Longterm Estuary Assessment Group

NOAA '07 Eco Impacts of Hypoxia

Developing biomarkers of reproductive health in fish and amphibians of the

Barataria-Terrebonne Estuary LaFleur, Pitre, Lasseigne, and Nelson nicholls state university

LaFleur Lab

Graphic from Restore or Retreat: www.restoreorretreat.org

The Barataria-Terrebonne Estuary is impaired

NOAA '07 Eco Impacts of Hypoxia

Having been isolated from freshwater inputs it is lacks flushing, sheet flow, and an ecological flood pulse

• lack of flow

• saltwater intrusion

• land-loss

The 46-year mean stage of the Atchafalaya still reflects an ecological flood pulse

USACE Butte La Rose Gage 03120 ('59-'04)

0

2

4

6

8

4/7/06 5/27/06 7/16/06 9/4/06 10/24/06 12/13/06 2/1/07

Dis

solv

ed O

xyge

n (m

g/L

)

In the Upper Barataria basin, Hypoxia can occur But it is usually associated with Rainwater

events, rather than flood stage

from Estay and Fontenot (thesis)

one project proposes diverting 300,000 cfs water into the BTE; up to 6 sq mi / year; at a cost of $25 / cubic yard

Graphic from Restore or Retreat: www.restoreorretreat.org

Our estuary is slated for largescale hydrologic modifications:

NOAA '07 Eco Impacts of Hypoxia

Pipeline Slurry System

A more recently presented scenario would utilize a sediment pipeline slurry system to build land even quicker: up to15 sq mi / year; at a cost of $4 /cubic yardNOAA '07

Eco Impacts of Hypoxia

MY OBJECTIVESIn preparation for hydrologic changes due to further deterioration or restoration activities in our estuary, my lab has begun a survey

•to monitor behavioral indicators of reproduction in amphibians of the estuary

•to monitor anatomical indicators of reproduction in amphibians of the estuary

•to monitor molecular indicators of reproduction in amphibians of the estuary

NOAA '07 Eco Impacts of Hypoxia

2 3

Site 1 Choctaw swamp; Site 2 Chacahoula swamp Site 3 Nicholls’ Environmental Ag Facility Site 4 Falgout Canal fw / brackish marsh Site 5 fish only at Isle Dernieres Barrier Islands

5

1

4

NOAA '07 Eco Impacts of Hypoxia

Reproductive behavior is being surveyed in three LAMP survey routes:

spanning Lafourche and Terrebonne Parishes

NOAA '07 Eco Impacts of Hypoxia

23-Jan7-Feb

6-Mar20-Mar

0

5

10

15

20

Cal

l Den

sity

Sco

re/

Tem

pera

ture

23-Jan

7-Feb

6-Mar

20-Mar

0

5

10

15

20

Cal

l Den

sity

Sco

re/

Tem

per

atu

re

temp

tempcalls

calls

In 2007, Spring Peepers reached peak calling in Feb

Northern Cricket frogs peaked in Jan, but are still calling

Confirmed Hyla avivoca

Choctaw Swamp, Lafourche Parish

Proposed Expansion of Geographic Range to the BTES,

south of the Mississippi River

Conant and Collins.1991

ovary

liver

Selected species are collected, dissected, and anatomical indicators are examined

NOAA '07 Eco Impacts of Hypoxia

Species Repro Behavior

Preserved Repro Anatomy Molec. Biomarker

Bufo valliceps X X X X

Acris gryllus X X

Hyla cinerea X X

Hyla squirella X X

Hyla versicolor/ chrysoscelis

X X

Hyla avivoca* X X

Pseudacris crucifer X X

Gastrophryne carolinensis X X

Rana catesbeiana X X X

Rana grylio X

Rana clamitans X X X

X

Rana utricularia

Amphiuma tridactylumNotophthalmus viridescens

X

X

X

X

X

X

X

X

X

X

X

Amphibian Survey Matrix

Approach todesigning biomarkers for

estrogen-induction

• Injection of estradiol into males

• Isolation of liver RNA

• RT-PCR for Vtg, Chg from homogenates using degenerate and heterologous primers

NOAA '07 Eco Impacts of Hypoxia

vitellogenins and choriogenins

of teleostchoriogenins and vitellogenins

HETEROSYNTHETIC ORIGIN OF

TELEOSTEANEGG PROTEINS

follicle cells

vitelline envelope precursor to the

chorion

yolk of oocyte

Estrogeninduced

Liver synthesis

micropyle

NOAA '07 Eco Impacts of Hypoxia

Choriogenin Primers

Row 78 TTCAGGTTCCAGAATTCTGAC

Row 81 CATTGGTTCATATCGCTGTCT

Vitellogenin Primers

Row 8 CGATATTGACATGTTTCCAA

Row 24 TACCAGCTTGGTTTCTACCT

Choriogenin and Vitellogenin primers derived from F. heteroclitus, the mummichog. Row 78 and Row 81 produce a 450 bp product while Row 8 and Row 24 produce a 729 bp product

NOAA '07 Eco Impacts of Hypoxia

a b c d

Choriogenin cDNAs indicating normal female reproductive activity.

choriogenin

450 bp

F.g. liver

F.c. liver

G.a. liver

A.t. ovary

NOAA '07 Eco Impacts of Hypoxia

(Estrogen-injected males)

a= Fundulus grandis liver, b= Fundulus chrysotus liver, c= Amphiuma tridactylum liver

vitellogenin

a b c

729 bp

NOAA '07 Eco Impacts of Hypoxia

(Non-injected males)

Three wild-caught males showing the presence of female specific RNA a= Bufo valliceps liver, b= Amphiuma tridactylum liver c= Gambusia affinis

a b c

vitellogenin

729 bp

NOAA '07 Eco Impacts of Hypoxia

Through collaborations between the labs of LaFleur, Ferrara and Fontenot we have established overlapping projects of the estuarine fauna, including monitoring of finfish, larval fish, and amphibians.

Fish and

Amphibians

NOAA '07 Eco Impacts of Hypoxia

Summary• Documented reproductive behavior

through calls of 12 local Anuran species.

• Established herpetology collection.• Tracking reproductive seasons by measuring GSI on 4

Anurans, 1 Salamander, 3 fish.• Molecular biomarkers were amplified using

RT-PCR on 3 anurans, 1 Salamander, 3 fish.

• Poised to implement reproductive survey to include water quality and expanded sampling regime.

• Proposed range extension for Hyla avivoca to include wetlands of the BTES.

LaFleur Lab

Post Katrina Directions

Amphibian and Fish reproduction will be utilized as a longterm assessment tool for

environmental health of the estuary

Nicholls has been awarded a grant for restoration planting by NOAA

Nicholls has signed an MOU with NRCS for restoration plant propagation at Nicholls Farm

LaFleur, Boopathy, and Zou are committed to longterm monitoring programs that will offer

consistency over timeNOAA '07 Eco Impacts of Hypoxia