What Cautionary Tales Can Lake Apopka Tell?
by Anna Maria Gillis
Louis J. Guillette, Jr., views Florida's Lake Apopka with more than a little suspicion. Its resident young alligators are not developing right sexually, a fact that troubles the University of Florida-Gainesville biologist, who specializes in reptiles and their endocrine, or hormone, systems. He and his colleagues are finding that Apopka's juvenile alligators from the time they hatch have abnormal testes and ovaries and thoroughly mixed-up hormone balances. "We think that the gonads of juveniles from Lake Apopka are permanently changed while they are in their eggs," says Guillette. "They aren't likely to recover and produce offspring when they reach adulthood."
Guillette thinks that contamination from chemical spills at Apopka has caused major disruptions in the endocrinology of the long reptiles. He wonders about the likelihood of similar reproductive peculiarities in the lake's birds and fish, particularly commercially raised catfish. Might the ever-growing human population near the lake also be affected in subtle ways? Surprisingly, even though many local residents know about the research being done by Guillette and other scientists at the university and in state and federal agencies, they haven't expressed much concern. "They've been more worried about how this might affect their land prices than their health," he says.
Hampering Hormones
Scientists have been evaluating pesticides, herbicides, plastics, petrochemicals, and detergents for decades to determine how they might threaten wildlife and human health. They have a good idea about which chemicals are highly toxic, and they know the doses that can kill. Some chemicals, like the synthetic estrogen diethylstilbestrol, or DES, clearly do reproductive harm. DES, for instance, had been used to prevent miscarriages until scientists discovered that daughters of women who had used it were highly susceptible to a rare vaginal cancer. But scientists are still largely in the dark when it comes to understanding how minuscule amounts of man-made chemicals in the environment might work on the hormone system of animals while they are still tiny embryos.
The endocrine system includes all the hormones and the glands that secrete them. Adrenaline, the product of the adrenal glands that prepares one to fight or flee, and thyroxine, which is manufactured by the thyroid to regulate growth and development, are important members of the hormone family. But as critical as these hormones are, researchers who study the effects of environmental contamination particularly follow the doings of estrogens and androgens. These sex hormones don't wait until puberty to kick in. Even before birth, they begin what will be a life-long balancing act to keep the sexes distinct. One of their first jobs is to help ovaries and testes form properly in embryos. After birth, the endocrine system periodically adjusts the ratio of the hormones--females and males produce both estrogens and androgens--so that animals properly progress in their sexual development.
But researchers worry that man-made chemicals might be mimicking or diluting natural hormones and, in the process, throwing a monkey wrench into the early sexual development of Apopka alligators, birds in several U.S. sites, and fish in North America and the United Kingdom. Some scientists have even suggested a link between environmental contamination and declines in human reproductive health. In the past few years, a spate of scientific reports and news accounts has documented decreases in sperm counts and increases in testicular cancer for men and increases in breast cancer for women, perhaps, some speculate, as a result of exposure to hormone-mimicking man-made chemicals.
The concerns are so great that the Environmental Protection Agency's (EPA) Health Effects Research Lab convened a meeting in April in North Carolina to develop a national research strategy, says Robert Kavlock, director of developmental toxicology for the lab. At the meeting, scientists from around the world with expertise in different fields of biology agreed that much more work had to be done before contamination could be directly linked to increases in human reproductive health problems. But, Kavlock says, "There are clear cases where exposure to something has affected animals. The Apopka alligators are the best example."
A Contaminated Lake
"Lake Apopka is a big chemical soup," says Michael Fry, a researcher from the University of California at Davis and an expert on the effects of contamination on birds. About a mile from the lake's shores is a Superfund site that is connected to the lake by a stream. On that site, the Tower Chemical Company had produced dicofol, a mixture of the pesticide DDT and DDE, a by-product of DDT. Periodic spills occurred there, but a particularly large accident in 1980 caused dicofol and sulfuric acid to spread from the stream into the lake, says Guillette. This Superfund site, along with agricultural runoff from citrus lands and vegetable farms, and the presence of the city of Winter Garden's sewage treatment plant, have combined to make the lake one of Florida's most polluted--so much so that the lake no longer produces enough fish to support the bass sport-fishing industry that thrived there before the 1960s.
It will be difficult to prove whether chemicals from pesticide spills long ago or those from more recent contaminations are the cause of current alligator reproductive abnormalities. Or, is some unknown factor responsible for these changes? Even though agricultural runoff is known to cause problems, Guillette and others think that the history of the lake before and after the spills from Tower Chemical tells a compelling story.
Nearly 15 years ago, a team of researchers led by Franklin Percival, now a National Biological Service scientist based at the University of Florida, was asked by the State of Florida to determine how many alligators could be taken from the wild without harming the population. The question had important species conservation and economic ramifications because a multimillion-dollar industry depends on the once-endangered animals. "They are commercially harvested for leather, and they appear on seafood menus and in grocery stores throughout Florida. I've even seen frozen alligator meat in specialty food shops as far north as Pennsylvania," says Allan Woodward, an alligator research biologist with the Florida Game and Freshwater Fish Commission, who works with Percival.
