Dear Mr. Slaughter,

Thank you for the opportunity to respond to your letter and white paper.

Firstly, with regards to Werner Kloas, my statements have been somewhat misrepresented in your letter. It is MY OPINION that Dr. Kloas’ involvement with Syngenta is inappropriate. Regardless of whether or not a deal was struck before the SAP or after, the very potential for a panel member to benefit from one outcome or another generates problems with the integrity of the process. Imagine if jurors in criminal trials could be hired by the accused (with lucrative contracts) by concluding “not enough evidence”. Again, I am not accusing anyone of anything, it is simply my opinion that it is inappropriate, especially given the many other influences that industry has over the process. This is also not a criticism of the EPA, SAP, or their processes. Syngenta can hire whomever they want. Similarly, Dr. Kloas can work for whomever he wants. In my opinion, Dr. Kloas made a poor decision. As you point out, this important “world-renowned expert” is now no longer eligible to serve on the SAP.

Secondly, with regards to the quality of the Syngenta-funded work, I have already addressed these issues in the Bioscience paper that you cite. Please note that even Syngenta said, “That study is crap, Tyrone, and you know it” (Tim Pastoor, Dec 13, 2005), regarding the EcoRisk studies that they funded (this statement was made to me during the very same APHA conference that you discuss in your letter). So, it is more than just MY opinion.

Thirdly, you must keep in mind that much of the criticisms you cite regarding my work are “expert opinions”, not facts. In some cases, my expert opinion is the same, in others I disagree. Some of the issues are important and some are not. For example, the suggested use of flow-through water is primarily based on practicality, not science. Atrazine is fairly stable and static conditions more closely mimic what tadpoles would experience in a pond in the wild. Static renewal is labor intensive, however, which is why flow-through is recommended. With regards to the other criticisms, you must understand that no scientific study is perfect…no matter who conducts it, no matter who funds it, no matter how much it costs. I used three pairs of animals in each of the two experiments published in the PNAS paper (so, six pairs and the animals in the two different experiments were from two different sources). What if I had used 10, or 100, or 1000, or 10,000? Different experts will have different opinions based on different considerations, but none would be perfect. Regarding measuring “critical parameters”, what if I measured 3, 5, or 1000 parameters? Or in my field studies, what if I measured atrazine 10 times, 1000 times? And should I collect from 100 sites, 10,000 sites? You see? These are all just opinions. However, when multiple laboratories generate similar data (especially given all of the variations and “flaws” in our studies), combined, these data strongly suggest that atrazine is a problem. It is a mistake to focus on any single study. It is also a mistake to focus on the differences and variations in experimental design between studies. The focus should be on the similarities in the findings, despite the differences in design. As James Carr (a member of the Syngenta-funded EcoRisk Inc.) wrote, “… the important issue is for everyone involved to come to grips with (and stop minimizing) the fact that independent laboratories have demonstrated an effect of atrazine on gonadal differentiation in frogs. There is no denying this.” (James Carr, Feb. 14, 2003).

Fourthly, and most importantly, the CRE, like the EPA, continues to focus on the wrong questions. The question is not whether or not Tyrone’s studies are correct (again there is no perfect study), nor is the important question whether atrazine affects frogs. The important question is whether the frog studies, despite their flaws, reflect results in other vertebrates, including humans. The important question is: “Is atrazine a potent endocrine disruptor that affects wildlife and humans?” The amphibian data simply support findings in every vertebrate class examined. Atrazine exposure is associated with demasculinization (chemical castration) and feminization in every vertebrate class examined (fish, amphibians, reptiles, birds, and mammals, including humans). Effects include reduced fertility and feminization in males and reproductive cancers in rodents and humans. The induction of aromatase (the enzyme that converts androgens into estrogens) and the decrease in testosterone and increase in estrogen has similarly been demonstrated. Even an EPA laboratory, which showed that atrazine causes a decrease in testosterone and an increase estrogens in rodents, concluded that “atrazine tested positive in the pubertal male screen that the EDSTAC (EPA Endocrine Screening and Testing Advisory Committee) is considering as an optional screen for endocrine disruptors” (parenthetical phrases mine) as early as 2000 (Stoker et al, 2000).

