ACHIEVING ECO-RELEVANT ORGANOCHLORINE PESTICIDE CONCENTRATIONS IN EGGS THROUGH MATERNAL EXPOSURE IN ALLIGATOR MISSISSIPPIENSIS

R. Heath Rauschenberger*^1,2, Jon J. Wiebe², Janet E. Buckland²,
J. Travis Smith², Maria S. Sepúlveda^1,2, and Timothy S. Gross^1,2

¹Department of Physiological Sciences,
College of Veterinary Medicine, University of Florida,
Gainesville, Florida, USA 32610

²USGS-BRD Florida Integrated Science Center,
Center for Aquatic Resource Studies,
Gainesville, Florida, USA 32653

Presented at the 12th International Symposium on Pollution Responses
 in Marine Organisms (PRIMO12), May 9-12, 2003, Tampa, FL.

Abstract

American Alligator (Alligator mississippiensis) eggs collected from contaminated sites in central Florida exhibit higher concentrations of organochlorine pesticides (OCPs) in yolk and higher rates of mortality compared to reference sites (p<0.05).  Potential hypotheses for increased embryonic mortality are maternal and/or embryonic exposure to OCPs.  Thus, the present study's objective was to use a captive breeding population of adult alligators to test the hypotheses that maternal exposure to OCPs results in increased OCP concentrations in eggs, and that increased exposure is associated with increased mortality / decreased viability. A total of 24 adult alligators were housed in eight pens (2 females: 1 male).  Female alligators in four pens each received an initial intraperitoneal and intramuscular injection in September 2001.  Oral dosing began April 2002 when animals resumed feeding, and the combined administrations resulted in each female receiving an average of 53.5 mgOCP / kg body weight prior to oviposition.  The dose was formulated to elicit elevated, eco-relevant OCP concentrations in eggs (range=3,000-28,000 ng/g yolk) without causing acute maternal toxicity.  Three females from each group successfully oviposited.  Treated females produced eggs containing increased OCP burdens (13,000 ng/g yolk) compared to control females (36 ng/g yolk).  In addition, treated females produced eggs that showed decreased viability (13 ± 22%) as compared to eggs of control females (45 ± 20%).  These preliminary results support that eco-relevant OCP concentrations can be delivered into eggs via maternal exposure and that such exposure is associated with decreased egg/embryo viability.  This work was supported by NIEHS Grant # P42-07375 to TS Gross.

The Problem

• In central Florida, alligator eggs from L. Apopka, L. Griffin, and Emeralda Marsh (a former agric. Property) exhibit significantly increased embryonic mortality compared to control sites (L. Lochloosa and L. Orange).
• Furthermore, alligator eggs from L. Apopka, L. Griffin, and Emeralda Marsh contain high concentrations of organochlorine pesticides (OCPs) [4,000–30,000 ng total OCPs /g yolk wet weight].
• These concentrations are several orders of magnitude greater than the control sites (L. Orange and Lochloosa) (100-350 ng total OCPs /g yolk wet weight).
   

Hypothesis:

1. Female alligators exposed to OCPs will produce egg yolks with increased [OCP concentrations].
    
2. Females alligators exposed to organochlorine pesticides will produce embryos that exhibit decreased viability.

Materials and Methods
 

Ø 4 Control Pens and 4 Treated Pens.
Ø 2 females and 1 male/pen.
Ø Pens were 30 m X 30 m and ~1.5 m in depth.
Ø Water level was controlled via onsite well and pond drains.
Ø Vegetation provided a visual barrier and nesting material.

 


Ø OCP exposures were initiated in Oct. 2001.
Ø 8 females received IP and IM injections of olive
       oil-OCP mixture.
Ø 8 females (controls) received equal volume of olive oil.
Ø OCP mixture consisted of p,p'-DDE, toxaphene,
       dieldrin, and chlordane.

Ø In April 2002, Feeding trials began as the activity
       level of the alligators increased as temperatures and
       photoperiod increased.
Ø Males and females began courtship behavior.
Ø Animals were conditioned to come to handler when
       called so they could receive weekly feed ration.

Ø Weekly feed ration consisted of a commercially
       purchased 2-2.5 kg rabbit.
Ø Each rabbit was injected with 0.25 kg of commercial
       fingerling fish feed (protein and vitamin enriched).
Ø Each rabbit was also implanted with a multi-vitamin
       designed for captive marine mammals.
Ø Rabbits were injected with an OCP mixture consisting
       of p,p'-DDE, toxaphene, dieldrin, and chlordane.

Ø In addition to the treated alligators housed as
       breeding group in the pens, 2 females were housed
       in concrete tanks.
Ø Health assessments included monthly monitoring
       of glucose, hematocrit, and total protein in blood.
Ø Behavior and feeding patterns were monitored
       on the day of feeding and the day after feeding.

Maternal Uptake and Transfer of Organochlorine Pesticides (OCPs)
 

Results
 

               3 Nests from treated females                                     3 Nests from control females

Organochlorine concentrations and blood chemistry values
(mean ± standard deviation) of captive, adult female alligators.
Blood chemistry values for control group taken from Mader (1996).
 

Conclusions

• Ecologically relevant OCP concentrations can be elicited in captive eggs via maternal exposure.
   
• Maternal exposure to OCPs is associated with increased embryonic mortality in captive alligator populations.
   
• Maternal transfer appears to be the major route of embryonic OCP exposure.
   
• First study to induce endogenous embryonic OCP exposure (via maternal transfer) in a crocodilian model.

Acknowledgements

This work was supported by NIEHS Superfund Basic Research Program Grant # P42-07375 to T.S. Gross. We thank Denise Gross, Kent Vliet, Nancy Szabo, and Alan Woodward for their important contributions. We also thank Eileen Monk, Shane Ruessler, Jesse Grosso, Carla Wieser, Nikki Kernaghan, Jennifer Muller, and Jessica Noggle for their technical assistance.