Hazards and Exposures Associated with DDT and Synthetic Pyrethroids used for Vector Control

World Wildlife Fund January 1999 ACKNOWLEDGMENTS

WWF appreciates the contributions and critical comments of the many experts who created and reviewed this report. Primary authors are Dr. Michael Smolen, WWF US; Dr. Susan Sang, WWF Canada; and, Dr. Richard Liroff, WWF US. Additional contributions came from Dr. Donald Mackay and his coauthors on the exposure model; Dr. Lizbeth Lopez-Carillo, Intituto Nacional de Salud Publica; and, Yvonne Martin Portugues-Santacreu. Special thanks are also due to Dr. Polly Hoppin for initiating this project while at WWF US. The editorial assistance of Stephen Leahy and Julia Langer is also acknowledged.

WWF would also like to thank the North American Commission for Environmental Cooperation and The J.W. McConnell Family Foundation (Canada) for their generous funding for this project and WWF’s work to conserve biological diversity.

Hazards and Exposures Associated with DDT and Synthetic Pyrethroids used in Vector Control is part of WWF’s project to gain a legally-binding agreement on Persistent Organic Pollutants (POPs). The exploration of DDT use in malaria control is aimed at documenting not only the hazards and exposures to humans and wildlife but also alternatives that protect both biodiversity and human health. WWF’s ultimate goal in this area is to have DDT banned globally.

Front cover photo credits: Peregrine falcon: US Fish and Wildlife Service; Mother and child from Manang District in Nepal: Galen Rowell

Other related publications: Resolving the DDT Dilemma: Protecting Biodiversity and Human Health WWF Canada and WWF US June 1998 52 pages Available in Spanish and English Also available on-line at www.worldwildlife.org

A Model and Assessment of the Fate and Exposure of DDT Following Indoor Application Katie Feltmate April 1998 125 pages

Disease Vector Management for Public Health and Conservation Dr. Patricia Matteson et al. In Press Approx. 200 pages

All publications available for $10 each (Cdn. or US) from: WWF Canada WWF US 245 Eglinton Ave. East, Suite 410 1250 24th Street NW Toronto, ON, M4P 3B7 Washington, DC, 20037-1175 EXECUTIVE SUMMARY reproductive failure.

This report summarizes the current state of knowledge regarding the health and The largest remaining legal use of DDT is for environmental effects of DDT and synthetic control of disease vectors. Although DDT is pyrethroids. Currently, DDT’s only official used for interior spraying of households, a use, as specified by the World Health WWF-commissioned mass balance model Organization (WHO), is for the control of shows that most of it ultimately ends up disease vectors in indoor house spraying – outdoors where it joins the pool of DDT in although other (illegal) uses are suspected. general circulation. Significant human Synthetic pyrethroids are increasing in exposure occurs from the DDT applied popularity among managers of vector control indoors which ends up in food. Infants and programs as alternatives to DDT, either for those responsible for house cleaning are indoor spraying or for impregnating bednets. particularly exposed to residues on floors and walls. Much of the traditional debate over DDT and DDT-alternatives has focused on effects such Concern about DDT has generally been as cancer (carcinogenicity), impacts on derived from its reproductive toxicity in reproduction, and gross birth defects. animals, as demonstrated by eggshell thinning. Traditional toxicological testing has attempted The mechanisms by which DDT (DDE) to discern these effects through tests that rely causes eggshell thinning are associated with on using high doses of chemicals. This report the inhibition of prostaglandin synthesis, an takes a broader view. In addition to looking at important initial step in shell creation. The these traditional effects, it examines current result—extremely thin eggshells that crack in knowledge about the effects of DDT and nests—has brought several species to the brink synthetic pyrethroids on the endocrine, of extinction. DDT’s estrogenic and/or anti- nervous, and immune systems and behavior, androgenic properties can contribute to and it emphasizes potential hazards from low feminization or demasculinization, resulting in doses of chemicals. altered behaviour, reduced fertility, and birth defects. Other developmental effects may Concentrations in humans of DDT and its involve incomplete urogenital development breakdown product, DDE, are clear and undescended testicles arising from the barometers of exposure. Although DDT prenatal disruption of testosterone. levels are decreasing in parts of the world, there are populations of people and wildlife The biological processes controlled by the that experience concentrations of DDT and endocrine system—including the immune, DDE above critical levels. For instance, nervous, and reproductive systems—are investigations in Mexico and South Africa common to all animals. As such, the adverse reveal that human breast milk contains DDE at health impacts identified in wildlife and concentrations that exceed the guidelines for laboratory animals from exposure to DDT and the acceptable daily intake by infants set by other classes of pesticides serve as indicators the WHO. Moreover, studies have shown that of potential hazards to humans. the length of lactation (milk production) decreases with higher DDE body burdens in Synthetic pyrethroids and DDT have been human mothers, thus depriving infants of associated with irreversible effects on the benefits provided by breast feeding. In developing nervous system. Specifically, addition, DDE concentrations in some bird DDT and some synthetic pyrethroids alter the species are still high enough to cause proportions of neuroreceptors in the developing brain of neonates, leading to effects whose causes are harder to discern. hypersensitivity and behavioural The impacts of chemicals on developing abnormalities. nervous systems, immune systems, and behavior must be taken into account with special attention given to exposure in the womb.

They can also reduce the efficiency of neural Synthetic Pyrethroids: The existing signal transmission. literature on use of synthetic pyrethroids for impregnating bednets and spraying houses By interfering with the endocrine system, fails to mention possible trans-generational specifically lymphocyte function, humoral consequences of chronic human exposures. response, and thymus weight, DDT and Laboratory studies describing such possible synthetic pyrethroids contribute to suppression hazards are summarized in this paper. WWF of immune responses. Symptoms of decreased urges pesticide manufacturers and public immune competency include, among others, agencies to conduct collaborative research to changes in antibody production and the time it analyze the possible hazards from chronic takes to respond to infections. These raise human exposure to synthetic pyrethroids. concerns about the vulnerability of certain portions of the population, such as the elderly In addressing DDT and pesticides which may and the very young. be proposed as alternatives to it, WWF urges application of the “precautionary principle” WWF recommends the following directions that already forms the basis of a growing for future research on endocrine-disrupting number of international treaties and chemicals such as DDT and synthetic agreements. According to the “precautionary pyrethroids: principle,” when substantial scientific evidence suggests good reason to believe that Low-Dose Testing: WWF recommends that an activity, technology, or substance may be traditional government-mandated tests of harmful, action should be taken to prevent chemicals, which historically have focused on harm. In other words, if an activity raises administering high doses of chemicals, usually credible threats of harm to the environment or to adult animals, must be revised to take human health, precautionary measures should account of the hazards associated with the be taken even if cause and effect relationships exposure of fetuses and embryos to extremely have not yet been fully established low doses of chemicals that disrupt the scientifically. hormonal systems of the body. Organisms frequently are exposed chronically to such The scientific findings summarized here doses in the environment. provide support for WWF’s view that DDT should be characterized by the WHO and Transgenerational Effects: Only recently international assistance agencies as a have attempts been made to assess more “pesticide of last resort,” to be used only when insidious and often overlooked efforts related no other vector control methods (including to pesticide exposure. Mushrooming scientific other pesticides) are available and likely to be understanding of the influence of hormonal effective. WHO and other organizations (endocrine) systems on the development and should take this step based on the additional health of humans and wildlife indicates that evidence about the human and biodiversity future consideration of DDT/DDT-alternatives impacts of DDT that has been gathered since must be broadened to address very significant the last major consideration of this issue by WHO’s scientific experts in 1993.

This change in WHO’s characterization should be an interim step en route to a global ban on production and use of DDT no later than 2007, under the auspices of the global treaty on POPs (persistent organic pollutants) that is now being negotiated (with mid-to-late 2000 as the targeted completion date). The 2007 deadline coincides with Mexico’s commitment, pursuant to the North American Regional Action Plan for DDT, to end its use. Mexico is one of the world’s few producers of DDT; if Mexico is willing to make such a commitment, other nations should also be willing and able to do so.

WWF’s initial report on DDT, “Resolving the DDT Dilemma: Protecting Biodiversity and Human Health,” published in June 1998, brings together in summary form the new information on the impacts of DDT and other pesticides used for vector control contained in this report, and six case studies of successful vector control projects which do not rely on DDT. This information provides the rationale for moving away from DDT and other pesticide-dependent malaria programs toward “bio-reliant” vector management techniques. That report and the complete texts of the case studies (to be published as “Disease Vector Management for Public Health and Conservation” in early 1999) are available from WWF.

TABLE OF CONTENTS

PART I – DDT AND OTHER CHEMICALS USED IN VECTOR MANAGEMENT PROGRAMS...... 1 A Brief History...... 1 Insecticides Currently in Use...... 1 Chemical Properties...... 5 Persistence and Transport Characteristics...... 5 DDT in the Arctic Food Web...... 5 Bioaccumulation in Organisms...... 6 The Role of the World Health Organization...... 9

PART II – HEALTH AND ENVIRONMENTAL EFFECTS...... 10 Introduction...... 10 Acute and Chronic Toxicological Effects...... 16 Reproductive Effects...... 18 Effects on the Nervous System...... 22 Effects on the Immune System...... 24 DDT and Cancer...... 31 Summary...... 32

PART III – EXPOSURE AND ITS IMPLICATIONS...... 33 Levels of DDT in Humans...... 33 Levels of DDT and Effects in other Species...... 34 Routes of Exposure...... 35 Synthetic Pyrethroids...... 38 Interpretation of Human Exposure Data...... 38

