
"'"'adult menhaden.Adult menhaden were captured by purse seining
in early summer on route to the warmer headwaters of Boston
Harbor and maintained in tanks at the New England Aquarium.
Supersaturation of the waters in experimental tanks was de-
.....Iiberately induced and the behavior and certain histological
indices studied at a range of temperature,salinity,and super-
saturation.
The occasion of menhaden mortality in the effluent
_of the Pilgrim Nuclear Power Plant in Plymouth,
Massachusetts (Marcelfo and Fairbanks,1974),
-prompted an investigation of the cause,and a
search for practical solutions to avoid future de-
struction of adult menhaden.If,indeed.supersatura-
tion of gases in the discharge canal of power plants
induces gas embolism in fish,a number of engineer-
_ing solutions are possible.An initial step in develop-
ing such solutions is to rigorously define the range
of environmental conditions that induce gas bubble
disease in a menhaden population.
Unfortunately,investigations of the environ-
mental conditions that induce gas embolism have
been hampered by an inability to hold adult men-
haden in captivity.This may seem strange in view
of the numbers of Atlantic menhaden caught and
processed each year,but realistic in view of the
fact that menhaden are considered a trash fishery
whose main use is for fertilizer and cat food.Thus,
there has not been any emphasis or effort to main-
tain or stock menhaden as a potential sports fishery,
nor even to hold adult menhaden for display or
research purposes.Consequently,a reliable capture
and maintenance program had to be developed.A
project was undertaken that incorporated three
phases:to capture and maintain a stock of men-
haden for testing purposes;to determine lethal
levels of supersaturation at various temperatures;
and ,to identify pathological symptoms at these
saturation levels.
The Atlantic menhaden,Brevoortia tyrannus,
is a pelagic,euryhaline species.It ranges along the
entire eastern United States coastline from Maine
to Florida.The general migration pattern includes a
northward migration in the spring and summer
months from the southern waters.In the early fall,
the schools begin to move from the northern areas
to the southern portions of the coast.Tremendous
schools appear off North Carolina in November and
December,where they remain until late spring
(Nicholson and Higham,1966).
CAPTURE AND MAINTENANCE
Adult Atlantic menhaden were captured by
commercial purse seiners in Boston Harbor during
July 1974.Since menhaden are an easily excitable
fish and once entrapped may become extensively
damaged by contact with the seine,observations-
were made aboard a carrier vessel to determine the
best means of collection.
The purse seine set consists of a carrier vessel
and a purse boat.A spotter plane locates the school
and directs the purse boat in setting the net.When
the school is encircled,lead weights are released to
close the net and trap the fish.The net is hauled
aboard the purse boat with the aid of a hydraulic
power block until the school is concentrated into
the heavily constructed pocket of the seine,called
the bunt.Then the carrier vessel comes alongside
the bunt,the fish are further "dried up,"and
brailed into the fish hold of the carrier vessel.
From our observations,it was determined that
the best time to transfer the menhaden was as soon
as the carrier vessel was secured adjacent to the
bunt.The fish were then transferred from the bunt
with a dip net into an aerated,2.4-m diameter tank
Clay,Barker,Testaverde:New England Aquarium Corporation,
Boston,Massachusetts;with Marcello and Mcleod:Boston
Edison Company,Boston,Massachusetts.
81
occurred in some fish,from bright silver to black,
possibly indicating thermal stress.Reduced feeding
behavior and sluggishness were also noted.Salinity
remained between 30 and 32 ppt,pH was maintained
between 7.5 and 7.8,and dissolved oxygen between
6 and 7 ppm.Ammonia levels rose for 3 weeks to a
maximum of 3.5 ppm until the bacterial population
in the biological filter was established.Values then
steadily declined to a stable level of 0.225 ppm and
the nitrate levels subsequently rose from 0.5 ppm
to a maximum of 2.5 ppm.Nitrite also rose from
0.006 ppm to 1.5 ppm during the same period.
Water changes were made as necessary to keep
nitrate and nitrite levels below 3 ppm NOr and
1.5 ppm NO~.
40 psi
COOLING
BATH
AI R \'RfT
~SATIJRATlON CHAMBER
'U 50 GAlLONS
PRESSURE
GAUGE
BAll VALVE
TESTTANK
145 GALLONS
TESTING PROCEDURE
Fish were removed as needed to perform tests,
and a control was run at each saturation.Twelve
fish were used in each experiment,six in the test
tank and six in the control.The test and control
tanks were 145-gal capacity with the test tank
attached to a 50-gal pressure chamber detailed
in Fig.2.The pressure chamber was constructed
of 60 cm diameter PVC pipe.A 3/4 hp pump provided·
water circu lation through the pressure chamber and
a ball valve on the return line to the test tank pro-
vided control of pressure and water flow.Original
experiments used a venturi method of aeration in
the pressure chamber;however,in later experi-
ments an air compressor was added-to allow for
higher saturations.
The temperature,dissolved oxygen,and dis-
solved gas pressure were measured at regular
intervals during the experiment in both the control
fiG.2 Supers.JItur.tion testing system.
RAPID SAND FILlER
60 GAUMIN
BIOLOGICAL
FILlERS
12 GAUMINJFI LTER
aboard our collecting vessel,R.V.Shrock.Approxi-
mately 150 fish were transported to the holding
facility at the New England Aquarium.
The holding facility (Fig.1)incorporated three
biological filters to break down nitrogenous waste
products and a mechanical filter to remove particu-
late matter.Water was gravity siphoned into the
biological filters,and then air lifted back into the
tank.The mechanical filter pumped water from the
bottom center of the tank through a sand filter and
back into the tank at about 60 gal/min flow.This
return line helps in aeration of the tank as well as
creation of a current which appears to aid in the
schooling behavior of the fish.The walls of the
holding facility were painted black,a technique
which reduces the likelihood offish colliding with
the walls (Hettler).The fish were fed finely grained
food 4 to 5 times a day;total daily intake was
approximately 5%of their body weight.Experimen-
tation with various food sizes indicated that Purina@
tropical fish food gave the most efficient feeding
since it remained in the water column long enough
for maximum consumption.
No immediate losses were noted due to handling
and transfer.However,during the first 2 weeks a total
of 34 fish,or 21:%,died,possibly from the effects of
net damage.Since that time only three additional
fish have died.Standard water quality parameters
were measured on a regular basis.During July and
August the temperature remained about 20°C,but
as cold weather set in the temperature steadily
dropped to noe,and an abrupt color change
FIG.1 ~holdins fKility.
82 Clay.Barker,Testaverde.Marcello,McLeod
-
AGE,YR
%Saturation Observations Pathology
N 2 Total OJ Appearance of gas
bubbles in:
100 95 70 None apparent.None apparent.
lOS 95 75 Some mucus,erratic None apparent.
swimming,body color
change noted in 1 fish,
no deaths in 96 hI.
110 107 102 Some mucus,erratic Eyes.intestines,
swimming,body color pyloric caeca,and
change noted in 2 fish,mesentery.
2 died within 96 hr.
120 110 85 Mucus,erratic Eyes,intestines,
'swimming,'death pyloric caeca,roof
in 24 hr.of mouth,bulbus
arteriosis,fins,
operculum.gill
arterioles.
TABLE 1 Etiology of Gas Bubble Disease in Menhaden
at nce
At 110%saturation nitrogen,behavioral changes
similar to those observed at 105%were noted.
Necropsies showed some evidence of emboli in the
intestines,the pyloric caeca,the operculum,and
the eye.Two fish died within 96hr,one during the
3rd day and one between the 3rd and 4th days.
At 120%nitrogen saturation,death occurred
within 24 hr with classic symptoms of gas bubble
disease displayed.Evidence of exophthalmia was
Gas Embolism in Adult Menhaden 83
RESULTS
The necropsies of control specimens showed
no apparent external or internal symptoms of gas
bubble disease.However,as can be seen in Table 1,
increased nitrogen saturation increased the evi-
dence of gas emboli at 22°C.At 105%saturation
nitrogen,all test specimens lived for 96 hr with no
pathological evidence of gas bubble disease.How-
ever,observations conducted during the course of
the experiment showed definite behavioral changes
in the test fish in comparison with the control fish.
The test fish were excreting mucus,swimming
erratically,and more excitable than the control fish.
Body color change was also apparent on some test
fish.
since capture,fish length ranged from 225 to
310 mm,the average weight was 304 g,46%were
males and 54%were females.The majority of fish,
60%,were age two,20%age three,17.3%age four,
and 2.7%age five.
J
5
o FEMILE
"MALE
T =observed temperature in 0C.
O 2 =oxygen concentration in mg/Q
B02 =Bunsen solubility coeffident for
oxygen
PH 0 =partial pressure of water vapor as
2 a function of temperature
where
anrl AP ==saturometer reading
Patm +AP
Total dissolved gas %saturation =Patm
where Patm =atmospheric pressure at time and
altitude of satu,rometer reading
FIG.3 Menhaden age determination.
%N2 sat ={(Patm +AP)_[02 l22.41 rnJlm mOle-)
[s02 \32.00 mg/m mole
and test tanks.Dissolved gas pressure was measured
with a Weiss saturometer made by Eco Enterprises,
and the nitrogen saturation and total dissolved gas
saturation calculated according to the equations;
As the test fish died,necropsies were performed
as soon as possible.Weight,fork length,sex and
sexual maturation were determined and age esti-
mated using a modification of a graph by C.E.
Richards (1969)(Fig.3).Of the 77 which have died
,~
-
-
,~
apparent in some of the fish.Emboli were also
apparent in all high saturation test specimens in
the pyloric caeca,intestines,eye,operculum,roof
of mouth,epithelium of dorsal fin,mesentery,and
gill arterioles.In two cases the bulbus arteriosis
was greatly distended,apparently by the presence
of an embolus.
CONCLUSIONS
Preliminary results indicated that adult men-
haden could be maintained,with care,for an
extended period of time.Results also showed that
gross symptoms of gas bubble disease could be
induced in menhaden at high levels .of nitrogen
saturation.A probable threshold toxic effect for
adult menhaden exists between 115%and 120%
nitrogen saturation at n°e.further testing at other
temperatures will be necessary to define a relation-
ship between nitrogen saturation,toxicity and tem-
perature.We also intend to explore the tolerance
and recovery capabilities of menhaden to the effects
of intermittent exposure to supersaturation levels
of 120%to 130%nitrogen.
84 Clay,Barker,Testa verde.Marcello,Mcleod
ACKNOWLEDGMENTS
This research project was funded by Boston
Edison Company,Boston,Massachusetts.The
authors gratefully acknowledge the technical advice
of Dr.Alan V.Nebeker of the Western fish.Tox.
lab.,Corvallis,Oregon,the assistance of Captain
John Russo and the crew of the Agatha &Patricia
in collecting the fish,and the assistance of Captain
Rodney Swift and the Research Vessel Shrock.
REFERENCES
Heltler,William F.Personal communication.
Marcello,R.A.,Jr.and R.B.Fairbanks.1974.Gas Bubble
Disease Mortality of Atlantic Menhaden.Brevoortia tyrannus,
al a Coastal Nuclear Power Plant Presented at Gas Bubble
Disease Workshop,Richland,Washington.
Nicholson.W.R.and J.R.Higham,Jr.1966.Age and Size
Composilion of lhe Menhaden Catch Along the Atlantic Coast
of the United Stales,1962;With a Brief Review of the Com-
mercial Fishery.U.S.Fish Wild!.Serv.,Spec.Sci.Rep.Fish.
(527),24 pp.
Reinljes,J.W.1969.Synopsis of Biological Data on Atlantic
Menhaden,8revoortia tyrannus.U.S.Fish.Wild!.Serv.Ore.
(320).
i3as Bubble IR.R.Rucker
-Disease of
r-Salmonids:Variation in Oxygen-
.Nitrogen Ratio with Constant Total
-Gas Pressure
J""':ABSTRACT
Coho salmon fingerlings were subjected to a total gas pressure 01
119%at 13.6°C with the O;;Nl varying from 50%/136%to 229%1
90%.The small fish {3.6 to 6 cm)were the most resistant and the
-larger fish {6 to 10 cm)the least resistant to gas bubble disease
at the gas concentrations used.A drastic decrease in lethal effect
of individual ratios of 02 to N2 occurred between 159%0~/109%
N2 and 173%Ol/105%N2 at the same total gas pressure (119%).
f".IJllll
A review of the literature regarding gas bubble dis-
-ease can be found in a recent publication by Rucker
(1972);one in press by the National Academy of
Science (1972);and an unpublished report by Weit-
""'"kamp and Katz (1973).Most discussions on gas bub-
ble disease have dealt with the inert gas,nitrogen;
oxygen was given a secondary role.It is important
to know the relationship of nitrogen and oxygen
""""when we are concerned with the total gas pressure
in water.Where water becomes aerated at da.ms or
falls,both oxygen and nitrogen are about equally
--saturated when expressed as a percentage.When
oxygen is removed from water by metabolic and
chemical action,or when oxygen is added to the
_water by photosynthesis,there is a definite change
in the ratio of oxygen and the inert gases (mainly
nitrogen with some argon,ett.).This present study
shows the effect of varying the oxygen and nitro-
-gen ratio in water on fingerling coho salmon
(Oncorhynchus kisutch)while maintaining a
constant total gas pressure.
GENERAL EXPERIMENTAL OBJECTIVES
AND METHODS
The primary purpose of these experiments
was to determine differenoes in lethality of various
gas ratios of oxygen and nitrogen at a constant
total gas pressure of 119"k.I also wished to deter-
mine whether there was al difference in suscepti-
bility between sizes and stocks of juvenile coho.
Also to be examined was the effect of reducing the
oxygen while holding the nitrogen constant.
juvenile coho salmon averaging 6 cm in length,
obtained from the Quilcene National Fish Hatchery,
Quilcene,Washington,and the Montlake Labo-
ratory of the National Marine Fisheries Service,
Seattle,Washington,were used during all the tests
concerning differences in lethality of 02/N2 ratios.
During these tests temperatures were 13.6°C.:!:.
0.1°C.Gas concentrations usually varied very
slightly from the desired ratios.The tank facility
consisted of six troughs,two of which were used to
hold experimental fish at normal saturation (100%)
and two pairs of troughs used to test fish at differ-
ent gas ratios.
Control of gas concentrations and the test
apparatus is described in a subsequent section.Dur-
ing initial testing of the gas control system,I deter-
mined that a ratio of 114%02 to 121%N2 could be
achieved by merely allowing air to be sucked into
the intake side of the recirculation pump.Since this
gas ratio did not require injection of either oxygen
or nitrogen,the resultant concentration (114%02
and 121%N2)was used as a quasi control for com-
parison with the other gas ratios.Several replicates
were completed at this concentration.This ratio
and concentration were also used to test for differ-
ences in size and stock and to provide base line
data in determining effect of reduced oxygen con-
centration while maintaining a constant nitrogen
level.
In all the tests free carbon dioxide was found
near normal,about 2 ppm.Oxygen is expressed as
"0/'and the inert gases as "Nt.
Rucker:Coastal Zone and Estuarine Studies Division,National
Marine Fisheries Service,Seattle.Washington.
85
"I
-----------------,,---------------------------------------
TABLE 1 Time 10 Deilth (OilYS)01 Juvenile Coho Sillmon (6 em)
Exposed to 119%TotoJ.l Gis SupersillluroJ.lion oJ.t Different
Leveh 01 ~oJ.nd N1 in 13.6°C Wilter
Rilnge ",,,erilge RoJ.nse Averilse
so 133 1.8 to 1.9 1.9 3.2 to 4.0 3.7
75 131 1.8 to 2.7 2.3 3.5 to 4.3 3.9
114 121 3.2 to 4.1 3.8 6.3 to 7.3 6.9
159 109 3.2 to 5.3 4.5 6.5 to 9.1 8.2
173 105 33.5 to 35.3 34.4
192 100 32.(}1
229 90
WS eoncenntion,
%SiltuRodon
0 1 N)
Effect of Variation in 02/N 2 Ratios
Times to death (lE 25 and LE so)of juvenile coho
salmon at various concentrations of 02 and Nz
during constant total gas saturation of 119%
appear in Table 1 and are shown graphically in
Fig.2.With one exception (192"'A.O 2 and 100%Nz),
all increases in ratio of Oz to Nz resulted in in-
creased tolerance to the total gas saturation.A
marked increase in tolerance to total gas pressure
occurred between concentrations of 159/109 and
173/105%saturation of Oz and Nz (Fig.3).
'Not reached,28%mortality in 39 days.
l()ne replicate reached,24%mortality in 30 days,
the other 25'%in 32 days.
lNot reached,test concluded at 33 days.
'Not reached,20%monality in 35 days.
Effect of Size and Stock
A number of tests were carried out in the water
containing 114%02 and 121%N z to determine effect
of size and stock of fish on susceptibility to gas
supersaturation (Table 2).Two groups of 3.8 cm
coho from the Montlake Laboratory which had just
added to the system.The spout (12)at the top
of the towers was to direct possible overflow water
back into the system.
