a

oo oe

\ v ee

eens, o—
re e. o-
—~e.
—*—~7._

pee ie

i a

b My. o—e—o—e—

WATER TEMPERATURE, C
+

we
T

Fs a a Ca a a

En a .
rN 4
= — a
Efe Ne
. > 7 “ oe. eee. -e
= E ~g~—9--% ~~. “ ° eo a te,
= é eo -e. 2 e.
a os Sa9-- a _
No
\
0-1 74 te iz ‘sie ~~—«920—~S~SC~«

TIME, b

Figure 4. Water temperatures and dissolved oxygen concentrations measured 5 cm
from the bottom by two Delta recorders in a northern pike spawning
marsh, A.-28 April 1974, day of highest water temperature. B.-Average water
temperature from 23 April-12 May 1974. C.-23 April 1974, day of lowest water
temperature. D.-24 April 1974, day of highest dissolved oxygen. E.-Average
dissolved oxygen from 23 April-12 May 1974. F.-Average dissolved oxygen from
1-4 May 1974, indicating an increase in dissolved oxygen before sunrise, fol-
lowed by a decrease at sunrise. G.-28 April 1974, date of lowest dissolved oxygen.

il


APPENDIX D (continued)

TABLE D-2. SURVIVAL OF SMALLMOUTH BASS SAC LARVAE EXPOSED 1 DAY AFTER HATCH-
ING FOR 6 h AT 17 C TO DISSOLVED OXYGEN CONCENTRATIONS LISTED
Ten Sac Larvae Initially in Each Chamber. Treatment Given on
30 May, and Survival Recorded 2 Days Later.

Duncan's Multiple Range Test®
Average Dissolved Oxygen

(mg/liter) 0.5 2.2 4.2 Control 9.0
Range of Dissolved Oxygen
(mg/liter) 0.3-0.6 2.0-2.5 3.5-4.5 Control 8.5-9.5
Percent Survival 0 53 73 77 83
-05
-01
Analysis of Variance”
sov df 88 ms F
Treatments 4 138.267 34.567 7.103**
Error 10 48 .666 4.867
Total 14 186.933

"Underlined values not different at 0.05 and 0.01 levels.

bpecause of zero readings, 1 was added to all values for computation
of ss. :

(continued)

32


TABLE C-1.

APPENDIX C
RESULTS OF NORTHERN PIKE OXYGEN TOLERANCE EXPERIMENTS

HATCHING SUCCESS OF NORTHERN PIKE EMBRYOS EXPOSED

7 DAYS AFTER FERTILIZATION FOR 8 h AT 10 C TO
DISSOLVED OXYGEN CONCENTRATIONS LISTED

Fifteen Embryos Initially in Each Chamber. Exposures
Made on 7 May, and Survival Recorded 5 Days Later.

Average Dissolved Oxygen

(mg/liter) 0.6 1.8 2.6 8.0 Control
Range of Dissolved Oxygen

(mg/liter) 0.5-0.7. 2.0-2.0 2.5-4.5 7.0-9.5 Control
Percent Survival 77 80 93 83 80
Days to First Hatch 10 10 10 10 10
Days to 90% Hatch 11 11 11 1 11
Analysis of Variance

s0V df 88 ms FE
Treatment 4 14.800 3.700 1.542 N.S.
Error 15 36.000 2.400
Total 19 50.800
TABLE C-2. SURVIVAL OF NORTHERN PIKE SAC LARVAE EXPOSED 1 DAY AFTER HATCH-

ING FOR 8 h AT 8-11 C TO DISSOLVED OXYGEN CONCENTRATIONS LISTED
Fifteen Sac Larvae Initially in Each Chamber. In Two Experiments
Survival Recorded 8 and 4 Days After Treatment.

Duncan's Multiple Range Test*
Average Dissolved

Oxygen (mg/liter) 0.3 2.0 Control 4.8 10.1 Control
Range of Dissolved .

Oxygen (mg/liter) 0.2-0.5 1.5-2.0 Control 3.0-6.5 9.0-11.0 Control
Average Length (mn) 10.63 10.85 10.73 10.87 10.83 10.73
Percent Survival> 51 87 91 91 92 97

05 _
01 _
(continued)

28


WATER
——_, a
LEVEL SAMPLE
TTLE
SUPPORT ROD WEIGHTED PAIL

SCREENED TYGON TUBING

OPENING ~~

TUBING CLAMP
CONNECTOR

Siphoning apparatus to collect water samples.
5

Figure 1.


Figure 6.

nee 0-0-0,
. te

° ‘.
| 2 1uLY
= + -0--0--0- -0--0- 0)
& ooo
= . AVERAGE - a —.,
Bim \
en, °
: 20, ob co
=

Se -e -0--0

17 JUNE,

15) De--0--0--0--

10|

DISSOLVED OXYGEN, mg/liter

a a ee ee SC SCC ey
O41 ue 1-8 11-12 15-16 19-20 23-24

TIME, &

Water temperatures and dissolved oxygen concentrations measured
5 cm from the bottom by two Delta recorders in bluegill spawning
areas from 9 June-5 July, 1974. Broken lines represent days of
highest and lowest temperatures and dissolved oxygen concentra-
tions.

15


APPENDIX D (continued)

TABLE D-3. HATCHING SUCCESS OF BLUEGILL EMBRYOS EXPOSED 1 DAY AFTER FERTI-
LIZATION FOR 4 h AT 25-28 C TO DISSOLVED OXYGEN CONCENTRATIONS
LISTED
Fifteen Embryos Initially in Each Chamber. Survival Recorded
1 Day After Treatment When Hatching was Complete.

Average Dissolved Oxygen

(mg/liter) 0.5 2.1 3.0 7.8 Control
Range of Dissolved Oxygen

(mg/liter) 0.5 1.5-2.5 2.5-3.5 7.5-9.5 Control
Percent Survival? 40 46 50 47 44

Analysis of Variance

Sov df ss ms FE
Subgroups 9 20.700 4.140
Treatments 4 7.533 1.883 0.824 N.S.
Experiments 1 4.033 4.033 1.491 N.S.
Interaction 4 9.134 2.284 0.845 N.S.
Error 18 48 .667 2.704
Total 27 69.367

*Percentages given are for combined survival rates of two experiments.

(continued)

33


FOREWORD

Our nation's freshwaters are vital for all animals and plants, yet our
diverse uses of water---for recreation, food, energy, transportation, and
industry---physically and chemically alter lakes, rivers, and streams. Such
alterations threaten terrestrial organisms, as well as those living in water.
The Environmental Research Laboratory in Duluth, Minnesota develops methods,
conducts laboratory and field studies, and extrapolates research findings

--to determine how physical and chemical pollution affects
aquatic life

--to assess the effects of ecosystems on pollutants

--to predict effects of pollutants on large lakes through
use of models

--to measure bioaccumulation of pollutants in aquatic

organisms that are consumed by other animals, including
man

This report provides measurements of fluctuations of dissolved oxygen and
water temperature adjacent to embryos and sac larvae of northern pike,
bluegills, and pumkinseeds in their natural spawning sites. It also
describes field experiments designed to determine the effects of brief
exposure of various concentrations of dissolved oxygen on early life stages
of northern pike, bluegill, and smallmouth bass in their natural habitat.

Donald I. Mount, Ph. D.

Director

Environmental Research Laboratory
Duluth, Minnesota

iii


APPENDIX B

DISSOLVED OXYGEN CONCENTRATIONS AND WATER TEMPERATURES IN
CENTRARCHID NESTS DURING EARLY-LIFE STAGES

MEASUREMENTS TAKEN AT 0800-0900 AND AT 1600-1700 CDST AT THE SURFACE AND AT
10 and 1 cm FROM THE BOTTOM AT UP TO 3 STATIONS IN 1973 AND 1974. MEAN
WATER DEPTHS WERE 0.7 m AND RANGED FROM 0.27 TO 1.07 m.

