intersex sturgeon in the missouri river? - the...

Shovelnose sturgeon -Scaphirhynchus platorynchus

Pallid sturgeon -Scaphirhynchus albus

INTERSEX STURGEON IN THE MISSOURI RIVER? - THE EFFECTS OF EMERGING CONTAMINANTS

ON A BIG RIVER

Photo courtesy of Aaron DeLonay

Overview:

• Our findings • Where intersex occurs • Historic occurrence • Intersex biology • Causes • Likely chemicals • What we (don’t) know

Photo Courtesy Aaron DeLonay

Sturgeon are Gonochoristic

Female Male

Intersex/Hermaphrodite/Testes-ova

Eggs Testis Eggs Testis

Intersex in MO River Sturgeon

Normal Early Stage Ovary

Presenter

Presentation Notes

Here you can see the folding better in this view of an early stage II ovary. The granular particles are oocytes.

Testis

Ovarian folds

Presenter

Presentation Notes

These gonads are predominately testicular. However, there are patches of early stage oocytes with the classical folding that I just described.

Microscopic View

Presenter

Presentation Notes

This final histology slides shows a normally developing perinucleor oocyte on the left and a nearly mature egg of the type we could see with the unaided eye adjacent to the normal testicular tissue. So, here we have a male fish, with seemingly normal mature testes producing sperm which we know requires elevated testosterone, able to support and produce eggs which we know requires elevated estrogens…… What can cause this apparent paradox???

Presenter

Presentation Notes

On the right panel the testis is slightly more advanced and we see an island of a couple of oocytes just under the surface. The tissue in the left panel shows advanced spermatogenesis with many lobules filled with early spermatozoa. Just under the tunica albuginea covering the testis, we see individual oocytes. In nearly all cases, oocytes occurred at or near the outside edge of the testis.

Presenter

Presentation Notes

Of the 22 abnormal hermaphrodites we observed, 17 were clearly visible to as such to the eye. Another 5 were identified as intersex by histological examination. Here we see developing stage II testicular tissue, with a stalk of fat in which is embedded previtellogenic oocytes. This association of oocytes with the fat tissue is what is typically seen in developing ovaries.

Historic occurrence of intersex

Presenter

Presentation Notes

Historically, we find reports of intersex in the MO River going back to the 1960s to the present at 6 locations

Distribution of Intersex Sturgeon

Male photo

Fix mdc results

RM 450 St. Joseph

RM 330 Napoleon

RM 250 Miami

RM 200 Franklin Isl

and

RM 170 Easley

RM 125 Mokane

RM 45 Weldon Sprin

gs

RM 298 Saverton

RM 149 Crystal C

ity

RM 52 Cape Girardeau

RM 847 Caruthersville

% of

Inters

exed

Male

s

0

5

10

15

20

25

Missouri RiverMississippi River

16

17

18

19

16

21

15

14

15 1413

Presenter

Presentation Notes

Cerc with MDC did a survey on the MO and MISS Rivers in 2008 from St Joseph to Crystal City. We tried to catch 15 – 20 males at each site and visually inspected them for intersex. Interestingly, we saw spikes in intersex below the major cities (i.e. KC, Columbia, St Louis). Highest result was 23% of the males.

Historic Occurrence

0

2

4

6

8

10

12

Perc

ent

'64-'71 '68-'69 '78-'79 '93-'94 '00-'01 07-'08

Year

of Catchof Males

Presenter

Presentation Notes

Reported historic accounts. A seemingly increasing trend

So What? • Not Normal • Reproductive consequences • May be increasing • Indicative of other problems? • Don’t know what the cause is • Could be related to water/sediment quality

Gametogenesis

Primary Germ Cell (PGC)

Eggs Sperm

Presenter

Presentation Notes

Before I list some of the suggested ways abnormal hermaphroditism can be induced, it is useful to briefly review how the male and female gametes are produced Gametogenesis is the process through which the primary germ cells or germ line stem cells become either sperm or eggs.

Genetics Hormonal

PGC

Eggs Sperm

Presenter

Presentation Notes

The PGC are bipotential and depending on the interplay of genetic factors and hormonal stimuli, they differentiate into either spermatogonia or oogonia

Hypothalmus

PGC

Eggs Sperm Gonad

Pituitary

Hormones Hormones

Presenter

Presentation Notes

But the gonads are not the only organs involved in this process; The hypothalmus and the pituitary transduce information from the fish’s environment and both send and receive signals from the gonad directing development of the gametes via the gonad’s sex steroid producing cells.