Inside dimensions of troughs in the fish hold-
ing area (13)were 104.5 x 23.5 x 20 cm high.Water
depth was maintained at 14 cm.Each trough could
be separated into three compartments with screens
-"A"was at the inflow end of the trough,"B"
middle,and "C"outflow end.In a few cases a com-
partment was dividedlongitudinal/y so that two
groups of fish could be subjected to almost identical
conditions.
86 Rucker
The number of days.to kill 25%of the fish at
the different gas levels is expressed as the lethal
exposure-LE2s,and to kill 50%-lEso •
FIG.1 ApsNqIul for subjedns 6sh 10 consunt-Iemperltul'e,
lIowins water wifola definite Clxyg_II'Id nitrosen contenL
TEST APPARATUS
Apparatus shown in Fig.1 was us.ed to produce
water with a definite oxygen and nitrogen content.
The tank (1)was divided so that two experiments
could be carried on simultaneously with the same
equipment.Water was circulated by a centrifugal
pump (2)with a valve (3)on the effluent side to
cause a controlled back pressure as read on a
gauge (4).This created a vacuum on the inflow side
(5)so that air could be introduced into the water
with either oxygen or nitrogen (6)through a "Y"
tube.Circulation of the water caused an increase
in temperature which was maintained at approxi-
mately 13.6°C by means of a refrigeration system
(7)and recorded on a thermograph (8).Water level
in the tank was maintained by float valves (9).
Each trough was supplied with 1 \:per min of water
regulated with flowmeters (10).The water used
was from the municipal supply,was quite soft,
and was passed through activated charcoal to re-
move the chlorine.A greater depth of water was
needed for absorption of the gases than was af-
forded by the tank (1),so two towers (11)were
-
-
.1
I.:
,.
A B C C,C',
Percent oxygen 113 110 105 105 105
Percent total
gas pressure 119 116 117 '17 117
LE zs in days 2.5 3.6 3.6 4.2 5.4
LE so in days 3.3 5.3 5.3 6.6 6.6
TABLE 3 Mortality of 8.5 cm Coho Salmon Subjected to
a Constant Temperature (13.6°qand 121%N,with
a Variation in Oxygen Tension
Trough compartments
produced LE 2s 's of 15.1 and 18.3 days.LE so was
reached in 24.7 and 30 days.Averages of the above
place lEis at 16.7 and lE Iso at 27.4 days.
Five groups of fish,approximately 10 cm long,
from the Montlake laboratory were then tested.
The average for all groups gave an lE'25 of 2.1 days
and an LI:50 of 2.6 days.
Three groups of 10 cm fish from the Quilcene
National Fish Hatchery were similarly tested.Aver-
ages were 2.9 days for lE 25 and 4.2 days for LE'so.
These results indicate that the larger fingerlings
are definitely more subject to harm from excess air
in the water than the smaller fish.These data agree
with those of Meekin and Turner (1974).
Although the data are limited,there appears
to be little difference between susceptibility of the
Montlake and Quilcene stocks.
Effect of Reduced O 2 Concentration on
Mortality Rate
Fish held in compartments in a trough utilize
oxygen so that the w::lter in compartment "c"(out-
flow end)would have less oxygen than in compart-
ment "A"(inflow end).Compartment "8"in the
central part of the trough would have 02 levels
somewhere between those in "A"and "C".Nitrogen
levels in these compartments,however,were the
same.To demonstrate the effect of reduced oxygen
in relation to gas bubble disease,48 coho of 8.5 cm
fork length were randomly distributed into com-
partments "A","B",and "C".Two additional repli-
cates of the "c"compartment tests were run using
32 coho (8.5 cm)in each trial.These are listed as C 1
and C 2 in Table 3.
Inspection of these da ta indicates that when
121'7'0 N 2 is maintained,oxygen plays a more sig-
nificant role above 110%than below 110%.
Some of the data obtained when the oxygen-
nitrogen ratio tests were done also illustrated the
Oxygen-Nitrogen Ratio Variation 87
2.9
4.2
2.1
2.6
'lo02 'N2
A 50 B8
8 15 m
C 114 121
0 1~9 109
E 113 105
F 192 100
C m 9D
27.4
16.7
Not reached
in 30 days
16.9
40
o lE so tJ.
30 tJ.1.£25
:r....
~I
Q
20 NOT RE~CHED-....'0....I
VI ,
>-
~I
I
10 ,
0-4O-~::::::6___6
tJ.-a
t N2 -O 138 131 121 109 105 100
'00 2 -0 so 75 114 159 173 192
TABLE 2 Resistance of Juyenile Coho Salmon to Gas
Supersaturation (114%O'a +121%fll,)in Relation to Size and
Stock of Fish.Lethal exposure leyels (LEu and LE',.)in days
to death,water temperahJre 13.6°C +O.l°C
and test duration 30 days
Days to deat1l1 by size and stock
3.8 em 4.6 em 10 em 10 em t
Montlake Quikene Montlake Quilcene
FIG.3 Relationship between O,ifN a leyels and time to death 01
6 cm coho salmon fingerlings at 13./jOC and total gas concentra-
tion of 119%.
DAYS
started feeding were initially tested.One group of
96 fish reached LE'2s in 22.9 days and LE 3Q after
30 days.The other group of 50 fish reached LE 25 .in
10.9 days.No fu rther losses occu rred until the 27th
day.Lossat 30 days was 34%.Averages of the two
groups place LE 25 at 16.9 days.Average loss at
30 days was 32%.
Two groups of 4.6 cm fish from the Quilcene
National Fish Hatchery were also tested.These tests
FIG.2 Mortality pattern of 6 cm coho salmon reared at different
.-Ol/N1leyeis at 13.6°C with a 119%toltal gas pressure.
....Lethal
exposure
LE 2s
LE so
~
.....
-
"""
......
--
effect of reduced oxygen on the mortality rate.This
was apparent in the experiment using 173%Oz and
105%Nz.At 173%O 2 there were losses of 26%and
30%in 39 days,whereas slightly larger fish at the
lower end of the troughs subjected to 169%O 2 had
losses of only 7%in 39 days.
PATHOLOGY
Generally,the fish died quite suddenly in the
higher nitrogen concentrations.Never was tissue
damage or any progressive pathology demonstrated.
The fish always seemed to die from gas embolism
restricting the flow of blood through the gills.When
the nitrogen was near normal and the oxygen quite
high,the fish were moribund for many days before
succumbing.These fish had blebs in the mouth
which interfered with feeding and caused
emaciation.
SUMMARY
Coho salmon fingeirlings were subjected to a
total gas pressure of 119%at 13.6°C with the Oz/N z
varying from 50%1138%to 229%/90%.The small
fish (3.8 to 6 cm}were the most resistant and the
larger fish (8 to 10 cm)the least resistant to gas
bubble disease at the gas concentrations used.A
drastic decrease in lethal effect of individual ratios
88 Rucker
of Oz to N2 occurred between 159%Oz/10go,-b Nzand
173%Oz/105%N2 at the same total gas pressure
(119%).
ACKNOWLEDGMENTS
Facilities for this work were furnished by the
Western Fish Disease laboratory,Bureau of Sport
Fisheries and Wildlife,Seattle,Washington.The
work was started when the author was an employee
of this organization.
The pathology was studied by William T.
Yasutake,pathologist,Western Fish Disease
laboratory.
REFERENCES
Anonymous.1974.Total dissolved gases (supersaturation).In:
Water Quality Criteria.National Academy of Science Publica-
tion.(In press)pp.135-139.
Meekin,T.K.and B.K.Turner.1974.Tolerance of Salmonid
Eggs.Juveniles,and Squawfish to Supersaturated Nitrogen.
Wash.Dept.Fish.,Tech.Rpt.No.12,pp.78-126.
Rucker.R.R.1972.Gas Bubble Disease of Salmonids:A
Critical Review.Technical Papers of the Bureau of Sport
Fisheries and Wildlife.Number 58.11 pp.
Weitkamp,D.E.and M.Katz.1973.Resource and Litera-
ture Review of Dissolved Gas Supersaturation in Relation to
the Columbia and Snake River Fishery Resources.To North-
west Utilities Cooperative.55 pp.
Eff f G 1M.H.Schiewe-.ect 0 Jas D.D.Weber
-Bubble Disease
on Lateral Line
Function in Juvenile Steelhead Trout
~,
-
..,
Schiewe and Weber:National Marine Fisheries Service,Seattle,
Washington.
FIG.1 Testins ~ppar~tU5 for monitoring qter~1 line response
of steelhead trout.The black disc .tdjacent.to the glass bulb
stimulator is a monitoring hydrophone;and the two electrodes
bene.tth the head 01 the fish are for volume conduction recording
of EKG.
89
MATERIALS AND METHODS
Experimental fish were hatchery-reared juve-
nile steelhead trout,Sa/mo gairdneri,from a
uniform population with a mean length and weight
of 202 mm and 74 g respectively.All fish were
acclimated and tested at a temperature of between
BOC and 15°C.The limited temperature range was
used because of the dependency of saturation on
temperature.
The fish support system,for monitoring both
control and stressed fish,is shown in Fig.1.Equili-
brated water was supplied from a dechlorinated tap
water source.Supersaturated water was generated
by passing water from the same source through a
high pressure pump operated under back pressure.
Mortality of fish resulting from gas bubble disease
has been extensively stud ied and documented (see
review by Rucker,1972).Death results from occlu-
sion of major vessels by gas emboli when the total
dissolved gas pressure of the water environment
(PN z +POz +PAr)exceeds approximately 110%of
atmospheric saturation.In addition,some recent
studies suggest the possibility of gas bubble disease
contributing indirectly to mortality (CoutQnt and
Genoway,1968;Newcomb,1974;Schiewe,1974).
Under close observation,the first visible sign
of gas bubble disease in fish is the presence of gas
emboli in the continuous scale pockets that form
the trunk lateral line.These scale pockets contain
the hair cell receptor mechanism of this sensory
system.Although controversy still exists concerning
the exact function of thl~lateral line system,it is
generally recognized that it responds to near field
water displacements (Harris and Van Bergeijk,
1962).Behavioral studies (Disler,1960;Dijkgraaf,
1962)tend to support the theory that the lateral
line serves as a "distant touch"sensory modality
for predator-prey relatlonsh ips,schooling,and
obstacle localization.
Impairment of a sensory system with such an
important role in fish behavior could adversely
affect survival.This paper reports eledrophysio-
logical studies designed to assess the effect of gas
bubble disease on lateral line function.
~ABSTRACT
Normal lateral line response of juvenile steelhead trout,Sa/mo
gairdneri,to a standardized set of stimuli was compared with
the response of fish affected by gas bubble disease.Electro-
physiological monitoring of individual afferent nerve fibers
showed that as gas emboli formed in the scale pockets of the
trunk lateral line of stressed fish,the ability to respond to
stimuli was either diminished or completely disappeared.Fur-
ther testing demonstrated that this sensory loss is reversible
and that upon return to equilibrated water,accompanied by the
disappearance of the gas emboli,normal function was regained.
This sublethal effect of gas bubble disease on the lateral line
sensory system may be an important element contributing to
indirect mortality.
-
•..''.
o
o
8Trade names are used to simplify description;no endorsement
is implied.
AG.2 Spontaneous activity of sinsle lateral Roe nerwe flberI-
and ~of scales innenated.Dil'"derived from"fish aa:fio.''
IIWltM utd tested ilt 13<><:to 15OC.Resression line (y =2.49'+,4.29X
N =21)p;1I'ilIMh!rs estimaam using reduced IIIiljor axis tech-
niques from Millet'mel Kohn (1962).
l2Dr-------------------,
o
100
20 control fish (maintained on equiHbrated wate~~
and 10 stressed fish which were maintained~'orit
equilibrated water for a short acclimation period<:,.
and then shifted to supersaturated water.·,Signs·Qf:':~
gas bubble disease began to appear in thestressedi f
fish within 2 to 6 hr.This protocol enabled us to'
characterize nerve fibers of fish,whoselateraUirle;
showed three'distinct stages of gas bubble occur.J"
rence:no bubble formation,partial bubble forma,"~
tion,and complete bubble formation:'>.'>'t'i
None of the control fish maintainec:lin equilil',
bra ted water developed gas emboli in lateral line:}
scale pockets.All spontaneously active fibers were?
responsive to light touch,surface waves,and;
pulsed water displacements generated by the glass,'
bulb stimulator.Corresponding sensorY,lulits \v~}~!:
easily located and sponta'ne(jus\iCtivlty,'cor~erated
with number of scales in the sensOry unit.,','':.:';-
The scale pockets of the trunk lateral line of,
stressed fish would progressively fill with gas
emboli until complete occlusion occurred.All fibers
isolated and characterized under a state of com-
plete occlusion did not respond to any form of
stimuli except firm pressure on a limited number
of scales.The number of scales reacting to stimula-
tion did not correspond to the observed spontaneous
activity (Fig.3).
90 Schiewe,Weber
RESULTS AND DISCUSSION,·
Our initialstudH!S indicated that individual
afferent lateral line nerve fibers would 'innervate
receptors in 1 to 26 scale poCkets to form a sensory
unit.The resting or spontaneous activity rate of an
isolated fiber was directly dependent on the num-
ber of scales comprising,that sensory unit (Fig.2).
This fact became a uSE~ful criteria in support of our
findings.
Activity from over 150 fibers was recorded in
Compressed air was metered into the intake side
of the pump to create:the desired saturation level.
Supersaturated water was maintained at 118 .:t.4%
of atmospheric saturation in the test range of HOC
to 15°C.Analysis procedures were the gas chromato-
graphic methods of Swinnerton et al.(1962)modi-
fied by Ebel (1969).
Anesthesia was accomplished with tricaine
methanesulfonate followed by gallamine tri-
ethiodide at the rate IOf 0.6 mg/10 g bOdy weight.
At'opercular arrest the fish was moved to the testing
apparatus where water (equilibrated or super-
saturated)was forced over the gills at the rate of
1 21min.If the heart rate,monitored by volume
conduction EKG,fell below normal for a given
temperature,the experiment waS terminated..'
Water level:in'the-testing apparatus was
adjusted to provide an unsubmerged area 2-to 3 cm
posterior to the operculum.In this area the lateral
line nerve bundle was dissected out and placed over
a silver,spoon-shaped indifferent electrode.Phos-
phate-buffered saline was used to both bathe the
exposed nerve and te>fill the depression of the
spoon.Individual nervle fibers were then teased out
and draped over an uninsulated,electrolytically
sharpened,stainless'steel hook electrode for
recording.
,Initial recording Qif nerve activity during isola-
tion was done with a Tektroni~model 565 dual
beam oscilloscope coupled with a loudspeaker.
Further study and documentation were accomplished
by transfer of the recording to a Tektronix'8
model 5103N storage oscille>st=Ope with a Polaroid'8
camera attachment.
Normal response pattem was based on the
abi,lity of the associated,receptors to'respond to
varying degrees of stimulatiOn.Once a fiber was
isolated,the corresponding :receptor response was
measured by application of light touch,surface
waves,and pulsed wa~er displacements.The pulsed
water displacements were generated by a 1 cm
glass bulb attached t<i.the,cone,of an'8-in.loud-
speaker driven by a WclYeteek8'rnodeI144 HF sweep
generator.During testing .the-bulbwciS',positioned
5 mm above the appropr~~,r~ptorS;.,
:.~:,:.:::,.-
-
,-
"""'
ANTERIOR SUPRASCAlAR PORE
CUPUlA
FIG.4 Juvenile steelhead trout trunk laten!nne showing!
complete gas bubble ocdusion.A..Anterior suprascaw pore.
B.Posterior infrascaqr pore..-
o
o PARTIAL OCCWSION
•TOTAL OCCWSION
4 8 12-16 211
NUMBER OF ACTIVE SCAlfS IN SENSORY UNIT
AG.3 SponQneous activity of single lateral line nene fiben
from gas stressed fish and corrll!5ponding number of scales
dkiting neur..1 ..ctivity.Regression line represents spontaneous
activity of fiben from control fish at 13°C to 15°C and is the
s..me as shown in fig.2.
·131--
100 0
Q
.~~u...lI}!I'"""~
'"...-~(i)~o·
~
'"o~5...«l I·"...~0
i:;8.'".2Il ..0
~·0/0
-
-
-
Twenty-two fibers WE~re isolated and charac-
terized under conditions of partial bubble formation.
Fifty percent responded normally in all respects,
and upon localization of the sensory units no
emboli were visible.The rremaining fibers behaved
as did those fibers isolated when emboli formation
was complete.In addition to the inability to match
spontaneous activity to the number of scales in the
sensory unit,the limited number of scales that did
respond to firm pressure either contained or were -
adjacent to scales with gas emboli._
We additionally isolated and characterized
nerve fibers from three fish that were maintained
3 consecutive days in the test apparatus.On days 1
and 3 support was with equilibrated water and day 2
was with gas supersaturated water.This procedure
provided for an evaluation of any residual effects
the gas emboli may have on the functioning of the
sensory units.Results showed a return to normal
response patterns as the emboli disappeared 16 to
20 hrafter return to equilibrated water.