Though the regulatory branch of the EPA has NEVER asked this important question, they have denied that the evidence even exists: “It has been claimed that research on frogs shows that atrazine causes changes in the production of aromatase, an enzyme that is involved in the conversion of testosterone to estrogen. It has also been claimed that other scientists have shown similar effects in other species … There is no direct scientific information to assess this hypothesis.” (Statement of Anne Lindsay before the Agriculture and Rural Development Committee of the Minnesota House of Representatives, Feb. 16, 2005). The “scientific information to assess this hypothesis” is represented by more than 40 publications, some of which are listed below (out of the 40+, I have only listed six of my own).

So, again, focusing on my work is a mistake. Can all of these studies, conducted by independent laboratories, all on different animal models, tissues and cell lines, under varying conditions, all come to these conclusions as a matter of coincidence? Or is atrazine a potent endocrine disruptor that both chemically castrates and feminizes exposed animals? THAT is the question that we should be asking.

PUBLISHED PAPERS DEMONSTRATING THAT ATRAZINE IS AN ENDOCRINE DISRUPTOR

Babic-Gojmerac T, Kniewald Z, J K. 1989. Testosterone metabolism in neuroendocrine organs in male rats under atrazine and deethylatrazine influence. Steroid Biochem 33: 141-146.

Carr J, Gentles A, Smith E, Goleman W, Urquidi L, Thuett K, et al. 2003. Response of larval Xenopus laevis to atrazine: Assessment of growth, metamorphosis, and gonadal and laryngeal morphology. Environ Toxicol Chem 22: 396-405.

Cooper RL, Stoker TE, McElroy WK. 1999. Atrazine (ATR) disrupts hypothalamic catecholamines and pituitary function. The Toxicologist 42: 60-66.

Cooper RL, Stoker TE, Tyrey L, Goldman JM, McElroy WK. 2000. Atrazine disrupts the hypothalamic control of pituitary-ovarian function. Toxicol Sci 53: 297-307.

Crain D, Guillette LJ, Rooney AA, Pickford D. 1997. Alterations in steroidogenesis in alligators (Alligator mississippiensis) exposed naturally and experimentally to environmental contaminants. Environ Health Perspect 105: 528-533.

Cummings A, Rhodes B, Cooper R. 2000. Effect of atrazine on implantation and early pregnancy in 4 strains of rats. Toxicol Sci 58: 135-143.

Du Preez LH, Solomon K, Carr J, Giesy J, Gross C, Kendall RJ, et al. 2005. Population structure characterization of the clawed frog (Xenopus laevis) in corn-growing versus non-corn-growing areas in South Africa. Afr J Herp: Accepted.

Eldridge J, Wetzel L, L T. 1999. Estrous cycle patterns of Sprague-Dawley rats during acute and chronic atrazine administration. Reprod Toxicol 13: 491-499.

Eldridge J, Tennant M, Wetzel L, Breckenridge C, Stevens J. 1994a. Factors affecting mammary tumor incidence in chlorotriazine-treated female rats: Hormonal properties, dosage, and animal strain. Environ Health Perspect 102: 29-36.

Eldridge J, Fleenore-Heyser D, Extron P, Wetzel L, Breckenridge C, Gillis J, et al. 1994b. Short-term effects of chlorotriazines on estrus in female Sprague-Dawley and Fischer 344 rats. J Toxicol Environ Health 43: 155-167.

Friedmann A. 2002. Atrazine inhibition of testosterone production in rat males following peripubertal exposure. Reproductive Toxicology 16: 275-279.

Hayes T. 2005. Welcome to the revolution: Integrative biology and assessing the impact of endocrine disruptors on environmental and public health. J Integrative Comp Biol 45: 321-329.

