CERP: F2 Experiment

Focused on exposing adult mummichogs to

• 4 levels of contaminated sediments

• Clean and contaminated food

njwrri.rutgers.edu/graduate/

• Mortality estimates

• Measure growth in length and weight

• Quantify egg production

II. Egg performance experiment (EP)

• Measure egg size, hatching success, and size at hatch for larvae

Objectives I. F2 experiment:

ehpnet1.niehs.nih.gov/docs/ 1996/104-9/innov.html

• Sediment treatment• Elizabeth River mud diluted to 6.25 % (ERL) , 12.5 % (ERM),

and 25 % (ERH) • Fishing Bay (FB) mud as control• Same as F1 experiment

• Clean/contaminated food treatment• Contaminated clams: Live clams were placed out in the ER

river on 2 occasions for 6-7 weeks (T)• Clean clams: live clams were bought from a local seafood

market (Chester and Ware Rivers) (C)• Clams were mixed with fish gel and water to make ‘clam jello’

• Example• ERMT =Elizabeth River 12.5 % (medium) dilution and toxic

food treatment

F2 Experiment

Characteristics of clean & contaminated food

0

1

2

3

4

5Control clam #1

ng

/g-w

et

0

100

200

300

400

500 Exposed clam #3

PAH Accumulation in Mercenaria(note scale change)

Control

Elizabeth River

F2 Experiment• 24 50-gallon flow-through tanks fitted with manifolds that released incoming flow just above the sediment surface to mix water column

• 3 replicates per treatment

• Extra aeration was added to each tank

• 56 fish were added to each tank ranging from 40-73 mm (~F1)

• Fish were fed 0.03 g/g/day of their body weight for the first 30 days then switched to 0.06 g/g/day after day 30 (same as low food from F1)

• 4 sampling days: Days 0, 30, 59, and 74• Experiment ran from 6/11/03- 8/22/03 • Sub-samples of fish removed for biomarker

expression, chemical analysis, bioenergetic partitioning, and growth

• Eggs collected from each tank

• Egg production (daily), hatchability (weekly), and egg/larvae size estimates (weekly)

F2 Experiment

• Fish were fed twice a day (by hand)

• Salinity, temperature, DO measurements taken daily

• Egg collectors and flow rates checked daily

• Tanks were checked daily for dead/dying fish

F2 Experiment

Environmental Parameters

Salinity: 7.5 –11.1 ppt2002: 11.3-20.9 ppt

Temperature: 20 – 24 oC2002 18.3-22.4 oC

DO: - 5.1-8.4 mg L-1

- No problems with hypoxia or anoxic mud this time!

Flow rate: 300 mL min –1 - same as higher flow in 2002

www.elizabethriver.org/monitoring/ mummichogfacts.htm

F2 Experiment

0

20

40

60

80

100

120

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

% m

ort

ality

0

20

40

60

80

100

120

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT%

mort

alit

y

B B B B A A A A

P<0.0001

A B A B A B A BC C B B A A AB AB

P<0.0001

Day 74

Day 59

Adult Fundulus mortality by treatment

SedimentFood

Sediment

F2 Experiment

0

10

20

30

40

50

60

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90

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110

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

% m

orta

lity

F1 Experiment

0102030405060708090

100110

FBL FBH ERLL ERLH ERML ERMH ERHL ERHH

% m

ortali

ty

Mortalitycomparisonsfrom the last day of samplingfor F1 and F2 experiments

• Disease prevalence: fish with red fins and nape• Problems with seawater system may have contributed– times when there was no water flow for half a day, several days in a row• Algal bloom?