In fish showing signs of gas bubble disease,
the reduced ability of the trunk lateral line sensory
units to respond to stimuli appears to be mechanical
in nature.Fig.4 is a photomicrograph of a section
of the trunk lateral line of a juvenile steel head trout
that exhibits complete occlusion with gas emboli.
A longitudinal section through this area would
appear as in Fig.5.The physical presence of the
gas emboli probably acts directly on the cupula
rendering it immobile.This cupular immobility pre-
FIG.5 longitudinal section through a tOQlly occluded liateral
line.
vents the shearing displacement action necessary to
stimulat~the hair cell receptors.Although we never
monitored nerve fibers from the numerous cephalic
branches of the lateral line,morphological and
visual observations indicate that these receptors are
equally affected.This being the case,the ability of
an affected fish to detect and localize predators,
or stationary objects may be impaired.This would
have a definite adverse affect on survival.
From mark and recovery experiments,Ray-
mond (1970)estimated that survival of downstream
migrating salmonidsin the Snake River basin
between 1964-68 was nearly 100%.Following dam
construction and resultant high concentrations of .
dissolved gas (during periods of heavy spilling·at
dams)in the water of the lower Snake River,sur~
vival of downstream migrants dropped to 30%.
Considering factors of fish tolerance,behavior.and
gas concentration,Ebel (1973)estimated that 60%
of the total 1970 mortality was due to direct effects
of gas ·bubble disease.Predation and mortality
associated with dam passage accounted for the
remaining 40%of the loss.We believe that a large
proportion of the mortality attributed to these non-
specific causes was probably an indirect effect of
gas bubble disease through impairment of the
lateral line sensory system.
,r
i
,~
Lateral Line Function 91
-
.....
REfERENCES
Coutant,C C.and R.G.Genoway.1968.Final Report on an
,Exploratory Study of Interaction of Increased Temperature and
Nitrogen Supersaturation on Mortality of Adult Salmonids.
Battelle Memorial Institute,Pacific Northwest'laboratories,
Richland,Washington.28'pp.(Processed).
Dijkgraaf,'S.1962.The functioning and significance of the
lateral line organs.BioI.R.ev.38(1):51-105.
Oisler,N.N.'1960.Organy chUVslVsistemy bokovoi linii i Utli
znachenie v'povedenii ryb (Lateral Line 5ense Organs and
Their Importance ;n,Fish Behavior);Izd.Akad.NAUK SSSR,
MoScow.(TransL Israel Program SCi.Transl.,19n).328 p.,
-avail.Nat!.Tech.lnt Serv.,Springfield,VA~as mQ.S4021.
Ebel,W.'J."1969.',,Supersaturation of nitrogen in the Columbia
River andits'effect on salmon and steelhead trotit.fish.Bull.:6B(1):1~11.,-:--,,':'c''7::"
£bel,W}J..1973/Relations between fish behavior,'bioassay
information,and dissolved gas concentrations on survival of
,jUvenile salmon and steelhead trout in Snake River;In:Western
Prog.53rdAnn;Cant..West Assoc.State Game and fish Comm.
.pp..5~6-527..~;.
Harris,G.G.and W.A.van Bergeijk.1962.Evidences that the
92 Schiewe,Weber
lateral line organ responds to near-field displacements of
sound in water.J.Acoust.Soc.Amer.34(12):1631·1841.
Miller and Kohn.1962.Statistical Analysis in the Geological
Sciences.John Wiley and Sons,Inc.New York.483 pp.
Newcomb,T.W.In Press.Changes in juvenile steelhead (Salmo
gain/neri)blood chemistry following sublethal exposure to
various levels of nitrogen supersaturation.J.Fish.R.es.Bd.,
Can.
'Raymond,H.l.1970.A Summary of the 1969 and 1970 Out-
migration of Juvenile Chinook Salmon and Steelhead from the
Snake R.iver.NOAA,National Marine Fisheries Service,Seattle,
Washington.Unpub/.Ms.14 pp (Processed).
Rucker,R.R.1972 Gas Bubble Disease of Sa/monids:A
Critical Review.U.s.Fish.Wild!.Serv.,Bur.Sport Fish.Wild\.,
Tech.Pap.58."pp.
SChiewe,M.H.1974.Influence of dissolved atmospheric gas
on swimming performance of juvenile chinook salmon.Trans.
Am.Fish.Soc.103:n7-721.
Swinnerton,J.W.,V.J.linnenbom,and C H.Cheek.1962
Determination of dissolved gases in aqueous solutions by gas
chromatography.AnaLChem.34:483-485.
I E.Casillas
l.Smith
B.G.D'Aoust
Effects of Stress
on Salmonid
Blood Clotting Mechanisms
-
ABSTRACT
Enhancement of blood dotting functions is a possible factor
which could k~i11 fish following stress resulting from exposure to
supersaturation.During examination of various ...hematological
parameters in rainbow trout which were subjected to stress from
exercise,blood coagulation timeswerefound to decrease to 45%
of the original pre-stress vallJles within a half hour after the
termination of the stress period.Thrombocyte counts were
found to increase three-to fourfold in the same period.Hemato-
crits and blood plasma glucose also rose significantly with
respect to the stress applied.Red blood cell and white blood
cell counts,however,did not increase in response to thestresser.
The degree of the responses observed were compared between
members of a wild trout population from Chester Morse,Wash-
ington,and hatchery-reared Donaldson-strain rainbow trout.
The wild strain showed a mOrE!rapid return to pre-stress condi-
tions than the hatchery-reared trout.Based on very preliminary
experiments,the responses of the clotting mechanisms in Pacific
salmon are similar and probably change even more rapidly than
in rainbow trout.This variability in the blood coagulation rate
is proposed as a mechanism to avoid disseminated intravascular
coagulation (D.I.C.)in the poorly perfused muscles of fish.
Experiments in this area are continuing.
Most of the bioassays regarding gas bubble disease
have ~reported that the incipient lethal dose ot
supersaturation for salmon ids kept in shallow water
is in the vicinity of 120%saturation for expoS,.ures of
more than 24hr (Meekin and Turner,1974).Work
reported by Beyer,D'Aoust,and Smith (this confer-
ence)indicated that 6 cm fish come to equilibrium
with the gas content of its environment in less than
2 hr.Based on these results,maximum lethality
should occur soon after 2 hr of exposure to any
supersaturation level,which however,is only the
case at high levelsuperscuuration.It therefore seems
reasonable to propose that some other mecha-
nism(s)in addition to or instead of simple bubble
formation operate·to cause mortality in fish during
long-term exposure to low levels of supersaturation.
One such factor-blood clots-will be presented in
this report as a possible contributing factor in
making low-level :supers,aturations eventually lethal.
METHODS
Rainbow trout from the University of Washing-
ton hatchery (Donaldson strain)and from Lake
Chester.Morse (an undisturbed watershed 30 miles
east of Seattle)were used in all of the experiments.
A typical trout from the hatchery was about 700 g
in weight,12 to 14 months of age.and reared 0rl a
commercial diet of Oregon Moist Pellets@.The trout
from Lake Chester Morse were of similar size and·
weight,but of unknown age and diet.They were
captured by hook and line and transported to our
.university laboratory in oxygenated tanks.They
'were kept in the lab in running lake water for at
least 2 weeks before being used in experiments.
To perform an experiment,a plastic tube was
inserted into the dorsal aorta and anchored in the
nasal cartilages using the method of Smith and Bell
(1964).MS-222 anesthetic,but no anticoagulants,
was used during the surgery,and then a recovery
period of 48 hr was allowed before beginning an
experiment.Stress was produced by placing a hook
in the caudal peduncle and producing vigorous
activity by the fish for 2 min,after which the fish
exhibited strong signs of fatigue.One blood sample
was taken before the stress period and additional
samples were taken for up to 5 hr afterwards.Blood
samples were replaced with an equal volume of
physiological saline,with the total blood volume
removed by sampling being less than 5%of the
fish's total blood volume.Blood samples used to
determine clotting time were collected into non-
heparinized capillary tubes,while those samples
used for other analyses were collected into hepa-
rinized tubes.Analyses included determination of
red and white cell counts,thrombocyte count,
hematocrit,and plasma glucose concentration.'.
Control fish were treated exactly the same as
the experimental fish except for the omission of the
period of stress.·
RESULTS
The decrease in blood clotting times for the
stressed fish are shown in Fig.1 and are nearly
the same in both strains of fish,except that the
----.,-.--..
Casillas and Smith:University of Washington,Seattle,Washing-
ton;and D'Aoust:Virginia Mason Research Center,Seattle,
Washington.
93
T
I
R=4--------
T
I
~LDSON STAAIN AAINBOW TROUT
lAKE QIESlER MORSE:RA INBOW'TlIQJT
llO
TIN(IMINJTESI
,STRfSS
1
fIG.]Effect of stress ;and repeilled Ampling on ,lhe number
of drculilling thromboc:ytesin the wild trout;_messed '
group;-'-,-control groUp.Each point represents ,the meiln i~
III
TIN(IMIMlTESl
3<>+211~;:
""<:;;+10
~
~0
~u
#,-10
0-
0:
~~-211
:I:
fiG.'4 Percent cMnse of hem;alocrits from initW levels in.
response to stress for iI hiltchery boUt;-stressed group;.
--control poup.E.Jch point represents the meiln .±.Le.
fiG.5 Percent dNtnse of hem;alocrits from 'initYI IeftIs in~;',:,
response to,stress '01 ..wild trout;---stressed group;;.);
--control poup.Uch point represents the moo .±.s.e.Li,i
Asterisk (-)indialtes sipifiunt difference from control
~~=D.05).
--------
IllR\l.J)SCIIj STRAIN RAINBOW TlIQJT
131
nr.t:IMlfIJID)
lID
nNE aullJ1(Sl
94 Casillas,Smith,D'Aoust
hatchery trout maintain a hypercoagulable condi-
tion for at least 5 hr.The corresponding increase
in numbers of thrombocytes is shown in Fig.2 and
3.While the hematocrits increased (Fig.4 and 5),
neither the counts of red cells nor white cells
increased,suggesting that the increased hemato-
crits resulted from increased cell volume.This
phenomenon has been observed before (Sovio and
Nyholm,1973)when red cells were exposed to low
Ozlevels.
Increases in plasma glucose have been identi-
fied as an indicator of the degree of stress in humans
and also in rainbow trout (Wedemeyer,1972).The
fiG.1 Effect of stress and repeilled Sillllpling on blood dotting
limes in iI hiltcherytl'OlIt (lop)ilnd iI wild b'out (bottom);
-stressed group;--conlrol group.E.Jch point represents
the mem .±.s.e.Asterisk ('")indiates signifkilnt difference from
initiill levels (P =0.05).
fiG.'2 Effect of stftss ;and repe.1Ited s.ampIins on the number
of circulatins dtromlJoqtles in iI hAtchery trout;--stressed
poup;_conbol poup.bdI point represents the mean.±.s.e.
Aslerisk (-)indicilks sipifiGlnt diffeleuce from initiiII Ieftls
[P =US).
DQN,f,LDSlW SlV.IN AAINBOW TROOT 81
210 Q lAKE O£SlER MORSE AAINBON TROOT
T §ro
u;190 I
Q n.3 -'CD 60~I ~
u 170 ---!-5!1...<!;!!!150 1'l 41~..;:III '"~110 s:~u0
90 CD:::eu0 10
Q ""§:I:0-0...Ill)
TIME tMlNJTESI
~-,
....
ACKNOWLEDGMENTS
Contribution number 408,College of Fisheries,
Seattle,Washington.Supported by NIH Grant
#Hl 16254-02.
REFERENCES
Meekin,T.K.and B.K.Turner.1974.Tolerance of salmonid
eggs,juveniles,and squawfish to supersaturated nitrogen.
In:Nitrogen Supersaturation Investigations.in the Mid-
Columbia River,Wash.Dept.Fish.Tech.Rpt.12,pp.78-126.
Smith,L.S,and G.R.Bel!.1964.A technique for prolonged
blood sampling in free-swimming salmon.J.Fish.Res.Bd.,
Can.21(4):711-717...
Sovio,A.and K.Nyholm.1973.'Notes on hematocrit deter-
minations on Rainbow Trout,Sa/mo gairdneri.Aquaculture 2:
31-35.
Wedemeyer,G.1972.Some physiological consequences of
handling stress in the juvenile coho salmon (Oncorhynchus
kisutch)and steelhead trout (Sa/mo gairdnerl).J.Fish.Res.
Bd.Can.29:1780-1783.
of the tissues supplied by the vessel and the dura~,.
tion of the bubble,the fish may die quickly,at the;:
one extreme,or not die at all,at the other extreme""
Once a blood vessel is blocked by a bubble,itis;
also highly.probable'that a clot will form there.':
The likelihood of clot formation and having the c10f
spread further once it has begun would be greatly
increased,we believe,by the presence of the clotting "
enhancement response induced by the shock of
bubble formation.The problem .caused by clotting
is further aggravated by the slow rate of blood flow
found in the white muscle of fish.Once a,clot is
formed,the severity of the resulting damage to
tissue due to intravascular'coagulation increases.
Such dots might take a week or more to be removed.
Once theblood vessel is blocked long enough;the
tissue downstream from the'blockage dies,with the
degree of importance depending'again on which
tissue is involved.We believe it is significant that
many of the juvenile coho which were used in our
decompression studies and which survived the
decompression with few or no symptoms of gas
bubble disease died a week later with severe tail
rot.
Finally our results suggest that the mechanisms·
involved in the pathology and mortality associated
with formation of bubbles in fish chronically
exposed to low-level supersaturation are more com-
plex than can be simply explained by simple bubble
formation.The role of sublethal changes by the fish
in response to supersaturation may eventually be
shown to be as important to the survival of Columbia
River salmon as the lethal effects which have been
the subject of most recent studies.
300
-==-f
lAKE CHESTER MORSE RAINBOW TROUT
lll)
1'111£lMINUTES)
010 30 60
70
60 !y[,L't-_l--_-,~
T .r
E so -·--...T'"-1-- - - - - - - - - - - ---- - --l~r-r ..cr-n=4 i§«l'----!-'-........_...J....__.........__--'-.l-J
~...on
:3 IlII
3
u 90
Q
9...
FIG.6 Effect of stress and repeated sampling on blood glucose
levels in a hatchery trout (tll)p)and a wild trout (bottom);
-stressed group;--control group.Each point represents
the mean .t.s.e.Asterisk (*)indicates significant difference from
initial level (P =0.05).
DISCUSSION
Our results clearly demonstrate the presence of
increased blood clotting rates in salmon ids after
periods of stress.The presence of increased num-
bers of thrombocytes at the same time was also
shown,but whether or not they cause the change
in clotting function is not clear.However,their
greater availability would enhance the ability of
the clotting system to IProduce the initial "plug"
of cells at a woiJnd site 1:0 minimize any blood loss.
The increases in blood glucose during the experi-
ments showed that the degrees of stress imposed on
the fish were physiologically significant.The
changes in blood '..glucose may also indicate that
hatchery fish could be Iless resistant to stress than
wild fish.,
,The significance of the blood clotting system in
relation to the gas bubble problems of fish is still
speculative and largely extrapolated from what is
known about clotting problems in people.It is
highly probable that the major immediate effect of
bubbles arising from external supersaturation is to
block blood vessels.Depending on the importance
changes in plasma glucose are shown in Fig.6 and
verify that a physiologically significant stress was
imposed on the fish.Another difference between
the hatchery and wild fish was that the blood glu-
cose in the wild fish returned to normal after 3 hr
and stabilized there,while the blood glucose for the
hatchery trout remained unstabilized for the dura-
tion of the sampling periiod (5 hr).
50
«l'--_...I....L"""-"_--l-__,--'-__---L .J...J
,J!I'IIllilIII
I~
Blood Clotting Mechanisms 95
Newcomb:National Marine Fisheries Service,Seanle,W
ington.,'
·B.G.D'Aoust,Reseaf(:h Biologist,Hyperbaric laborato
Virginia fl,1ason Research Center,Seattle.Washington.
MATERIALS AND METHODS '".,
The fish used in these experiments were,st
head trout,Sa/mo gairdneri,provided .by the;W
ington State Department of Game Hatche,'
point potential long-range problems encountered .
by the juvenile sa,lmonid such as ingestion and'
assimilation of food,predator avoidance,locomo-"
tion,homing behavior,fecundity,and the quantity'"
and quality of subsequent fertilized eggs andfry~~
In addition,measuring these blood chemistry char:.."<!·
acteristics may allow in situ evaluation of nitrog~tr·
gas supersaturation stress response both singIY'ilnct~:
in combination with other normally-occurringi!
stressors (e.g.passage through turbines)which·
occur to the juvenile salmonid on its downstr~~.'
~•:-.'_J'~t;..migration.5'1.'