TABLE B-1. DISSOLVED OXYGEN CONCENTRATIONS (mg/liter) IN BLUEGILL
AND PUMPKINSEED NESTS DURING EMBRYO AND SAC LARVA
STAGES, AND THE FREQUENCY AT WHICH VARIOUS DISSOLVED
OXYGEN CONCENTRATIONS OCCURRED

Dissolved Oxygen

Morning Afternoon

Depth No. Average S.D. Range No. Average S.D. Range

Surface 145° 7.7 1.31 4.8-13.2 144? 12.1 1.76 7.1

10 145 7.5 1.35 4.6-13.5 144 12.2 1.99 6.8

1 145 1.57 2.4-12.6 144 11.5 2.53 4.8-
4.8

7.1
Total 435 7.5 1.43 2.4-13.5 432 12.0 2.13 -8-

Frequency of Dissolved Oxygen

Morning Afternoon
Dissolved lo Total lo Total
Oxygen
fug/liter) No. % No. _% No. % No. _%
2.1- 3.0 2 1.4 2 0.5 0 0 0 0
3.1- 4.0 3 2.1 3 0.7 0 0 0 0
4.1- 5.0 8 5.5 12 2.8 3. 2.1 3 0.7
5.1- 6.0 17. «11.7 43 9.9 2 «21.4 2 0.5
6.1- 7.0 39 26.9 109 9.25.1 3 2.1 5 1.2
7.1- 8.0 34 23.4 11l (25.5 1 0.7 3 0.7
8.1-:9.0 30 420.7 109 -25.1 5 3.5 12 2.8
9.1-10.0 8 5.5 32 7.3 16 «11.1 34 7.9
10.1-11.0 2 1.4 8 1.8 25 17.4 74 17.1
11.1-12.0 1 0.7 3 0.7 28 «19.4 95 22.0
12.1-13.0 1 0.7 1 0.2 20 13.9 75 17.4
13.1-14.0 0 0 2 0.5 13° (9.0 59 13.7
14.1-15.0 0 0 0 0 17°«:11.8 40 9.3

(continued)

26


temperatures for combined depths 10 and 1 cm from the bottom in 1974 ranged
from 3.5-12.8 C during the embryo, 5.0-11.2 C in the sac larva, and 5.0-9.0 C
in the larva stages; in 1975 the ranges were 3.5-8.5 C, 6.2-14.5 C, and
8.8-16.2 C. Water temperature fluctuations from morning to afternoon were
greater in 1974 than in 1975.

Embryo

Because embryos and sac larvae were observed on vegetation, dissolved oxygen
concentrations and water temperature measured in 1974 and 1975 at 1 and

10 cm from the bottom are used to describe conditions experienced by these
stages. Dissolved oxygen concentrations during the northern pike embryo
stage ranged from 0.3-16.6 mg/liter 1 cm from the bottom and 1.2-15.0 at

10 cm (Appendix A, Table 1). Mean dissolved oxygen concentrations at 10 and
1 cm, respectively, were 6.0 and 3.5 mg/liter in the morning and 9.3 and 6.9
mg/liter in the afternoon. When measurements at the 10 and 1 cm levels were
combined, concentrations less than 1.0 mg/liter occurred in 10% (6 out of 60)
of the morning measurements or 5 of the 15 mornings; concentrations of
1.1-2.0 mg/liter occurred in 18% of the morning measurements or 7 of the 15
days. Supersaturation occurred in 14% (9 out of 64) of the afternoon
measurements or 6 of 16 afternoons.

Water temperatures from combined measurements 10 and 1 cm from the bottom
ranged from 2.5-16.0 C (Appendix A, Table 2). Mean water temperatures at 10
and 1 cm, respectively, were 6.5 and 5.6 C in the morning and 8.0 and 6.6 C
in the afternoon.

Sac Larvae

During northern pike sac larva stages in 1974 and 1975, dissolved oxygen
concentrations ranged from 0.0-13.4 mg/liter (Appendix A, Table 1). Mean
dissolved oxygen concentrations at 10 and 1 cm, respectively, were 4.4 and
2.2 mg/liter in the morning and 7.6 and 5.6 mg/liter in the afternoon. Of 74
morning measurements at 10 and 1 cm, concentrations of 1.0 mg/liter and less
occurred in 24% of the measurements (13 of the 18 days); concentrations of
1.1-2.0 mg/liter occurred in 9.5% of the morning measurements (9 of the 18
days). Of 72 afternoon measurements supersaturation occurred in 9.8% (6 of
the 18 days). .

Water temperatures in 1974 and 1975 from combined measurements 10 and 1 cm
from the bottom ranged from 4.0-17.0 C (Appendix A, Table 2). Mean water
temperatures at 10 and 1 cm, respectively, were 8.5 and 8.0 C in the morn-
ings and 10.9 and 9.7 C in the afternoons.

Larvae

In 1974 and 1975, dissolved oxygen concentrations ranged from 0.0 at 1 cm
from the bottom to 12.0 mg/liter at the surface (Appendix A, Table 1). Mean
dissolved oxygen concentrations at 10 and 1 cm, respectively, were 3.8 and
1.9 mg/liter in the morning and 6.4 and 4.2 mg/liter in the afternoon; at the
surface means were 5.8 (morning) and 8.3 mg/liter (afternoon). Although
concentrations of 2.0 mg/liter and less occurred in 25% of the morning

12


Figure 8.

L __
80) ‘ ° tn
L Y eer oo)
eon
L . y
4
col Yo
%
L ; SHALLMOUTH BASS
el 1
+ /
/ .
L fa
4
me fa
/
, Lfe
= 4
z ee
3
.
= wm,
3
=
= be
aL
-e
ro tase 2 ee
ol =} °
i
L & ' e-
5 ee
a 7 °
~~
foot
‘
|
'
re
'
i Se a
12 3 «4 8 © 7 8 § 10 Comme

Survival of smallmouth bass and bluegill embryos and sac larvae
exposed for 6 h and for 4h, respectively, to various concentra-
tions of dissolved oxygen. A.-Hatching success of smallmouth bass
embryos exposed 2 days after fertilization. B.-Survival of
smallmouth bass sac larvae exposed 1 day after hatching. C.-
hatching success of bluegill embryos exposed 1 day after fertili-

zation. D.-Survival of bluegill sac larvae exposed 1-4 days after
fertilization. Stars indicate significant differences from
controls. Detailed information in Appendix D.

18


91

»
&

<
= Jf
=
> |“
= 4 al.
& | Z
a *
3 20, “AN
= /
= Nw
% 16,
Fo fd \ . La Fore
18, r
|
. t fj
15, t
ry
: . if /
5 i
= o\ VV ‘
= 12h , ¢
oa of L
z | Xs sel . \
3s f \, b
o | 5 J¥
\ d .
5 Le ‘. ; . 1s fy 4, :
of i HK . 7 “A 1 h
> Pr aly toe | rg | it
2 1 yy wily ‘, ‘ \ it
af . ! .'. \ iH 1 ee
B 6 af er . . be e aw ‘ease |S
a wy Bs . vy ° H
, r ‘.
t b
10 20 30 10 1 10 we 20 30 yy 10
JUNE 1973 JULY Jul 1974

Figure 5.

TIME, days

Mean water temperatures ana dissolved oxygen concentrations 1 cm trom the bottom in bluegill
and pumpkinseed nests. Broken line represents measurements at 0800-0900 (CDST), and solid line

represents measurements at 1600-1700.

Each point represents the mean determined from 1-3 nests.


APPENDIX B - TABLE B-1 (continued)

Frequency of Dissolved Oxygen

Dissolved
Oxygen
{mg/Liter)
15.1-16.0
16.1-17.0
17.1-18.0
18.1-19.0
19.1-20.0

Sum

Morning Afternoon
lo Total Loa Total
No. _% No. _% No. _% No. %
0 ti) 0 0 7 4.9 18 4.2
0 0 0 0 1 0.7 6 1.4
0 0 0 0 2 1.4 4 0.9
0 0 0 0 1 0.7 1 0.2
(0) 0 0 0) 0) 0 1 0.2
145 100 435 100 144 100 432 100

499 mornings from 4 June to 5 July in 1973 and 31 mornings from
7 June to 8 July in 1974.

bos afternoons from 4 June to 5 July in 1973 and 28 afternoons from
7 June to 8 July in 1974.

TABLE B-2. WATER TEMPERATURES (C) IN BLUEGILL AND PUMPKINSEED NESTS
DURING EMBRYO AND SAC LARVA STAGES

Water Temperature

Morning Afternoon
Depth No. Average S.D. Range No. Average S.D. Range
Surface 145 19.7 2.60 14.0-25.0 144 23.7 2.73 18.0-29.0
10 145 20.0 2.59 15.0+26.5 144 23.3 2.42 18.5-27.5
1 145 20.0 2.54 15.0-25.5 144 23.3 2.44 18.5-27.5
Total 435 19.9 2.58 14.0-26.5 432 23.4 2.53 18.0-29.0

27


Figure 2.