In the late 1970s, scientists considered Lake Apopka a prime place to harvest eggs and hatchlings, and they began collecting there to see how baby alligators would fare in captivity. In 1980 and 1981, Woodward says, the scientists counted populations of 1,200 to 2,000 alligators in a single night on the lake. By the late 1980s, they found 150 a night. "We hadn't seen such a population crash," says Woodward, who continues to monitor the population. There was nothing quite like it elsewhere, even on lakes with comparable agricultural runoff, adds Guillette. In the years just after the 1980 chemical spill, the number of hatchlings dramatically dropped, and Woodward recalls some deaths among the large alligators. "We now know that the crash was linked to poor egg viability," says Woodward. Recently, there seems to be a bit of recovery in hatching rates, he adds.
Systematic studies of thousands of eggs found in nests near several lakes indicated Apopka's egg profile was abnormal. Alligators lay clutches of 30 to 45 eggs and well over half the eggs are likely to hatch in the wild. (Under ideal conditions in the lab, nearly all baby alligators emerge from healthy clutches.) At Apopka, less than a fifth of the eggs produced live young. In some nests, only a couple of eggs hatched. Of the animals that hatched, half died shortly after birth, says Guillette. Many of the eggs collected had levels of DDE that were higher than those known to harm eagles.
Although many Apopka eggs were duds, the scientists could not be sure that all surviving clutchmates would also be biological failures. Given the lifespan of alligators, it will take years to get a complete picture, researchers say. Sturdy specimens can live 50 years, though most animals die by age 40. They do not reach sexual maturity until ten, and females may lay eggs for 20 to 30 years. Females that had reached or that were approaching sexual maturity in 1980 could still be producing offspring. Animals born shortly after the spill are entering what should be their reproductive prime. But Guillette has little hope for the young adults given findings by himself, University of Florida reproductive endocrinologist Timothy Gross, and their colleagues in state wildlife agencies.
A few years ago, when the scientists compared juvenile alligators from Apopka with alligators from Lake Woodruff National Wildlife Refuge, a relatively pristine area for Florida, they found Apopka's animals abnormal at all stages of early sexual development. From the time alligators are embryos, both sexes produce estradiol, a kind of estrogen, and testosterone, a type of androgen. Normal animals have both these hormones in a particular balance that changes over time. At Apopka, female hatchlings and sixmontholds had elevated levels of estradiol."They were superestrogenized," says Guillette. Further, the Apopka males had quite high levels of estradiol relative to testosterone.
Apopka's juveniles grow at the same speed as Lake Woodruff's young, and, by outward appearance, they look like any other suppleskinned alligator. But they are sexually malformed. Studies of twoyearold to eightyearold animals showed some males with penises onequarter to onehalf the size of normal. The males also formed a structure something like a follicle, the body where eggs develop in females. Females with misshapen ovaries had follicles containing three or four eggs instead of the usual one. Neither sex was making appropriate amounts of reproductive hormones.
Still, these findings don't prove that chemicals like those found on the Superfund site feminize the animals. Even though the lake is about as close as one can get to a controlled experiment in the wildthe scientists have a good idea what was put into the lake and when it was put there, and they have a relatively pristine site for comparisonit isn't a lab where variables can be carefully manipulated. So in the summer of 1993, Gross and Percival collected eggs from nests at uncontaminated Lake Woodruff and brought them into the lab for a controlled chemical study. The eggs were put in incubators packed with sphagnum moss to imitate as much as possible the nests built by mother alligators. Then, to guarantee a nest full of malesthe researchers wanted males in order to highlight any feminization that occurredthey used a trick of nature. For many reptiles, including alligators, the temperature experienced by eggs at a key period determines whether a male or a female animal will emerge from the egg. By incubating at 34 degrees Celsius, the scientists could produce allmale clutches. During incubation, some of the eggs were painted with DDT, DDE, or estradiol. Others were left unpainted.
All the unpainted eggs hatched, but only threequarters of the ones painted with chemicals did. More important, DDE levels and hormone balances in the egg-painted hatchlings were comparable to those seen in animals that had been born at Lake Apopka. The finding supports the hypothesis that man-made chemicals can significantly alter embryos, says Guillette.
The scientists are now evaluating older animals. Gross, Woodward, Percival, and Ken Rice of the University of Florida began trapping adult females near their nests last summer. They aim to collect fat samples from the animals' abdomens and some of their eggs. Wildlife biologists know from studying many species that a mother animal may pass contaminants she has stored in her fat reserves on to her eggs. Gross, who will do the hormone and chemical evaluation, says the researchers hope to determine whether the mother alligators have accumulated significant amounts of chemicals that might be feminizing their offspring. "This is an important piece that is still missing from the puzzle," Gross adds.
Beyond Alligators
Do any of the other species that share the Apopka alligator habitat have problems? Preliminary research suggests they do.