This blood chemistry experiment represe:nts;\
an extension of a bioassay by Dawley and Ebel·(M$)j;:
which was designed to provide a variety of lethal;:?
and sublethal biological criteria which would her'
form the basis for an EPA provisional water qua~,
standard.The primary goal was the establishm,"
of a series of lDso values.Secondary goal.s:W
changes in voluntary swimming performance,·::
blood chemistry characteristics which migh(',g
some estimate of sublethal stress response to"
bubble disease.'"~
•T.W.Newcomb
96
One major water pollution problem of the Colum-
bia and Snake Rivers is atmospheric gas super-
saturation which produces gas bubble disease in
fish.
Gas emboli form in the fish's supersaturated
blood in some as yet undefined mannerJD'Aoust,·
personal communication).Gas emboli·may then
lodge at a variety of sites throughout the fish's body.
These emboli can then limit delivery of oxygen
and removal of camoodioxide and toxic metabo-
li,es from the.affected tissues and organs.Persis-
tence of this physiological condition will lead to the
ultimate death of the fish.
A number of mortality studies with nitrogen
gas supersaturation have been made on juvenile
salmon ids,but little work has been published on
the sublethal effects (Dawley and Ebel,MS).Sub-
lethal effects from gas supersaturation may cause
alterations in the fish's blood chemistry which can
be used as tools to determine the degree of stress.
These blood chemical measurements may help pin-
ABSTRACT
Groups of juvenile steelhead trout (Salmo gairdneri)were exposed
for 35 days to various (103.105.110,and 116%sublethal
nitrogen plus argon saturations.Pooled serum samples were
analyzed for Ca,Na.PO.,K.CI,albumin,total protein,choles-
terol.alkaline phosphata~~.glucore,urea,uric add,total bili-
rubin,lactate dehydrogenase and serum glutamic oxalacetic
transaminase.An increase in serum potassium and phosphate,
and a decline in serum albumin.calcium,cholesterol,total
protein and alkaline phosphatase were noted in steelhead ex-
posed to 116%nitrogen (N z +Ar)saturation (total atmospheric
gas saturation 110%).No major changes in blood chemistry were
obrerved at nitrogen satufOltions of 110%or less.
Changes in
Blood Chemistry
of Juvenile
Steelhead,Sa/rna gairdneri,
Following Sublethal Exposure
to Nitrogen Supersaturation
"""
,~-
....
Aberdeen,WA.They weighed 20.S i 6.3 g and mea-
sured 132 .!:.17 mm in fork length at the completion
of these tests.Fish were acclimated to water tem-
peratures of 1S.0iO.SoC (SOC per week with 2 weeks.
at 1S°C)in acclimation tanks described by Ebel
et al.(1971),and Dawley and Ebel (MS).Fish were
fed once each day a maintenance ration of Oregon
Moist Pellets®.
Fish were exposed to various nitrogen super-
saturation levels in shallow rectangular tanks
(Dawley and Ebel,MS)with water 23 em deep Jrom
the same source as used in the acclimation tanks.
Tests were performed at 15.0.!:.0.S6°C at nitrogen
gas (N 2 +Ar)saturations of 103.5,105.4,110.1,and
116.0 .!:.2.0%as calculated from Weiss (1970).These
nitrogen saturations corresponded ta~total ga~
pressure (TGP)Saturations (N 2 +Ar +O 2)of .99.9,
102.0,105.9 .md 110.2%.Mean oxygen saturations
ranged between 87.5 and 90.5%.fish were continued
on the same maintenance ration of Oregon Moist
Pellets®during these tests.
Sixty fish were taken concurrently from the
acclimation tanks and placed in each of three
experimental and three control tanks (102.0 versus
99.9;105.9 versus 99.9;and 110.2 versus 99.9%TGP).
Tests at each saturation level were continued until
35 days had elapsed (the 3S-day'test period was
chosen to parallel the average exposure time of
seaward migrant steelhead passing downstream
through the nitrogen srupersaturated waters of the
lower Snake and Columbia Rivers).At this time,
each fish was given an individual voluntary swim-
ming performance test (2 to 5 min)in an inclined
vee trough against a water current of 1.28 m/sec
(15.0 .!.0.5°C)(Schiewe,in press). A variety of
experimental limitations dictated that these fish
must be used in the !iwimming performance test
before being used in the present blood chemistry
work.Therefore,to minimize short-term blood
chemistry changes due to the swimming challenge,
each experimental and'control group was placed
in a control tank for 24 hr before blood sampling
(Grant and Mehrle,1972;'WeUs,1932;Nakano and
Tomlinson,1967;and Hill and Fromm,19GB).
Fish were then individually anesthetized with
neutralized .MS-222 .(tricaine methanesulfonate)
(Wedemeyer,1970a)'~ina concentration sufficient to
reach plane three anesthesia (Klontz and Smith,
1964),[in about S min).The fork length of each fish
was measured.Blood ,/Vas then collected from the
severed caudal peduncle and processed after the
method of Miles and Smith (1968)except that plain
hematocrit tubes were used to obtain serum.These
fish were then weighed and examined for signs of
gas bubble disease (Dawley and Ebet MS).Indi-
vidual serum samples were pooled within each ex-
perimental or control group and immediately sealed
and frozen at O°C until chemical analysis was
performed.Pooling was necessary to make up the
minimum sample volume needed for the following
analysis.
A Technicon®SMA 12-60 (Sequential Multiple
Analyzer)was used to determine serum calcium,
phosphate,glucose,urea,uric acid,cholesterol,
total protein,total bilirubin,alkaline phosphatase
(AP-ase),lactate dehydrogenase (lDH)and glutamic
oxalacetic transaminase (SGOT).All samples were
run sequentially with a reference synthetic serum
(General Diagnostic's Calibrate Automated
"lock-in"),while three control synthetic sera
(General Diagnostic's Calibrate)were used to cali-
brate the output of each of the 12 channefs.Mea-
surement error was less than 1.0%with the excep-
tion of cholesterol and LDH which·were 1.2%.
Serum calcium was determined by the method of
Kesler and Wolfman '(1964)as modified by Techni-
con®.Inorganic phosphate was detected by a
,Technicon®modification of the method of Fiske and
.Subbarow (1925).Glucose was detected by a
modification of the procedures of Brown (1961),
and Bittner and McCleary (1963).Urea nitrogen was
determined by a modification of the method of
Marsh et al.(1965).Uric acid is measured by a
Technicon®modification of the cupric-neocuproine
procedure originally described by'Bittner et al.
(1963).A Technicon®modification of the Lieberman-
Burchard reagent was used in the direct determina-
tion of serum cholesterol.Total protein was
quantitated by the Technicon®modification of the
biuret reaction.The HABA anionic dye {[(2-(4'
-hydroxyazobenzene)benzoic acid)]J used in the
Technicon®SMA 12160 was unusable for salmonid
albumin,and a manual procedure using the brom-
cresol green method of Doumas et al.(1971)was
substituted in these experiments.The method for
the estimation of total bilirubin is based on a Tech-
nicon®modification of the procedure of Jandrassik
and Grof (1938).Alkaline phosphatase was deter-
mined by a modification of the King-Armstrong
procedure developed by Marsh et al.(1959).lDH
was measured by a method based on the procedure
of Hochella and Weinhouse (1965).SGOT was
quantitated after the procedure of Morgenstern
et al.(1966).
Serum sodium'and potassium were measured
ona Corning Model 170 digital flame photometer,
while serum chloride was determined by the method
of Schales and Schales (1941).Correlation coeffi-
cients were calculated for all of the above blood
characteristics along with oxygen,nitrogen and
TGP saturations,and mean wet weight and fork
length.
.Water analysis for n itrogenand oxygen were
performed daily in each test tank using the methods
Changes in Blood Chemistry 97
DISCUSSION
It is important to distinguish between blood
chemistry changes due to chronic as opposed to
acute (short-term)stressors (Wedemeyer,1970b).
Much of the literature on salmonid blood chemistry
is concerned only with acute stressors;those applied
over 1 hr to 1 week (Cardwell et aI.,1970;Miles
and Smith,1968;and Wedemeyer,1970,19n,and
1972).Because no mortality occurred over the 35-day
test at 116%nitrogen (110%TGP),the author only
considered chronic stressor responses in this discus-
sion.The chronic stress noted at 110%TGJ1is
particularly significant because 110%TGP saturation.
is the level presently suggested by EPA as apro~
visional water quality standard (Rulifson,·personal
communication).,"
Two mechanisms have been invoked in an
attempt to explain gas bubble disease.These
mechanisms are gas embolism of the heart,gills
or other vital portions of the blood system (Marsh
and Gorham,1905;and Dawley and Ebel,MS},a.nd
starvation due to blebs in the lining of the mouth
preventing ingestion of food (Dawley and Ebel.M~l.,(,~
A 50%reduction in serum glucose was indicat~\~:
by Robertson et at,1962,when rainbo.....troutw~*.:'
starved.Similarly,albumin (Booke,1964)and total
protein (PhiJIips,et,al.,1960)decreased in ;prook "
trout,Sa/ve/imJs fontina/is.,,_-'1'"'-"".
Emboli may be enviSioned to block arteries::~!:l,(r-',
veins in a variety of critical tissues and'or8~~,
which would result in local or systemic hy~~,<
respiratory acidosis and necrosis from the buildup ,.>'
of toxic wastes.The gills,brain,heart and kidney C:':',
are four such critical sites.Short-term hypoxia was
In addition to potassium,a decrease in albumin
was negatively correlated with an increase in
phosphate (r=-0.62;P <0.05),and positively corre-
lated with decreases in cholesterol (r =0.97;P <0.01)
and total protein (r =0.90;P <0.01).'
No correlation (P <0.5)could be drawn between,
oxygen,nitrogen or TGP saturation and mean weight
or length..
A 46%incidence of external gas bubble disease
signs were noted in steelhead exposed to 116.0%"
nitrogen saturation,while no external signs were
observed at lower saturations.These signs were,
largely in the form of -lateral line bubbles with a'
lower incidence of blebs appearing in the dorsal-
and caudal fins.No mortalities occurred
these tests (Dawley and Ebel,MS).
~~~~,;;~~~Ii<on
s!!!!gi §~§
i:illIWlJ #K5
~1(,0
~
~1.31 ~us
'"'"~tOO
0
iE 0.0
!LO ~U.,.;z-
go 10.0 E (,0
::e ::e
::>:::>
§9.0 ;;:;3.S'"<<u 6&.0 ~10
RESULTS
Alteration from control mean values (>30)were
observed in 7 of 16 blood characteristics in juvenile
steel head exposed'to 116.0%nitrogen saturation,
while meaningful changes _in these 16 substances
were ,not noted'illt .rOwef:Saturations (Fig.1).'An
increaSe in concentratiOns,of potassium arid phos-
phate were noted at 116.0%nitrogen saturation,
while decreases in concentration were noted in
albumin,calcium,cholesterol,AP-ase and total
protein.Decrease iin serum calcium was positively
correlated with delcreases in cholesterol (r =0.86;
P <0.05)and albumin (r =0.95;P <0.01),and
negatively correlated with an increase in potassium
(r =-OJ?;P <0.05).
#15e.,.
z 3.0~g:2.5
gz.o l-J",......I-JL..-L~~_.....-
of Beiningen (197:3)and Ebel (1969).Weekly mea-
surements of total alkalinity (16 to 20 ppm),total
hardness (10 to 21 ppm),carbon dioxide (1.2 to
2.0 ppm),chlorine~T chloramine «0.02 ppm)and
pH (6.8 to 7.3)were made according to Tarus et al.
(1971).Calcium (6.4 to 6.7 ppm)and potassium
(0.3 to 1.0 ppm)were determined monthly by flame
spectrophometer (Dawley and Ebel,MS).
.-
L
-
l
1
1
1
1
1
1
1
I""'"
I
HGo 1 ,..."serum c1"-'isay ~with i~
N 2 +N salurMions.Conlb'Ol (1~N 2 +Al)...1IeS _mHII
and nnce.
98 Newcomb
--~.'".
-
,~
-
-
-
shown to cause severe hemoconcentration,a dou-
bling of serum phosphate,a 600-b increase in serum
glucose and an uncomp1ensated respiratory acidosis
in menhaden,Brevoortiill tyrannus (Hall et aI.,1926).
Extreme hypoxia in the brown bullhead,Ictalurus
nebulosus,and the American eel,AnguiJ/a rostrata,
in fresh water,resulted in severe albuminemia
(Bieter,1931).Hunn (1969)found that rainbow trout
subjected to hypoxic stress showed a doubling of
plasma phosphate.
Juvenile steelhead stressed for 35 days at 116.0%
nitrogen saturation show no significant decrease
in mean weight or length when compared to other
test groups.These data would tend to support the
hypoxia argument at thl~expense of the starvation
proposition.Hypoxia should show anjncrease in
serum cholesterol (Wedemeyer,1970a),but the
present data suggest that cholesterol in fact declines
in steel head exposed to 116.0%nitrogen saturation.
While steelhead exposed to 116.00-6 nitrogen satura-
tion alter their serum concentrations of phosphate
(Hunn,1969)and calcium (Wedemeyer,1971)in a
manner consistent with the hypoxia thesis,compar-
able data for the starvation interpretation is missing.
In reciprocal fashion,decreases in albumin and total
protein are known (Phillips et aI.,1960)following a
period of starvation,but comparable data for
chronic hypoxia is needed.Steelhead serum potas-
sium and AP-ase levels need to be determined for
both chronic hypoxia and starvation.
Further work will be necessary to ascertain
whether these signs reflect hypoxia and tissue
necrosis,starvation or some generalized.stressor
response.
REfERENCES
Beiningen,K.T.and W.].Ebel.1971.Dissolved Nitrogen,
Dissolved Oxygen and Related Water Temperatures in the
Columbia and Lower Snake Rivers,1965-1969.NOAA,National
Marine fisheries Service,Data Rpt.No.56,60 pp.
Bieter,R.N.1931.Albuminuria in glomerular and aglomerular
fish.J.Pharm.and Exper.TIlerap.43(3):407-412.
Bittner,D.l.,S.G;Hall and M.L.McCleary.1963.A method
for determination of uric acid,using the curpic-phenanthro/ine
indicator system.Am.J.Clin.Path.40:423-24.
Bittner,D.L.and M.L.McCleary..1963.The cupric-phenan-
throline chelate in the determination of monosaccharides in
whole blood.Am.j.Clin.Path.40:423.
Booke,H.E.1964.Blood serum protein and calcium levels in
yearling brook trout.Progressive Fish Cull.26:107-110.
Brown,M.E.1961.Ultracmicro sugar determination using 2,9-
dimethyl -1,10-phenanthroline hydrochloride (neocuproine).
Diabetes.10:60-62.
Cardwell,R.D.,J.B.Saddler and L.S.Smith.1970.Hemato-
logical effects of Dennison tagging upon juvenile pink salmon
(Oncorhynchus gorbuscka)..Compo Biochem.Physio/.38A:
497-508.
Dawley,E.M.and W.J.Ebel.Lethal and Sublethal Effects of
Various Levels of Nitrogen and Argon Supersaturation on
juvenile Chinook Salmon and Steelhead Trout.National Marine
Fisheries Service.(Ms.in preparation).
Doumas,B.T.,W.A.Watson and H.G.Biggs.1971.Albumin
standards and the measurement of serum albumin with
bromcresol green.Clin.Chim.Acta.31 :87.
Ebel,W.J.,E.M.Dawley and B.H.Monk.1971.Thermal
tolerance of juvenile Pacific salmon and steelhead trout in
relation to supersaturation of nitrogen gas.Fish.Bull.69(4):
833-843.
Fiske,C.H.and Y.5ubbarow.1925.The colorimetric deter-
mination of phosphorus.}.Bioi.Chern.66:375-400.
Grant,B.F.and P.M.Mehrle.1973.Endrin toxicosis in rain~
bow trout (Salmo gairdneri).).Fish.Res.Bd~Can.30:31-40.
Hall,F.G.1928.Blood concentration in marine fishes.}.Bioi.
Chern.76(3):623-631.
Hill,C.W.and P.O.Fromm.1968.Response of the interrenal
gland of rainbow trout (Sa/mo gairdneri)to stress.Gen.Compo
Endocrino!.11:69-77.
HocheIla,N.J.and S.Weinhouse.1965.Automated assay of
lactate dehydrogenase in urine.AnaL Biochem.13:322-35.
Hunn,J.B.1969.Chemical composition of rainbow trout
urine following acute hypoxic stress.Trans.Amer.Fish.Soc.
98(1):20-22.
Jandrassik,l.and P.Grof.1938.Vereinfachte photometrische
methoden zur bestimmung des blutbilirubins.Biochem.Z.297:
81-89.
Kesler,G.and M.Wolfman.1964.An automated procedure
for the simultaneous determination of calcium and phosphorus.
Clin.Chem.10:686-703.
Klontz,G.W.and L.S.Smith.1968.Methods of using fish as
biological research subjects.In:Methods of Animal Experi-
mentation,Vol.Ill.Ed.by W.J.Gay,New York:Academic
Press,pp.333-385.