PART IV – RECOMMENDATIONS FOR RESEARCH...... 38 Low-Dose Testing...... 39 Testing for Transgenerational Effects...... 39 Assessment of Synthetic Pyrethroid Exposure to Children and the Developing Fetus from Bednets...... 40 In Closing – The Precautionary Principle...... 40 I. DDT AND OTHER CHEMICALS USED IN The effects of DDT on wildlife reproduction and VECTOR MANAGEMENT its residues appearing in food products that had been sprayed with DDT became evident in the PROGRAMS 1960s. Long term studies showed that DDT was found at alarming levels in many animal species A Brief History including fish, birds, and mammals. Many birds such as peregrine falcons, California condors, DDT (dichlorodiphenyltrichloroethane) is an and bald eagles with high levels of DDT in their organochlorine insecticide used mainly to bodies began producing weak eggshells, which control mosquito-borne malaria. DDT’s were crushed upon incubation. The result was a insecticidal properties were discovered in the decline in the bird populations and a threat to 1930s by Swiss chemist Paul Müller. Considered their very existence. These findings led to DDT harmless to mammals this odorless, tasteless, use restrictions and bans in the U.S., Canada, white crystalline chemical was used during the and most European countries in the early 1970s. Second World War for crop protection as well as DDT is now banned in 34 countries and severely protection of troops from malaria and typhus. restricted in 34 (IEM on POPs, Annex II). DDT’s characteristics of insolubility in water, persistence, long half-life of 10-35 years and high-contact toxicity made it appear to be the Insecticides Currently in Use ideal insecticide. As a consequence, Müller was awarded the Nobel Prize in 1948. Only a few The World Health Organization (WHO) years later, Swiss scientists confirmed the approves use of DDT in controlling malaria, connection between unborn and functionally- provided several conditions are met, including impaired calves whose mothers had been limiting its use to indoor spraying, taking grazing on pastures that had been sprayed with appropriate safety precautions, and using DDT. Previously, U.S. agricultural researchers materials that meet WHO specifications. Four had linked similar severe impairments in calves major groups of insecticides are available for whose mothers had been eating feed salted with indoor spraying: organochlorine chemicals DDT for pest control (IEM on POPs, Annex II). (DDT), organophosphates, carbamates, and the Still others had found that young roosters treated synthetic pyrethroids (Table I-1). The with DDT had severely underdeveloped testes undesirable effects of DDT are widely known; and failed to grow the normal combs and wattles they have driven the restrictions on DDT that roosters use for social display (Colborn et al., have occurred to date and are responsible for 1996). DDT being targeted in international POPs negotiations. The organophosphates and Regardless of these effects, DDT’s efficacy and carbamates are acutely toxic to humans, and low-production costs made it the most widely pose a high hazard in particular to those who used agricultural insecticide in the world from work with them (Herath, 1995). The synthetic 1946 to 1972. Total world production of DDT pyrethroids are not as toxic as the carbamates or during this period has been estimated from 2.8 organophosphates, and are widely used as an million tonnes to more than 3 million tonnes alternative to DDT or used to impregnate (IEM on POPs, Annex II). bednets. Because most reports of wide-scale applications of pesticides for vector control involve DDT or the synthetic pyrethroids, the discussion that follows focuses mainly on these pesticides. Table I-1: Toxicity of Vector Control Insecticides † Insecticide LD50* Toxicity to Environmental Toxicity Source Oral Humans/Mammals (Rat) Organochlorines DDT 113-800 Can affect liver, kidneys, Toxicity very low to birds, very 1-4 mg/kg immune system. high to fish and aquatic Neurotoxicant, probable invertebrates, nontoxic to bees. carcinogen, teratogen, Chronic effects (eggshell reported reproductive/ thinning, etc) may be endocrine disruptor. significant. Extremely persistent and bioaccumulative. Dieldrin 37-87 Can affect liver. Extremely persistent and 2-8 mg/kg Neurotoxicant, probable bioaccumulative. carcinogen, reported endocrine/reproductive disruptor. Endosulfan 18-160 Can affect kidneys, liver, Toxicity moderate-high to birds, 1-3 mg/kg blood, parathyroid gland. very high to fish and aquatic Neurotoxicant, suspect invertebrates, moderate to mutagen, possible bees. Persistence moderate in teratogen, reported soil, varied in water, low in endocrine/ reproductive plants. Bioaccumulation may be disruptor. significant in aquatic organisms. HCH (lindane) 88-190 Can affect liver, kidney, Toxicity extremely low- 1-4 mg/kg pancreas, testes, nasal moderate to birds, high-very mucous membrane. May high to fish and aquatic affect immune system. invertebrates, high to bees. Neurotoxicant, probable Persistence high in most soils carcinogen, reported and in water, varied in plants. endocrine/reproductive Bioaccumulation significant in disruptor. aquatic organisms. Organophosphates Chlorphoxim >5000 Possible neurotoxicant. No sufficient data found. 5, 8 mg/kg Chlorpyrifos 95-270 Can affect cardiovascular Toxicity moderate-very high to 1 mg/kg and respiratory systems. birds, very high to fish and Neurotoxicant. other aquatic organisms. Poses serious hazard to honeybees. Persistence moderate in soil. Bioaccumulation in aquatic organisms. Fenitrothion 250-800 Neurotoxicant, reported Toxicity low-high to birds, 1, 3, 9 mg/kg endocrine/reproductive moderate to fish, high to disruptor. crustaceans, aquatic insects, and bees. Not persistent. Moderately bioaccumulative. Malathion 1000- Can affect immune Toxicity moderate to birds, very 1-3 10,000 system, adrenal glands, low-very high to fish, high to mg/kg liver, blood. Neurotoxicant, aquatic invertebrates and suspect mutagen, reported honeybees. Low persistence. endocrine/ reproductive disruptor. Table I-1 Continued † Insecticide LD50* Toxicity to Environmental Toxicity Source Oral Humans/Mammals (rat) Pyraclofos 237 No sufficient data found. No sufficient data found. 5 mg/kg Phoxim 300- Neurotoxicant. Short residual life. 5, 8, 10 >2000 mg/kg Temephos 1226- Can affect liver. Potential Toxicity moderate-high to birds, 1 13,000 to cause significant moderate-very high to fish and mg/kg neurotoxic effects with other aquatic organisms, high long-term exposure. to bees. Persistence low- moderate in soil, low in water, high in plants. Potential to bioaccumulate in aquatic organisms. Carbamates Bendiocarb 34-156 Neurotoxicant. Toxicity moderate to birds, 1 mg/kg moderate-high to fish, high to bees. Persistence low in soil. Not bioaccumulative. Carbosulfan 91-250 Neurotoxicant. Toxicity moderate-high to birds, 19 mg/kg very high to fish and other aquatic organisms. In soil rapidly transformed to carbofuran, which is moderately persistent. Low potential to bioaccumulate. Propoxur 100 Can affect liver. Toxicity high-very high to birds, 1, 4 mg/kg Neurotoxicant, probable low-moderate to fish and other carcinogen, teratogen. aquatic species, high to honeybees. Persistence low- moderate in soil. Low bioaccumulation. Synthetic Pyrethroids Bifenthrin 54-70 Neurotoxicant, possible Toxicity moderate to birds, very 1, 3, 4 mg/kg carcinogen, reported high to fish and other aquatic endocrine/reproductive species, high to bees. disruptor. Persistence moderate. Possible bioaccumulation. Cyfluthrin 869- Can affect kidney. Toxicity low to birds, high to 1, 9 1271 Neurotoxicant fish, other aquatic organisms, mg/kg and bees. Virtually non- persistent to moderately persistent. Moderate bioaccumulation. ß-Cyfluthrin 450 No sufficient data found. No sufficient data found. 11 mg/kg -Cyhalothrin 56-144 Neurotoxicant, reported Toxicity very low to birds, very 1, 3 mg/kg endocrine/reproductive high to fish, other aquatic disruptor. organisms, and bees. Persistence moderate in soil. Bioaccumulation unlikely. Table I-1 Continued † Insecticide LD50* Toxicity to Environmental Toxicity Source Oral Humans/Mammals (rat) Cypermethrin 150- Can affect liver, thymus, Toxicity very low to birds, very 1, 4 (includes ) 4123 adrenal glands, lungs, high to fish and other aquatic mg/kg skin. Neurotoxicant, organisms, high to bees. possible carcinogen. Persistence moderate in soil. Moderate potential to bioaccumulate in aquatic organisms. Deltamethrin 31-5000 Neurotoxicant, reported Toxicity low to birds, high to 1, 3, 9 mg/kg endocrine/reproductive laboratory fish, very high to disruptor. bees. Moderate potential to bioaccumulate. Ethofenprox (or >42,880 Can affect liver, kidney, No sufficient data found. 3-5, 12 etofenprox) mg/kg thyroid. Possible carcinogen, reported reproductive/ endocrine disruptor. Permethrin 430- Can affect liver, immune Toxicity very low to birds, high 1, 4, 9 4000 system. Possible to fish and other aquatic mg/kg carcinogen organisms, very high to bees. High potential for bioaccumulation.

* LD50 indicates the amount of toxicant necessary to kill 50% of the organisms being tested. LD50 is used to measure the acute oral (and dermal) toxicity of a chemical. The lower the LD50 the more poisonous the chemical. Different sources frequently reported different LD50s (rat, oral) for the same chemical. In general, we used range values from the most recent source. When values differed greatly between two sources, we gave range values incorporating all LD50s given in both sources. † Sources varied in their interpretation of the environmental toxicity of each pesticide. Rather than making our own interpretation, we quoted directly from the most recent or most reliable source. 1. Extension Toxicology Network (EXTOXNET): A Pesticide Information Project of Cooperative Extension Offices of Cornell University, Oregon State University, the University of Idaho, and the University of California at Davis and the Institute for Environmental Toxicology, Michigan State University (http://ace.ace.orst.edu/info/extoxnet/). July, 1999 (access date). 2. Colborn T. Endocrine disruption from environmental toxicants. In: Environmental and Occupational Medicine, Third Edition (Rom WN, ed). Philadelphia, PN: Lippincott-Raven Publishers, 1998;807-816. 3. Brucker-Davis F. Effects of environmental synthetic chemicals on thyroid function. Thyroid 8(9):827-856, 1998. 4. United States Environmental Protection Agency, Office of Pesticide Programs. Pesticidal chemicals classified as known, probable or possible human carcinogens (http://www.epa.gov/pesticides/carlist/). July, 1999 (access date). 5. Meister RT, Sine C, eds. Farm Chemicals Handbook ‘99. Willoughby, OH: Meister Publishing Company, 1999. 6. United States Environmental Protection Agency, Integrated Risk Information System (IRIS) (http://www.epa.gov/iris/). July, 1999 (access date). 7. Spectrum Laboratories Inc (http://www.speclab.com/). July, 1999 (access date). 8. Hayes WJ, Laws ER, eds. Handbook of Pesticide Toxicology. San Diego, CA: Academic Press, Inc., 1991. 9. Andersson L, Hemming H, Johnson A, Kling L, Tornlund M. Hazard assessments of chemical alternatives to POP pesticides. Annex 1 to Chapter 3, pp. 202-279 in: Alternatives to Persistent Organic Pollutants (KEMI Report 4/96). Solna, Sweden: Swedish National Chemicals Inspectorate and Swedish Environmental Protection Agency, 1996. 10. Lu FC. A review of the acceptable daily intakes of pesticides assessed by WHO. Regulatory Toxicology and Pharmacology 21(3):352-364, 1995. 11. Chavasse DC, Yap HH, eds. Chemical methods for the control of vectors and pests of public health importance (WHO/CTD/WHOPES/97.2). World Health Organization, 1997. 12. Food and Agriculture Organization/World Health Organization, International Programme on Chemical Safety. Etofenprox (JMPR 1993). Pp. 215-232 in: Pesticide residues in food—1993: Part II—toxicology (WHO/PCS/94.4). World Health Organization, 1994. Chemical Properties DDT sprayed on a field can remain in the soil 10 to 35 years after its last application. For DDT is available in several different forms: example, an Oregon (U.S.) orchard still had 40 aerosol, dustable powder, emulsifiable per cent of the original DDT used 20 years later. concentrate, granules, and wettable powder. DDD has also been shown to be even more Technical grade DDT is actually a mixture of persistent in soils, sediments, and waters, lasting three isomers of DDT, including the p,p’-DDT as long as 190 years (IEM on POPs, Annex II). isomer (85%) with the o,p’-DDT and o,o’-DDT isomers present in much lesser amounts These compounds do not remain in the soil, but (ATSDR, 1994). The content of these isomers is are transported into the general environment by important because the o,p’(ortha-para) isomer is the processes of volatilization, through wind and said to be five to nine times less toxic in tests water erosion. Although more than 20 years with rats than the p,p’(para-para) isomers. While have passed since the last applications of DDT, DDT is highly resistant to degradation, some cotton soils are estimated to be volatilizing 110 microbes can degrade DDT into a variety of tonnes of DDT and its metabolites annually into metabolites. Among the more important of these the atmosphere. These small particles are is DDE and TDE (DDD). The latter is also transported long distances on air currents, and manufactured as a commercial product (IEM on are returned to the land surface by precipitation. POPs, Annex II). DDT in the Arctic Food Web Persistence and Transport There has been very little local use of DDT in Characteristics the high arctic, therefore the presence of DDT in At present, most of the millions of tonnes of arctic biota is indicative of the global or DDT that have been produced in the past hemispherical transportation of this compound. continue to be transformed and redistributed DDT has been found at various concentrations in throughout the environment. DDT and its all trophic levels of the arctic food chain. Table metabolites have been detected in virtually all I-2 is a summary of DDT concentrations found media throughout the world. An extremely in the lower trophic levels of the arctic marine stable chemical compound, 50 per cent of the food web. Table I-3 shows concentrations of DDT in the blubber of arctic mammals.

