et al. demonstrate that at exposure levels far beneath
those found in the lakes, rivers, streams, drinking water and even
rainwater, atrazine causes frogs to mature with multiple, mixed
gonads and to become demasculinized. These effects occurred at exposure
levels 10,000 - 30,000 times beneath levels previously identified
as non-toxic to frogs.
impact on frogs appears to be caused by this herbicide's ability
to promote the conversion of testosterone to estrogen via activity
of the enzyme aromatase. This mechanism is found not just in frogs,
but other vertebrates as well, including mammals. Their study raises
important, and as yet unanswered, questions about the possible role
of atrazine in world-wide frog population declines, and about the
potential for atrazine to affect human health via the same enzymatic
is one of the most widely and heavily used agricultural chemicals
in the world. Each year, American farmers alone apply 60 million
pounds to U.S. farmland to prevent growth of weeds competing with
corn and other crops. The US EPA considers short-term exposure to
200 ppb acceptable for people and allows 3 ppb atrazine in drinking
water. In rainwater in regions where atrazine is applied, atrazine
contamination has been measured up to 40 ppb. It has even been found
at 1 ppb in rainwater in regions where atrazine is not
used. Hence it is virtually certain that frogs (and people)
living in the real world are regularly exposed to atrazine at levels
many times greater than what Hayes et al. report is sufficient
to harm frogs.
did they do? Hayes et al. used two different experimental
designs to study the effect of atrazine on the laboratory rat of
the amphibian world, the African Clawed Toad Xenopus laevus.
In both designs, Hayes took great care to make sure that the scientists
measuring the exposed animals were unaware of which treatment they
had received. Only after all the measurements were taken did a coding
system reveal what treatment each animal experienced.
sample sizes used in the experiments were quite large (90 animals
per treatment); these experiments were repeated four times; and
experiments with atrazine at these low levels were replicated 51
times in Hayes' lab with similar results.
1: expose developing tadpoles to different levels of atrazine and
examine individuals for morphological effects after metamorphisis.
The tadpoles were exposed to concentrations of atrazine from 0.01
ppb to 25 ppb.
2: expose adults directly to 25 ppb atrazine and measure testosterone
and estrogen levels. Adults were used because they found it not
possible to obtain enough serum from tadpoles to do the hormonal
did they find?
Experiment 1: When exposed as tadpoles to as little as 0.1 part
per billion atrazine, individuals mature with hermaphroditic deformities
in the reproductive tract. Normal adult frogs have two gonads, either
two testes if male or two ovaries if female. Between 16% and 20%
of animals treated with atrazine at 0.1 ppb and above had either
two many gonads and/or mixtures of ovaries and testes. This is not
normal. Hayes et al. comment that they had never seen these
effects in 6 years of study of Xenopus involving over 10,000
figures below, from Hayes et al. shows the deformed reproductive
tract of one hermaphroditic frog. This particular individual has
three testes and three ovaries. The microscopic photograph on the
left shows the entire complex of the animals gonads and kidneys.
The microphotograph on the right shows cross-sections through gonadal
material at places specified by the arrows on the left. T= testis;
tadpoles were exposed to atrazine at 1 ppb and above, Hayes et
al. noted a demasculinization of the adults secondary sexual
characteristics, specifically a reduction in the size of the males'
larynges (vocal chords). Normally male larynges are larger than
female. The atrazine-treated males had larynges that were intermediate
in size between normal male and normal female larynges. There was
no effect on the female larynges size.
2: Adult males exposed to 25 ppb atrazine showed a 10-fold
decrease in testosterone compared to controls. This effect
is highly significant statistically. Treated male testosterone
levels were indistinguishable from control females.
Hayes et al.
does it mean?
This research eliminates any doubt about atrazine as an endocrine
disruptor at extraordinarily low levels of exposure. It puts endocrine
disruption high onto the list of plausible factors contributing
to frog population declines. And because of the apparent mechanism
of action, via enhancement of aromatase conversion of testosterone
to estrogen, it raises important concerns about atrazine disrupting
the hormonal control of development in other organisms, including
Hayes et al. summarize in their paper, atrazine contamination
is extremely widespread , including, via atmospheric transport,
in rainwater in regions where atrazine is not used for agricultural
crops. The level of atrazine sufficient to cause reproductive abnormalities
in Xenopushermaphroditismis 1/30th of the level allowed
by the US EPA in drinking water:
recommended application level of atrazine ranges from 2,500,000
29,300,000 ppb, the allowable contaminant level for atrazine
in drinking water is 3 ppb, and short-term exposures of 200
ppb are not considered a health risk. Atrazine can be as high
as 21 ppb in ground water, 42 ppb in surface waters, 102 ppb
in river basins in agricultural areas, up to 224 ppb in Midwestern
streams, and up to 2,300 ppb in tailwater pits in Midwestern
agricultural areas. Atrazine can be found in excess of 1 ppb
in precipitation in localities where it is not used and up to
40 ppb in rainfall in Midwestern agricultural areas."
in many places in the world are undoubtedly exposed to atrazine
at these levels. It remains to be determined whether other frog
species are as sensitive to atrazine as is Xenopus, and whether
the effects are similar. Hayes' work should reinvigorate efforts
to understand the contribution of chemical exposure to frog declines.
has strongly implicated infection by a chytrid fungus as an important
factor driving frog extinctions, other causes, including contaminants
and the introduction of exotic species, remain plausible contributors
to what is most likely a multi-factoral process. And given the ability
of some contaminants to reduce
disease resistance, it is not implausible to hypothesize that
contaminants are involved in the rapid pace at which amphibians
have succumbed to the fungal infection. No research has assessed
the impact of low-level atrazine exposure on the development of
frog immune competency.
added in August 2002: new research
confirms atrazine impairs frog immune systems
research by Hayes et al. also points out the crucial need
to carry out experiments assessing toxicological impacts at environmentally-relevant
levels. Prior work had seemingly dismissed atrazine as an endocrine
disruptor affecting frogs, but the research used exposures that
were 10,000 to 30,000 times higher than the dosage found by Hayes
et al. to produce developmental disruption:
teratogenesis,growth inhibition, and mortality in amphibians
in response to atrazine were not considered environmental concerns
of the high doses required to produce these effects. Effects
in the current study, however, occurred at levels 10,000 times
lower than the dose required to produce effects in amphibians
in these previous studies. Allran
and Karasov (2001) reached the conclusion that atrazine
was probably not a significant factor in amphibian declines
based on their studies of toxicity, deformities, and effects
on feeding and ventilation in leopard frogs that did not produce
noticeable effects below 3 ppm. The current data show that negative
effects on sex differentiation occur at doses 30,000 times lower
than effective doses reported by Allran and Karasov."