Marsh,M.C.and F.P.GOTham.1905.The gas disease in
fishes.In:Report of the Bureau of Fisheries,1904,p.343·376.
Marsh,W.H.,B.Fingerhut,and E.Kirsch.1959.Adaptation
of an alkaline phosphatase method for automatic colorimetric
analysis.Clin.Chem.5:119-126.
Marsh,W.H.,B.Fingerhut,and H.Miller.1965.Automated
and manual direct methods for the determination of blood
urea.Clin.Chem.11 :624-27.
Miles,H.M.and L.5.Smith.1968.Ionic regulation in
migrating juvenile coho salmon,Oncorhynchus kisutch.Compo
Biochem.Physiol.26:381-398.
Morgenstern,S.,M.Oklander,J.Auerbach,J.Kaufman,and
B.Klein.Automated determination of serum glutamic:oxal-
acetic transaminase.Clin.Chem.12:95-111.
Nakano,T.and N.Tomlinson.1967.Catecholamine and carbo-
hydrate concentrations in rainbow trout (Sa/mo gairdneri)in
relation to physical disturbance.j.Fish.Res.Bd.•Can.24(8):
1701-1715..
Phillips,A.M.,Jr.,H.A.Podoliak,D.L Livingston,R.F.Dumas,
and R.W.Thoesen.1960.Calcium metabolism of brook
trout.Fisheries Research Bul/.No.23 (Cortland Hatchery Rept.
No.28).Conservation Dept,New York State,Albany.
Robertson,O.H.,S.Hane,B.C.Wexler,and A.P.Rinfret.1963.
The effect of hydrocortisone on immature rainbow trout.Gen.
Compo Endocrinol.3:422-436.
Schates,O.and S.S.Schales.1941.A simple and accurate
method for the determination of chloride in biological fluids.
}.Bioi.Chem.140:879-884.
Schiewe,M.In press.The influence of dissolved atmospheric
gas on the swimming performance of juvenile chinook salmon.
Trans.Amer.Fish.Soc.
Taras,M.J.,A.E.Greenberg,R.D.Hoak,and M.C.Rand (Eds.).
1971.Standard Methods for the Examination of Water and
Wastewater.Washington:Amer.Pub.Health Assoc.874 pp.
Wedemeyer,G.A.1970a.Stress of anesthesia with MS-222
and benzocaine in rainbow trout (Sa/mo gairdneri).}.Fish.
Res.Bd.,Can.27(5):909-914.
Wedmeyer,G.A.1970b.The role of stress in the disease
Changes in Blood Chemistry 99
resistance of fishes.In:A Symposium on Disease of Fishes
and Shellfish,Ed.by S.F.Sniezko,Amer.Fish.Soc.Special
Pub!.No.5.Sept.,1970.pp.30-34.
Wedemeyer,G.A.1971.The stress of formalin treatments in
rainbow trout (Sa/roo gairdneri)and coho salmon (Oncorhyn-
chus kisutch).}.Fish.Res.Bd.•Can.28(12):1899-1904.
Wedemeyer,G.A.1972.Some physiological consequences of
handling stress in the juvenile coho salmon (Oncorhynchus
100 'Newcomb
kisutch)and steelhead trout (Sa/mo gairdneri).}.Fish.
Res.Bd••Can.29(12):1780-1783.
Weiss,R.F.1970.The solubility of nitrogen,oxygen and argon
in water and seawater.Deep Sea Res.17:721-735.
Wells,N.A.1932.The importance of the time element in the
determination of the respiratory metabolism of fishes.Proc.
Nat.Acad.Sci.,Wash.18:580-585.
,-
Continuous IT.F.Jenkins
Monitoring
of Total Dissolved Gases}
a Feasibi Iity Study
-
ABSTRACT
A preliminary investigation was undertaken to determine jf a
continuous analyzer could be configured to monitor dissolved
gases in natural waters.A three-component system was designed
consisting of a pumping system,a continuous stripper,and a
detector.Prototypes of the first two components were assembled
and evaluated under field conditions.Based upon these results,
it is possible to configure an unattended,near-continuous
monitor to measure total dissolved gas concentration in natural
waters.
The symptoms of gas bubble disease were first
observed in fish by Gorham (1901)at Woods Hole,
Massachusetts.These symptoms include the forma-
tion of gas bubbles in the blood vessels,behind the
eyes,and in the fins.Death can be caused directly
by gas blockages within the circulatory system or
indirectly by blindness or infections that enter the
system through associated breaks in the skin.
In a later paper,Marsh and Gorham (1905)
explored the cause of this disease and associated it
with supersaturated levels of dissolved gases in the
aquaria waters.Gas bubbles occurring in diseased
fish were found to contain primarily nitrogen.They
concluded that supersaturated levels of nitrogen
gas were primarily responsible for the development
of the aforementioned symptoms.Subsequent
research,has generallyi substantiated their conclu-
sions.Under certain unusual circumstances,how-
ever:.an extremely high level of oxygen (Plehn,
1924;Woodbury,1941)i~;also capable of producing
these effects.It is the common view today that,
while nitrogen is undoubtedly the major contributor
to this problem,its effect will be felt only when both
it and the total gas concentration·are in
supersaturation.
Recently the Environmental Protection Agency
has published its proposed water quality guidelines
(Vol.1,1973).The proposed maximum acceptable
value for dissolved gas pressure is 110%of existing
atmospheric pressure.While this value will be dis-
cussed and may be changed,EPA's concern in this
area is clear. The Army Corps of Engineers'recent
water quality directive will require all districts to
maintain adequate surveillance on all pertinent
water quality parameters.This will undoubtedly
apply to dissolved gas levels in the Pacific North-
west where potential problems have already been
'identified.
The three state-of-the-art methods currentlyus~d to measure dissolved gases are:the Van Slyke,
method,gas chromatography using a Swinnerton
strippi ng chamber (Swinnerton,1962),and the
saturometer.The Van Slyke method is a well-known
technique in which water samples are introduced,
the dissolved gases liberated by a combination of
chemical and physical means,and the pressure
change measured with a manometer.From the
combination of this pressure change and the known
volume of the system,a total dissolved,gas concen-
tration can be calculated.Individual dissolved gases
can also be found by the addition of chemicals which
specifically remove individual components of the
released gas,thereby reducing the pressure.
The gas chromatographic method involves the
use of a Swinnerton stripping chamber.A known
volume of water is injected into the chamber,a con-
tinuous supply of helium gas strips the dissolved
gases from the water,and the stripped gases are
directed onto a gas chromatographic column.The
column most frequently used is molecular sieve SA,
which separates the individual components by molec~,
ular size.Each component elutes'individually from
the column and passes through a thermal condu·c~
tivitydetector whose output is recorded on a strip
chart recorder.The area under each peak on the
Jenkins:Earth Sciences Branch,Research Division"Cold Regio~s
Research Engineering Laboratory,U.S.Army Corps of Engi-
neers,Hanover,New Hampshire.
-Because of the equilibrium solubility of atmospheric gases,
tOlal gas concentration may be closely approximated by con-
sidering only nitrogen and oxygen.
101
'1'1',I
IIiI,
HlJ
:1I:
!!
I
1
1
1
1
L
L
I",c,;L
1'-'
[
i':'"-:-
~:
chart is proportional to the concentration of each
component in the original water sample.
The saturometer,unlike the other two methods,
is designed to measure total gas pressure in situ
and consists of a length of semipermeable silastic
tubing which is attached to a pressure gauge.The
tubing is inserted in the river and after gas equili-
bration across the tubing is achieved,the gas
pressure is measured directly.
While the Van Slyke and gas chromatographic
methods of measuring dissolved gas levels are
appropriate for small-scale research studies,they
are not suitable for long-term monitoring of dis-
solved gas levels.Water samples must be collected,
returned quickly and carefully to the laboratory,
and "analyzed by highly trained individuals at a
relatively sophisticated laboratory.Routine moni-
toring at a large number of locations using either
of these methods would be both inconvenient and
expensive.
The saturometer,however,has been designed
primarily for routine monitoring.It is manually
operated,relatively inexpensive and designed for
use by individuals with limited training.However,
investigators are somewhat skeptical about the
quality of the data acquired using this device.This
skepticism results primarily from the variability of
the results obtained using different devices.This
may be due in part to allowing an insufficient
amount of time for this system to come to equilib-
rium.Fifteen to 30 min is commonly required,with
the last small change in pressure being the most
time consuming.High pressure teaks are also fre-
quent occurrences and are difficult to detect and
eliminate.If these problems are satisfactorily
solved,this instrument may be the most useful of
the currently available devices for routine monitor-
ing.The three methods described above are manual
techniques which do not lend themselves to
unattended operation as would be required to inter-
face with automatic data collection equipment.No
device capable of unattended monitoring of dis-
solved gases is currently available.Should such a
device become available,it wO!lld certainly be the
most convenient and perhaps the most cost effective
way of maintaining a large monitoring program.As
a result of this need,the Seattle District,Corps of
Engineers,asked CRREl to investigate the possi-
bility of configuring a monitoring device capable of
unattended operation and incorPoration in a data
collection network.
EXPERIMENTAL APPROACH
Basicatly,two different approaches can be
envisioned with regard to constructing an
unattended monitoring system.One could either
measure total gas content of the water in situ or the
102 Jenkins
gas could be removed quantitatively from the water
and analyzed in the gaseous state.While the firSt
approach would be ideal,the number of applicable'
methods are quite limited.Since any device used
must be capable of measuring both oxygen and'
nitrogen the in situ dissolved oxygen electrodes in '
common use are not sufficient.In addition the
possibility of monitoring dissolved nitrogen in i a :
similar manner is very remote.In fact,the only
current in situ approach which merits consideration
for unattended operation is one which physically,
measures total gas pressure,like the saturometer.';
On the other hand,if the gases were removed;;:!';
from the water matrix,many current state-of-the-art.';
chemical detection techniques are applicable.These';(
include many common techniques such as;"g!~;1
chromatography,mass spectroscopy,and",optiql:;::2
spectroscopy,as well as various less expensive ..
specific gas detectors.However,the use of any of
these methods requires a satisfactory solution to,
two problems:1)River water must be supplied on
a continuous basis to an external device while sam-
ple integrity is maintained with respect to total gas
concentration;2)Dissolved gases must be quanti-
tatively stripped from the continuously supplied
water sample.
Faced with two alternatives,a limited time
schedule and a small budget,a decision was required'
on whether to pursue the in situ or gas phase
monitoring approach.While the in situ method of
directly sensing total gas pressure as'in the
saturometer has some obvious advantages,the
problems identified with manual operation of the '
saturometer would be even more significant for
unattended use.While these problems are potentially
solvable,the more novel second alternative,with a
wider choice of detectors,seemed to have a higher
probability of success and was thus chosen for addi:-·'
tional study.With this in mind,the FY-74 feasibility
study was directed toward developing prototype'
systems to solve the two problems previously me~:
tioned.Although time did not allow for the develop:",c;
ment of a prototype detector system,it was hoped,{'
that a specific detector,which was both applicable
and relatively inexpensive,could be identifiedfof .'
future development.
SYSTEM CONFIGURAliON
Pumping'System
Initial discussions with Corps personnel"at
libby Dam,Montana,indicated that significant
problems had been encountered in attempting'to
pump river water supersaturated with dissolved,,
gases.Bubble formation within the pump lines was
so intense that the water supplied became efferves-
cent.After discussions with CRREl engineers:,'a:
RESULTS AND DISCUSSION
Following construction,preliminary testing of
the stripper was conducted at CRREl.Tap water
containing 10 to 12 ppm of dissolved oxygen was
continuously passed through the device.Water at
the outflow was collected under an inert atmosphere
and analyzed for residual dissolved oxygen content
by the classical Winkler method.levels consistently
less than 0.5 ppm were found.While this trace of
dissolved oxygen was always present,it could have
been introduced by dissolved oxygen within the
reagents or explained by problems in maintaining
a totally oxygen-free environment during the 10-min
period required to collect the sample.
The pumping system was also configured within
the laboratory and under visual inspection did not
seem to produce bubbles within the pump lines.
Complete checkout was impossible,however,since
a large supply of water supersaturated with gases
was not readily available.
Final checkout of both systems was conducted
in the field at the National Marine Fisheries Service
Environmental Research laboratory at Prescott,
Oregon.This facility (Fig.2)is located on a barge in
the Columbia River,35 miles downstream from
Portland,with easy access to river water having
high gas levels.The field configuration included
a submersible pump which was maintained about
1 ft beneath the water,the stripping chamber,and
a portable gas chromatograph to be used as a detec-
tion system (Fig.3 and 4)<
While the system was in operation,water sam-
ples were withdrawn with a liquid syringe from a
septum cap (Point A,Fig.3)located on the system
just upstream from the water control valve,and
from the river -near the location of the submersible
pump.The dissolved gas content of these samples
Water
In
·6-!-"-
Water Out
pumping system was configured based upon a cen-
trifugal type submersible pump.The system included
a control valve at thl~farthest downstream point
where sample integrity was required.The back
pressure in the lines caused by this control valve
would maintain a high equilibrium gas solubility
and thus reduce the tendency toward bubble forma-
tion.This valve would also serve to maintain -control
over the flow rate of water continuously supplied
by the pump.
Int.rna~MoIMt
T.UOIl
8ean ll9,
Stripper Unit
While attempts were being made to devise a
system to remove dissolved gases quantitatively
from a continuously supplied water sample,a device
developed by Dr.D.Williams and R.J(Miller (1962)
at the Naval Research laboratory was discovered.
This device was an adaptation of the spinning disc
oxygenator used in the medical field to oxygenate
blood during open heart surgery.A device con-
figured for medical application was obtained from
Mary Hitchcock Hospital,Hanover,New Hampshire,
and tested.The results were so encouraging that a
stripper _device spedfk to our application was con-
structed based on the design parameters published
by Williams and Miller (1962)(Fig.1).
Motor'
-
fiG.1 DiOlgrOlm of stripper unit
_-The principle of operation is as follows.A con-
tinuous water sample is introduced to the device
(Fig.1).A water level approximately 1/3 of capacity
is maintained in the device prior to outflow.A con-
tinuous -supply of helium carrier gas circulates
through the stripper.During operation,the motor,
magnetically coupled to the internal shaft,rotates
the internal Mylar discs at several hundred rpm.
As the discs rotate through the water,a thin film
is spread over the surface and carried into the helium
headspace.This thin film rapidly equilibrates with
the headspace,which is essentially devoid of air.
Since this thin film and the air involved are con-
tinuously removed,ra.pid and efficient gas stripping
is achieved.FIG.2.Prescott enyironmental field station.
Continuous Monitoring 103
FIG.6 Proposed monitor configuration.
I
1•l
!
il
,1
.r~~He',.",;--L-J --Cctrie.r Go'S
,Gos FlOW
..Co"I,,,1 Unft
rh.rtnGI ConduCllvirV
Ott.ctor.\
Somple i=1eferel'lCF
Cell Cell
'..ren'
I
i
Flow Conl",l IVa/".__
I
ZeroGas--~
Aulom01ic
3-WQ1 VQllle
REFERENCES
Gorham,F.P.1901.The gas bubble disease of fish and its
cause.In:Bul/.U.S.fish.Commission,1899,vol 19,pp.33-37.
Marsh,M.C.and F.P.Gorham.1905.The gas disease in
fishes.In:Report of the Bureau of fisheries,1904,pp.343-376.
Plehn,M.1924.Praktikum der Fischkrankheiten.Handbuch
der Binnen fisherei Mittel Europas,1 :301-479..
Proposed Criteria for Water Quality,vol 1,October 1973,U.S.
Environmental Protection Agency,pp;102-103.
Swinnerton,J.W.,A.J.linnenbom,and C.H.Cheek.1962
Determination of dissolved gases in aqueous solutions by gas
chromatography,Anal.Chem.,34:483-485.
Williams,D.D.and R.R.Miller.1962.An instrument for
on-stream stripping and gas chromatographic determination
of gases in liquids,Anal.Chem.34:657-659.
Woodbury,l.A.1941.A sudden mortality of fishes accom-
panying a supersaturation of oxygen in lake Waubesa,Wise.,
Trans.Am.fish.Soc.,n:112-117.
1 Water Flow '---~
I
·C.n".,Un"jL--o--._-Veo'
Wote ...Level
Control Unit
From
Submersible Pump
cation of this system the following areas must be
investigated:1)The detector design must be
refined,a prototype assembled,and the system
validated under field conditions;2)The electronic
systems required for detector operation,tempera-
ture control,flow cycling,and signal processing
must be designed and configured;and 3)Systems to
maintain a higher degree of control on flow rates
within the stripper unit must be designed and
implemented.
Should a system be required to quantitate a
single component of the gas matrix,a more specific
detector would be required but the remainder of
the system would be largely applicable.