Table I-2: DDT Concentrations (ppb lipid wt.) in Marine Biota in Various Locations in High Arctic [adapted from Jensen, J., K. Adare, and R. Shearer (eds.) Canadian Arctic Contaminants Assessment Report. (Ottawa, Ontario, Canada: Department of Indian Affairs and Northern Development, 1997)] Biota Region Total DDT

Epontic Particles Ice Island 20-70 Barrow Strait 150-360 Zooplankton Ice Island 8-150 Barrow Strait 2-20 Amphipods Pelagic Ice Island <350 Pelagic Barrow Strait 3-60 Benthic Arctic Ocean 2,200-25,900 Benthic Barrow Strait 15-1,590 Biota Region Total DDT

Fish Arctic Cod Lancaster Sound 66-120 Barrow Strait 15-255 Turbot Cumberland Sound 626-1,044 Beaufort Sea 659-1,1251 Four-horn Sculpin Wellington Bay 93 Cambridge Bay 1,225 Hall Beech 135 Bivalves Clams Sanikiluaq 34 Septentrion sp Manitounuk Sound 13

Table I-3: Mean Concentrations (ppb wet wt.) of Total DDT in Blubber of Arctic Mammals [adapted from Jensen, J., K. Adare, and R. Shearer (eds.) Canadian Arctic Contaminants Assessment Report. (Ottawa, Ontario, Canada: Department of Indian Affairs and Northern Development, 1997)] Species Female Male

Ringed Seal 473 959 Harp Seal 486 NA Beluga Whale 1,940 4,974 Narwhal NA 3,232 Walrus 744 1,744 Adult Polar Bear (Bernhoft et al., 1997) 372 340

Water runoff provides another mode of Bioaccumulation in Organisms transportation. DDT sticks to soil particles by the process of adsorption. These particles are Bioaccumulation Potential transported to lakes and rivers and are the Bioaccumulation reflects the relationship principal route by which lakes and streams between how much is taken into an organism by become contaminated. In an experimental plot of exposure versus how much is lost through cotton, runoff waters transported 2.8 per cent of metabolism and excretion. The key in pesticide the DDT applied in six months. exposure scenarios is whether the rates of Under tropical conditions, residues continue to metabolism and excretion remove enough of the be detected in major water bodies in the substance to prevent a gradual increase in the Philippines despite DDT’s restricted-use status. organism. If the rates of metabolism and Fish, as well as duck eggs, from lake areas also excretion are not rapid, an organism will show residues (IEM on POPs, Annex II). accumulate ever-increasing concentrations, While DDT will evaporate and photo-oxidize adding to the concern about chronic, low-dose from soil surfaces to a certain degree, it is a exposures. robust and long-lived chemical compound. Even Chemicals that are water soluble are more easily when its use is banned globally, DDT and its excreted, as well as more easily mobilized to various metabolites will continue to travel in the sites responsible for metabolism of the winds and waters and accumulate in the bodies compound. On the other hand, a chemical with of the world’s organisms for decades to come. high solubility in lipids (fats, oils, or waxes) has bioaccumulation potential. Such lipophilic America’s Great Lakes basin despite restrictions chemicals easily move into cells and are on its use in the United States and Canada. It sequestered in fat where they can become more appears that much of the DDT currently being persistent. DDE is an example of a lipophilic deposited in the basin is atmospherically chemical that resists enzymatic degradation and, transported from Central and South America. therefore, rapidly bioaccumulates. DDT is also Even though the concentration of DDT in lipophilic, however, it is more readily degraded plankton is 1/100 part per million, the flesh of a and excreted from the body. fish-eating bird in the same lake system may contain 630 times that concentration (Colborn et Bioaccumulation in the Great Lakes Food al., 1990). Web DDT continues to be deposited in North

Table I-4: Bioaccumulation of DDT in Lake Ontario Food Web (Colborn et al, 1990) Species Concentration (ppm wet weight)

Plankton 0.01 Mysis 0.03 Pontoporeia 0.10 Sculpin 0.40 Smelt 0.40 Lake Trout 1.10 Herring Gull 6.30

Synthetic pyrethroids are also lipophilic though (0.75%), cypermethrin (10%), or cyhalothrin they are more like the isomers of DDT in that (4.5%) to dairy cows was detectable in both the they can be metabolized to more water soluble cows’ blood and milk for 28 to 35 days forms that can then be excreted. Furthermore, (Bissacot et al., 1997). In these situations, the sites where they can be metabolized are not bioaccumulation results in much lower peak limited to the liver and therefore, metabolism is concentrations since the differences between much quicker. For example, the elimination exposure and intake are not widely different half-life for deltamethrin in plasma of the rat is from metabolism and excretion. The concern 33 hours (Anadon et al., 1996) with almost would be if the exposure is periodic, with a span complete elimination from the body by day 4 shorter than the rates of excretion. Chronic, low- (Ruzo et al., 1978). Cypermethrin is more dose exposures may lead to slightly increased resistant to elimination; 90% is lost in the first concentrations in the body. There is little known four days, however, total elimination may take about the pharmacokinetics of the synthetic as long as 17 to 26 days (WHO Working Group, pyrethroids. 1992). Synthetic pyrethroids (permethrin, deltamethrin) DDT Bioaccumulation in Humans are rapidly distributed in the body (Anadon et In surveys around the world of human blood, fat al., 1991, 1996). The primary sites of deposition tissue, and breast milk, DDT and its metabolites are the central nervous and peripheral nervous are found in substantial quantities (Thomas and systems, which can have concentrations of Colborn, 1992; Jensen, 1990). For example, permethrin ranging from 1.5 to 7.5 times higher Table I-5 reviews concentrations of two isomers than those observed in plasma. In another study, that are known endocrine disruptors. Since DDT a single topical application of deltamethrin is very lipophilic, it accumulates in all fats, including the 3% fat found in breast milk source (breast milk) and a lower total body fat (Rogan et al., 1986). The quantity of DDT and content in the baby. As the baby matures, fat DDE varies with the age of the individuals with accumulations increase the available pool which young individuals having higher concentrations in effect dilutes the DDT/DDE enriched fetal fat than older individuals. This is probably the reserve. result of a combination of a pesticide-rich food

Table I-5: Concentrations of o,p’-DDT or p,p’-DDE (endocrine-disrupting isomers of DDT) in Breast Milk of Women (standardized to ppm fat) Country # Year o,p’- p,p’-  Citation Women DDT DDE DDT Canada 497 1995 0.003 0.22 Newsome et al., 1995 U.S.A, New York 7 1985-87 - 0.54 - Schecter et al., 1989 Mexico, Veracruz 43 1994-95 0.27 5.02 6.44 Waliszewski et al., 1996 Mexico, Mexico-City 50 1994-95 0.14 0.59 0.93 Torres-Arreola et al., 1998 Germany 150 1985-87 - 0.75 - Schecter et al., 1989 Spain, Madrid 51 1991 - 0.60 0.66 Hernandez et al., 1993 Norway 20 1988 0.97 Skaare et al., 1988 United Kingdom 193 1989-91 - 0.40 - Dwarka et al., 1995 France 20 1990-91 - 2.18 - Bordet et al., 1993 Slovakia 50 1994 1.20 Prachar et al., 1996 Yugoslavia, Krk Island 33 1986-87 - 1.10* - Krauthacker, 1991 Labin 20 1986-87 - 0.55* - Krauthacker, 1991 Croatia 50 1981-82 1.90 Krauthacker et al., 1986 Nigeria 10 1987 0.99 Atuma and Okor, 1987 Nigeria, Benin 35 1981-82 - 1.1 - Atuma and Vaz, 1986 Kenya 68 1983-85 1.73 Kanja et al., 1986 New Guinea, Papua 41 1990 - 0.45 0.89 Spicer and Kereu, 1993 Uganda 143 1992-93 0.06 2.35 Ejobi et al., 1996 Zimbabwe, Kariba 39 1994 13.60 25.26 Chikuni et al., 1997 Australia, Victoria 60 1995 0.96 Quinsey et al., 1995 India 60 1985-86 1.43 7.28 Zaidi et al., 1989 India 25 1988 2.00 Tanabe et al., 1990 Jordan, Amman 15 1989-90 0.23 2.04 3.31 Alawi et al., 1992 Saudi Arabia 115 1995-96 - - 0.27 Al-Saleh et al., 1998 Turkey 104 1995-96 - 2.01 2.36 Cok et al., 1997 Thailand, Bangkok 3 1985-87 3.61 Schecter et al., 1989 Vietnam 7 1985-87 6.70 Schecter et al., 1989 Note: Methodologies used to quantify isomers varied, however they allow for comparisons of geographical differences. * Median