Hayes T, Haston K, Tsui M, Hoang A, Haeffele C, Vonk A. 2002a. Feminization of male frogs in the wild. Nature 419: 895-896.

Hayes T, Haston K, Tsui M, Hoang A, Haeffele C, Vonk A. 2002b. Atrazine-induced hermaphroditism at 0.1 ppb in American leopard frogs (Rana pipiens): Laboratory and field evidence. Environ Health Perspect 111: 568-575.

Hayes T, Collins A, Lee M, Mendoza M, Noriega N, Stuart AA, et al. 2002c. Hermaphroditic, demasculinized frogs after exposure to the herbicide atrazine at low ecologically relevant doses. Proc Natl Acad Sci USA 99: 5476-5480.

Hayes T, Stuart A, Mendoza G, Collins A, Noriega N, Vonk A, et al. 2005a. Characterization of atrazine-induced gonadal malformations and effects of an androgen antagonist (cyproterone acetate) and exogenous estrogen (estradiol 17b): Support for the demasculinization/feminization hypothesis. In press.

Hayes T, Case P, Chui S, Chung D, Haefele C, Haston K, et al. 2005b. Pesticide mixtures, endocrine disruption, and amphibian declines: Are we underestimating the impact? In press.

Heneweer M, van den Berg M, Sanderson J. 2004. A comparison of human H295R and rat R2C cell lines as in vitro screening tools for effects on aromatase. Toxicol Letters 146: 183-194.

Keller J, McClellan-Green P. 2004. Effects of organochlorine compounds on cytochrome P450 aromatase activity in an immortal sea turtle cell line. Marine Environmental Research 58: 347-351.

Kettles MA, Browning SR, Prince TS, Hostman SW. 1997. Triazine exposure and breast cancer incidence: An ecologic study of Kentucky counties. Environ Health Perspect 105: 1222-1227.

Kniewald J, Mildner P, Kniewald Z. 1979. Effects of s-triazine herbicides on 5 -dihydrotestosterone receptor complex formation, 5 -reductase and 3 -hydroxysteroid dehydrogenase activity at the anterior pituitary level. J Steroid Biochem 11: 833-838.

Kniewald J, Osredecki V, Gojmerac T, Zechner V, Kniewald Z. 1995. Effect of s-triazine compounds on testosterone metabolism in the rat prostate. J Appl Toxicol 15: 215-218.

Kniewald J, Jakominic M, Tomljenovic A, Šimic B, Romac P, Vranešic Ð, et al. 2000. Disorders of male rat reproductive tract under the influence of atrazine. J Appl Toxicol 20: 61-68.

Maclennan P, Delzell E, Sathiakumar N, Myers S, Cheng H, Grizzle W, et al. 2002. Cancer incidence among triazine herbicide manufacturing workers. JOEM 44: 1048-1058.

Matsushita S. 2003. Atrazine effect on the reproductive tract in the chicken, In: 6th Annual EDC Meeting, Sendai, Japan, 2003.

Mckoy KA, Sepulveda MS, Gross T, S. 2002. Atrazine exposure and reproductive system abnormalities in field collected Bufo marinus, In: Soc Environ Toxicol Chem, 23rd Annual Meeting in North America,, Salt Lake City, UT, 2002.

Miyahara M, Oka T, Mitsui N, Sagoe C, Kashiwagi A, Shinkai T, et al. 2003. Evaluation of atrazine on Xenopus laevis in a partial life test, In: 6th Annual Meeting of Japan Society of Endocrine Dirsuptor Research, Sendai, Japan, 2-3 December 2003 2003; 259.

Moore A, Waring C. 1998. Mechanistic effects of a triazine pesticide on reproductive endocrine function in mature male Atlantic salmon (Salmo salar L.) parr. Pesticide Biochem Physiol 62: 41-50.

Narotsky M, Best DS, Guidici DL, Cooper RL. 2001. Strain comparisons of atrazine-induced pregnancy loss in the rat. Reprod Toxicol 15: 61-69.