RECOMMENDATIONS
The results of this investigation confirm the
feasibility of developing an unattended monitoring
system for dissolved gas1es.The next step is to
actually configure a prototype system for.field
validation.A diagram of a specific system,which
utilizes a thermal condu1ctivity detector for total
gas measurement,is shown in Fig.6.Prior to fabri-
Instrument Development,Model 512)was used.
Quantitative results were obtained using a sampling
loop and a thermal conductivity detector and com-
paring the output to that obtained when running
a calibration gas.
The major problem encountered in obtaining
accurate measurements involved maintaining pre-
cise knowledge of the wa,ter and gas flow rates in
the stripper unit.Specifilcally the value obtained
from the output of the gas chromatograph must be
adjusted using a factor based upon the ratio of the
water and gas flow rates.If the gas and water flow
rates can be maintained at an identical value,the
concentration of dissolved!gas found in the carrier
stream is equivalent to that in the river water
~~~-
The difficulty encountered maintaining con-
stant flow rates resulted from changes of resistance
in the gas flow system.These changes occurred as
the drying tube picked up water,resulting in a
higher gas head pressure in the stripper and a lower
water flow rate through the system.The second
problem was encountered when the valve on the
gas chromatograph sampling Ipop was switched,
sending a gas sample into the GC column.When
this was done,the resistanlce on the gas flow system
was also changed,resulting in a different head
pressure in the stripper.This caused a change in
water and gas flow rates as well as a change in the
water level in the stripper.
In spite of these difficulties,when the system
was manually operated with extreme care,reason-
ably constant flow rates could be maintained.Values
obtained under these conditions were compared
with those found by injecting river water samples
into the Swinnerton chamber of the gas chromato-
graph.The values obtained for dissolved nitrogen
were 21.1 ppm by the stripper system and 20.7 ppm
by the Swinnerton method.While we were unable
to compare these methodologies over an extended
period,the results are quite encouraging.
.....
.....
.....
--Continuous Monitoring 105
Sample Tak""From
Point A On Pump
Line
FlOW Control
Valve Helium
r--~J--CarrierGas
'------8
Water With
Dissolved Gases
Removed
River Sample
194 196 /97 199 198 186
Nitroqen
92 94 94 94 93 93 ,
O.ygeo
Flow rates used in these tests were approximately
8 ml/min of water and 22 mllmin of helium.When
the system was operating under these conditions,
samples were taken from the output of the stripper
(Point B,Fig.3)and analyzed for residual dissolved
gas content with the Hewlett Packard8 gas chromato-
graph previously described.Generally,no peaks
were detectable on the strip chart.In a few casesj
an extremely small peak was observed but was too
small to be detected by the digital integrator.These
results indicate that the stripper was quantitatively
removing dissolved gas from the continuous water
sample and transferring this gas to the helium'
carrier stream.
The final phase of the field test was to deter-
mine whether a detector could sense the amount
of dissolved gases in the carrier stream and from
this accurately quantitate the dissolved gas level
in the river.The majority of the experimental diffi-
culties were encountered in this phase.Since time
did not permit fabrication of a detector specific for
this task,a portable gas chromatograph (Analyti«:al
RG.5 Chromiltogrilm oi dissolved pses in river wilter i1nd
continuously pumped Silmple.
Drying
Tulle
A
Septum Cop
For Woter
Withdrawal
From
Submersible Pump
Flow Control
Valve
GC Gas StreamSampiing--"'-=:""=":':"=:":":':""--l:.:H---{
Loop Containing
Dissolved
Gases
Re.3 Field instrumental configuriltion•
RG.4 System in operillion.
Vent
Standard
Gas
was analyzed with a gas chromatograph (Hewlett
Packard Model 5700A8)equipped with a Swinnerton
stripping chamber (Swinnerton,LinnenbOm,and
Cheek,1962).The results are shown in Fig.5.Both
the.strip charts and a digital integrator on-line to
the gaS chromatograph indicated the samples to be
identicil with:resped to dissolvedl1itrogenand
oxygen.Hence the pumping'system·was indeed
supplying water to the stripper with dissolved gas
concentrations equal to those in the'river itself.
The next step in the field test was to determine
the stripper's efficiency in dissolved gas removal.
Published information (Williams and Miller,1962)
regarding operation of the stripper unit indicated
that 100%stripping efficiency was found when the
flow rate ratio of heliUm/water was maintained
between 10 and 0.5 for flow rates up to 100 cc/min.
104 Jenkins
....
.....
.....
....
.....
....
....
#602-105)is particularly suitable for this application._.
since its small diameter provides optimal surface,:
.';.::~"/;~7ta::~~:~d,~fti~~e~~;~s~f :Ub~~bl:~de~:~i~~L~•.:~"j~
other communicating with the diaphragm or sensi~,"
tive element of a low dead-space pressure trans-
ducer,the pressure of all gases and vapors in the
water contacting the tubing will,at equilibrium,be
measured.A most important consideration in
design,then,is elimination of dead-space.This,
involves both the choice of the appropriate pressure
transducer and the design of a low dead-space
connection of the tubing to the pressure transducer.
DESICN
The unit described has been designed for
minimal expense,simplicity of fabrication, and ,,"
ease of operation.Its basic design also allows
individual modifications according to the needs·
and/or preferences of the user..,
Fig.1a indicates the complete parts and mate-
rials necessary for construction of the unit.Cat~
logue sources and numbers are given as the part is
described.
Pressure Transducer and Probe Assembly
A National Semiconducto~LX 1601 AF inte';'
grated-circuit pressure transducer which measureS
from 10 psia to 20 psia appears ideal for the rcll:,ge
in dissolved gas pressure likely to be encountere(B
in the field.However,there are many additional
models and makes to choose from.The pressure,>
transducer is shown (Fig.1b)mounted on a 3/16 in.,
brass Swagelock8 O-seal connector fitting (B-0300-'
1-20R)using two O-rings (Parker 1/16 in.-0(8)'
instead of the metal ferrules which are normally7.
used.The Swagelock fitting (Fig.13)is adapted to
the pressure transducer nipple diameter by drilling
with a #7 (0.201)drill.In addition it is advisable,
D'Aoust,White and Seibold:Virginia Mason Research Center,
Seattle.Washington.
I,B.G.D'Aoust
R.White
H.Seibold
ABSTRACT
The environmental and biomedical problem of supersaturation
of dissolved gas and the research related to it has produced a
need for a more effident means-of measuring and monitoring
total dissolved gas pressur,e than those now in use.A modification
of the Weiss saturometer is described which equilibrates within
8 min.is portable and can be operated remotely in a recording
mode.The basic unit is inexpensive.easily constructed out of
available components and allows many options in design so
that such units can be custom-made to specific needs.It has
been field-tested and is oJrrently in use.
The environmental problem of supersaturation of
rivers, lakes and streams due to naturally or man-
induced excess runoff and/or temperature increases
has created an urgent need for a less expensive
and more efficient and reliable means of monitoring
total dissolved gases then is presently available.In
this paper we describe the design and function of
a basic instrument which involves several modifi-
cations of the original Weiss saturometer which has
been used and evaluated for the past 3 yr in the
northwestern United States.It is based on the use
of a watertight plastic tube across which dissolved
gases exchange according to their partial pressures
and the total pressure is measured manometrically
(as described by Enos et aI.,1965,in studies of the
effect of hydrostatic pressure on dissolved gases).
We have replaced the large dead-space Bourden
tube gauge with a low dead-space solid state elec-
tronic pressure transducer to facilitate both porta-
bility and remote monitoring.This ability to monitor
a particular location inexpensively should provide
more useful and meaningful information at much
less public and private expense.
An Ele<:tronic
Monitc)r
for Tot;al Dissolved Gas Pressure
PRINCIPlE OF OPERAnON
Since the devic1~measures total dissolved gas
tension or partial pressure,we propose the term
"tensionometer"as .a more accurate description of
what is actually being measured.The basic prin-
ciple uses the fact thelt most gases and vapors diffuse
rather quickly through thin layers of most plastics,
one of which (silicon rubber in the form of Dow-
Corning medical grade silastic8 tubing,catalogue
~.
.l ~
"
~
......
-
1
1
106
~,
,."..
-
FIG.1 a Complete-parts and materiah to cons~et electronic
tensloOOmeler.-VOMmeter on right can be used to read output
(Vo')of deyke.Parts and suJ'Plies are described Indiyidually In ,
lext.
AG.1c Unit completed with approximately 30 It of cable.R1
and R2 (see text)are on lower two comers of aluminum box;
switch (51)is in center below readout (V0)jacks.Hair curler
spool to hold silastic:is ShOWlI1 without perforated metal protector.
though not essential,to machine a part of the
Swagelock fitting insert to extend into the nipple
of the pressure transducer as shown in Fig.2a so
it occupies most of the dead space of the pressure
transducer nipple.The other end of the machined
rod holds a 2 in.length of #25 hypodermic stock
to connect to the siilastic tubing.Care must be
taken when constructing and assembling this part
of the unit to avoid the risk of stressing or punc-
turing the silastic fluid-isolating membrane of the
pressure transducer (Fig.la).
The Swage lock fitting is screwed into a brass
insert (Fig).1b and 2a)which is constructed of
FIG.:1b PVC threaded pipe union which senes •submefsible ,
probe housing shown with brass insert containl..SwagelOc:k'
fitting In which pressure transducer (LX 1601A.f)Is mounted."
FIG.1d Same as 1C but with digital YOM meter connected to
output Yo'
1/8 in.plate 2-3/4 in.in diameter,soldered or brazed
to a 1 in.length of 1-5/80.0.brass bar stock which
has been machined to an 1.0.of 1 in.to receive the
Swagelock fitting.
This brass adapter fits inside a schedule 80 PVC
1-1/2 in.threaded plastic pipe union as shown in
Fig.2a and 1d.Approximately 10 ft of silastic tube
is wound around a plastic hair curler (TipTop@)
#5840/X,Faberge Inc.,Omaha,Nebraska)and one
end is tightly knotted and/or plugged with a short,
smooth length of nylon fishing line.The other end
of the silastic tubing is strain-relieved on the side
of the hair curler by a loose knot or RTV adhesive
Electronic Monitor 107
-IMIN-r 7.5 MIN
RE.wJVAL I
136~SAT'N ---+-t
FIG.2d Reeordingof appl'Oilch to maximum read"mg taken in
the N.M.f.5.Mon~lce Laboratory in Seattle.Final pn!55Ufe mea-
sured (referred to surfxe)was 136%Aluration reached in 7.5 min
and WillS reproduced in three consecutive triilh.Notice the more
rapid retum 10 equilibrium folowing removal of the probe
because of the physically c&fferentO(ps/membrane/ps)diffusiOn
sitwdons.
FIG.2b Circuitry showing bitttery operation:Reg..1M 340-15
National Semiconductor 15 volt regulator.51,three position
switch for on/off and bitttery or regulilted voltage reading;R1,'
2000 ohm 10 tum helipot.~SOOO ohm 10 lum heIipot.~,
2000 ohm (1/4 watt)resistor.PO'pressure transducer.Yo,jacks
where output or bitttery voltage is measured.
r=Ii"_~_1_!1-.-l--L._+-
R.:...1---I.--".h-.-1
The terminals of the pressure transducer are>
connected to the waterproof three-wire neoprene-c'
covered cabte (Belden 19229,18-3 5JO Rancho.;,,
prenel8l )by constructing a small five-wireclip.<
from one side of an in-line integrated circuitlC:
socket.Only the ground (pin 2),supply (pin 5)and
output (pin 1)terminals of the chip are used in this,.
configuration and are connected to the cable by
5 in.of small flexible #20 hookup wire.The cabte
attachment to the PVC union is both waterproofed
and strain-relieved by means of a plastic cord grip,,;
FLUID ISQATOR
LX 160lAF
PRESSURE
TRANSDUCER
SWAGROCK O-SEAl mE
CONNECTOR B-3QO-I-20R
o.S LO L5
50 +VQTS
100
150
200
mm Hg
V ,:
:C ci •
&/B ./A
:~/,:.
I P /~//.IN 10 mV
.~.dP •mm I-t:J
-,-......~_.---+---
/
l5 lQ/1t
!faI•
A<:~:
B<:~~;
C <R l 948
R2 933
AMBIENT PR£SSURE
,
L625"BAR STOCK-'-----,
UP/'R..ATE BRASS
O.R.
SOLDER
0.020 S. S.'\
FIG.h Detail of mounting of pressure trmsducer in Swagelock
fitting and brass insert.The ~tter is most NSiIy constructed by
first mKhining theilNr stock to correct dimensions mel then
adding the 01/1 in.plate.The ceotel'~er of the 0Swagelock
fitting can be _mined from 1/4 in.brus rod and the 0.020 s.s.
apil~ry soldered inside it.
which ,does not occlude the lumen of the tube and
the other end is attached to the protruding end of
the staintess steel capitlary.The hair curler is
fastened (with a hose damp)to the machined end
of a pvc pipe ptug which threads directly into the
PVC union.A cylinder of perforated metal is in
turn hose-clamped directly to the latter to protect
the silastic tube.This construction facilitates carry-
ing several prewound "spare membranes"in field
studies so that replacement,if necessary.can be
accomplished in minutes.
108 D'Aoust,White,Seibold
FIG.2c A~tiYe CalibAtion slopes for Yo',t,to read in
mm Hg where 1 mm Hg ,=10 mY.a.to read in inches of mercury
where 1 in.mercury =UO mY .et inches of mercury =Yo I(10.
C,to read in percent 5UpeI'Y.IuAIion where 10%SUpersatuAtion
=100 mY,the pen:entase extes of undersatUration =Yo I(100.
The latter cMibration "throws _ay"most of the preuure semi-
me output and is iIlUordiiI'lsly less KOIrate.
I
,.,..l
.I'
j:
-.
connector (1/2 in.male pipe,catalogue #2672,
Thomas and Betts,Elizabeth,New Jersey)which
screws into the 1-1/2 in.to 1/2 in.PVC reducing
bushing (Fig.1)and Ifirmly strain-relieves the heavy
cable.Teflon tape is used in all of the threaded con-
nections to assure a watertight seal.
ELECTRON ICS
There are many options available in the power
supply and the readout circu itry used.That which
is described is offered only as a starting point but
provides a reasonably inexpensive,efficient unit.
Fig.2b indicates the circuitry used,all of which,
save the cable and the pressure transducer,is con-
tained in a cast aluminum box (BUD #CU-247)as
shown in Figs.1a,1c,and 1d.The uJlit is powered
by a DC source bet.lVeen 18 and 30 volts.This can
be from two -to four 9-volt transistor batteries or
two larger 12-volt batteries.A DC power supply is
desirable for laboratory calibration since it saves
battery life.
The DC source shown in Fig.2b supplies a regu-
lator (LM 34015 National Semiconductor)which
maintains a constant 15 volts.This voltage is fur-
ther dropped by resistors R3 and R1 to vary the
offset voltage of the transducer against which the
output voltage of the pressure transducer-itself
attenuated by R2-is measured.Any VOM multi-
meter of at least 20,000 ohms/volt will suffice to
measure Vo 'Ten-turn small helipots are used for
R1 (2K)and R2 (5K).It is desirable to have either
the regulator voltage or the battery voltage read-
able by ":leans of the two position switch 51.
Calibration
Fig.2c is a curve showing several different ways
the device can be calibrated.The voltage measured
can accurately indicate (by sh ifting a decimal)either
millimeters of mercury pressure,inches of mercury
pressure,or percent saturation,referred to baro-
metric pressure (B);a reading of zero volts is equiva-
lent to ambient pressure.Approximate helipot dial
settings are given for R1 .and R2 associated·with
the three calibration graphs assuming B =760 mm
Hg.These will vary slightly with the altitude and
ambient pressures prevailing during use,and it is
recommended that the instrument be thoroughly
calibrated with a number of different slope and
pressure settings prior to use.The pressure trans-
ducer itself is sensitive enough to act as a barom-
eter and/or altimeter.
Maximum accuracy is obtained when dV/dP
is maximal;that is,maximum voltage per unit
pressure.In the experience with the unit thus far
it appears most feasible to calibrate in terms of
10 mV/mm of mercury so that millimeters of mer-
cury can be read directly by multiplying by 100.It
is of course feasible to integrate into the unit a digi-
tal voltmeter for readout.However,this involves
considerably greater expense not only for the DVM
but also for the more sophisticated power supplies
to run it.Applications oriented toward monitoring
and telemetry over long time periods may justify
such further sophistication.
Use of the Unit
Since the measurement of supersaturation
always suffers from the uncertainty that a physically
unstable situation is being measured,several notes
of caution are in order for the use of these instru-
ments.As with other units,agitation is necessary
to remove bubbles from the silastic tubing to pro-
vide the pressure reading.However,when super-·
saturation is not present,equilibrium Occurs without
agitation within 8 min.Fig.2d is a curve showing
the titne course of pressure buildup in the tube
following immersion of the probe in approximately
30 ft of supersaturated water in the tower operated
by the National Marine Fisheries Service in Seattle.