months of breast feeding. The levels of DDT in A breast feeding baby can acquire the blood begin to decline at about 3 years of concentrations of lipophilic chemicals at age, again probably reflecting the shift in diet to extraordinary rates. Mes et al. (1984) estimated a less contaminated food, and an increase in new that babies could acquire 1.8 micrograms of fats. p,p’-DDE per gram body fat (or 1.8 ppm) by the 14th week of breast feeding from breast milk alone. Furthermore, the infant’s DDT levels Based on observations in South Africa of DDT could reach those of the mother in the first three (and DDT derivatives) in breast milk, Curtis (1994) estimated that a baby fed entirely by Lactational transfer is not limited to DDT or breast milk exceeds the allowable daily intake DDE. Any pesticide that enters the body can be (ADI) for DDT (0.02 mg/kg), as determined excreted in breast milk (Rogan and Ragan, by FAO/WHO (1985), by 5 to 18 times. Rogan 1994). Synthetic pyrethroids have also been and Ragan (1994) estimated that over a nine- reported in the milk of dairy cows when month period of breast feeding, an infant can pesticides were applied as a part of an acquire 21.5 mg of DDE based on the 90th ectoparasite control program. Deltamethrin, percentile level. Such estimates identify breast cypermethrin, or cyhalothrin were reported in feeding as a principal source of exposure to milk within 24 hours of application (Bissacot DDT and DDE. It must be pointed out that this and Vassilieff, 1997), with concentrations of exposure of the newborn coincides with 0.51, 0.36, and 0.19 ppm, respectively. The development of their brains (Eriksson, 1997), so organophosphate pesticide, chlorfenvinphos, such exposure has implications for neural another topical treatment for ectoparasites in development, behaviour, and susceptibility to cattle, was recorded to vary from 1.18 to 10.40 insecticides later in life (Johansson et al., 1996). ppb in milk from cows in Kenya (Kituyi et al., This concern for exposure during early 1997). Only recently has attention been focused development will be discussed in more detail on the transfer of synthetic chemicals in human later in the paper. or cow milk, and therefore, it is not known what the magnitudes of transfer are for commonly The DDT and DDE a nursing baby acquires used pesticides. from breast milk come directly from the low background exposures and accumulation over The fact remains that this lactational transfer is the years in the mother. In one study, levels of the rule of pharmacokinetics of synthetic DDE were found to be 17% lower in women chemicals, and not the exception. The concern who had breast fed previously as compared to increases as the transfer rate and concentration those who had not. This was confirmed in of the chemicals increases, which is related to studies of women who were followed through both the application rate, frequency, and the two pregnancies, where there was a 23% bioaccumulation potential of the chemicals. difference in DDE levels between their first and second child. The Role of the World Health The quantity also changes with the length of the Organization nursing period – declines in DDE in milk of 20% at six months, and a 40% decline by the National health authorities determine the nature 18th month of nursing (Rogan et al., 1986). The of their malaria control efforts, including amount of DDT and DDE in the bodies of the whether or not DDT is to be used, based on women participating in this study was the result specific national circumstances, priorities, and of low background exposures beginning early in political considerations. However, they do their lives. It was not the result of accidental or receive and rely on guidance from the WHO. agricultural exposures. The WHO’s Expert Committees collect and analyze information and the WHO supports and The persistence of lipophilic chemicals is cause participates in the development of new methods for concern because exposure to low and strategies for disease control, issuing concentrations over an extended period of time recommendations and technical guidelines (Bos, may lead to substantial burdens later in life. In pc). an analysis of DDT exposures associated with indoor application of DDT for malaria control in In 1992, governments signed a WHO-sponsored KwaZulu, Africa, Bouwman and colleagues World Declaration on the Control of Malaria and (1991, 1993, 1994) found that household endorsed a Global Strategy for Malaria Control. members in regions that used indoor pesticide Prior to this time, DDT was considered the applications for vector control had significantly insecticide of choice and it was thought that higher DDT levels in their sera than those in there were no acute toxic health impacts to regions where no spraying occurred. applicators nor any significant risks to humans. However, the new Global Strategy recognized addition to scientists’ and regulators’ knowledge the need for an integrated approach that involved of more traditionally recognized acute and early diagnosis and prompt treatment; selective chronic effects. Cancer, reproductive, and and sustainable preventive measures, including teratogenic endpoints have historically been the vector control; prevention, early detection, and basis for public health use approvals; the containment of epidemics (WHO, 1993). growing body of evidence of effects on immune function, the nervous system, and certain aspects By 1993, against the backdrop of a report on of reproduction in both humans and wildlife, has high levels of DDT in breast milk of women and not been considered or examined thoroughly. the occurrence of cancer, a WHO expert group Such effects are not the result of genetic, re-examined DDT. They concluded that the behavioural, or immune competence. These evidence on the adverse effects of DDT as a health endpoints are primarily mediated result of indoor residual spraying was mutations, but arise from problems in gene insufficient. However, because new and safer expression. These result in functional changes insecticides such as synthetic pyrethroids were that affect future performance, whether it be available, the experts indicated that “DDT no physiological, reproductive, neural, through the longer merits being considered the only neural and endocrine systems. This section insecticide of choice” (WHO Report Series 857, outlines the latest information regarding the key 1995). mechanisms by which pesticides used in vector The group also urged closer examination of such control can cause endocrine disruption with health issues as cancer and exposure of babies to potential for reproductive, neural, immune contaminated breast milk. A background paper impairment/dysfunction and cancer. for the group had concluded that “it can no longer be confidently stated that DDT anti- Overview of The Endocrine System malarial spraying is harmless to human health”(Curtis, 1994). The endocrine system in humans and other organisms consists of glands that secrete In 1997, reflecting growing interest at various hormones into the bloodstream to regulate body international forums in the reduction and processes. Components of the endocrine system elimination of POPs, the World Health that control development and function include Assembly adopted a resolution to reduce the ovaries and testis, thyroid, adrenals, and reliance on insecticides for control of vector- parts of the pancreas gland and their many borne disease by promoting Integrated Pest hormones, including estrogen, testosterone, and Management and ensuring that DDT was used thyroxine. Hormones are specialized molecules only within programs that take an integrated that travel through the bloodstream and interact approach (WHA, 1997). with a specific target cell (Table II-1). These In late 1998, an Expert Committee will develop chemical messengers trigger a complex series of recommendations for consideration at the events that choreograph development and January 1999 meeting of the WHO Executive function and are involved in: Board (Bos, pc, 1998).  sexual differentiation: regulating the rates of cell division leading to the construction of II. HEALTH AND tissues and organs that eventually determine ENVIRONMENTAL future function, such as sperm production EFFECTS and ovulation;  controlling the development of cells that Introduction make up the immune system, thus affecting A new set of concerns is emerging regarding the future ability to combat disease; and, impact of DDT and other pesticides on the endocrine system. These are a significant  influencing neural development, such as that required for constructing the hypothalamus, chemical concentrations. In the womb, for cerebellum, and other regions of the brain. example, natural hormones cause effects at concentrations of trillionths of a gram. The endocrine system operates at very low

Table II-1: Selected Hormones: Sources and Effects Hormone Source Effects on

Estrogen ovary Reproductive tract development and function; Expression of secondary sexual characteristics; brain development and behaviour Testosterone testis Reproductive tract development and function; Expression of secondary sexual characteristics; brain development and behaviour Thyroxine thyroid Brain development and behaviour; growth; metabolism Mullerian-Inhibiting Substance (MIS) testis Development of reproductive tract

The release of hormone molecules is carefully throughout life, it is “highly conserved.” In other regulated and balanced by a variety of feedback words, distantly related groups like reptiles, mechanisms. In each case, the response depends insects, birds, mammals and humans share upon the binding together of a hormone almost identical hormones and receptors, as well molecule with a specific receptor (protein). as similar biological responses. As a result, Together, they initiate specific responses in the certain effects observed in the development of cell through the expression of genes that are particular species can convey potentially coded on the cell’s DNA, yielding enzymes that important lessons regarding the hazards of influence specific biochemical pathways. This chemicals to other species, including humans. results in changes in rates of cellular activity, These similarities among conserved systems shifts within reproductive cycles, or increases in form the basis for the testing of chemicals on rates of cell divisions. animals in laboratory settings and the drawing of inferences from those tests about hazards to The endocrine system is extraordinarily human health. They also raise concerns about complex. For example, cells in different tissues observed effects in wildlife and the implications respond differently to the same hormone. In for human health. brain tissue, estrogen can alter behaviour, while the same hormone, when “plugged into” Endocrine Disruption: The Newly- receptors in cells lining the reproductive tract, Emerging Science can initiate changes in a reproductive cycle such as cell proliferation. Moreover, cells of the same Human-made chemicals can interfere with or tissue may respond differently to the same perturb the endocrine system at both high and hormone at different stages of development. low doses. Such perturbations have been More importantly, hormone-receptor reported in numerous laboratory studies and in interactions occurring early in development field studies involving exposure of wildlife to frequently lead to a cascade of developmental chemicals released into the environment. The events that are irreversible (Colborn and well-documented story of diethylstilbestrol Clement, 1992). (DES), a pharmaceutical administered to pregnant women to increase the probability of Because of the critical role the endocrine system successful births, exemplifies the plays in directing development and maintaining “transgenerational” effects of estrogen-like the physiological homeostasis of organisms compounds, including human-made chemicals released into the environment (Bern, 1992). The except for disruption of neurotransmitters, these effects of the confirmed estrogen-mimic DES on processes will not be discussed in this paper. mothers were minimal, and their babies were The developing offspring is the most sensitive born “healthy.” However, some of the exposed target of endocrine disruption since much of the children developed a variety of anomalies later neural, reproductive, and immune development in life which subsequent research has linked to occurring in the womb continues into early the disruption of their endocrine system in utero childhood (Bern, 1992). Many human-made by DES. chemicals can cross the placental barrier, thereby allowing the mother’s body burden of Endocrine disrupting chemicals work by a chemicals to be shared with her developing variety of mechanisms : offspring. Doses of chemicals during embryonic, 1. As agonists, they impersonate natural fetal, and early post-natal development can be hormones by binding to receptors and the highest exposures encountered throughout initiating a new cellular response. life. 2. As antagonists, they bind and block the Chemicals vary in their endocrine-disrupting receptor, thereby making these regulatory effects since they interact with different switches unavailable to signals from the receptors and target cells, accumulate at body’s naturally produced hormone different rates, and have different binding messengers. affinities. In field and laboratory studies, 3. By altering concentrations of natural perturbations to the endocrine system leading to hormones through promoting or interfering developmental, reproductive, behavioural, with the breakdown of the hormone by the immunological, and physiological changes have liver’s enzyme system. been demonstrated. Examples include thyroid 4. By altering the number of receptors in dysfunction in birds (Moccia et al., 1986), fish developing tissue types, thereby (Moccia et al., 1981), and mammals (Brouwer et predisposing these tissues to abnormal al., 1989); growth or developmental responses later in life. irregularities in fish (Jobling et al., 1996, Gray and Metcalfe, 1997; Leatherland, 1993), reptiles The net result of any of these disruptive (Guillette and Crain, 1996; Guillette et al., mechanisms is a perturbation to systems that are 1994), turtles (Bergeron et al., 1994), and critical for the creation and maintenance of the mammals (Gray and Kelce, 1996; Gray et al., body plan that has been molded by natural 1994); behavioural abnormalities in birds selection over countless generations. These (Barron et al., 1995); demasculinization and perturbations are not from a genetic mutation but feminization of males in invertebrates (Ellis and from confused chemical messenger systems that Pattisina, 1990), fish (Davis and Bortone, 1992; alter how and when genes are “expressed.” Gimeno et al., 1996), birds (Fry and Toone, These changes in expression can lead to 1981), and mammals (Facemire et al., 1995); “functional deficits” – changes in how well an defeminization and masculinization of females organism’s immune, reproductive, and other in fish (Monosson, 1997), and compromised systems perform. immune systems in birds (Grasman et al., 1996) In addition, endocrine disruption is not limited and mammals (Ross et al., 1996). to interference with receptors. Other mechanisms involve alterations in Key Considerations for Health Effects neurotransmitters (nervous system), cytokines Determinations (immune system), and expression of liver DDT and its metabolites have received enzymes whose presence alters hormone considerable attention in part because of their concentrations in the blood. However, little is persistence and ability to accumulate in fat. known about the mechanisms involved with Concentrations are easy to quantify, and DDT’s these pathways, including possible effects from association with eggshell thinning in wild-bird pesticides used in vector-control programs and, species has given it a high visibility. Research to identify the pathways and mechanisms through Chemical exposures during development can which DDT and several other similar create a lifetime legacy. organochlorine compounds act has led to the Alterations to the function and fate of cells or discovery of steroid hormone agonistic and tissues during development are irreversible. antagonistic receptor binding behaviour, the Once a specific cell type is differentiated and the more subtle effects on nerve performance, and cells are calibrated to perform at a level set by alterations to developmental processes. the embryonic environment, the fate of the Furthermore, there is growing interest in organism is set. All of the age classes from possible linkages to immune suppression, developing fetus through young adolescent are behavioural effects, and breast cancer. Less is delicate stages during which chemical exposure known about some of the newer pesticides used can undo the balance designed by generations of in vector-control programs. evolution. For instance, there has been research into For instance, if chemical exposure occurs during synthetic pyrethroids to determine LD50s and a critical pre-natal/neonatal developmental high-dose cancer screening but only recently has window it can cause alterations to an interest been directed at determining their individual’s neural and chemical messenger potential for interacting with the immune systems and have deleterious long-term effects. system. Preliminary work with neural These may include diminished performance of development further indicates that synthetic physiological, reproductive, or immune pyrethroids behave like DDT in causing subtle processes and therefore may not become changes during the development of the apparent unless there is an unusual stress. Food cerebellum, neural transmissions, and behaviour deficiencies, extremes in weather or climate, and (Eriksson, 1992, 1997). other environmental stressors exacerbate an otherwise tranquil physiology. In order to arrive at some conclusions concerning the environmental and public health safety of DDT or its substitutes, much more Biologically active materials have many research needs to be done to understand the ‘targets’ and produce a range of effects. more subtle ‘health endpoints’ discussed in this Multiple targets and varying mechanisms are not paper. However, the scientific literature offers unusual with biologically active molecules. The compelling evidence of effects resulting from isomer o,p’-DDT provides a good example. It exposure to these pesticides that deserve serious alters the sodium/potassium channel in a neuron, consideration. These effects reflect impingement alters calcium balance in gap junctions, and is an on aspects of biological function that are very estrogen agonist. Effects documented involve different from the mechanisms associated with anatomical and morphological changes to the cancer and teratogenesis. The latter are extreme brain and reproductive system, aspects of and easily detectable endpoints, whereas immune suppression, increased or decreased perturbations involving subtle shifts to neural sensitivity to stimuli, behavioural performing neural, endocrine, and changes, and possible increased susceptibility to developmental pathways are conceptually some types of cancer. DDT’s modes of action different. While not ignoring the traditional and effects should not be considered acute and chronic (cancer, teratogenesis) effects exceptional. with which DDT and other pesticides have been associated, this paper deals primarily with the All living organisms share biochemical effects related – in one way or another – with strategies. disruption of the endocrine system. This biological reality means that an adverse effect on the ion pump of an insect should be Some important points that must be expected to similarly affect an ion pump of a considered when evaluating the health risks mammal. Many structures involved in cellular involved: and intercellular functions are shared even among distantly related species. For example, mitochondria power cells in all animals and animals reached adulthood. But no trace of DDT sodium/potassium channels pump ions in nerve was found in their brain tissue one month after cells in invertebrates and all vertebrates. Subtle administration. If investigators were looking for differences are more often related to alternate a cause/effect relationship involving DDT, no pathways or processes that arise with increasing connection would be made. complexity than with alterations of basic Exposure to DDT early in development has also structures. been found to modify the sensitivity of the In addition, similar biological processes are at neurons. When offspring were exposed to play throughout an organism. For example, paraoxon, the metabolite of permethrin, neurons are distributed throughout the body and hyperexcitability occurred at unexpected and perform a variety of specialized and isolated seemingly unexplainable low doses although no functions. Chemicals that alter basic biological DDT residues were detected (Johansson et al., functions, like polarization of neurons or 1996). destruction of synaptic messengers, can lead to Even if a chemical is rapidly metabolized and various effects. Likewise, estrogen mimics excreted, it cannot be assumed there is no health acting through the estrogen receptor have risk, especially when there is chronic exposure. different effects in different tissues and stages in For instance, cypermethrin is an example of a the life cycle. Recent research indicates that rapidly metabolized synthetic pyrethroid that, if estrogen receptors appear in a wide variety of briefly present at a critical time during tissues, e.g. brain, testis, lung, bone cells, and development, can lead to measurable changes in kidney. the immune system later in life.