15 SURFACE

Empryo

% 10 CM FROM BOTTOM

DISSOLVED OXYGEN, mgytiter

2, 1 CM FROM BOTTOM

2 2020 ape 3 ; 1© way 20

TIME, days

Dissolved oxygen concentrations in a northern pike spawning
marsh. Each point represents the mean determined from measure-
ments at two stations at the surface, and at 10 and 1 cm from
the bottom. Broken line represents measurements at 0800-0900
(CDST), and solid line represents measurements at 1600-1700.


SECTION 1

INTRODUCTION

In their extensive review of dissolved oxygen requirements for fishes,
Doudoroff and Shumway (1) reported that the contradictory results in many of
the studies could be lessened through controlled experiments in which natural
conditions are closely simulated. Our purpose was to contribute to the design
of such experiments at the Environmental Research Laboratory--Duluth,
Minnesota by providing measurements of fluctuations of dissolved oxygen and
water temperature adjacent to embryos and sac larvae of northern pike (Esox
lucius), bluegills (Lepomis macrochirus), and pumpkinseeds (Lepomis gibbosus)
in their natural spawning sites. We studied embryos and sac larvae because
they are relatively immobile and therefore unable to avoid exposure to
extremes, and because there is a lack of detailed information on dissolved
oxygen concentrations and water temperatures encountered by these stages.

To assess how low dissolved oxygen concentrations observed in the field
influenced hatching success and survival we conducted field experiments with
embryos, sac larvae, and larvae of northern pike, and embryos and sac larvae
of bluegills and smallmouth bass (Micropterus dolomieui). Except for the
brief exposure times to various levels of dissolved oxygen, all fish were
kept in their natural surroundings where they were exposed to fluctuating
environmental factors; differences in survival from controls were then
attributed to effects of dissolved oxygen concentrations used in the treat-
ments.


APPENDIX A - TABLE A-1 (continued).

Frequency of Dissolved Oxygen in the Marsh

Morning Afternoon
Dissolved
Oxygen Embryo Sac Larva Larva Embryo Sac Larva Larva
(mg/liter) No. % No. % No. h No. % No. % Now %
0.0- 1.0 6 6.7 18 16.2 10 14.5 1 1.0 2 1.9 0 0
1.1-2.0 11 12.2 7 6.3 7 10.1 4 4.2 4 3.7 3 4.5
2.1- 3.0 4 4.4 9 8.1 6 8.7 5 5.2 6 5.6 6 9.1
3.1- 4.0 7 7.8 12 10.8 10 14.5 2 2.1 392.8 4 6.1
4.1- 5.0 10 11.1 10 9.0 16 23.2 1 1.0 6 5.6 6 9.1
5.1- 6.0 11 12.2 20 18.0 9 13.0 6 6.3 11 10.2 10 15.2
6.1- 7.0 8 8.9 27 24.3 7 10.1 9 9.4 9 8.3 12 18.2
7.1- 8.0 7 7.8 #7 6.3 3 4.3 9 94 12 11.1 7 10.6
8.1- 9.0 11 12.2 1 0.9 2-L 1.4 6 6.3 19 17.6 11 16.7
9.1-10.0 11 12.2 0 0 60 1) 17°:17.7—) 20 18.5 3 4.5
10.1-11.0 2 2.2 .0 0 0 0 12°12.5 10 9.3 1 1.5
11.1-12.0 1 1.1 0 0.698 0 7 7.3 2 51.9 3 4.5
12.1-13.0 1 1.1 0 0 Oo Y) 3° 3.1 2 =#1.9 0 0
13.1-14.0 0 0 (0 0 oO 0 3 3.1 2 °=#1.9 #0 0
14.1-15.0 tt) o Oo o 90 0 7 7.3 ce) 0 Oo 0
15.1-16.0 0 0 Oo 0 oO 0 3 3.1 (0) 0. 60 0
16.1-17.0 0 o 0 o 0 0 2 10 0 o 0 0
Totals 90 100 111 100 69 100 96 100 108 100 66 100

87 mornings from 23-30 April in 1974 and 8 mornings from 25 April -
2 May in 1975.

bg afternoons from 23-30 April in 1974 and 8 afternoons from 24
April - 1 May in 1975.

a) mornings from 1-8 May in 1974 and 10 mornings from 3-12 May in
1975.

43 afternoons from 1-8 May in 1974 and 10 afternoons from 2-12 May
in 1975.

*7 mornings from 9-16 May in 1974 and 4 mornings from 13-16 May in
1975.

£ afternoons from 9-16 May in 1974 and 4 afternoons from 13-16 May
in 1975.

24


Centrarchids

In Crane Lake, where centrarchids were studied, dissolved oxygen concentra-
tions and water temperatures during the spawning seasons 1974 and 1975 were
similar and were characterized by low day-to-day fluctuations, little strat-
ification with depth, and generally high concentrations of dissolved oxygen;
supersaturation occurred on 90% of the days sampled. Lows of less than 4.0
mg/liter of dissolved oxygen were found on 10% of the mornings. We found
dissolved oxygen concentrations on cloudy days to be lower than on clear
days, but there were too few cloudy days to make statistical comparisons.

No differences were found in dissolved oxygen inside and outside of bluegill
nests guarded by male fish.

OXYGEN TOLERANCE EXPERIMENTS

For 8 h exposures to various levels of dissolved oxygen, survival was simi-
lar to that in the controls for northern pike embryos treated at concentra-
tions of 0.6 mg/liter, for sac larvae treated at concentrations of 2.0
mg/liter and greater, and for larvae treated at concentrations of 4.0
mg/liter and greater. Apparently, tolerance to low dissolved oxygen con-
centrations decreases from embryo to larva stages. Even the age of sac
larvae influenced tolerance to low dissolved oxygen concentration as no
3-day-old sac larvae survived a 6 h exposure to a concentration of 0.8
mg/liter, whereas 51% of 1-day-old sac larvae survived an 8 h exposure at
0.3 mg/liter. Survival of 3-day-old sac larvae was drastically reduced

by exposure to 0.5 mg/liter of dissolved oxygen for only 2 h.

Siefert, Spoor, and Syrett (5) found that constant exposure to dissolved
oxygen concentrations of about 5.0 mg/liter would permit good survival of
northern pike from the embryo state through first feeding of larvae. Be-
cause of short exposure times to low dissolved oxygen concentrations used in
our experiments, it is difficult to make comparisons with results from other
studies. However, during embryo through larva stages, dissolved oxygen
concentrations at combined depths 1 and 10 cm from the bottom were less than
5.0 mg/liter in 70% of the morning measurements and in 30% of the afternoon
measurements indicating that naturally-occurring dissolved oxygen concen-
trations are frequently lower than concentrations reported necessary for
good survival determined from laboratory experiments.

Hatching success of smallmouth bass embryos exposed for 6 h at concentrations
of 1.0 mg/liter and greater of dissolved oxygen was similar to controls, as
was survival of smallmouth bass sac larvae treated at 2.2 mg/liter and
greater. Smallmouth bass embryos died when exposed to 0.5 mg/liter for 6 h.
Siefert, Carlson, and Herman (6) found that after constant exposure of small-
mouth bass embryos and larvae to various levels of dissolved oxygen, hatch-
ing took place at 2.5 mg/liter but death occurred by the fifth day of expo-
sure; embryos treated with 1.2 mg/liter failed to hatch.

Hatching success of bluegill embryos exposed for 4 h at concentrations of
0.5 mg/liter was similar to controls; for sac larvae, survival was similar to
controls at 1.5 mg/liter and greater. Survival of bluegills sac larvae was
severely reduced by exposure to 0.5 mg/liter of dissolved oxygen for only 2 h.

21


Foreword ..... .
Abstract ......
Figures ......
Acknowledgments .

CONTENTS

1. Introduction . sone . oe
2. Conclusions . . wee .
3. Recommendations oe :
4. Methods. ... ona see
5. Results. ... . woe
6. Discussion . . eae oe
References
Appendices
A. Dissolved Oxygen Concentrations and Water Temperatures
in a Northern Pike Spawning Marsh. .........-
B. Dissolved Oxygen and Water Temperatures
in Centrarchid Nests ..... . wee ee
C. Northern Pike Oxygen Tolerance Experiments .....
D. Centrarchid Oxygen Tolerance Experiments .....