• Food differences?• Pellet food vs clam jello• No energy effect

• Brood effects?• Other expts using fish from same collections experienced higher than expected mortalities• Other expts using different fish experienced high morts as well

0.0

1.0

2.0

3.0

4.0

5.0

6.0

7.0

Pellet food Oven driedclean clam

jello

Oven drieddirty clam

jello

Freeze driedclean clam

jello

Freeze drieddirty clam

jello

kcal

g-1

Differences between F1 and F2 experiment:

40

45

50

55

60

65

70

0 20 40 60 80

Sampling date

Le

ng

th (

mm

)

ERHC

ERHT

ERMC

ERMT

ERLC

ERLT

FBC

FBT

Adult Fundulus mean length by treatment

Adult Fundulus mean absolute growth rate in length (+/- SD) for day 59 and 74

Day 74

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

mm

d-1

Day 59

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

mm

d-1

No sediment or food effect on growth

F1 Experiment

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

FBL FBH ERLL ERLH ERML ERMH ERHL ERHH

mm

d-1

F2 Experiment

0.00

0.02

0.04

0.06

0.08

0.10

0.12

0.14

0.16

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHTm

m d

-1

Mean growth on the last day of sampling for F1 and F2 experiment

Slightly slowergrowth in 2003than 2002

Mean egg production by week throughout the F2 experiment

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50

100

150

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250

0 2 4 6 8 10 12

Week

Num

ber o

f egg

s/w

eek

ERHC

ERHT

ERMC

ERMT

ERLC

ERLT

FBC

FBT

F2 Experiment

0

50

100

150

200

250

0 2 4 6 8 10 12 14 16

Week

Num

ber o

f egg

s/w

eek

ERHC ERHT

ERMC ERMT

ERLC ERLT

FBC FBT

F1 Experiment

0

50

100

150

200

250

0 2 4 6 8 10 12 14 16

Week

No.

of e

ggs/

wee

k

ERHH ERHL

ERMH ERML

ERLH ERLL

FBH FBL

Mean egg productionfor F1 and F2experiments

Total egg production (+/- SD) from F2 experiment

0

200

400

600

800

1000

1200

1400

1600

1800

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

# of

egg

s

B A B A B A B A

Food effect: p=0.0096Sediment effect: p=0.0527

F1 Experiment

0

200

400

600

800

1000

1200

1400

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FBL FBH ERLL ERLH ERML ERMH ERHL ERHH

No

. of e

gg

s

F2 Experiment

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800

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1200

1400

1600

1800

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

# of

egg

s

Total egg production for F1 and F2 experiments

• Eggs were collected daily from collectors in each tank. Eggs were subsequently grouped by week # of the F2 experiment

• Eggs incubated for 14 days in standard culture media, exposed to air for 2 hours and then placed into highly aerated seawater

for 30 minutes

• This hatching process occurred for 3 days, eggs unhatched on the 3rd day of the hatching process were termed “unhatched” or “dead”

www.miljolare.no/virtue/img/Species/

pages/Fish%20eggs.php

Egg Performance (EP) experiment:

Weeks 1-4

1.5

2.0

2.5

3.0

3.5

4.0

4.5

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHTm

m2

1

2

3

4

Weeks 5-8

1.5

2.0

2.5

3.0

3.5

4.0

4.5

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

mm

2

5

6

7

8

EP experiment:Mean area (+/- SD) of eggs by week collected from F2 experiment

Weeks 1-4

3.0

4.0

5.0

6.0

7.0

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

Tota

l Len

gth

(mm

)

1

2

3

4

Weeks 5-8

3.0

4.0

5.0

6.0

7.0

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

Tota

l Len

gth

(mm

)

5

6

7

8

EP experiment: Mean length (+/- SD) of larvae at hatch by week

Weeks 1-4

0

20

40

60

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100

120

140

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

%

week 1

week 2

week 3

week 4

Weeks 5-8

0

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120

140

FBC FBT ERLC ERLT ERMC ERMT ERHC ERHT

%

week 5

week 6

week 7

week 8

EP experiment:Mean % hatching success (+/- SD) of eggs by week collected from F2 experiment

Egg and larval traits from EP experiments

F2 showed similar results with F1 that there appears to be no difference among sediment or food treatments:

• Larval length

• Egg area

• Hatching success

Conclusions:

• Growth appeared to be affected by the lower food level at the start of the expt and the presence of disease

• Egg production • Higher and longer time of egg production than F1• Egg production was significantly higher in the toxic tanks• Dose response curve with higher total egg production

in the FB tanks (marginally significant)

• There were no apparent differences among sediment or food treatments with larval and egg size or traits

www.whoi.edu/fishtox/ bonyfish.html