Chemical culprits may not leave “footprints Alterations in neural or chemical messenger in stone.” systems can lead to many subtle changes in Causal relationships between chemicals and the body. effects are difficult to establish when dealing Alterations in neural or chemical messenger with complex animals that have long life spans, systems can lead to permanent change in cells diverse lifestyles, and multiple chemical and tissues resulting in diminished capacities or exposures. The time from the initial damage to altered outcomes. For instance, exposure to the manifestation of the effect may take many DDT during development can perturb the years or even skip a generation and only be processes which give an individual the ability to expressed in offspring exposed in utero. If the fight pathogens or parasites later in life; chemical is present along with an observed enzymatically metabolize foreign chemicals in effect, then the chances increase of establishing the liver; rally the immune system to contain and a causal link. For instance, when DDT or a destroy foreign materials; respond to synthetic pyrethroid, like deltamethrin, are environmental cues to coordinate reproduction; administered to an adult animal, disruption to its and assimilate a wide variety of stimuli to neural performance is observed in a dose- coordinate behavioural reactions to the dependent fashion. Furthermore, as the chemical environment and the other individuals in it. is metabolized, sequestered in fat, or excreted from the body, the effects diminish. However, effects on the offspring that result from Even low-dose exposure is biologically developmental events may not be so easily relevant. linked. Developmental windows are briefly opened during which time chemical messengers signal Eriksson and colleagues (Eriksson, 1992; the developmental fate of cells and tissues. Eriksson et al., 1992) conducted an elaborate Neither the natural nor impostor chemical cues series of experiments that demonstrated that required to trigger these changes need to be DDT and some synthetic pyrethroids altered overwhelming. For instance, mice exposed brain structure and function. Neural tissue perinatally to a single dose of 0.5 mg/kg DDT appeared to become more hyperactive when the (Eriksson et al. 1992, 1997) experienced level implications ranging from reduced changes in brain development at an estimated 15 effectiveness of immunization programs, parts per trillion in the brain tissue. This is in persistence of pathogens in reservoirs in the contrast to the (high) dosing regimes of 10, 20, population from which new outbreaks can rise, 40, or 100s of mg/kg that are typical of increased parasite loads, and illnesses of longer experiments conducted to assess acute and duration. chronic health effects of DDT using rodents. While behaviour has much plasticity of Very little research has been done using low, expression and is a difficult parameter to assess, environmentally relevant and biologically the kinds of effects that pesticides may have meaningful doses. New research to determine include those that lead to decreases in memory, hormone-receptor-mediated effects of synthetic learning, tolerance of stress or changes in chemicals emphasizes that the effects observed surroundings, and alterations in aggression at low doses usually reflect alterations to the levels. normal functioning of biological systems that are not observed when the system is Endocrine Disrupting Pesticides overwhelmed by doses hundreds of thousands of Many pesticides have been identified as times more concentrated (vom Saal et al., 1997). endocrine disrupting chemicals, including While such extremely high doses are useful in dicofol, vinclozolin, chlordecone, toxaphene, identifying some behaviours of chemicals and 2,4-D, 2,4,5-T, atrazine, dieldrin, heptachlor, hazards from cancer, they miss the impacts on mirex and chlordane. Various pesticides used in functional responses through modifications to vector control are also considered endocrine regulatory systems. Low-dose effects need to be disruptors, namely DDT and its metabolites, considered when identifying possible threats parathion, methoxychlor, lindane, endosulfan, from pesticides. chlorpyrifos, malathion, diazinon, carbaryl, and Adverse effects at the individual level have some of the synthetic pyrethroids. However, population-level implications. since current regulations governing pesticides Traditional assessment of health effects focuses the world over do not require screening of on the effects to the individual and rarely pesticides for endocrine disruption, per se, it is extends to populations or ensuing generations. not known for certain how many currently-used However, considering the potential for pesticides are endocrine disruptors. Those developmental effects to alter behavioural, identified thus far are so labeled because of the reproductive, and immune systems, more far- basic research efforts of independent reaching implications need to be considered. For laboratories. instance, immune suppression has population- Endocrine Disruption by DDT and Methoxychlor Concerns about the effects of DDT and its metabolites on the health of wildlife and humans have a long history. Besides well-documented effects on eggshell thinning, a variety of abnormalities are seen in male sexual development. These effects were often described as estrogen-like. One of the chemicals in technical-grade DDT, the isomer o,p’-DDT, is an estrogen hormone mimic that competes for the estrogen receptor. It binds to the receptor and activates biochemical pathways as if it were estrogen. Exposure has been shown to lead to the early appearance of the first ovulation and loss of fertility. Target tissues where effects have been seen include the mammary gland, uterus, cervix, vagina, prostate gland and seminal vesicles. Recently, Kelce and coworkers (1995) have shown that the primary metabolite of DDT, p,p’-DDE, is a potent androgen receptor blocker (antagonist). It binds to the androgen receptor, blocking a switch critical for the development of normal males. Male mice exposed to DDE in utero are feminized and demasculinized. Methoxychlor and its metabolites provide an example of how a pesticide can disrupt the chemical messenger system. Methoxychlor is an estrogen agonist, but is weak compared to its metabolite. Methoxychlor itself has a very low binding ability to the estrogen receptor. However, methoxychlor is metabolized in the liver to 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE) (Cummings, 1997). When HPTE binds to the estrogen receptor, it initiates the same biochemical pathways as estrogen, resulting in increases in specific enzyme activity and changes in both uterine weight and fertility. The effects of methoxychlor vary with concentration (ATSDR, 1994b; Cummings, 1997; Cummings and Metcalf, 1995a; Martinez and Schwartz, 1991; Eroschenko and Cooke, 1990) in a fashion that highlights the chimerical nature of endocrine disruption. At high doses, where most laboratory testing is done, one finds cellular degeneration in the liver and kidneys and an increased incidence of hepatocellular carcinomas in the liver of mice and rats. At lower doses of exposure during prenatal/early-postnatal development, females develop an increased incidence of precocial vaginal opening, hypertrophied uterine and oviductal epithelium, persistent vaginal estrus, decreased implantation rates, and an acceleration in the transport of embryos through the reproductive tract. Exposure of adult males to methoxychlor decreases spermatogenesis, while changes in uterine weight and increases in atretic follicles are common in females. Within six hours of exposure, there is a significant increase in uterine weight indicating the quick responses to methoxychlor. Changes in adult behaviour have also been demonstrated. When pregnant mice are exposed from day 11 to 17 of pregnancy, their male offspring display increased territorial markings when they grow to adulthood. Such alterations occur in a dose- dependent manner and emphasize potential developmental effects during brain development (vom Saal et al., 1997). Methoxychlor has been linked to endometriosis (Cummings and Metcalf, 1995b), a disease in women where uterine endometrial tissue grows outside the uterus. At low doses, methoxychlor sustains endometriosis, as does estrogen. At higher doses, methoxychlor exacerbates the development of endometriosis and can potentially contribute to its recurrence. Endometriosis, once instigated, is maintained by the estrogen-mediated endocrine system, and an agonistic chemical mimic would serve as another stimulus to the maintenance or growth of the tissue. For those women currently suffering from endometriosis, the relative importance of exposure to synthetic estrogen agonists, as compared to other causal factors, has not been determined.

Acute and Chronic Toxicological health have been evaluated using information Effects from controlled laboratory studies with human volunteers, non-human primates, and other Acute Toxicity mammalian species. Overall, these studies indicated that DDT may cause a number of The toxicological effects of DDT on human adverse effects in humans ranging from acute toxicity to cancer (Hayes, 1982). births, absorbed fetuses and lower birth weights. For instance, in California Sea Lions living on Symptoms of acute poisoning include the Channel Islands, one of the world’s most paraesthesia, tremors, confusion, headache, DDT-contaminated areas, early termination of fatigue and vomiting. Dermal exposure does not pregnancy and premature births have been seem to be associated with illness and irritation observed. Premature pups had two to four times (Hayes, 1982). higher concentrations of DDT than full-term DDT is highly toxic to fish and aquatic pups (DeLong et al., 1973). invertebrate species, slightly to partially non- There are indications that long-term exposure to toxic to birds, and moderately to slightly toxic to DDT may reduce sperm counts and lower mammalian species (See Tables II-2 & II-3.). fertility rates, and may cause other reproductive DDT has also been associated with premature problems.