ABSTRACT

Fluctuations of dissolved oxygen concentrations and water temperatures in
their natural spawning sites were measured during embryo through larva
stages of northern pike (Esox lucius), and during embryo and sac larva
stages of bluegills (Lepomis macrochirus) and pumpkinseeds (Lepomis
gibbosus). At northern pike sites, dissolved oxygen concentrations from
combined measurements 1 and 10 cm from the bottom ranged from 0.0-16.6
mg/liter, and water temperatures from 2.5-23.0 C; average daily fluctua-
tions were 3.0 mg/liter and 1.6 C. For bluegill and pumpkinseed nests,
dissolved oxygen concentrations 1 cm from the bottom ranged from 2.4-18.2
mg/liter and water temperatures from 15.0-27.5 C, with average daily
fluctuations of 4.4 mg/liter and 3.3 C. In field experiments to deter-
mine acute effects of a single exposure to low dissolved oxygen concen-
trations, tolerance decreased from embryo to larva stages for northern
pike and from embryo to sac larva stages for bluegills and smallmouth

bass (Micropterus dolomieui).

This report was submitted in fulfillment of Grant Number R801976, by
the Department of Zoology, North Dakota State University, under the
partial sponsorship of the Environmental Protection Agency. Work was
completed August 1976.

iv


2 SURFACE

L oat]
EMBRYO 'SAC LARVA
1

WATER TEMPERATURE, C

16 1 CM FROM BOTTOM

10° may 20

'

1

H red bevels ‘
20 apa «30 10 way 7020 apr 3° gz,

TIME, days

Figure 3. Water temperatures in a northern pike spawning marsh. Each
point represents the mean determined from measurements at two
stations at the surface, and at 10 and 1 cm from the bottom.
Broken line represents measurements at 0800-0900 (CDST), and
solid line represents measurements at 1600-1700.

10


ACKNOWLEDGEMENTS

The authors thank the Minnesota Department of Natural Resources, Detroit
Lakes Fish Hatchery for permitting us to conduct studies at Crane Lake and
the spawning marsh at Silver Lake; the U.S. Fish and Wildlife Service for
use of hatchery ponds at the Valley City National Fish Natchery; and both
the above as well as the North Dakota Game and Fish Department, Spiritwood
Lake Fish Hatchery for providing fish embryos and larvae.

vii


Appendix C, Table 1). Hatching began 10 days after fertilization; ail
embryos had hatched by the 12th day following fertilization.

Sac Larvae

Mean survival of sac larvae, exposed 1 day after hatching for 8 h to dis-
solved oxygen concentrations of 0.3 mg/liter in two experiments was 51%.
This was significantly lower (P < .01) than the 87-97% survival in the high-
er concentrations tested (Fig. 7; Appendix C, Table 2). Mean total lengths
of sac larvae measured 8 days after hatching were similar (P < .05) in the
first experiment; no lengths were measured in the second experiment.

Because survival was reduced when we exposed sac larvae to 0.3 mg/liter of
dissolved oxygen for 8 h, we tried to assess effects of various durations of
exposure to low dissolved oxygen concentrations. No sac larvae treated 3
days after hatching in dissolved oxygen concentrations of 0.8 mg/liter sur-
vived exposure for 6 h, 6% survived for 4 h, 14% survived for 2 h, and 88%
survived in the controls (Appendix C, Table 3). Survival in the controls was
higher (P < .01) than in all treatments; survival in the treatments was not
different at the .01 level, but at the .05 level, survival after 2 h exposure
was greater than after 6 h.

Larvae

All larvae that had just reached the feeding stage died after 8 h in dis-
solved oxygen concentrations averaging 1.6 mg/liter (Fig.7; Appendix C,
Table 4). Survival in controls and in treatments where dissolved oxygen
ranged from 3.5-9.3 mg/liter was 93-100%. High mortality of larvae in con-
trols prevented assessment of survival for more than 1 day following treat-
ment.

OXYGEN TOLERANCE EXPERIMENTS~-CENTRARCHIDS

Smallmouth Bass--Embryo

Two-day-old embryos exposed to 0.5 mg/liter of dissolved oxygen for 6 h died
before hatching (Fig. 8; Appendix D, Table 1). Mean hatching success of
69-91% was similar (P < .01) in higher concentrations tested of 1.8-7.9
mg/liter.

Smallmouth Bass--Sac Larvae

No sac larvae survived at a dissolved oxygen concentration of 0.5 mg/liter
when treated 1 day after hatching for 6 h (Fig. 8; Appendix D, Table 2).
Mean survival of 53-83% was similar (P < .01) in the higher concentrations
tested of 2.2-9.0 mg/liter. In another experiment, survival of sac larvae
treated with dissolved oxygen levels of 1.0-2.0 mg/liter for 6 h was 85%;
this was not significantly different (P < .01) from the 95% survival in con-
trols.

16


EPA-600/3-76-113
December 1976 Ecological Research Series



SECTION 3

RECOMMENDATIONS

Because marked stratification of dissolved oxygen concentrations and water
temperatures occurred in spawning sites of northern pike, we found it
necessary to take samples manually at depths near and just above the bottom
to represent conditions to which embryos and sac larvae were exposed; marked
stratification was not found in bluegill or pumpkinseed nests. We found 20
ml water samples for dissolved oxygen determinations were the smallest we
could take and still retain accuracy. Delta dissolved oxygen recorders
provided useful information regarding daily and seasonal trends and durations
of extremes, but we did not find them consistent enough to provide accurate
measurements. The continuous movement of water caused by the stirring
mechanism affected micro-stratification.

It is feasible to use field-based experiments to determine effects of dis-
solved oxygen concentrations on survival of early life-stages of northern
pike, bluegills and smallmouth bass. However, we found it difficult to
maintain fish from embryo to early feeding stages. Future experiments
should be designed to provide a minimum handling of fish, particularly during
daily counts, in order to minimize shock.


SECTION 2

CONCLUSIONS

In a northern pike spawning marsh, average dissolved oxygen concentrations
and water temperatures measured at combined depths of 1 and 10 cm from the
bottom, increased from morning to afternoon, respectively: 4.8 to 8.1
mg/liter and 6.1 to 7.3 C during the embryo stage, 3.3 to 6.6 mg/liter and
8.3 to 10.3 C during the sac larva stage, and 2.8 to 5.3 mg/liter and 8.2
to 9.8 C during the larva (free swimming) stage. Dissolved oxygen concen-
trations 1 and 10 cm from the bottom were 1.0 mg/liter and less in 7% of the
measurements made during embryo stages and in 18% of the measurements during
sac larva stages; their maximm duration was 10 h in a 24 h period.

In bluegill and pumpkinseed nests, average dissolved oxygen concentrations
and water temperatures 1 cm from the bottom during embryo through sac larva
stages increased from morning to afternoon and were, respectively, 7.1 to
11.5 mg/liter and 20.0 to 23.3 C; values measured at 10 cm from the bottom
and at the surface were similar. Dissolved oxygen concentrations during
embryo through larva stages were 4.0 mg/liter or less on only 5 of 53 days;
supersaturation occurred in the afternoons on 48 of 53 days. The presence
of males guarding the nests did not appear to affect oxygen levels; oxygen
concentrations adjacent to the nests were similar to those inside.

For all species tested, tolerance to low dissolved oxygen concentrations
decreased from embryo to larva stages. While northern pike embryos exposed
for 8 h to 0.6 mg/liter survived as well as controls, larvae required con-
centrations of 2.0 mg/liter and greater, and exposure for only 2 h to 0.8
mg/liter drastically reduced their survival. Larvae exposed for 8 h re-
quired oxygen concentrations of 4.0 mg/liter and greater. For smallmouth
bass exposed for 6 h, embryo hatching success at 1.0 mg/liter and greater
and sac larva survival at 2.2 mg/liter and greater was similar to controls.
Compared with controls, bluegill embryos exposed for 4 h tolerated concen-
trations as low as 0.5 mg/liter, and sac larvae tolerated levels as low as
1.8 mg/liter; survival of sac larvae was drastically reduced by exposure
for 2 h to 0.5 mg/liter of dissolved oxygen.


APPENDIX D (continued)

TABLE D-5. SURVIVAL OF BLUEGILL SAC LARVAE EXPOSED 1-4 DAYS AFTER HATCHING
AT 23-28 C TO LOW DISSOLVED OXYGEN CONCENTRATIONS OF 0.5 mg/liter
FOR TIMES LISTED
Ten Sac Larvae Initially in Each Chamber. Survival Recorded
1 Day After Treatment in Five Experiments.