The reading shown in Fig.2d is 136%saturation and
required 7.5 min to reach this value,which was
reproduced on three consecutive measurements.It
is clear that if the depth of water which is to be
measured is at least 10 ft it should be possible to
measure supersaturations as high as 33%relative
to the surface without haVing to agitate the probe.
It appears that in most field situations it would be
sufficient to measure at a depth of 15 ft and,pro-
vided the assumption of vertical mixing was valid,
agitation should not be necessary.This should give
data of greater reliability than that afforded by
using existing units in a large sample of water.
Further,these units are not efficient because of the
large amount of tubing necessary to counteract
their large dead space.Since the tubing also acts
as growth sites for bubbles,adding more of it to
counteract a large dead volume is of questionable
benefit.Comparison of the present unit with that
commercially available on single samples of super-
saturated tap water has shown that the latter can
under-read by as much as 40%of the maximum
value shown by the un it described here.
It is important in comparing these devices to
other quantitative modes of analysis such as the
Van Slyke and/or gas chromatography to keep in
mind that,where supersaturation exists in the water
being analyzed,tensionometers can underestimate
the total dissolved gas content and quantitative
methods can overestimate the dissolved gas tension
because of the possible presence of microbubbles.
Use of either measurement to estimate the other
with solubility tables (Weiss,1970)must be inter-
preted with caution.It is also emphasized that the
only reliable measurements where supersaturation
Electronic Monitor 109
I
j
J
J,.'
J'.'.
:.,.'
1
1
is present are those taken under conditions of
steady-state where the readings stabilize and are
reproducible.For purposes of natural water manage-
ment it appears most desirable to more continuously
monitor dissolved gas tension at different locations
to accurately quantify and delimit the problem.
ACKNOWLEDGMENTS
This resear<:h was supported by USPHS grants
#Hl 12015,#Hl 14801,to the Virginia Mason
Research Center,Seattle,Washington,and Career
Award K04 Hl 70543 to B.G.D'Aoust.
We thank Bruce Monk of the National Marine
Fisheries Service,Montlake Laboratory,for the use
of the supersaturation test tower.The .author's
110 D'Aoust,White,Seibold
original interest in this problem was sparked by a
meeting in January 1971 between Dr.Robert Rucker
(1972)of the Western Fish Disease Lab and Dr.Mer-
rill Spencer,then director of Virginia Mason
Research Center.
REFERENCES
Enns,T.,P.F.Scholander and E.D.Bradstreet.1965.Effect
of the hydrostatic pressure on gases dissolved in water.}.Phys.
Chern.19:389.
Rucker,R.1972.Gas Bubble Disease:A Crititcal Review.
Bur.Sport Fish.and Wildl.Tech.Paper No.58.U.S.Dept.of
the Interior.
Weiss,R.F.1970.The solubility of nitrogen,oxygen and argon
in water and sea water.Deep Sea Res.17:721.
Round Table
Discussions
WATER QUALITY
STANDARDS
DISSOLVED GASES ANALYSIS ~~:~~NMENTAL
~ELECTROLYTIC
STANDARDIZATION
l
,GAS CHROMATOGRAPH
SATUROMETER I \";WINKLER
\~VANSLYKE
DISSOLVED GAS ~
ANALYSIS
RESEARCH NEEDS IN
DISSOLVED GAS STUDIES
(MITIGATION)"...,/...............
"STEAM ELECTRIC
THERMAL DAIMS ./NATURA~/NTS
EFFECTS +,\
1
"""AIR ENTRAINMENT
""\THERMALSUB-LETHAL INiREMENT
LETHAL 1 FIELD __DISSOLVED'~A~>::.A~'"..:"1:''"i1~~00_
DEI'TH 1 THEORY
DISTRIBUTION
~CRITICAL
EXPOSURE
INTERMITTENT ,,/""'-.
EXPOSURE 'It..I POPULATION EFFECT
GAS TRANSFER "'"
RATE _'\.,
Informal round table discussions were held dealing
with several topic areas related to gas bubble
disease.The co-chairmen of each discussion were
asked to establish an agenda and lead the discus-
sion with goals of determining the current state of
knowledge and identifying research needs in the.
subject area.A diagram of research areas (Fig.1)
was presented as a basis for discussion.
Participants were asked to select from four
topic areas:biological studies,physical and
engineering studies,analytical problems,and
water quality criteria.The biological studies were
subdivided into groups dealing with field and
laboratory oriented studies that were held concur-
rently as were sessions on analytical problems and
physical and engineering stud ies.
The following summaries of each of the dis-
cussions were prepared from notes and tapes of
the sessions and we hope they accurately represent
a generalconcensus of the participants'opinions.
.~
FIG.1 Research areas in dissolved gas studies
1
111
"""
Biological Studies:
Laboratory
Orient<ltion
laboratory studies of the effects of dissolved gas
supersaturation on aquatic organisms have been
designed primarily to determine acute tolerance
using death as an endpoint.
Gas supersaturation is an immediate and
urgent problem in the Columbia River system,
particularly with regard to salmonid resources,
requiring management decisions on a time scale
which precludes long-term or basic studies of the
effects of dissolved gas supersaturation on aquatic
biota.We have by necessity focused on short-term
experiments designed to answer questions of
lethal levels of gas saturation,in most cases during
acute experiments.These tests assume that once
exposed to excess dissolved gas an organism
becomes and remains supersaturated.However,
two other contingencies exist:the gas solubility
may be increased (i.e.,by sounding to increase
hydrostatic pressure or by decreasing the tempera-
ture)or the tissue dissolved gas content may be
decreased (by re-equilibration with water having
a lower dissolved gas tension).If neither of these
occurs,the excess gas will leave solution and form
gas-phase bubbles causing gas bubble disease.
Hydrostatic pressure increases gas phase
partial pressures such that the gas saturation level
of an organism decreases as it moves to a deeper
location.Most river systems subject to artificial
supersaturation are deep enough to permit this
pressure compensatio1n,but the behavior of fishes
exposed to supersaturation is poorly understood.
Factors other than gas levels (e.g.,light,tempera-
ture,prey density,pressure)may cause a depth-
selective response.Trauma due to gas bubble
disease may be another stimulus affecting depth
distribution.Diadromous species are forced to
surface when passing over dams,decompressing,
and perhaps producing gas bubble disease when
the hydrostatic pressure decreases.
Dissolved gases move down a gradient of gas
tension:the external gas tensions must be less
than the internal tension at the exchange surface
for a net outward flux of gas to take place.
There is some evidence that the rate of equili-
bration between organisms and water is rapid,with
112 Round Table Discussion
I Co-Chairmen
A.V.Nebeker
D.H.Fickeisen
equilibration nearly complete in a matter of 1 to 2
hr.Re-equilibration is also rather rapid;however,
in the Columbia River system re-equilibration
during the spillway operating season ,is not feasible:
under present operating schemes as 'essentially the '
entire system is artificially supersaturated for ex-
tended periods.
During acute tolerance tests we have recog-
nized that additional information on swimming
depth distribution,behavioral responses to gas
supersaturation,and effects of intermittent expo-
sure to excess dissolved gases is needed.long-
term and basic studies dealing with sublethal and
chronic effects at the population and ecosystems
level rather than limited to the individual level
should also be undertaken.Specific research needs
identified during the discussion include:
•Sublethal and chronic effects of exposure
to gas supersaturated water.Effects on fecundity,
predator-prey relationships,and sensory physiology
require further study.'
•Combined stressor ,effects.More informa-
tion is needed detailing effects of temperatur~,
fish disease,and operations of hydroelectric facili-
ties acting with gas supersaturation.It is quite
possible that these and other factors act syner-
gistically to increase the magnitude of their indivi-
dua I effects.
•Gas bubble formation trigger mechanisms.
Initial experiments and a review of available data
indicate that mechanisms triggering bubble forma-
tion are multiple and differ in acute and chro'nic
tests.Basic physiological research is required to,'
define trigger mechanisms,effects of internal body"
pressures and nucleation sites,and factors resulting,"
in bubble size stabilization.Differences in'indivi-
dual gases also require attention.
•Detection of and response to gas super-
saturation.Evidence regarding the ability of fishes
to respond to supersaturation is conflicting.Some
Nebeker:U.S.Environmental Protection Agency,Corvallis.
Oregon;and Fickeisen:Battelle-Northwest,Ecosystems De-
panment,Richland.Washington.
,-
.....
"....
estuarine species appear to avoid water containing
dissolved oxygen tensions in e>;.cess of 110%of
atmospheric pressure,while other species do not
appear to respond to elevated gas tensions.Simple
selection chambers should be employed in addi-
tional tests with several species.
In addition,the following acute tests are
needed:
•Intermittent exposure.Selective spilling
such that only a portion of a river system is super-
saturated might alleviate the effect of gas super-
saturation on diadromous fishes.About 2 yr ago
the Nitrogen Task Force recommended that the
feasibility of spilling "slugs"of water and the
effect of the anticipated intermittent exposure be
investigated.Additionally,diadromous species
are subject to decreases in hydrostatic pressure
when they pass over dams.Although some inter-
mittent exposure data is being collected,additional
studies are needed.
•Acute tolerance.Data are available des-
cribing the tolerance of freshwater fishes to dis-
solved gas supersaturation;however,tolerances
of aquatic invertebrates and marine organisms
to acute exposure are poorly defined at present.
The session closed with a plea for basic re-
search aimed at determining long-term population
and ecosystems effects and for some immediate
cooperative solution to maintain the Columbia
River system's salmonid resources.
Round Table Discussion 113
Biological Studies:
Field ()rientation I Co-Chairmen
W.Ebel
R.McConnell
J
J
1
J
1
L.~ji.
During the review of past research and throughout
the discussion of current research,a consensus of
opinion was developed concerning several points.
A general summary of these points is as follows:
•Both adult and juvenile fish populations
suffer substantial mortalities if exposed for a suffi-
cient duration at levels exceeding 120%saturation
even though they have the option to sound and
compensate for supersaturation.
•Adult mortalities that have occurred at
Bonneville Dam since 1955 have been related to
gas bubble disease and,in 1968,20,000 adult
chinook were estimated lost in the vicinity of John
Day Dam.
•Spawning ground surveys in the Columbia
and Snake Rivers have indicated a recent decline in
numbers of redds and these can be directly related
to years when supersaturation was high.
•Percentage adult return of steelhead and
spring and summer chinook to the Snake River
have steadily declined since 1970,in spite of in-
creased production by hatcheries.
•Mortality of juvenile populations of sal-
monids varies between dams and the cause of
,mortality changes,diepending on the flow and the
type of dam the fIsh must pass through.During
high flow years the majority of the mortalities
can be attributed to N 2 supersaturation.However,
in low flow years such as 1973,all mortality had to
be the result of delays in migration rate,predation,
and passage through turbines.It was the consensus
that an extremely low flow year such as 1973
created much great~~r mortality to juveniles than
high flow years when supersaturation is present.
For example.Raymond (NMFS)estimated the
juvenile loss of chinook from the salmon River in
Idaho to Ice Harbor Dam was about 50%in 19n
and 70%in 1972 (both high flow years)while in 1973
{a low flow year with no supersaturation}mortality
was about 88%.
•Resident fish species and invertebrate
population also are affected by gas bubble disease.
Eighteen species of fish in the lower Columbia
River have been obs.erved with gas bubble disease
and invertebrate species diversity below libby Dam
has been reduced Ileaving primarily one species
(Chironomus sp.).
•Knowledge of depth distribution of fish
114 Round Table Discussion
species is important when attempting to assess
mortality that might be caused by exposure to
supersaturation of N2 •When levels of 130%are
recorded at surface pressures,juvenile salmon in
the Snake River are still subjected to levels of at
least 118%,even after accounting for the hydro-
static compensation indicated by their average
depth distribution.Studies of depth distribution by
Mains and Smith (1954-55)indicated that 44%of
the outmigrant chinook were in the top 2.5 ft of
water and 68%were near shore.Studies of depth
distribution at lower Monumental by Smith (1973)
indicated 34%of the juvenile chinook and 27%of
the steelhead were in the top 5 ft of water.Prelimi-
nary tests to determine depth distribution in the
lower Columbia by use of a sonic fish detector
indicated 58%were in the top 5 ft.
All three studies indicated more fish move-
ment occurred during the night hours.
•live cage studies done by Parametrix were
in agreement with past live cage studies.No mor-
tality occurred in a vertical cage even though levels
rose to 125%saturation.It was pointed out that
other live cage studies done earlier resulted in sub.;,
stantial mortalities in the volitional cage (as high
as 60%in the Snake River)but levels of satura-
tion were as high as 135 to 140%.
•Intermittent exposure of juvenile salmon
increases survival over that recorded for constant
exposure to high levels of supersaturation.
•Recovery of exposed juveniles with obvious
symptoms of gas bubble disease does occur but
nonlethal exposure of adults usually results in
death from secondary infection from some other
disease such as Columnaris.
•Studies to evaluate devices for .reducing
N 2 concentrations indicate that spillway deflectors
are the best solution to date.All data obtained
indicate that they reduce N2 levels substantially
from what would occur with the standard spil1way
and no adverse effects to either juveniles or adults
passing through or in the vicinity of the devices
could be detected.
Ebel:National Marine Fisheries Service.Seattle.Washington;
and McConnell:Nalional Marine Service Fisheries.Longview.
Washington.
.-
•Transportation studies indicate that sur-
vival of chinook and steelhead can be increased
from 10 to 400%by collecting juveniles at the upper
dam and transporting them to locations below
Bonneville Dam.
The following areas were indicated by the
group as areas where research should be continued
or,if not already in progress,should be started.
•Transportation studies should continue.
•Survival data should be continually ob-
tained to determine changes in survival rates that
occur at various flows as spillway deflectors are
installed.
•Additional hodzontal and vertical distribu-
tion data are needed for both salmonid and resident
species to estimate what happens to ,the food chain
and predafor-prey rE!lations when the biomass is
subjected to Varying degrees of exposure to super-
saturation of N 2 •
•Additional data on tolerance of adult sal-
monids to supersaturation of N 2 are needed.Several
species have not been tested and additional bio-
assays should be conducted.
•Observations of the Kootenai River below
libby Dam should continue to obtain data on degree
of recovery of the ecosystem after N2 levels are
reduced.
•Studies to define effect of long-term sub-
lethal exposures on survival and productivity are
needed.
•Synergistic effects of other factors on toler-
ance to supersaturation are unknown and some
investigation is needed.
A final general comment was made and gener-
ally agreed upon,and that was:"We have enough
information now to proceed with correction of the
problem and spillway deflectors should be installed
as soon as possible.There is little need to delay
installation while additional nebulous information
is obtained."
Round Table Discussion 115
F""""'-
Analytical
Metho-ds I Co-Chairmen
M.J.Schneider
B.G.D'Aoust
For total gases excluding water vapor relative to
moist air,use
water
barometric +saturometer vapor
pressure pressure pressureSaturation=-,;"..-_._--,;"..---....._--
barometric
pressure
Methods of dissolved gas quantification were dis-
cussed and compared.Presently available methods
report data for a single discrete sample and each
has unique features making the selection depen-
dent on intended use of the data,operator skill,
and equipment cost.In addition,the two methods
being developed for continuous and semi-continu-
ous unattended monitoring were discussed as was
the need for such capabilities.Each of the methods
will be treated separcltely below.
The Weiss saturometer or tensionometer mea-
sures total dissolved gas tension directly.Dissolved
gases are permitted to diffuse across a semi-
permeable membrane into a closed,gas-filled
space connected to a pressure sensor.The design
most commonly used consists of a coil of Silastic@l
tubing inside a protective cage which is submerged
in the water being analyzed.The tubing is con-
neded to a pressure gauge providing total dis-
solved gas pressure including water vapor relative
to the surface atmospheric pressure.A positive
pressure,therefore,represents a supersaturated
condition.
Three different Irormulae have been used to
compute the percentage of equilibrium saturation.
If the degree of total dissolved gas saturation (in-
cluding water vapor pressure)relative to moist air
is desired,use the formula:
barometric
pressure
(3)
water
vapor
pressure
water
vapor
pressure
barometric
pressure
x 100
barometric saturometer
pressure +pressure
Saturation =
And for the degree of total gas saturation excluding
water vapor relative to dry air,use:
Equation (2)correlates with values normally re-
ported in the literature for other methods;however,
the appropriate formula to use depends on the
intended use of the data.In any case,the baro-
metric pressure and water temperature should be
reported with data obtained with the saturometer
to permit conversion between values.
Advantages of the saturometer include low
,cost and capability of use in the field.It is readily
portable and provides data on site.It does not,
however,provide data for individual gases and
requires running a simultaneous Winkler titration
to determine oxygen concentration so that sepa-
rate gas data may be computed.Several partici-
pants were critical of the instrument due to prob·
lems arising from the relatively long equilibration
time (approximately 20 min)and the need to vigo-
rously agitate the instrument during this equilibra-
tion time.It was agreed that the operator must be
familiar with potential problems including leaks
and the equilibration time and must conscientiously
use the saturometer.