Table II-2: Toxicity of DDT and its Derivatives to Selected Aquatic Organisms (adapted from Ritter et al., 1995)

Species Compound Temperature 96-h LC50 (C) (µg/L)

Estuarine/Marine Organisms Crangon setemspinosa DDT 20 0.40 (shrimp) Mysidopsis DDT 25 0.45 (mysid shrimp) Cyprinodon variegatus DDT 15 2.00 (48-h) (sheephead minnow) Morone saxatillis DDT(77%) 17 0.53 (striped bass) DDE 2.50

Freshwater Daphnia magna DDT 20 1.10 (48-h) (water flea) Palemonetes kadiakensis DDT 21 2.30 (glass shrimp) TDE 0.68 Pteronarccys california DDT 15.5 7.00 (stonefly naiad) TDE 380.00 Salmo gairdneri DDT 20 42.00 (rainbow trout) DDE 12 32.00 TDE 12 70.00 Pimephales promelas DDT 18 13.20 (fathead minnow) Lepomis macrochirus DDT 18 6.30 (bluegill) DDE 17 240.00 TDE 24 42.00 Table II-3: Acute Toxicity of DDT and its Derivatives to Selected Avian Species (adapted from Ritter et al., 1995)

Species Compound/Purity % Age LD50 (mg/kg body weight) Callipepla californica DDT (TG) 6 months 595 (California quail) TDE (>95) >760 Anas platyrhynchos DDT (77.2) 3 months >2240 (mallard duck) TDE (>95) >2000 Phasianus cochicus DDT (>99) 3 months 1334 (pheasant) TDE (>95) 3-4 months 386 in embryo deaths. Researchers found an inverse relationship between eggshell thickness and the Chronic Toxicity p,p’-DDE residue in the eggs of various species The pathways over which DDT is metabolized of birds. While DDT is often accompanied by are similar among rodents and humans. other pollutants such as PCBs, dioxins, and However, in controlled studies where adult mercury, laboratory tests and comparisons humans were given DDT, DDE, or DDD, the among the effects of various pollutants in the DDT to DDD pathway accounts for the vast eggs of 14 species of birds have concluded that majority of the principal urinary metabolite p,p’-DDE showed the greatest correlation with (DDA). This further highlights the fact that the eggshell thinning. Also, investigations of the primary source of DDE detected in humans is mechanism of action of p,p’-DDE-induced that acquired in the diet as DDE. Little, if any, eggshell thinning found little evidence of other DDE is produced from the metabolism of DDT pollutants, in particular PCBs, having similar in humans (ATSDR, 1994). effects (see How DDT Weakens Eggshells, page 19). In adult experimental animals, chronic exposure to DDT has led to effects on the liver, kidney, and nervous and immune systems. Effects on the nervous system include tremors (in rats at doses of 16-32 mg/kg/day over 26 weeks, in mice at doses of 6.5-13 mg/kg/day over 80-140 weeks), changes in cellular chemistry in the central nervous system (monkeys at doses of 10 mg/kg/day over 100 days) and loss of equilibrium (monkeys at doses of 50 mg/kg/day for up to 6 months) (ATSDR, 1994). The toxic effects of DDT on the liver in adult study animals include changes in liver cell physiology, increased liver weight, and increased liver enzyme activity. Kidney effects include adrenal gland hemorrhage. The immunological effects include reduced antibody formation and reduced levels of immune cells (ASTDR, 1994).

Reproductive Effects

Eggshell thinning In North America, widespread declines in predatory and fish-eating bird populations including peregrine falcons, brown pelicans, bald eagles, white eagles, eagle owls, sparrow hawks, gulls, terns, and osprey, became known in 1960s. The cause is attributable to eggshell thinning and the subsequent effect on reproductive success. Thinner eggshells crack easily under normal nesting conditions, resulting ion transport and thereby retard calcium transport (Lundholm, 1994). How DDT Weakens Eggshells Eggshell formation in birds involves the transfer of large amounts of calcium from the blood to the eggshell gland lumen. Concurrently, an equal amount of carbonate ions is required for A catastrophic drop in bald eagle population the shell calcification to occur. Carbonate ions recruitment – from 1.26 young per breeding area are predominantly produced by carbonic in 1966 to as low as 0.46 in 1974 in anhydrase activity in the shell gland mucosa northwestern Ontario – led to the eagle’s and, to a lesser extent, are derived from blood designation as an endangered species in Canada (Skimkiss and Taylor, 1971). Calcium transport and the United States. Reproductive success as from the blood to the shell gland lumen is stimulated by the presence of sodium and measured by chicks per nest increased not long bicarbonate in the gland lumen. During the after Canada and the U.S. banned DDT in the eggshell formation process, the calcium level is early 1970s. By 1981 nest success improved to regulated by a calcium-binding protein, 1.12 young/nest in northwestern Ontario (Grier, calmodulin. Calmodulin and other calcium- 1982). In the U.S., where there were fewer than binding proteins are critical intercellular 500 pairs of bald eagles in the lower 48 states in regulators that act as receptors for calcium ions. 1963, bald eagle population size dramatically Various enzymes can interact with the calcium- rebounded to more than 5,000 pairs by 1996. In calmodulin unit to initiate a biochemical or 1995, the eagle was down-listed from physiological response. The activity of endangered to threatened status, under the U.S. calmodulin and other calcium-dependent 3’,5’cAMP phosphodiesterase was found to be Endangered Species Act. strongly inhibited by DDT during an in vitro However, despite the recovery of bald eagles in assay (Hagmann, 1982). some parts of the U. S. and Canada, there are This finding provided support for the hypothesis areas where bald eagles still experience poor by Lundholm (1987) that an inhibition of reproductive success. A study of the bald eagle calmodulin is involved in DDE-induced eggshell population along the Columbia River estuary, in thinning. This hypothesis correlates with many the states of Washington and Oregon, found that toxic properties of this chemical class since eagles in only 30% of the occupied breeding other organochlorines such as DDT, PCBs, and territories were successful in fledging young. dieldrin can also inhibit calmodulin activities Also, the analysis of egg contents revealed the (Feyk and Giesy, 1998). However, it does not correspond with specificity of eggshell thinning presence of DDE along with other of DDE. The most current hypothesis is that the organochlorines and metals. The concentration mechanism of DDE-induced eggshell thinning of DDE ranged from 4 to 20 ppm wet weight involves an inhibition of prostaglandins (PGs) by (these concentrations are within the range of 15- the shell gland mucosa. PGs synthesis is 20 µg DDE/g, known to cause reproductive stimulated by progesterone, and plays an failure in predatory birds, and well above the 5 important role in the control and regulation of ppm critical level). The examination of eggshell reproduction in birds (Lundholm and Bartonek, thickness showed that on average 10% of the 1992). The synthesis of PG is inhibited by p,p’- eggs had thinner eggshells than pre-DDT. In DDE in duck shell gland mucosa, both in in vitro experiments and following in vivo experiments. some cases, the eggshell thickness was up to But p,p’-DDT, o,p’-DDT, and o,p’-DDE did not 44% thinner than pre-DDT average thickness. inhibit the synthesis of prostaglandins. p,p’-DDE The analysis of prey fish samples from the may induce eggshell thinning by inhibiting PG estuary showed that all had detectable DDE synthesis and thereby reducing carbonate ion concentrations ranging from 0.1 to 0.5 ppm wet secretion to the shell gland lumen. Prostaglandin mass (Anthony et al., 1993). E2 stimulates the transport of carbonate ions from shell gland mucosa to the gland lumen. If North America’s peregrine falcon suffered this is the case, inhibition of prostaglandin similar declines in reproductive success. Heavily synthesis by p,p’-DDE would hamper carbonate contaminated with organochlorine residues, DDT in particular, populations of peregrine falcons in Rankin Inlet, NWT, declined to 35% pelicans, double-crested cormorants, and other of their pre-DDT numbers by the 1970s (Kiff, birds resulted from a reduced number of adult 1988). Like bald eagles, peregrine falcon males (a highly skewed sex ratio: 3.8 females for populations have recovered as a direct result of each male), and female-female pairing (Gress et the DDT ban. However, due to DDT and its al., 1973; Anderson et al., 1975). In a related metabolite’s long half-life and persistence, they laboratory experiment, when gull eggs were continue to affect reproduction. Even in remote injected with DDT at concentrations comparable areas like Rankin Inlet in Northwest Territories, to those found in contaminated seabird eggs, peregrine eggshell fragments collected from 54 abnormal development of ovarian tissue and clutches were 15% thinner (0.306 mm) between oviducts in male embryos was induced (Fry and 1991 and 1994 than the average pre-DDT shell Toone, 1981; Fry et al., 1987). thickness (i.e. 0.360 mm). Also, 28% of all For a short period after conception, embryos samples showed thinning equal to or greater than have the potential to become male or female critical levels associated with reproductive depending on whether a developmental ‘switch’ failure and population decline in this species is thrown or not thrown. Approximately 15 days (i.e., 17% of average pre-DDT shell thickness). after conception for rats and after 56 days in Analysis in 1991 showed that 10% of the humans, a slight pulse in testosterone signals the population had eggs with DDE concentrations male reproductive organs to begin developing exceeding the critical levels (Johnstone et al., and the existing tissues fated for the female to 1996). Peakall et al. (1975) noted that eggs with self-destruct. The “default route” of DDE residues of 15 to 20 g/g would experience µ development – if the developmental switch is reproductive failure. In addition to North not thrown – is for the organism to develop as a America, peregrine falcon eggs collected in female. The testosterone pulse is triggered by 1990 in Zimbabwe had 10% thinner shells the Y-chromosome in mammals. The resulting compared to pre-DDT values (Hartley et al, chorus of endocrine messengers begun by 1995). Other raptors such as hawks, eagles, and testosterone directs the elaboration of the falcons in Zimbabwe experienced reproductive structure (anatomy), shape (morphology), failure as well. In 1981, Thomson predicted that function (physiology) and behaviours necessary many raptor species in Zimbabwe would for the genetically-determined sex. Disruption of become extinct within 15 years unless the use of hormone messengers during this stage of DDT was curtailed (Thomson, 1981, cited in development can cause feminization and Hartley, 1994). demasculinization of males or defeminization Long-term studies of brown pelicans in the Gulf and masculinization of females. This can result of California, Mexico, identified reproductive in functional abnormalities, altered behaviour, problems under conditions of both DDE reduced fertility, and birth defects such as exposure and food stress. Some years, pelicans incomplete urogenital development failed to breed or after mating they deserted (hypospadias) and undescended testicles nests and abandoned eggs and young. DDE (cryptorchidisms). residues in the adipose tissue of breeding The period of gestation to shortly-after-birth has pelicans ranged up to 2050 ppm (Keith and a large number of windows where hormone Mitchell, 1993). signals are critical for proper development. For instance, at various times during this period, the Direct Feminization/Demasculinization differentiation of regions of the brain coincides with the establishment of adult sexual Along with the immediate effects on behaviours and is guided by hormone secretion. reproductive success, i.e., eggshell For example, the Sexually Dimorphic Nucleus thinning/breakage and abortion/premature birth, of the Preoptic Area (SDN-POA) is a region of DDT exposure can result in feminization and the hypothalamus that, in rats, develops in a demasculinization of the offspring. One example span of a few days. The size of the SDN-POA is comes from the southern coast of California two or more times larger in males than females. where the poor breeding success of brown Once a female is born, the size of the SDN-POA the developing testis disperses readily becomes fixed. Perturbations of hormone signals throughout the fetus, enters cells, and is between day 16 to 22 of prenatal development converted to estrogen by aromatase, an enzyme can alter the size of the SDN-POA, the future specific for this purpose. Aromatase is produced frequency of lordosis (mounting behaviour), and in select cells in the hypothalamus, so synthesis luteinizing hormone (LH) concentrations (Rhees of estrogen is extremely localized. But it is this et al., 1997; Rhees et al., 1990a, 1990b; Davis et estrogen that masculinizes the preoptic area and al., 1995). ventromedial nucleus. Thus, masculization can be affected by estrogen or estrogen agonists, In humans, the preoptic area has a window of such as o,p’-DDT or DES (Tarttelin and Gorski, development associated with the LH-releasing 1988; Dohler et al., 1984). In addition, the hormones (Kandel et al., 1995) that is believed estrogen receptor can be blocked by anti- to open near the ending of the first trimester with estrogen compounds such as tamoxifen, which is changes in size evident early in postnatal used in cancer treatments. If tamoxifen or development (Kaplan et al., 1976). another anti-estrogen is present during The SDN-POA is one of the regions of the development, estrogen hormone messages are hypothalamus whose neurons manufacture and blocked, simulating an absence of estrogen and secrete luteinizing-releasing hormone and the presence of the female state (Vancutsem and gonadotropin-releasing hormone. These Roessler, 1997). releasing hormones travel to the pituitary where Since this process is mediated through estrogen- they signal the release of LH or follicle receptor processes, concern is raised that the stimulating hormone (FSH). The nuclei presence of estrogen mimics during these early differentiate in late fetal life and their form stages of development can masculinize and (morphology) is determined by the ratio of defeminize female brain function and behaviour. estrogen to testosterone concentrations in the Laboratory experiments underscore the exquisite fetus during a critical moment during sensitivity of developing embryos to low doses development (Dohler and Jarzab, 1992). of estrogen, testosterone, and chemicals that Another developmental window opened during mimic or otherwise interfere with them. For this same period involves the ventromedial example, masculinization and defeminization of nucleus of the hypothalamus. This region is female mice has been reported when these associated with the expression of sexual fetuses are developing beside two male pups in behaviour. In rats, exposure to estrogen through utero (vom Saal et al., 1992). The source of aromatase activity on testosterone establishes testosterone in this in utero example is the male mating behaviours (Kandel et al., 1995). natural diffusion that occurs across the Female rats exposed to testosterone during this embryonic membranes that enclose each embryo period became more aggressive and physical and is estimated to be parts per trillion in during bouts of play. concentration. Such behavioural differences are the result of a Studies of the population of western gulls very unusual hormone triggering system. nesting in the Channel Islands, off the coast of Estrogen is typically thought of as a feminizing southern California and herring gulls from the hormone and testosterone as masculinizing in its Laurentian Great Lakes in the 1960s and 1970s, effects. This is true regarding the development showed a high incidence of supernormal of the reproductive tract and expression of clutches and female-female pairing (Hunt and secondary sexual characteristics. However, Hunt, 1977; Shugart, 1980). This is an abnormal during development of certain regions of the phenomenon since gulls are long-lived brain, estrogen acts to masculinize. During the monogamous species that usually lay 2 or 3 eggs early stages, free estrogen occurs at very low every year. Further studies revealed that these concentrations as a protective measure to limit supernormal clutches were a result of multiple estrogen-mediated pathways during females sharing the same breeding space. Fry development. However, testosterone secreted by and co-workers studying female-female pairing of western gulls, California gulls, and herring neurotransmitters longer than the others, thereby gulls concluded that the increased number of influencing how signals are passed. The blend of females and decreased population of the gulls the two types establishes the sensitivity of the could be a reflection of the fact that the majority nerve; if the balance is not correct, the nerve of male gulls in 1970s were feminized and may be activated or fired too easily, providing incapable of reproduction and also very few hypersensitivity, or may require more time to eggs in supernormal clutches were fertile. pass messages in the neural tissues. Wingfield et al. (1982) and Hunt et al. (1984) reported that there are no significant hormonal Neural Effects of Pesticides or behavioural differences between females A majority of the pesticides used for vector paired with females and those paired with males. control achieve their results through alterations However, embryonic feminization of males may in the nervous system of pest and non-target result in suppression of sexual behaviour and species. These pesticides include DDT, self-exclusion from the breeding colonies. methoxychlor, the synthetic pyrethroids, carbamates, and organophosphates (Ware, 1994; Effects on the Nervous System Narahashi, 1992; Casida et al., 1983). While these chemicals may cause mortality in the pest Overview of the Nervous System with no visible neural damage to non-target species, other more subtle developmental effects The nervous system functions by the have been recently reported. For example, DDT, transmission of electrical impulses along the bioallethrin, and deltamethrin can shift the nerve cells (neurons) that comprise it. As the proportion of the two receptor types mentioned neurons connect during development, synapses previously, resulting in conditions that can lead or junctions are formed where two neurons to hyperactivity of the nerves. When mice were connect. Neurotransmitting chemicals convey exposed to DDT or deltamethrin on the tenth signals across these junctions. Receptors for the day after birth, permanent changes were seen, neurotransmitters form and become permanently which persisted in the adult four months later attached to the surface of the nerve cell at the (Eriksson, 1992; Eriksson et al., 1992). Table II- synapse. These receptors become the site of 4 also outlines other effects from laboratory attachment of the neurotransmitter messenger studies of [DDT and] deltamethrin. released by the neighboring neuron that is passing its electrical message. When a sufficient Pesticides, including those commonly used in number of receptors in the synapse have vector-control programs, have not been routinely neurotransmitters attached, a threshold is screened for alterations in synaptic receptor reached and the neuron receiving these development. Studies with mice and rats further transmitters begins conveying its own electrical document gross anatomical, morphological, and signal to the next neuron beyond it. functional changes in the brain with exposure in Acetylcholine is a common form of utero. They also show linkages between such neurotransmitting chemical in nerve tissues. changes and other end points, such as behaviour. There are two types of receptors that respond to These effects involve brain structure and as this chemical. One type holds the such, direct effects on humans are difficult to assess. Table II-4: Neurological Effects from Neonates’ Exposure to Low Doses of Pesticides Species Exposure Response Effect Reference