Duncan's Multiple Range Test®

Treatment Time 4h 3h 2h lh Control
Percent Survival> 0 1 11 71 79
.05
01

Analysis of Variance”

sov df ss ms FE
Subgroups 24 1001.013 41.708
Treatments 4 923.147 230.787 53.320 **
Experiment 4 8.631 2.153 1.217 N.S.
Interaction 16 69.253 4.328 2.447 *
Error 49 86.667 1.769
Total 73 1087 .680

4Underlined values not different at 0.05 and 0.01 levels.
bpercentages given are for combined survival rates of five experiments.

“Because of zero readings, 1 was added to all values for computation
of ss.

35


APPENDIX A

DISSOLVED OXYGEN CONCENTRATIONS AND WATER TEMPERATURES IN A NORTHERN PIKE
SPAWNING MARSH DURING THE EMBRYO, SAC LARVA, AND LARVA STAGES

MEASUREMENTS TAKEN AT 0800-0900 AND AT 1600-1700 CDST AT THE SURFACE AND AT
MEAN WATER

10 AND 1 cm FROM THE BOTTOM AT 2 STATIONS IN 1974 AND 1975.

DEPTHS WERE 0.5 m AND RANGED FROM 0.46 TO 0.58 m.

TABLE Awl. DISSOLVED OXYGEN CONCENTRATIONS (mg/liter) IN A NORTHERN PIKE
SPAWNING MARSH DURING THE EMBRYO, SAC LARVA, AND LARVA STAGES,
AND THE FREQUENCY AT WHICH VARIOUS DISSOLVED OXYGEN CONCENTRA-
TIONS OCCURRED
Dissolved Oxygen
Embryo
Morning Afternoon
Depth No, Average S.D. Range No, Average S.D. Range
Surface 30° 7.4 2.58 2.4-12.4 32 10.3 2.23 5.6-15.8
10 30 6.0 2.76 1,.2-10.4 32 9.3 3.24 8.5-15.0
1 30 3.5 2.65 0.3- 9.6 32 6.9 4.62 0.8-16.6
Total 90 5.6 3.08 0.3-12.4 96 8.9 3.75 0.8-16.6
Sac Larva
Surface 37° 6.4 0.96 3.3- 8.2 364 9.1 1.21 7.4-13.1
10 37 4.4 1.56 1.3- 7.4 36 7.6 2.17 2.8-12.6
1 37 2.2 2.03 0.0- 7.0 36 5.6 3.65 0.6-13:4
Total 111 4.3 2.33 0.0- 8.0 108 7.4 2.91 0.6-13.4
Larva
Surface 23° 5.8 1.24 3.6- 8.4 22F 8.3 1.74 5.5-12.0
10 23 3.8 1.62 1.0- 6.8 22 6.4 1.63 2.9- 9.1
1 23 1.9 1.56 0.0- 4.8 22 4.2 2.34 1.0-11.0
Total 69 3.8 2.14 0.0- 8.4 66 6.3 2.56 1.0-12.0
(continued)

23



REFERENCES

Doudoroff, P. and D, L. Shumway. Dissolved Oxygen Requirements of
Freshwater Fishes. Food and Agriculture Organization of the United
Nations, Rome, Italy. Technical Paper Number 86. 1970. 291 p.

Magnuson, J. J. and W. E. Stuntz. A Siphon Water Sampler for Use
Through the Ice. Limnol. Oceanogr. 15:156-158, January 1970.

Standard Methods for the Examination of Water and Wastewater.
American Public Health Association, New York. 1971. p. 477-481.

Steel, R. G. D. and J. H. Torrie. Principles and Procedures of
Statistics. McGraw-Hill Book Company, Inc., New York. 1960. p. 132-156.

Siefert, R. E., W. A. Spoor, and R. F. Syrett. Effects of Reduced
Oxygen Concentrations on Northern Pike (Esox lucius) Embryos and Larvae.
J. Fish. Res. Board Can. 30:849-852, June 1973.

Siefert, R. E., A. R. Carlson, and L. J. Herman. Effects of Reduced
Oxygen Concentrations on the Early Life Stages of Mountain Whitefish,
Smallmouth Bass, and White Bass. Prog. Fish Cult. 36 (4):186-190,
October 1974.

22


FIGURES

Siphoning apparatus to collect water samples

Dissolved oxygen concentrations in a northern pike spawning
marsh

Water temperatures in a northern pike spawning marsh

Water temperatures and dissolved oxygen concentrations
measured 5 cm from the bottom by two Delta recorders in a
northern pike spawning marsh

Mean water temperatures and dissolved oxygen concentrations
1 cm from the bottom in bluegill and pumpkinseed nests

Water temperatures and dissolved oxygen concentrations
measured 5 cm from the bottom by two Delta recorders in
bluegill spawning areas

Survival of northern pike embryos, sac larvae and larvae
exposed for 8 h to various concentrations of dissolved
oxygen

‘Survival of smallmouth bass and bluegill embryos and sac

larvae exposed for 6 h and for 4 h, respectively, to
various concentrations of dissolved oxygen

10

11

14

15

17

18


measurements, mostly at 1 cm from the bottom, higher oxygen concentrations
were present at other depths.

Water temperatures ranged from 4.0-22.5 C (Appendix A, Table 2). Mean water
temperatures for the three depths ranged from 8.0-8.3 C in the mornings and
from 9,1-13.2 C in the afternoons.

DISSOLVED OXYGEN AND WATER TEMPERATURE--CENTRARCHIDS

In general, dissolved oxygen concentrations and water temperatures measured
in nests of bluegills and pumpkinseeds in 1973 and 1974 were higher at a
given depth in the afternoon than in the morning; measurements taken at the
surface in water depths of about 0.7 m differed only 0.6 mg/liter and 0.4 C
from measurements taken 1 cm from the bottom (Appendix B), and daily
fluctuations and mean values were similar in both years (Fig. 5).

Because, unlike the northern pike studied, embryos and sac larvae were found
only at the bottom, and because no stratification occurred we will limit our
discussion here to measurements 1 cm from the bottom. The mean dissolved
oxygen concentration 1 cm from the bottom was 7.1 mg/liter in the morning and
11.5 mg/liter in the afternoon with a range of 2.4-18.2 mg/liter (Appendix B,
Table 1). In spite of this large range, 71% of the morning measurements were
6.1-9.0 mg/liter and 61% of the afternoon measurements were 9.1-13.0 mg/liter.
Supersaturation occurred on 18 of 53 mornings and on 48 of 53 afternoons.

The low of 2.4 mg/liter of dissolved oxygen was measured in one pumpkinseed
nest found in very shallow water of 0.3 m. Both maximum and minimum
dissolved oxygen concentrations were recorded at stations on the lee side of
the lake. Mean water temperatures at 1 cm from the bottom were 20.0 C in
the morning and 23.3 C in the afternoon with a range of 15.0-27.5 C (Appen-
dix B, Table 2).

Mean dissolved oxygen concentrations measured 5 cm from the bottom with Delta
recorders at 2 stations from 9 June to 5 July 1974, were lowest at 0800-0900
and highest at 1700-1800 CDST (Fig. 6). The lowest dissolved oxygen concen-
tration, 5.0 mg/liter, was recorded on 21 June, and highest, 16.0 mg/liter,
on 26 June.

To determine if nest-guarding activities of male fish affected dissolved
oxygen levels, we measured dissolved oxygen with manual methods in 8 nests
and 1 m outside each of these nests in 1974. Average dissolved oxygen con-
centrations 1 cm from the bottom were 7.5 mg/liter inside the nests (morn-
ings); this was not significantly different (P < .01, N = 43) from 7.2
mg/liter outside the nests. Means of the afternoon measurements were 11.9
mg/liter (N = 39) both inside and outside the nests.

OXYGEN TOLERANCE EXPERIMENTS--NORTHERN PIKE

Embryo

A 77% hatching success occurred with embryos exposed for 8 h on the 7th day
after fertilization to dissolved oxygen of 0.6 mg/liter, which was similar to
hatching success of 80-93% in the higher concentrations tested (Fig. 7;

13


APPENDIX C (continued)

TABLE C-4. SURVIVAL OF NORTHERN PIKE LARVAE, EXPOSED AT EARLY FEEDING STAGE
FOR 8 h AT 10-14 C TO DISSOLVED OXYGEN CONCENTRATIONS LISTED
Ten Larvae Initially in Each Chamber. Treatment Given on 20 May
and Survival Recorded 1 Day Later.