Some birds on the lake have laid thinshelled eggs, says Fry. He begins a study this year to look for the sorts of reproductive abnormalities in Apopka birds that he has seen elsewhere. He recently discovered feminized sex organs in some male embryos of common terns collected near a toxic waste site in Massachusetts. To learn how fisheating anhinga and glossy ibis are faring, Fry is collecting eggs from Lake Apopka and other Florida lakes. He'll look for pollutants in the yolk sac and signs that the birds' livers are responding to them. In development, the liver of embryonic birds will produce certain enzymes only when contaminants are present. "If the enzymes are there, it will indicate which birds have been exposed," Fry says.
Gross
and Guillette also have seen abnormalities in redbellied
sliders, a turtle common in the Lake Apopka area. No turtle
hatchlings from eggs collected at Lake Apopka produced normal
amounts of androgens. And, says Gross, when he and his researchers
treat animals sliders collected from clean sites with DDT,
DDE, or the herbicide atrazine, they find alterations.
Gross also has begun a chemical treatment study of largemouth
bass with funding from the National Institute of Environmental
Health Sciences. Some of Apopka's male bass have elevated
levels of vitellogenin, a substance used by egg-laying animals
like fish in the production of yolks. Although in many species
both sexes produce vitellogenin, males normally produce tiny
amounts compared to females.
Unusually high levels of vitellogenin in male fish in Apopka raises red flags in light of recent findings by researchers in Great Britain. John Sumpter, a biologist at Brunel University in Uxbridge, England, started studying the effects of sewage-treatment effluent on trout after anglers reported catching fish that were partially male and partially female. When he and a team of researchers from the United Kingdom's Ministry of Agriculture, Fisheries, and Food placed caged, healthy male trout near the discharge points of sewage treatment plants, they found that the animals produced vitellogenin at levels that were 500 to a million times higher than normal for males. Sumpter suspects that breakdown products of detergents are disrupting the endocrine systems of the fish. Studies are being done in the lab to verify this idea and field studies of freeranging fish in U.K. rivers are underway.
How Does Disruption Happen?
With so many environmental chemicals suspected of causing reproductive disruption, may the substances cause mischief in hundreds of different ways? Probably not, say researchers. From what endocrinologists and toxicologists already know about hormones, they propose the following general scenario for disruption. The chemical culprits dock at special sites called receptors, which are located in an animal's cells. Receptors usually are picky about what they allow to bind. For instance, the estrogen receptor accepts estrogens; the androgen receptor takes on androgens. Receptors that would normally accept only estrogens or androgens floating around in the body might accidentally bind a chemical that mimics the receptor's true match. The attachment of the foreign chemical prevents the resident hormones from doing their normal jobs. In the case of the alligators, Guillette suspects that DDE binds to the animals' androgen receptors and prevents androgens from doing their masculinization work. Guillette feels confident that DDE is the culprit--the anti-androgen--based on a recent study by scientists at EPA's Reproductive Toxicology Branch in Research Triangle Park and at the University of North Carolina.
The findings, published in the British scientific journal Nature in June, demonstrate that DDE is indeed an antiandrogen, say William Kelce and Earl Gray, EPA researchers who led the study. They found that DDE prevented androgens from binding to human androgen receptors in cultured cells, and male rats exposed to DDE had physical abnormalities in reproductive parts. DDE's binding also kept certain genes that depend on androgens from turning on in the rats. These genes, which control the development of male tissues, would generally be activated when androgen was bound to its receptor. Hundreds of genes might be affected with an antiandrogen present, explains Kelce.
In Perspective
That small amounts of chemicals might permanently organize an animal's anatomy and physiology in ways that prevent it from reproducing has profound implications for animal and human health. Researchers worry that chemical contamination may thwart attempts to recover endangered species and could threaten now healthy populations.
But imagining the worst-case scenarios for humans is an inclination Sumpter cautions against. He says he regularly gets letters from people who make unreasonable leaps from a few animal studies to their own cases. One letter claimed, "My son has undescended testicles because he fell into a contaminated river," says Sumpter. A woman asked whether a possible exposure to environmental estrogens while in the womb might explain her son's homosexuality. People with breast cancer contact him, grasping at straws, trying to find a reason for their disease. Sumpter points out that, "There are few studies that actually show a tie between exposures and effects. In the case of alligators and the fish in England, we have strong circumstantial evidence. But even there, we may be five to ten years away from knowing the full story on endocrine disrupters."
In studies of human populations with reproductive problems, it is hard to know what these people were exposed to 30, 40, or 50 years, says Sumpter, who adds that lowered sperm counts might just as likely be the result of "too tight underwear and too many hot baths. We won't know until we do the studies."
Like Sumpter, Guillette wants solid information from the field and lab to build the case against endocrine disrupters. He thinks the alligators and other species at Apopka and similar contaminated sites will continue to provide convincing evidence of the threat some man-made chemicals pose to the reproductive future of diverse wildlife.
Anna Gillis is features editor of BioScience, a Washingtonbased publication of the American Institute of Biological Sciences.
(ZooGoer 24(4) 1995. Copyright 1995 Anna Maria Gillis. All rights reserved.)