mice DDT Altered proportions of receptor increases Eriksson, neonate 0.5 mg/kg types sensitivity of 1992 Day 10 (15 ppt in brain) Spontaneous behaviour changed nerves age 10 days when adults (habituation) (irreversible)

mice Deltamethrin Altered proportions of receptor increases Eriksson, neonate 0.5 mg/kg types sensitivity of 1992 Day 10 (15 ppt in brain) Spontaneous behaviour changed nerves age 10 days when adults (habituation) (irreversible) rats Deltamethrin Changes in weight and enzyme changes in Husain et adult 7.0 mg/kg activity among regions of brain behaviour al., 1996 for 15 days Increased spontaneous motor activity and aggressive behaviour Decrease in maze learning rats Deltamethrin. Brain & body weight decreased neurogenesis Patro et al., neonates 0.7 mg/kg Neural development delayed and final brain 1997 day 9 - 13 for 5 days including proliferation and anatomy migration altered Reduced blood flow in brain

Nerve cells also establish and pass electrical switches that is responsible for neural impulses by manipulating the balance of two functioning. chemicals – sodium and potassium – inside the Pesticides can block the receptors or destroy the cell. DDT and synthetic pyrethroids interfere enzymes whose job it is to remove with and slow the actions of the cellular pumps neurotransmitters. For example, that change levels of sodium and potassium, organophosphate and carbamate pesticides can thereby slowing the frequency with which destroy the enzyme acetylcholinesterase that is neurons transmit electrical signals. responsible for destroying the neurotransmitting Other pesticides alter additional facets of the chemical acetylcholine. Destroying this enzyme communication process in the nervous system. leads to uncontrolled discharges. When a large When an electrical nerve impulse reaches the number of neurons are so influenced, classic end of a nerve, specialized vesicles release pesticide poisoning effects are experienced neurotransmitter molecules (such as including twitching, tingling sensations, tremors, acetylcholine, noradrenalin, dopamine, convulsions, seizures, and possibly even death. serotonin, or gamma-aminobutyric acid) into the Nervous systems vary within an individual, synaptic junction that connects two neurons. The between individuals of the same species, and neurotransmitter molecules diffuse across the between species. Neurons in regions of the brain gap and bind to specific receptors in the have subtle differences in the numbers of membrane of the next link in the neural receptors in the synapses, the types of pathway. After the neurotransmitters do their neurotransmitters used, and the methods used to jobs, they are chemically destroyed by enzymes. restore synaptic conditions. Neuromuscular This resets the receptor for reception of another junctions, the neural connections that carry wave of neurotransmitters. It is this sequential signals to muscles, differ from the typical passing of electrical discharges by chemical synapses in the brain and thus have different susceptibilities to neurotoxic chemicals. throughout the body so that they can provide the Susceptibility also differs among species and is wide diversity of cell types that comprise the more related to increased complexity and immune system. alternative methods by which an organism can Granulocytes (neutrophils, eosinophils, degrade or sequester nerve poisons. basophils) comprise one class of these highly specialized immunological cells. They are Effects on the Immune System produced in the bone marrow and released during inflammation. Neutrophils and Overview of the Immune System eosinophils actually engulf, in a process called phagocytosis, foreign material and debris which The immune system provides the primary are then digested inside the cell using defense against invasions by foreign materials specialized enzymes. Basophils also release such as microbes and parasites. It also protects histamines that increase inflammation at the site the body from aberrant or damaged cells, such as of the infection; this acts as a beacon to call in tumors. The immune system is a complex more granulocytes to the site. assemblage of cells and organs (spleen, lymph nodes, thymus, bone marrow) that have highly Lymphocytes are a second major subdivision of specific and, to a degree, overlapping functions. the immune system, and are composed of B- and No single type of cell is sufficient, however, to T-lymphocytes. There are approximately two provide all of the defenses necessary for the trillion B-lymphocytes in the body. Their wide variety of possible assaults. The immune progenitor cells are located in bone marrow and system is tied to the endocrine and nervous are the source of all B-lymphocytes an systems, both in its development during individual will ever have. Each B-lymphocyte is gestation and early postnatal life, as well as capable of manufacturing a unique and highly during its operation throughout life. Many of specific antibody that is used to bind to antigens these interconnections are only now being (pathogens and other foreign matter) found elaborated. outside cells. One B-lymphocyte can release as many as 3,000 antibodies per second. These An abundance of literature describes immune antibodies drift in body fluids and when they suppression from exposure to pesticides meet the antigen, they bind and facilitate its (reviewed by Repetto and Baliga, 1996). Much rapid ingestion by the white blood cells of the attention has been focused on agricultural (phagocytes). Since the antibodies passively applications of organochlorines, drift throughout the body, it is necessary to organophosphates, carbamates, and produce them in vast quantities. Millions of thiocarbamates and only recently have pesticides lymphocytes producing antibodies may seem commonly used in vector-control programs been like a formidable defense, however, because studied. Technical grade DDT and the synthetic each B-lymphocyte remains dedicated to pyrethroids cypermethrin, deltamethrin, and producing an antibody specific to a single permethrin, have effects on the immune systems antigenic particle, a large number of such cells is of mice, rats, rabbits and goats (Table II-5). required in order to maintain an adequate repertoire or “library” to fight a broad range of infections. Immune System Development T-lymphocytes are another critical component of The different cell types that make up the the immune system and arise from the thymus immune system arise from primordial stem cells gland. During the 11th day of development in the that form early in development. These cells mouse or the 8th week in the human, stem cells differentiate into a variety of more specialized in the bone marrow migrate to the thymus where cells that ultimately populate the bone marrow they differentiate into progenitor cells that give and thymus. The successful development of the rise to mature T-lymphocytes. These cells immune system requires that the primordial cells migrate from the thymus early in life and multiply, migrate, and become established circulate in blood, acting as coordinators of immune responses (T-helper lymphocytes) or they acquire a limited array of “non-self” killers of virus-infected cells (T-cytotoxic recognition abilities and thus when a foreign lymphocytes and Natural Killer [NK] cells). molecule enters the system only a small Since the thymus becomes smaller as an component of the total complement of individual matures, the thymus of a mature lymphocytes can identify it as foreign and individual can be removed with no visible respond. detrimental effects. However, if the thymus is Immunization programs utilize this recognition removed or disturbed during development, this ability by introducing benign or “killed” parts of line of defense can be adversely affected. the pathogen to “teach” the immune system how Thymus weight, an indicator of the health of this to recognize a specific pathogen. The organ, decreases when exposed to DDT or appropriate B-lymphocytes locate and recognize cypermethrin during gestation or early these as a foreign threat, which triggers the rapid development (Santoni et al., 1997; Queiroz, cellular multiplication of these specific 1993). lymphocytes. When immunity is acquired in this In contrast to B-lymphocytes, which work fashion, a large population of primed B- outside cells, T-lymphocytes specialize in lymphocytes with a ‘memory’ are the sentinels clearing an infection that is already established that offer the first critical response in an within the cells of the body. T-helper cells eventual exposure situation. provide an important function in coordinating The response component of the immune system the various components of the immune system is the defense against invasions of pathogens and through secretions of cytokines, such as includes many cell types that are specialists in interleukins, interferons, and colony-stimulating defense against specific kinds of foreign bodies factors. The cytokines are chemical messengers like bacteria, viruses, and parasites in body through which the lymphocytes coordinate the fluids; target cells that have been invaded by activity of other cells in the immune system. T- pathogens; and debris associated with infections helpers also destroy cells by perforating the cell and wounds. Hormonal response involves the surface. NK cells play a critical role in production of antibodies which come in contact destroying tumor cells and for this reason the with the antigens, bind, and cause them to immune system plays a critical role in the precipitate, dissolve, or to stick together forming prevention of spontaneous cell proliferations that agglutinated masses. Then phagocytic cells may lead to cancer. destroy them. “Cell-mediated” response relies on the activity Pathogen Recognition and Response of specific cell types to aid in locating antigens, binding to them, and ingesting viruses and The immune process involves two basic bacteria or encapsulating parasitic organisms. activities: recognition of foreign material and Furthermore, some of the cell types in the response to its presence. Recognition hinges on immune system serve coordinating functions as lymphocytes’ ability to detect subtle differences they move to sites of infection and release between “self cells” and foreign material. homing chemicals that call in masses of white Furthermore, differences among foreign blood cells. materials can be identified and highly specific responses can be directed against them. For A lymphocyte has its own small repertoire of example, lymphocytes can recognize as antigens it can recognize. An effective immune “different” a long protein in which just a single system relies on a very large population of these amino acid has changed. This identification is cells to increase the likelihood of recognition. incorporated into lymphocyte ‘memory’ from When a particular lymphocyte finds an antigen which future invasions can be quickly identified invader, it springs into intense activity since it and even quicker future responses provided. Not must be responsible for producing both the every B-lymphocyte can respond in this fashion chemical defense as well as more lymphocytes to each specific foreign molecule. When created, to help with the task. That translates into a few specific cells giving rise to a lineage of Pesticides have been demonstrated to lead to thousands of clones designed for this one immune suppression (Repetto and Baliga, 1996). purpose. It can take about five days for the cells Outcomes include detectable changes in to undergo these divisions. Disease resistance antibody production, lymphocyte proliferation, depends on the ability of the body to both mount phagocytosis rates and white blood cell counts, an adequate defense relying on the two trillion as well as increases in the time it takes to B-lymphocytes and to quickly mobilize a small respond to infections. Much of the attention has subset of appropriate cells to combat a particular focused on organophosphates, carbamates, and infectious assault. The rates of cell divisions are organochlorine pesticides. A pattern of effects key to quickly stemming the assault especially if has been documented for DDT and such the pathogen itself is dividing during its attack. synthetic pyrethroids as cypermethrin, deltamethrin, and permethrin (Table II-5). The specific immunological effects include changes to: 1) lymphoproliferation (production of lymphocytes), 2) the humoral response rate, 3) Effects of Pesticides on the Immune thymus weight, and 4) overall system System performance.