Duncan's Multiple Range Test*

Average Dissolved
Oxygen (mg/liter) 0.5 1.6 8.9 Control Control 4.0

Range of Dissolved
Oxygen (mg/liter) 0.5 1.0-2.0 8.2-9.3 Control Control 3.5-4.5

Percent Survival 0 0 93 93 97 100

-05

-01
Analysis of Variance”

sov af 88 ms F

Treatments 5 368.278 73.656 441 .885**
Error 12 2.000 0.129
Total 17 370.278

"underlined values not different at 0.05 and 0.01 levels.

»pecause of zero readings, 1 was added to all values for computation
of ss.

30


APPENDIX D
RESULTS OF CENTRARCHID OXYGEN TOLERANCE EXPERIMENTS

TABLE D-1. HATCHING SUCCESS OF SMALLMOUTH BASS EMBRYOS EXPOSED 2 DAYS AFTER
FERTILIZATION FOR 6 h AT 20-23 C TO DISSOLVED OXYGEN CONCENTRA-
TIONS LISTED

Fifteen Embryos Initially in Each Chamber. Treatment Given in
Two Experiments, and Survival Recorded When Hatching was Complete.

Dunean's Multiple Range Test®
Average Dissolved Oxygen

(mg/liter) 0.5 1.8 3.5 Control 7.9
Range of Dissolved Oxygen
(mg/liter) b 0.5 1.0-2.0 3.0-5.0 Control 7.5-9.0
Percent Survival 0 69 87 89 91
05
01

Analysis of Variance”

sov af ss ms F
Subgroups 9 976.326 108.408
Treatments 4 943.314 235.829 57. 967#*
Experiment 1 14.860 14.860 3.118 N.S.
Interaction 4 18.152 4.538 -952 N.S.
Error 24 114.377 . 4.766
Total 33 1090 .703

4underlined values not different at 0.05 and 0.01 levels.
Percentages given are for combined survival rates of two experiments.

Because of zero readings, 1 was added to all values for computation
of ss.

(continued)

31


Bluegill--Embryo

Mean hatching success of embryos, exposed for 4 h in two experiments to 0.5
mg/liter of dissolved oxygen was 40%; this was not significantly different
(P < .05) from 44-50% hatched in concentrations of 2.1-7.8 mg/liter (Fig. 8;
Appendix D, Table 3).

Bluegill--Sac Larvae

In four out of five experiments no sac larvae, 1-4 days after hatching, sur-
vived when exposed for 4 to 6 h to a dissolved oxygen concentration of

0.5 mg/liter. Mean survival ranged from 65-74% in the higher concentrations
tested of 1.8-8.0 mg/liter (Fig. 8, Appendix D, Table 4). Exposure for 6 h
rather than 4 h in one experiment did not reduce survival, which in this
experiment was second highest of the five experiments. In concentrations of
more than 1.8 mg/liter of dissolved oxygen, mean survival in water tempera-
ture of 26 C was 89% and was significantly higher (P < .01) than the mean
survival of 56% in water temperature of 21-22 C. Mean survival of sac larvae
exposed to dissolved oxygen concentrations of 0.5 mg/liter for periods rang-
ing from 1-4 h was 0, 1, 11% for the 4, 3, 2 h exposures, respectively, and
was lower (P < .01) than the 71, 79% survival in the 1 h exposure and in the
control (Appendix D, Table 5). Although there was a significant interaction
between experiments and treatments, this was probably due to the low survival
of the controls in one of the five experiments. When this experiment is
eliminated from the analysis, the interaction was non-significant.

19


DISCLAIMER

This report has been reviewed by the Environmental Research Laboratory -
Duluth, U.S. Environmental Protection Agency, and approved for publication.
Approval does not signify that the contents necessarily reflect the views
and policies of the U.S. Environmental Protection Agency, nor does mention of

trade names or commercial products constitute endorsement or recommendation
for use.

ii


TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)

T. REPORT NO. 2. 3. RECIPIENT'S ACCESSION NO.
EPA-600/3-76-113
4. TITLE AND SUBTITLE iS. REPORT DATE
Dissolved Oxygen, Temperature, Survival of December 1976 (Issuing Date)
Young at Fish Spawning Sites 6. PERFORMING ORGANIZATION CODE
7, AUTHOR(S) is. PERFORMING ORGANIZATION REPORT NO.

John J. Peterka and James S. Kent

‘9, PERFORMING ORGANIZATION NAME AND ADDRESS 10, PROGRAM ELEMENT NO.

Department of Zoology
North Dakota State University 77 en RRotaRaNT NO.

Fargo, North Dakota 58102

R 801976
12. SPONSORING AGENCY NAME AND ADDRESS 13. TYPE OF REPORT AND PERIOD COVERED
Environmental Research Laboratory - Duluth, Minn. Final
Office of Research and Development 14. SPONSORING AGENCY CODE
U.S. Environmental Protection 4£ency
Duluth MN 55804 EPA/600/03

15, SUPPLEMENTARY NOTES.

16. ABSTRACT

Fluctuations of dissolved oxygen concentrations and water temperatures in

their natural spawning sites were measured during embryo through larva

stages of northern pike (Esox lucius), and during embryo and sac larva stages
of bluegills (Lepomis macrochirus) and pumpkinseeds (Lepomis gibbosus).

At northern pike sites, dissolved oxygen concentrations from combined
measurements 1 and 10 cm from the bottom ranged from 0.0-16.6 mg/liter, and
water temperatures from 2.5-23.0 C; average daily fluctuations were 3.0 mg/liter
and 1.6 C. For bluegill and pumpkinseed nests, dissolved oxygen concentrations
1 cm from the bottom ranged from 2.4-18.2 mg/liter and water temperatures from
15.0-27.5 C, with average daily fluctuations of 4.4 mg/liter and 3.3 C. In
field experiments to determine acute effects of a single exposure to low
dissolved oxygen concentrations, tolerance decreased from embryo to larva stages
for northern pike and from embryo to sac larva stages from bluegills and
smallmouth bass (Micropterus dolomieui).

(V7.

KEY WORDS AND DOCUMENT ANALYSIS

b DESCRIPTORS b.1DENTIFIERS/OPEN ENDED TERMS |c. COSATI Field/Group
*Missolved gases, *Temperature,
*Embryos, *Larvae, Bass, Fishes, Piseclved oxygen, 06/F
>
Mortality, Fresh water, Bioassay Spawning, Esocidae,
Centrarchidae,
Natural habitat
HN. DISTRIBUTION STATEMENT 19. SECURITY CLASS (This Report) 21. NO, OF PAGES
Unclassified 44
Release to Public [20. SECURITY CLASS (This page) [22. PRICE
Unclassified
EPA Form 2220-1 (9-73) 36

‘US. GOVERNMENT PRINTING OFFICE: 1977-757-056/S504


oxygen determinations in small samples, duplicate macro samples (200 ml) and
small samples (8.2 ml) were prepared from water collected at the surface.
Any differences of dissolved oxygen in macro and small samples were used to
adjust dissolved oxygen values for all small samples, assuming the dissolved
oxygen in macro samples to be correct. The mean difference between duplicate
macro samples was 0.03 mg/liter (N=30).

To reduce disturbance from sampling of any dissolved oxygen or water tempera-
ture stratification, siphon tubes extended 1-2 m from each station. A
tubing connector at the intake end of each siphon tube was cut at a 45°
angle. Because tests in laboratory aquaria using methylene blue crystals as
tracers showed water was drawn to the tip of the connector we positioned

the connector 10 cm from the bottom to draw water from above the connector
tip, and the connectors 1 cm from the bottom to draw water from below the
comector tip to help prevent disturbance of any oxygen stratification due
to slight currents created by siphoning (Fig. 1). Each tubing connector-end
was covered with 3161 Nitex netting to prevent entrance of detritus.

Water temperatures to the nearest 0.5 C were measured with thermistors at
times and depths.dissolved oxygen samples were collected. As with siphon
tubes, thermistor probes were left in place to prevent any disturbance of
water during sampling.

Two Delta (Model 3610) recorders were used in 1974 to provide a continuous
record of dissolved oxygen and water temperatures. Probes were placed 5 cm
from the bottom, and dissolved oxygen was recorded to the nearest 0.2
mg/liter and water temperature to the nearest 1C. The meters provided a
good indication of daily trends and duration of extremes, but problems with
the need for frequent calibration led us to discontinue their use after one
year.