Pintér A, al e. 1980. Long-term carcinogenicity bioassay of the herbicide atrazine in F344 rats. Neoplasma 37: 533-544.

Reeder A, Foley G, Nichols D, Hansen L, Wikoff B, Faeh S, et al. 1998. Forms and prevalence of intersexuality and effects of environmental contaminants on sexuality in cricket frogs (Acris crepitans). Environ Health Perspect 106: 261-266.

Roberge M, Hakk H, Larsen G. 2004. Atrazine is a competitive inhibitor of phosphodiesterase but does not affect the estrogen receptor. Toxicol Letters 154: 61-68.

Sanderson JT, Seinen W, Giesy JP, van den Berg M. 2000. 2-chloro-triazine herbicides induce aromatase (CYP19) activity in H295R human adrenocortical carcinoma cells: A novel mechanism for estrogenicity? Toxicol Sci 54: 121-127.

Sanderson JT, Letcher RJ, Heneweer M, Giesy JP, van den Berg M. 2001. Effects of chloro-s-triazine herbicides and metabolites on aromatase activity in various human cell lines and on vitellogenin production in male carp hepatocytes. Environ Health Perspect 109: 1027-1031.

Sass, J. 2003. Letter to the Editor. JOEM 45(4): 1-2.

Shafer TJ, Ward TR, Meacham CA, Cooper RL. 1999. Effects of the cholorotriazine herbicide, cyanizine on GABAA receptors in cortical tissue from rat brain. Toxicology 142: 57-68.

Šimic B, Kniewald Z, Davies J, Kniewald J. 1991. Reversibility of inhibitory effect of atrazine and lindane on 5 -dihydrotestosterone receptor complex formation in rat prostate. Bull Environ Contam Toxicol 46: 92-99.

Spano L, Tyler C, van Aerle R, Devos P, Mandiki S, Silvestre F, et al. 2004. Effects of atrazine on sex steroid dynamics, plasma vitellogenin concentration and gonad development in adult goldfish (Carassius auratus). Aquatic Toxicology (Amsterdam) 66: 369-379.

Stevens JT, Breckenridge CB, Wetzel LT, Gillis JH, Luempert III LG, C EJ. 1994. Hypothesis for mammary tumorigenesis in Sprague-Dawley rats exposed to certain triazine herbicides. J Toxicol Environ Health 43: 139-154.

Stoker T, Laws S, Guidici D, Cooper R. 2000. The effect of atrazine on puberty in male Wistar rats: An evaluation in the protocol for the assessment of pubertal development and thyroid function. Toxicol Sci 58: 50-59.

Stoker TE, Robinette CL, Cooper RL. 1999. Maternal exposure to atrazine during lactation suppresses suckling-induced prolactin release and results in prostatitis in the adult offspring. Toxicol Sci 52: 68-79.

Swan S, Kruse R, Liu F, Barr D, Drobnis E, Redmon J, et al. 2003. Semen quality in relation to biomarkers of pesticide exposure. Environ Health Perspect 111: 1478-1484.

Tennant MK, Hill DS, Eldridge JC, Wetzel LT, Breckenridge CB, Stevens JT. 1994. Chloro-s-triazine antagonism of estrogen action: Limited interaction with estrogen receptor binding. J Toxicol Environ Health 43: 197-211.

Trentacoste S, Friedmann A, Youker R, Breckenridge C, Zirkin B. 2001. Atrazine effects on testosterone levels and androgen-dependent reproductuve organs in peripubertal male rats. J Andrology 22: 142-148.

Wetzel LT, Luempert III LG, Breckenridge CB, Tisdel MO, Stevens JT, Thakur AK, et al. 1994. Chronic effects of atrazine on estrus and mammary gland formation in female Sprague-Dawley and Fischer-344 rats. J Toxicol Environ Health 43: 169-182.

Yukinori K, Place A, Trant J. 2004. Effects of endocrine disrupting chemicals on the expression of CYP19 genes in zebrafish (Danio rerio) juveniles. Aquat Toxicol (Amsterdam) 69: 25-34.