Some Causes of Abnormal Hermaphroditism in Fish

• Senescence • Genetic abnormalities • Hybridization • Radiation • Diet • Temperature • Environmental disturbances • Chemicals

Presenter

Presentation Notes

With that in mind, I will now go through the various proposed inducers of abnormal hermaphroditism. They include: Senescence, the aging of the fish and change in hormone levels, genetic abnormalities caused through mutations such as might occur through inbreeding, hyrbidization, radiation and chemical exposure, the composition of the diet or diet restriction, exposure to certain temperatures, and a variety of environmental disturbances disrupting the normal reproductive processes. These causes have either been demonstrated in the laboratory to result in ovotestes or there is good correlative data from the field.

Age and Size Characteristics

FL WT Age

mm

or g

ram

s +

SE

0

200

400

600

800

1000

Year

s +

SE

0

2

4

6

8

10

12

14

16

18

20

intersex normal

Some Proposed Causes of Abnormal Hermaphroditism in Fish

• Senescence • Genetic abnormalities • Hybridization • Radiation • Diet • Temperature • Environmental disturbances • Chemicals

Presenter

Presentation Notes

With that in mind, I will now go through the various proposed inducers of abnormal hermaphroditism. They include: Senescence, the aging of the fish and change in hormone levels, genetic abnormalities caused through mutations such as might occur through inbreeding, hyrbidization, radiation and chemical exposure, the composition of the diet or diet restriction, exposure to certain temperatures, and a variety of environmental disturbances disrupting the normal reproductive processes. These causes have either been demonstrated in the laboratory to result in ovotestes or there is good correlative data from the field.

AN ENDOCRINE DISRUPTOR IS:

A compound that interferes with, or changes, normal endocrine (hormone) function.

17 alpha-Ethinyl Estradiol

OH

C CH

HO OH

nonylphenol

HO OH

Bisphenol A

HO

Cl

Cl

Cl

Hydroxy PCBs

Cl Cl

H

C-Cl3

DDT

Cl

C2H3N H

NCH(CH3)2 H

N N

N

Atrazine

(C4H9)

Sn

(C4H9)

(C4H9)

Tributyl Tin

Reproductive Characteristics, Functions, Processes that Can be Altered with

Endocrine Disrupting Chemicals:

• Sexual differentiation and development • Offspring sex ratio • Gonad and duct development • Secondary sexual characteristics • Fertility • Fecundity • Mating and sexual behavior • Gametogenesis • Hormone levels

WHERE DO EDCS COME FROM?

Personal care products Commercial products Detergents Industrial discharge Residential wastewater Municipal WWTP Agriculture

and where do go? 223

MAPS Courtesy of Gust Annis, TNC

9

Contaminant

Water Exceedances Sediment Exceedances Tissue Exceedances Preliminary Risk Conclusion

Trace Elements

Arsenic 0 of 302 1 of 21 No Data COC

Cadmium 6 of 151 2 of 21 0 of 43 COC

Chromium 2 of 151 0 of 20 No Data COC

Copper 5 of 151 1 of 25 No Data COC

Lead 36 of 151 2 of 26 0 of 41 COC

Mercury 0 of 90 0 of 15 25 of 43 COC

Nickel 0 of 151 5 of 27 No Data COC

Selenium 31 of 302 No Data 1 of 2 COC

Zinc 3 of 151 1 of 28 No Data COC

Nutrients

Ammonia 6 of 268 No Data No Data COC

Total Nitrogen 320 of 320 No Data No Data COC

Total Phosphorous 346 of 346 No Data No Data COC

From forthcoming report on Contaminants of Concern in the MOR

Pesticides

2,4-D 0 of 1 No Data No Data DG

Carbaryl 1 of 110 No Data No Data DG

Chlordane No Data 1 of 4 29 of 141 COC

Chlorpyrifos 0 of 206 0 of 4 No Data DG

Diazinon 0 of 206 0 of 1 No Data DG

Dieldrin 0 of 205 0 of 4 14 of 140 COC

DDT No Data 0 of 4 21 of 141 COC

DDD No Data 0 of 4 0 of 139 NL

DDE 0 of 202 4 of 4 26 of 139 COC

Metolachlor 0 of 206 No Data No Data DG

Triazine Herbicides 73 of 206 No Data No Data COC

Industrial Organics

Dioxins/Furans No Data No Data No Data DG

Nonylphenol No Data No Data No Data DG

PAHs No Data 0 of 4 No Data DG

PBDE No Data No Data No Benchmark (94 samples)

DG

PCBs No Data 4 of 4 133 of 141 COC

Other

Chlorophyl A 198 of 198 No Data No Data COC

Estrogens No Data No Data No Data DG

pH 0 of 361 No Data No Data NL

Water Sediment Tissue

“The Missouri Department of Health and Senior Services (DHSS) recommends that all consumers be aware of the positive benefits of

eating fish and the potential adverse health effects of contaminants that may be found in fish.”