We measured water movement near the bottom of spawning sites of northern pike
by timing the drift of plumes from methylene blue crystals which were dropped
on a metal grid placed on the bottom. Observations of the plumes were made
with a glass-bottomed pail.

Durations of early stages of northern pike development were estimated from
field observations of hatching of embryos and development of sac larvae
and larvae.

OXYGEN TOLERANCE EXPERIMENTS

In field experiments, early stages of northern pike, smallmouth bass and blue-
gills were exposed to dissolved oxygen concentrations ranging from no oxygen
to saturation for periods ranging from 1-8 h (8 h was the longest period
during which dissolved oxygen concentrations measured remained below 1.0
mg/liter in the northern pike spawning marsh) .

Northern pike, obtained from the National Fish Hatchery, Valley City, N.D.

and from N.D. State Hatchery, Spiritwood, ranged from 1-day-old embryos to
larvae. Smallmouth bass embryos and sac larvae were obtained from nests in

6


SECTION 5

RESULTS

DISSOLVED OXYGEN AND WATER TEMPERATURE--NORTHERN PIKE

Dissolved oxygen in 1974 and 1975 generally was lowest at 1 cm from the
bottom and highest at the surface. Dissolved oxygen concentrations were
lower in the morning than in the afternoon at all water depths measured, and
decreased at a given depth from embryo to larva stages (Fig. 2; Appendix A,
Table 1). Daily fluctuations in dissolved oxygen and differences between
morning and afternoon measurements during the embryo and sac larva stages
were less in 1975 than in 1974.

Dissolved oxygen concentrations at all depths fluctuated markedly from day to
day and from morning to afternoon (Fig. 2). The maximum day-to-day decreases
in 1974 were from 14.7 to 3.8 and from 10.2 to 2.7 mg/liter at 10 and 1 cm
from the bottom, respectively, from the afternoon of 26 April to the after-
noon of 27 April. In 1975, a decrease at 10 cm from 9.8 to 6.2 mg/liter and
at 1 cm from 6.5 to 2.9 mg/liter from the afternoon of 6 May to the afternoon
of 7 May was the largest measured. The greatest differences between morning
and afternoon concentrations on the same day were 3.5 to 15.0 mg/liter at 1
cm on 23 April 1974, and 3.2 to 9.4 mg/liter at 10 cm on 5 May 1975.

Mean dissolved oxygen concentrations measured 5 cm from the bottom with Delta
oxygen recorders at two stations from 23 April to 12 May 1974, were lowest
at 0800-0900 and highest at 1600-1700 CDST (Fig. 4). Because daily patterns
of dissolved oxygen fluctuations were similar, we chose 2 days to illustrate
data from continuous recordings: 24 April, when highest concentrations of
dissolved oxygen occurred, and 28 April, when lowest concentrations occurred
(Fig. 4). Dissolved oxygen was 0.0 mg/liter for 6 h on 28 April and was less
than 0.6 mg/liter for 8 h. We observed that oxygen increased slightly before
and then decreased at sunrise on 8 mornings from 27 April-13 May at one of
the recorders and on 6 mornings from 24 April-8 May at the other recorder
(Fig. 4). The weather from 24 April-13 May was windy and cloudy with much
rain which could have caused the pre-sunrise increases of dissolved oxygen.
Dissolved oxygen concentrations measured by the recorders were similar to
averages of manual measurements made 10 cm from the bottom. Means of dis-
solved oxygen determined with recorders for embryos, sac larvae, and larvae,
respectively, were 7.4, 6.5, and 5.5 mg/liter; those for manual measurements
were 7.6, 6.0, and 5.1 mg/liter (Appendix A, Table 1).

Average daily water temperatures generally increased from April to May in
1975, whereas in 1974 they remained rather uniform (Fig. 3). Water

8


EPA~600/3-76-113
December 1976

DISSOLVED OXYGEN, TEMPERATURE, SURVIVAL OF YOUNG AT FISH SPAWNING SITES

by

John J. Peterka
James S. Kent
Department of Zoology
North Dakota State University
Fargo, North Dakota 58102

Grant No. R801976

Project Officer

Richard Siefert
Environmental Research Laboratory
Duluth, Minnesota 55804

ENVIRONMENTAL RESEARCH LABORATORY--DULUTH
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
DULUTH, MINNESOTA 53804


APPENDIX C = TABLE C-2 (continued).

Analysis of variance

sov dt ss ms FE
Subgroups 11 219.417 19.417
Treatments 5 192.917 38.583 10 .445*
Experiment 1 8.028 8.028 1.428 N.S.
Interaction 5 18.472 3.649 0.657 N.S.
Error 23 129.333 5.623
Total 34 348.750

‘Underlined values not different at 0.05 and 0.01 levels.
percentages given are for combined survival rates of two experiments.

“Because of zero readings, 1 was added to all values for computation of
ss.

TABLE C-3. SURVIVAL OF NORTHERN PIKE SAC LARVAE EXPOSED 3 DAYS AFTER
HATCHING AT 12-13 C TO LOW DISSOLVED OXYGEN CONCENTRATIONS
OF 0.8 mg/liter FOR TIMES LISTED
Fifty Sac Larvae Initially in Each Chamber. Treatment Given
on 14 May, and Survival Recorded 3 Days Later.

Duncan's Multiple Range Test®

Exposure Time 6h 4h 2h Control

Percent Survival 0 6 14 88
+05 _
01 —

Analysis of Variance?

sov df ss ms F

Treatment 3 2530 843.333 187 .407**

Error 4 18 4.500

Total 7 2548

®Underlined values not different at 0.05 and 0.01 levels.

Because of zero readings, 1 was added to all values for computation of
88.
(continued)

29


PER CENT SURVIVAL

Figure 7.

—-
L OA , = en 8 eee 8
sf 1
7
80. ee. / —_—___,
/
| /
7) ‘
sol | /
/
l% \ !
8
40,
U
b og /
20, \I
LF xt nN L 1 1 1 1 L 1 1 j

1 2 3 4 5 6 7 8 9 10 11 CONTROL
DISSOLVED OXYGEN, mg/liter

Survival of northern pike embryos, sac larvae and larvae exposed
for 8 h to various concentrations of dissolved oxygen. A.
-Hatching success of embryos exposed 7 days after fertilization.
B.-Survival of sac larvae exposed 1 day after hatching. C.-Sur-
vival of larvae exposed in early feeding stage. Stars indicate
significant differences from controls. Detailed information
found in Appendix C.

17


RESEARCH REPORTING SERIES

Research reports of the Office of Research and Development, U.S. Environmental
Protection Agency, have been grouped into five series. These five broad
categories were established to facilitate further development and application of
environmental technology. Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in related fields.
The five series are:

1. Environmental Health Effects Research
2. Environmental Protection Technology
3. Ecological Research

4. Environmental Monitoring

5. Socioeconomic Environmental Studies

This report has been assigned to the ENVIRONMENTAL PROTECTION
TECHNOLOGY series. This series describes research performed to develop and
demonstrate instrumentation, equipment, and methodology to repair or prevent
environmental degradation from point and non-point sources of pollution. This
work provides the new or improved technology required for the control and
treatment of pollution sources to meet environmental quality standards.

This document is available to the public through the National Technical Informa-
tion Service, Springfield, Virginia 22161.


APPENDIX A (continued)

TABLE A-2. WATER TEMPERATURES (C) IN A NORTHERN PIKE SPAWNING MARSH DURING
THE EMBRYO, SAC LARVA, AND LARVA STAGES
Water Temperatures
Embryo
Morning Afternoon
Depth No. Average S.D, Range No. Awerage S.D. Range
Surface 30 7.2 3.21 3.5-15.0 32 121.1 5.27 5.0-22.0
10 30 6.5 2.46 3.5-14.0 32 8.0 2.94 4.7-16.0
1 30 5.6 2.22 2.5-12.0 32 6.6 2.65 3.5-14.0
Total 90 6.4 2.71 2.5-15.0 96 8.6 4.22 3.5-22.0
Sac Larva
Surface 37 8.8 2.29 5.0-13.0 36 14.4 4.46 7.0-23.0
10 37 8.5 2.38 5.0-12.5 36 10.9 3.04 6.0-17.0
1 37 8.0 2.39 4.0-12.5 36 9.7 2.50 5.0-15.0
Total 111 8.4 2.36 4.0-13.0 108 11.7 3.95 5.0-23.
Larva

Surface 23 8.2 3.14 4.0-14.0 22 13.2 5.39 6.0-22.5
10 23 8.3 3.36 5.0-14.5 22 10.0 4.18 5.0-17.5
1 23 8.0 3.08 5.0-14.0 22 9.1 3.39 5.0-15.0
Total 69 8.2 3.15 4.0-14.5 66 10.8 4.68 5.0-22.5

25


APPENDIX D (continued)

TABLE D-4. SURVIVAL OF BLUEGILL SAC LARVAE EXPOSED 1-4 DAYS AFTER HATCHING
FOR 4 h AT 21+26 C TO DISSOLVED OXYGEN CONCENTRATIONS LISTED
Ten Sac Larvae Initially in Each Chamber. Survival Recorded 1
Day After Treatment in Five Experiments.

