CHLORINATED PESTICIDES AND GROWTH AND DEVELOPMENT OF AMERICAN ALLIGATORS

María Sepúlveda, Jon Wiebe, James Basto, and Timothy Gross

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

USGS - Florida Integrated Science Center
Gainesville, FL 32653

Abstract

This study evaluated American alligator (Alligator mississippiensis) clutch, embryo, and hatchling parameters in relation to exposure to organochlorine pesticides (OCPs) in Florida Lakes. Clutches were collected from lakes Apopka and Griffin, and from reclaimed agricultural properties (Emeralda Marsh).  Orange Lake served as a reference site. Clutches were monitored for fecundity, number of unbanded eggs, hatch rates, and embryonic mortality.  Nonviable eggs were opened to estimate the stage of embryonic loss, and neonatal mortality classified as either pre or post-hatch.  Several morphological parameters were measured from eggs and hatchlings. [OCPs] were measured in over 100 yolks representing different stages of development and viability conditions.  Yolk [OCPs] were ranked in the order: Apopka > Emeralda > Griffin > Orange, and the primary pesticides observed were: DDT and derivatives, toxaphene, and dieldrin.  Fecundity was increased and hatch rates were reduced in Apopka, Emeralda, and Griffin clutches when compared to Orange Lake. Low hatch rates were due to increased neonatal mortality, mainly during the first 35 days of development. Lower hatch rates, however, were not related to increased yolk [OCPs]. In addition, although live eggs had similar [OCPs] when compared to nonviable eggs (unbanded and early embryonic mortality), and higher [OCPs] did not result in eggshell thinning nor in the production of smaller hatchlings, some [OCPs] in this study fell within ranges known to affect survival, growth, and/or reproduction of fish. Environmental contaminants may be playing a role in the observed decreased egg viabilities by affecting normal reproductive function (vitellogenesis) in females. In addition, it remains unknown whether these pesticide mixtures might be acting in more than just an additive manner, exerting direct toxicity to the developing embryos (funded by NIEHS-SFBRP).

Objective

The overall objective of this project was to:

   Conduct field epidemiologic studies to evaluate embryonic development and mortality of the American alligator (Alligator mississippiensis) as a function of parental exposure to organochlorine pesticides (OCPs) in Central Florida.

Materials and Methods

Study Design:

Between June and July 2000, alligator egg clutches (24 to 35 per site, total of 111 clutches) were collected from lakes Apopka, Griffin, and from the reclaimed/flooded agricultural properties adjacent to lakes Griffin (Emeralda Marsh).  Orange Lake served as the reference site for these studies (Fig. 1).  In the laboratory, eggs were candled to determine whether they were fertile or "banded", and if so, nested in moist sphagnum moss and incubated at 30.5C and 91-95% humidity in an artificial incubation building.  One egg/clutch was opened to determine stage of development.  The following parameters were recorded from all clutches: fecundity (clutch size); percentage of "unbanded" eggs (which probably result from very early embryonic mortality, see Poster # PT008; [(number of unbanded eggs)/(fecundity) x 100]; percent hatchability [(number of eggs that hatched)/(fecundity) x 100], and neonatal mortality.  Eggs were opened when diagnosed as nonviable to estimate the stage of embryonic loss, and neonatal mortality was classified as either pre-hatch (early: 1-35 days of development; late: 36 - hatching), or post-hatch. Eggshell thickness and hatchling parameters (body weights, total length, snout-vent length, and head lengths) were also measured in a subset of embryos/hatchlings. A total of 109 yolks representing different sites and stages of egg development and mortality, were analyzed for [OCPs] via GC-MS.

Statistical Analyses:

Analysis of variance (ANOVA) followed by a Tukey's test was used to evaluate differences on clutch parameters across sites. Since the concentration of some OCPs in yolk was affected by age of development (Fig. 2), differences in contaminant levels across sites were determined using analysis of covariance (ANCOVA) using age in days as the covariate.  Linear regressions between clutch and hatchling parameters and yolk [OCPs] were also performed. Statistical significance was declared at p<0.05.

Conclusions

     Yolk [OCPs] were ranked in the order: Apopka > Emeralda > Griffin > Orange, and the primary pesticides observed were: DDT and derivatives, toxaphene, and dieldrin (Fig. 3).

   Fecundity was increased and hatch rates were reduced in Apopka, Emeralda, and Griffin clutches when compared to Orange Lake clutches.  Low hatch rates in these sites were due to increased neonatal mortality (Fig. 4).

   Live eggs had similar [OCPs] when compared to nonviable eggs (unbanded and early embryonic mortality) (Fig. 5).

   Higher [OCPs] in yolk did not result in lower  hatch rates (Fig. 6) nor in an increase in the incidence of unbanded eggs (data not shown).

   Similarly, higher [OCPs] in yolk did not affect length of incubation and size of hatchlings (Fig. 6) and did not result in eggshell thinning (data not shown).

   [OCPs] in this study fell within ranges known to affect growth, survival, and/or reproduction in different fish species (Table 1).

   Environmental contaminants may still be playing a role in the observed decreased egg viabilities by affecting normal reproductive function (vitellogenesis) in females.  In addition, it remains unknown whether these pesticide mixtures might be acting in more than just an additive manner, exerting direct toxicity to the developing embryos.

References

1. Jarvinen A.W., and G.T. Ankley.  1999. Linkage of effects to tissue residues: development of a comprehensive database for aquatic organisms exposed to inorganic and organic chemicals. Pensacola FL: SETAC. 364 pp.
2. Beyer, W.N., G.H. Heinz, and A.W. Redmon-Norwood.  1996. Environmental contaminants in wildlife: interpreting tissue concentrations.  Pensacola FL: SETAC.  494 pp.

 Acknowledgements

• NIEHS, Superfund Project: "Chlorinated pesticides and developmental mortality in wildlife".
• University of Florida, Department of Physiological Sciences, College of Veterinary Medicine.
• USGS - Florida Integrated Science Center.
• Florida Fish and Wildlife Conservation Commission.