Year

ng/g

egg

0

200

400

600

800

1000

1200

1400

1600

1800

Year

ng/g

fille

t

0

500

1000

1500

2000

2500

'84 '86 '88 '96 '04 '08

Egg Concentrations

Fillet concentrations

Data: Courtesy Mike McKee, MO Dept. of Conservation

PCBs, Chlordanes

Photo courtesy of Aaron DeLonay

Unlikely Legacy Chemicals:

ng/g

Lip

id-2000

0

2000

4000

6000

8000

10000

12000

14000

Normal Intersex Normal IntersexGonad Brain

Organochlorine Pesticides

ng/g

Lip

id

0

20000

40000

60000

80000

100000


Total PCBs

Presenter

Presentation Notes

CERC analysis of chemicals that bioaccumulate in tissues and that can affect the endocrine system and therefore are believed to potentially cause intersex were analyzed in brains and gonads of 24 males and 24 intersex shovelnose sturgeon. There were no differences in chemical concentrations, and all concentrations were elevated in both intersex and normal males.

Flame Retardants: PBDEs ng

/g L

ipid

-2000

0

2000

4000

6000

8000

10000


Chemical Suspects?

Ethinyl estradiol Estradiol Atrazine

Nonylphenols Mercury

Heavy metals

How do we link exposure to

MOR EDCs to intersex in sturgeon?

Testis

A small incision in the belly of a pallid sturgeon to implant a transmitter revealed it was intersex

Eggs

Must Consider:

• Exposure • Uptake • Metabolism • Excretion • Mechanism and Site of

action

Vulnerability of Sturgeon

• Benthic feeders/dwellers • Long-lived • Age at first maturity • High lipid content • Spawning frequency • Other?

DEPARTMENT OF THE INTERIOR U.S. FISH AND WILDLIFE SERVICE

REGION #6

FY11 ENVIRONMENTAL CONTAMINANTS PROGRAM OFF-REFUGE INVESTIGATIONS SUB-ACTIVITY

Evaluation of Endocrine Active Chemicals and Synthetic Hormones in Shovelnose Sturgeon (Scaphirhynchus platorynchus): Implications for the reproduction and recovery of

an endangered species

Testicular Malformations

Reduced Size of Testes

1000 10000 100000-2

0

2

4

6

8

10

ng OCPs/g Lipid

Test

icul

ar G

onad

osom

atic

Inde

x

R2=0.57

Egg Parasites

Teratomas

R² = 0.30

0

20

40

60

80

100

120

0 100 200 300 400 500 600

Surv

ival

(cum

ulat

ive

%)

dl-PCB (ng/g lipid)

2 dpf (p<0.01)

Embryological Survival (2-3 dpf) (From Buckler, J. Thesis 2011)

Presenter

Presentation Notes

(WALK THROUGH FIRST GRAPH) I then looked at embryological survival. I assumed that a percentage of eggs dead on day 2 were due to not being fertilized so stocking numbers and mortalities on day 2 were corrected to use only “viable” eggs for analyses. As you can see cumulative survival at 2 dpf was correlated with dl-pcb concentration. (CLICK) Cumulative survival at day 3 post fertilization was also correlated with dl-pcb concentration. Although both regressions explain less than than 30% of the variation in survival.

Intersex in other MOR species?

Egg

Sperm

Environmental Stressors

• Over harvest • Channelization

• Dams • Loss of habitat • Contaminants ?

Photo courtesy of

Robb Jacobson

Intersex Male Immature Females Mature Females

Mea

n Vi

tello

geni

n (n

g/m

L) ±

SD

0

1

2

200400600800

100012001400160018002000

14

2

2

1

Mimic endogenous hormone Antagonize hormone effects

Disrupt hormone synthesis/metabolism Disrupt receptor synthesis/metabolism/function

Estrogen or Mimic

Plasma Binding Protein

Receptor

Hormone-Receptor Complex

mRNA

DNA Target Cell Altered

Functional Response

Protein Synthesis

Nucleus

intersex sturgeon in the missouri river? - the...

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