Duncan's Multiple Range Test®
Average Dissolved Oxygen

(mg/liter) 0.5 3.7 8.0 1.8 Control
Range of Dissolved Oxygen
(mg/liter) 0.5 2.5-4.5 7.4-9.0  1.5-2.5 Control
Percent Survival 1 65 69 70 74
-05
-01
Analysis of Variance
SOV df 838 ms F
Subgroups 24 707.187 29.466
Treatments 4 561.587 140.397 33.224%%
Experiments 4 77.987 19.487 6.131
Interaction 16 67.613 4.226 1.329 N.S.
Error 49 155.833 3.180
Total 73 863.020

"Underlined values not different at 0.05 and 0.01 levels.

percentages given are for combined survival rates of five experiments.

“Because of zero readings, 1 was added to all values for computation of

(continued)
34


SECTION 6

DISCUSSION

DISSOLVED OXYGEN AND WATER TEMPERATURE
Northern Pike

We measured dissolved oxygen concentrations and water temperatures in a
northern pike spawning site during 2 years of extremely different weather
conditions. In 1974 warm water temperatures during spawning were followed
by cooler temperatures, and there were extreme day-to-day and morning-to-
afternoon fluctuations of dissolved oxygen concentrations and water tempera-
ture. In 1975 there was a gradual warming trend from spawning to larva
stages, and fluctuations of temperature and of dissolved oxygen were less
than in 1974. In both years dissolved oxygen concentrations decreased from
embryo to larva stages. The decrease was probably due to decomposition of
grasses that had grown when the marsh bottom was dry during the preceding
summer and fall.

Natural reproduction was measured from counts of numbers and wet weights of
fingerlings released to Silver Lake when the marsh was drained in the summer
(Detroit Lakes Fish Hatchery, Minnesota Department of Natural Resources).
Estimated numbers of fingerlings resulting from 12 to 15 female northern pike
in 1974 and 1975, respectively, were 3190 and 2500; the average wet weights
were 1.6 and 2.7 g. The estimate for 1975 is believed to be low because
large flows of water over the retaining wall dam during the spring permitted
fingerlings to escape before the census was taken.

The two stations where manual measurements were taken had different expo-
sures to wind, sunlight, and water currents which affected dissolved oxygen
concentrations and water temperatures. Station A was sheltered from the wind
and shaded in the afternoon by trees and cattails; water currents near the
bottom on a calm day were 0.05 cm/sec. Station B was exposed to wind and
afternoon sunlight; located near the main channel draining the marsh, its
water current was 0.23 cm/sec. The faster movement of water at Station B
was probably the reason the mean dissolved oxygen concentration at that
station, 5.6 mg/liter, was greater than the 4.9 mg/liter mean at Station A.
Both means are of measurements at combined depths 1 and 10 cm from the
bottom. Exposure to afternoon sunlight was responsible for warmer average
water temperatures from the combined depths 1 and 10 cm from the bottom of
9.0 C at Station B compared to 7.5 C at Station A. The faster water currents
and afternoon sunlight also caused the frost to leave the bottom mds of
Station B sooner than at Station A.

20


SECTION 4

METHODS

DISSOLVED OXYGEN AND WATER TEMPERATURE

Dissolved oxygen and water temperatures were measured in a northern pike
spawning marsh, Silver Lake, Clay County, and in bluegill and pumpkinseed
nests, Crane Lake, Ottertail County, Minnesota. Northern pike were studied
from mid-April, when spawning began, until mid-May when fry were observed,
1974 and 1975. Adult northern pike were stocked and marsh water levels
were controlled by fisheries personnel of the Minnesota Department of
Natural Resources. For bluegills and pumpkinseeds, dissolved oxygen and
water temperatures were measured from the time a nest was first seen until
the male fish abandoned the nest. Measurements were made from early June
until mid-July, 1973 and 1974.

In 1973, we determined, from samples collected every 2 h for several days
in Crane Lake, that lows of dissolved oxygen occurred at 0800-0900 (CDST)
and highs at 1600-1700. We confirmed these times both there and in the
Silver Lake marsh in 1974 with continuous records of dissolved oxygen con-
centrations. Daily samples were taken during these periods to provide
measurements of extremes in dissolved oxygen concentrations. Dissolved
oxygen was measured from water samples collected at the surface with a

van Dorn sampler and from those siphoned 1 and 10 cm from the bottom
through Tygon tubing (inside diameter of .24 cm) into 20 ml glass-stoppered
weighing bottles. To develop siphons, we placed sample bottles in a
weighted plastic pail to allow the bottles and tips of siphon tubes to be
lowered below the water's surface [Magnuson (2)] (Fig. 1). Sampling depths
of 1 cm from the bottom were chosen to represent dissolved oxygen concen-
trations adjacent to embryos and larvae of northern pike on the bottom;
those at 10 cm to represent conditions adjacent to embryos and sac larvae
on vegetation above the bottom.

At each depth duplicate samples were prepared by thoroughly flushing the
siphon tubes and replacing the volume of the sampling bottles at least twice.
Samples were fixed using the azide modification of the Winkler method
[A.P.H.A. (3)]. Chemicals for fixing the 20 ml samples were used in reduced
volumes (0.1 ml); 8.2 ml of fixed sample was titrated with 0.002 N NajS003 in
a buret with graduations of 0.05 ml. Dissolved oxygen was measured to the
nearest 0.1 mg/liter. Where discrepancies greater than 0.3 mg/liter between
duplicate samples occurred, a third titration was run using the mixed re-
mainders of the two samples; means were determined from the closest two out
of three measurements. To provide a check on the accuracy of dissolved

4


spawning ponds at the National Fish Hatchery, Valley City. Bluegill embryos
and sac larvae were obtained from nests at Crane Lake, Minnesota.

Chambers containing embryos, sac larvae or larvae were returned to natural
surroundings following each treatment where they were elevated on racks to
prevent disturbance of the bottom and avoid siltation. Survival was measured
daily by counting live and dead fish in the chambers. For experiments with
embryos, survival was recorded until hatching was complete. Survival of sac
larvae and larvae was recorded until most in control chambers died. Northern
pike embryos were treated in solutions of 1:600 formalin for 16 min about
every two days (method used at the National Fish Hatchery, Valley City, N.D.)
when fungused embryos were observed. Fungus was not a problem with centrar-
chids because of short incubation periods.

Chambers were constructed of PVC pipe with an inside diameter of 4.3 cm and a
length of 2.3 cm with covers of 3161 opening Nitex netting. To permit
development of northern pike sac larvae to the feeding stage where any
delayed exposure effects could be observed, we used large holding chambers
constructed of 13.2 liter (3 gal) and 22.0 liter (5 gal) polyethylene tubs
with openings covered with 7501 Nitex netting.

During treatment, chambers were placed in 22.0 liter polyethylene buckets
filled with lake water in which dissolved oxygen was controlled by bubbling
various mixtures of compressed nitrogen and air. The buckets were kept in
the lake to maintain natural temperatures. Gas mixtures were adjusted by
flow meters and a gas manifold. Dissolved oxygen and water temperatures were
recorded at about 1 h intervals during the experiments. The change in pH due
to bubbling nitrogen gas in the northern pike marsh water was from 8.5 to 8.7
at a concentration of 0.3 mg/liter of dissolved oxygen; the pH of water in
Crane Lake changed from 8.4 to 8.9 at a concentration of 0.3 mg/liter
dissolved oxygen. We did not consider these changes in pH to affect sur-
vival of fish.

Analysis of variance with replications and with treatments fixed was used in
the experiments. When zero values occurred, 1.0 was added to each value
for computation of sums of squares. Duncan's Multiple Range Test was used
to determine significant differences among means [Steel and Torrie (3)].