Dr.Brian 0'Aoust suggested that the concept of
the saturometer has not been thoroughly explored
through radical design changes including reducing
the volume of the dead space in the tubing and
pressure sensor.As little as 2 in.of tubing may be
(1)x 100
saturometer+pressure
barometric
pressure
Saturation =
--
....
-
x 100.(2)
SChneider:Battelle-Northwest.Ecosystems Department,Rich-
land,W~hington;and D'Aoust:Virginia Mason Research
Center,Seattle.Washington.
116 Round Table Discussion
-
-
.-.
-
successfully used rather than the 100 to 300 ft com-
monly used.Tubing dead space may be reduced by
inserting stainless steel wire.Use of a pressure
transducer in place of the gauge adapts the satu-
rometer for continuous monitoring (see paper by
D'Aoust,this conference).In addition,a technique
for pre-pressurization of the saturometer to the
expected final reading was discussed.This aids in
reducing the equilibration time.
The microgasometric method of Scholander
et al.(Bioi.Bult.109:328-334;1955)was mentioned
as being intermediate in difficulty between the
saturometer and Van Sryke methods.The apparatus
is portable and can be used in the field;however,
it does not appear to have been widely used in gas
bubble disease rese,arch and may deserve further
investigation.
The Van Slyke manometric method has been
widely used in conjunction with a Winkler titration
to report data on oxygen and nitrogen saturation.
It is generally agreed to require high-level compe-
tence to operate and is not readily portable.In
addition,mercury is used in relatively large quan-
tities presenting a health-safety problem.Partici-
pants agreed,however,that it is a highly accurate
method and is useful for calibration of other
methods,as well as for primary data collection.
Gas chromatography offers several advantages
including rapid analysis of multiple samples.It is
not portable and is relatively costly although it
was reported that Carle offers a suitable chromato-
graph for Jess than $1000 (without integration).
Use of electronic peak area integration greatly
increases accuracy and precision.A disadvantage
is the requirement for calibration with each use;
however,development of a simplified electrolytic
calibration is continuing and a recently developed
ultrasonic detector offered by Tracor is claimed to
eliminate external calibration for gas analysis.
Finally,a technique of continuous monitoring
devices indicated that additional research in this
area is needed and that the capability would be
useful for determining the dissolved gas regime in
the river.
Round Table Discussion 117
L
L Physics of ICO~~~~i~:~~rdson
R.BacaDissolvedGases
and Engineering Solutions
PHYSICS OF NITROGEN
SUPERSATURATION
The condition of supersaturation occurs when water
falling over a spillwiily entrains large volumes of
air as it plunges into the stilling basin.Atmospheric
gases are driven into solution by the high pressure
of the impacting water.Because the condition of
supersaturation is a chemically unstable condi-
tion,a natural degasification process occurs which
releases the excess g.IS in solution.The rate of gas
release across the air-water interface is generally
controlled by atmospheric pressure and water tem-
perature.The degasification rate can be estimated
from the relation:
MODELING OF GAS REGIMES
There are two distinct gas regimes which require
modeling,namely the near-field and far-field re-
gimes.The near-field program involves describing
the processes of entrainment and supersaturation
which occur in the stilling basin.This problem
represents a considerable challenge to any modeling
effort because it involves a description of a highly
turbulent phenomenon.The far-field problem refers
to the problem of describing motion and distribu-
tion of nitrogen-supersaturated water given the
level of supersaturation at the stilling basin.This
problem has been modeled with considerable
success using one-dimensional transport models.
C s =23 -0.55808 T +0.00763 T 1 (2)
where the temperature,T,is given in degrees
centigrade.The rate coefficient,k,is a function of
temperature,the degree of turbulence and the
interfacial area over which the gas transfer occurs.
One of numerous correlations for k proposed in
the literature is the relation:
where C is the concentration of dissolved nitrogen
(N 2 ),Cs.the equilibrium saturation level and k is a
rate coefficient.The saturation concentration is
principally a function of temperature.At 1 atm
pressure,the nitrogen solubility data is adequately
described by the relatiion:
-
Sr =k(C-C s)
k l zU
1.028 (T-20)
hJ
(1)
(3)
MITIGATION ACTIVITIES
Consideration of methods to reduce the levels
of dissolved gas supersaturation in the Columbia-
Snake River systems caused by operation of hydro-
electric facilities led the Corps of Engineers to two
approaches:A reduction in spill volume and modifi-
cation of spillways to reduce the degree of air
entrainment to stilling basin depths.Several methods
of achieving these goals were discussed,including:
•Additional upstream storage reservoirs to
reduce the flow during the run-off season and re-
lease the water later in the year.However,there is
much opposition to construction of additional dams
due to their anticipated environmental impact.
•Increase water flow through power houses,
thereby reducing the volume passing over the spill-
way.There are two methods to do this;either by
installing additional generators in skeleton bays of
the Snake River dams or by passing water through
the skeleton bays which have no turbines.Both
approaches have been used,but present plans are
to install a full complement of turbines by 1979.
The use of slotted bulkheads to break the force of
the water was tested and successfully aided in re-
where Dz is the film diffusion coefficient,U is the
average flow velocity and h the mean water depth.
118 Round Table Discussion
Richardson:Corps of Engineel'5.Walla Walla Washington;
and Baca:Battelle-Northwest,Richland,Washington.
..-
dudng levels of supersaturation;however,fish pas-
sage through the bulkheads resulted in a high (50%)
mortality rate and their use was discontinued
during periods of downstream salmonid migration.
•Spillway deflectors to reduce plunging
were designed and tested.These have resulted in a
substantial reduction of saturation levels below
the spillways,and the deflectors are planned for in-
stallation at several dams in the next few years.The
deflectors are designed to break the fall of spilling
water and deflect the water and entrained air across
the surface of the stilling basin rather than per-
mitting it to plunge to depths where hydrostatic
pressure forces the air to dissolve into the water.A
constraint on the design was to permit normal
plunge basin operation during very hjgh flow years
to dissipate the energy of the spilled water and pre-
vent structural damage to the dam.Thus,the de-
flectors were designed for a 1-in-10-yr flood.Tests
indicated up to a 15%reduction in saturation values
when the forebay was normally saturated and a 10
to 20%degasification when the forebay was super-
saturated at Lower Monumental Dam.Tests of coho,
chinook,and steelhead survival passing over
normal spillway bays and those with deflectors
indicated results ranging from no significant dif-
ference to greatly improved survival with the
deflectors.
A final method of mitigation which was dis-
cussed was transportation of salmon ids around the
dams.In this scheme,which has been tested with
good results,screening devices are installed before
the turbines of an upstream dam and downstream
migrants transported by truck to a location below
Bonneville Dam.Initial tests are favorable with a
greatly increased rate-of-return.Estimates are that
40%of the migrants could be captured at a given
Snake River dam,meaning that about two million
chinook and two to three million steel head
would be transported.Research on screening con-
figuration is continuing to minimize capture dam-
age (primarily scaling).
Round Table Discussion 119
.~..
J;
J
.f
H.'.'.'.JJ
Water Quality
Standards
Based on a review of 20 papers dealing with the
biological effects of total dissolved gas supersatu-
ration on aquatic organisms,a group consisting
of Rob RuJifson,Ron Pine.Ed Quan,and Gene
Ralston representing EPA and state regulatory agen-
cies of Washington,Oregon and Idaho,respec-
tively,concluded that total dissolved gas super-
saturation is only one of many factors affecting
the fisheries of a river system where hydroelectric
projects are involved.During high flow years
supersaturation is a predominant factor causing
juvenile mortalities.Prototype tests with spillway
deflectors indicate they contribute minimally to
total losses;however,it is generally agreed that
the 110%level of total dissolved gas supersatura-
tion will probably never be reached.A level of 115%
supersaturation can be tolerated by juvenile sal-
monids considering the compensatory effects of
hydrostatic pressure and travel time in the river
system which will act to minimize mortality.There-
fore,the group conciluded,"the total dissolved gas
standard in the Columbia and Snake Rivers could
be raised,to 115%without causing significant
mortal ities."
Comments and discussion were solicited from
the round table participants and the resulting dis-
cussion raised several questions concerning the
appropriateness of the value 115%as a criterion to
provide reasonable protection of the most desir-
able water use and the riverine ecosystem as well
as attributes of a strong enforceable water quality
standard.
There was no !iubstantial opposition to the
recommendation of "115%as a criterion for protec-
tion of juvenile salmcmids from direct lethal effects,
but it was suggested that the proposed value would
not adequately protect shallow water resident
organisms,in partlicular,invertebrate benthos
which are important food chain organisms,and
that 110%would provide reasonable protection
for them.In addition,levels below 115%may re-
sult in as yet undefined sublethal effects.On the
other hand,several suggested that even higher
levels would protect fishes in some locations given
sufficient hydrostatic pressure compensation,but
detailed knowledge IOf depth distribution of fishes
is not yet available.
120 Round Table Discussion
I Co-Chairmen
R.L.Rulifson
R.Pine
Provisions suggested for inclusion in an en-
forceable standard included:
•Methodology:to be employed in measuring
dissolved gas levels.This should include computa-
tions,sampling locations,and exclusion of water
vapor pressure from total gas tension calculations.
•The area or extent of permitted levels of
excess dissolved gas tension should be defined,
including the zone allowed for mixing of power-
house and spillway waters.
•Deviations from a basic enforcement level
might be permitted for a specified duration.
•Temperature might be a factor involved in
a standard if studies of combined effects indicate
a synergistic effect of supersaturation and tem-
perature.
• A flexible system might be developed which
would provide a separate enforcement level for
each site based on species composition,water
depth,and minimum practicable dissolved gas
levels.
•Relevancy to regions beyond the Pacific
Northwest should be considered carefully as the
standard or criteria may become nationally
accepted.
A general consensus was that a level of 115%
would protect salmonid fishes migrating through
the Columbia-Snake River systems but that a lower
level (110%)should be adopted as a criterion for
protection of shallow-water benthos.As an en-
forceable standard,115%total gas saturation
appeared to be agreeable for the Columbia-Snake
River system as a realistically enforceable value
except during particularly high flow years.
Rulifson:United States Environmental Protection Agency,Water
Quality Management Section,Seattle,Was~ington;.and Pine:
Washington Department of Ecology.Olympia,Washington.
List of
Participants
Dick Allen Ron Bush Duane H.Fickeisen
Washington Dept.of Fisheries Corps of Engineers Battelle-Northwest-Wenatchee,WA Seattle,WA Richland,WA
Bob D.Anderson Alexander Ciesluk Ronald R.Garton
Washington Water Power Texas Instruments Environmental Protection
~Spokane,WA Buchanan,N.Y.Agency-~
Corvallis,OR
Chuck Apts M.J.R.Clark
Battelle-Northwest,Water Resources Service Chuck Gibson
Sequim,WA Victoria,B.C Battelle-Northwest
Sequim,WA
Henry Aus Alice M.Clay
Washington State University New England Aquarium Earl E.Gjelde
Albrook lab Boston,MA Bonneville Power Admin.
Pullman,WA Portland,OR..-Brian G.D'Aoust
Robert Baca Virginia Mason Research Robert H.Gray
Battelle-NorthwE!st Center Battelle-Northwest
.-Richland,WA Seattle,WA Richland,WA
C E.Bagnall Donald Danielski Floyd Hall
Grant County PUD Yankee Atomic Electric Co.Corps of Engineers
,1Il1~Ephrata,WA Westboro,MA Portland,OR
Constance von Barloewen larry Davis R.W.Hanf,Jr.
,-B.C Fish &Wildlife u.s.National Marine Fisheries Battelle-Northwest
I Victoria,B.C longview,WA Richland,WA
Ron Bates John Douglas Donald Hauck
Seattle City light Washington Game Dept.Chelan PUD
Seattle,WA Olympia,WA Wenatchee,WA
..-CD.Becker Paul F.X.Dunigan William F.Hettler
Batte II e-Northwest U.S.Atomic Energy Commission U.S.National Marine Fisheries
Richland,WA Richland,WA Atlantic Estuarine Fisheries
Center
~Kirk Beiningen Wes Ebel Beaufort,N.CI
Oregon Fish Commission U.S.National Marine Fisheries
Clackamas,OR Seattle,WA Thomas Jenkins•U.S.Army,.-
W.W.Bentley Mike Erho Cold Regions Research &
U.S.National Marine Fisheries Douglas County PUD Engineering lab
.-Pasco,WA Wenatchee,WA Hanover,N.H .
Donald Beyer R.B.Fairbanks Charles Junge
Fisheries Research Institute Mass.Division of Marine Fisheries Commission Oregon
~University of Washington Fisheries Clackamas,OR~
Seattle,WA Sandwich,MA
121
~,
,..-~
Stan Katkansky Bruce Monk Lynwood S.Smith
Portland General Electric U.s.National Marine Fisheries Fisheries Research Institute
Portland,OR Seattle,WA University of Washington
Seattle,WA
Steve Kiesser Jerry C.Montgomery,-
Battelle-Northwest Battelle-Northwest Wendell E.Smith
Sequim,WA Richland,WA Idaho Power Co.
Boise,10
Kenneth B.Kral Alan V.Nebeker
Tacoma City Light Environmental Protection George R.Snyder
Tacoma,WA Agency U.s.National Marine Fisheries
..-Corvallis,OR Seattle,WA
Morris Larson
Corps of Engineers Tim Newcomb Richard Stroud
Portland,OR U.S.National Marine Fisheries Dept.of Veterinary Medicine
~Seattle,WA Oregon State University
David l.legg Corvallis,OR
Corps of Engineers Thomas l.Page
~Portland,OR Ba ttelle-Northwest John S.Thompson
Richland,WA Corps of Engineers
Bernie Leman Seattle,WA
Chelan PUD Ron Pine
Wenatchee,WA Washington Dept.of Ecology Burton E;Vaughan
Olympia,WA Battelle-Northwest
Rocco A.Marcello Richland,WA
~Boston Edison Co.G.C.Richardson
Boston,MA Corps of Engineers Charles R.Weaver
Walla Walla,WA U.S.National Marine Fisheries
Bruce May Portland,OR
Montana Dept.of Fish &Game Robert R.Rucker
libby,MT u.s.National Marine Fisheries Douglas Weber
Seattle,WA U.s.National Marine Fisheries.-Daniel Mayercek Seattle,WA
Texas Instruments Robert l.Rulifson
Buchanan,N.Y.Environmental Protection Edward Weiss
Agency Pacific Power &Light
Robert McConnell Water Quality Management Portland,OR
U.S.National Marine Fisheries Section
Longview,WA Seattle,WA Donald Weitkamp
Parametrix,Inc.
Tom Meekin Bill Sandeen Seattle,WA
Washington Dept.of Fisheries Federal Power Commission
Olympia,WA San francisco,CA
John W.Meldrim Mark J.Schneider
Ichthyological Associates Battelle-Northwest
Middletown,MD Richland,WA
,~
122 Participams
Index
Avoidance,11,75
Blood chemistrY',96
Blood dotting mechanisms,93
Cascading bubble effect.66
Condition factor,1
Continuous monitoring,gCls stripping,
101
tensiometer (saturometer),106
Degasification rate,'1
Depth compensation,chinook
salmon,1,11,24
crappie,11
cutthroat trout,11
smelt,11
steel head trout,1,11
Detection and avoidance,1',75
Echogram,20
Equilibration rate,fish-water,47
water-air.11
Exercise,effect on toleranc:e,66
Gas bubble formation dynamics,47
Intermittent exposure,chinook
salmon,",24
coho salmon,"
cutthroat trout,'1
largemouth bass,11
rainbow trout,l'
steelhead trout,11
whitefish,11
Jntestinal motility,66
lateral line function,89
live cage bioassay,24
Naturally occurring super-
saturation,37
Oxygen-nitrogen ratio,effect on
tolerance,85
Pathogenesis,aquatic invertebrates,
51
chinook salmon,24
menhaden.75,8'
steel head,66
Power plant discharge,gas
supersaturation,75
Predation,41
Pumping supersaturated water.101
Recovery from gas bubble disease,
1,24
Size,effect on tolerance,85
Smoltification,1
Stomach content analysis,41
Temperature.effect on tolerance.72
Tolerance of,black bullhead.72
chinook salmon,1.11,24
coho salmon.'1
crappie.11
crayfish.51
cutthroat trout,11
daphnids,51
largemouth bass,l'
menhaden,a1
rainbow trout,l'
smelt,'1
sQuawfish,4'
steel head trout,"11,51
stoneflies,51
whitefish,11
Vertical distribution of fish,1.20
NOTICE
This book was prepared as an account of work sponsored by the United States
Government.Neither the United States nor the United States Energy Research
and Development Administration,nor any of their employees,nor any of their
contractors,subcontractors,or their employees,makes any warranty,express or
implied,or assumes any legal liability or responsibility for the accuracy.
completeness or usefulness of any information,apparatus,product or process
disclosed,or represents that its use would not infringe privately owned rights.
123
1
L
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