Table II-5: Reports of Effects of DDT, Cypermethrin, Deltamethrin and Permethrin on the Immune System of Mice, Rats, Rabbits, and Goats Species Exposure Response Effect Reference mice DDT Humoral response not altered h Banerjee et al., 20, 50, 100 ppm (antibody titer and plaque- 1997 4 weeks forming cells) Humoral response decreased when animal stressed (handling or temperature) mice DDT Humoral response decreased c.m. Banerjee et al., 20, 50, 100 ppm Plaque-forming cells decreased 1986 3-12 weeks rats DDT/DDE/DDD Humoral response suppressed h Banerjee et al., 200 ppm 1996 6 weeks Cell-mediated response c.m. suppressed mice DDT Humoral response decreased h Banerjee, 1987a 20, 50, 100 ppm 3-12 weeks Plaque-forming cells decreased c.m. rats DDT Humoral response decreased h Banerjee, 1986 20, 50, 100 ppm 8-22 weeks Lymphocyte mobility altered c.m. rats DDT Humoral response decreased in h Banerjee, 1987b 20, 50, 100 ppm highest dose 8-22 weeks Immunoglobulin concentration decreased with tetanus toxoid challenge Decline in migration inhibition factors c.m. rats DDT At 50 or 100 ppm a 3% protein h Banerjee et al., Species Exposure Response Effect Reference

20, 50, 100 ppm diet leads to suppression of 1995 4 weeks humoral and cell-mediated c.m. responses mice DDT Lymphoproliferative response to m Rehana and 0.0316, 0.316, 3.16 LPS decreased at highest doses Rao, 1992 mg/kg/day T-Cell plaque-forming decreased for 6 months at highest dose. c.m. mice DDT Lymphoproliferative response m Rehana and (offspring 0.0316, 0.316, 3.16 decreased which continued over Rao, 1992 of exposed mg/kg/day time mothers) for 6 months Plaque-forming cells decreased c.m. in two highest mice Cypermethrin Lymphoproliferative response m Stelzer and 1x10-5 M to 5x10-5 M decreased in both T- and B- Dordon, 1984 lymphocytes mice Cypermethrin Tuberculin Skin Test decreased c.m. Tamang et al., 50 mg/kg/day response 1988 for 26 days rats Cypermethrin Dose dependent decrease in h Varshneya et al., 5, 10, 20, 40 mg/kg/day delayed hypersensitivity 1992 for 90 days Lower leukocyte count in highest dose Depression in cellular response rats Cypermethrin No effects seen _ Madsen et al., 4, 8, 12 mg/kg/day 1996 for 28 days rats Cypermethrin Peripheral NK-cells increased c.m. Santoni et al., (prenatal 50 mg/kg/day Antibody-dependent cytotoxicity h 1997 exposure) for 10 days over Day 7 to increased Day 16 of gestation Lymphocytes: more in plasma, fewer in thymus gland rats Cypermethrin Two highest doses: decreased Desi et al., 1986 1/40, 1/20, 1/10 LD50 Complement Binding and hl Agglutination Test Plaque-formation in spleen cells c.m. rabbits Cypermethrin Dose dependent reduction in: Desi et al., 1986 1/40, 1/20, 1/10 LD50 Agglutination test h Complement Binding h Tuberculin Skin Test c.m. goats Cypermethrin Tuberculin Skin Test decreased c.m. Tamang et al., 41.6 mg/kg/day response 1988 for 30 days Plaque-forming response h decreased mice Permethrin Lymphoproliferative response m Stelzer and 1x10-5 M to 5x10-5 M decreased in both T- and B- Gordon, 1984 lymphocytes mice Permethrin Lymphoproliferative response Blaylock et al., 0.4, 0.04, 0.004 mg/kg not altered 1995 for 10 days Mixed lymphocyte response (1%, 0.1%, 0.01% LD50) reduced Lysis activity decreased c.m. Species Exposure Response Effect Reference

NK cell activity to tumor cells c.m. decreased c.m. mice Deltamethrin Significant decreases: Lukowicz- 6 mg/kg for 84 days Agglutination Test h Ratajczak and or Hemagglutinin Test h Krechniak, 1992 15 mg/kg for 14 days Plaque-forming response c.m. mice Deltamethrin Bone marrow stem cells colony Queiroz, 1993 5 mg/kg for 3 days formation increased Thymus weight decreased rats Deltamethrin Highest doses: significant c.m. Madsen et al., 1, 5, 10 mg/kg/day enhancements of 1996 for 28 days SRBC-Plaque-forming cells and NK-cell activity Significant increase in lymph node weight; reduction in thymus weight h = humoral; c.m. = cell mediated; m = mitogen

Effect on Lymphocyte Function groups, one receiving DDT and the other unexposed. All these offspring showed DDT, cypermethrin, and permethrin decrease the decreased lymphoproliferation. Furthermore, by division rates of T- and B-lymphocytes in 12 weeks of age the rate continued to decline laboratory cultures, suggesting a functional (Rehana and Rao, 1992). impairment of these cells (Rehana and Rao, 1992; Stelzer and Gordon, 1984; Blaylock et al., Effect on Humoral Response 1995). Additional effects were identified from exposure of these lymphocytes to pesticides. For There have been few studies involving exposure example, the rates at which foreign bodies were to DDT and effects on the immune system. dissolved decreased when T-lymphocytes were Work on adult mice and rats (Banerjee et al., exposed to permethrin (Blaylock et al., 1995). It 1986; Banerjee, 1987) indicates that exposure to is not known whether a decreased DDT leads to the suppression of primary and lymphoproliferative rate was sufficient to secondary humoral responses. When mice were account for this decrease or if additional effects exposed to 100 ppm there were significant of permethrin resulted in this decline. Likewise, alterations in antibody titers and the numbers of NK cells also showed decreased cytolytic plaque-forming cells. This response was affected activity (Blaylock et al., 1995). It is not known by the duration of the exposure, thus suggesting if this effect was due to a suppressed rate of a threshold level of exposure is necessary for early mitotic divisions or is a manifestation of a these observed effects. Banerjee (1987) functional impairment of these or other cells of hypothesized that the reductions in humoral the immune system. response are mediated through effects on B- lymphocytes. Mice exposed to DDT in utero and during lactation had a decrease in the rate of Rats also showed reductions in humoral lymphoproliferation (as high as 37%), regardless responses when exposed to DDT (Banerjee et of the pesticide concentrations administered in al., 1986). Exposure to 100 ppm for 18-22 the study (Rehana and Rao, 1992). These effects weeks or to 50 ppm for 22 weeks led to a are transgenerational. Female mice were significant decrease in humoral response. This exposed over six months and mated to control observation coincides with the effects seen in males. The offspring were divided into two mice where duration of exposure suggests a threshold level for effects. Banerjee further decreased numbers of T-lymphocytes in the noted that rats exposed to either 50 or 100 ppm gland. There was an increased lymphocyte count and later immunized with a tetanus toxoid had a in the plasma, which may reflect an accelerated decrease in mobility of the lymphocytes, release of lymphocytes from the thymus. What possibly further reducing the immune defenses. effect this may have had on the performance or Additional work is necessary to test the fate of the released lymphocytes is not known significance of the threshold effect seen in the since such tests were not conducted. Banerjee studies and to determine if lower dose exposures over longer periods of time will Effect on Overall Immune System provide comparable reductions in humoral Performance responses. Furthermore, the ramifications of a change in mobility in the B-lymphocytes and the Observing individual components of the immune degree that this may contribute to potential system may provide benchmarks of effects. health effects are unknown. However, it becomes difficult to predict if these are sufficient to lead to undesirable health Cypermethrin and deltamethrin have been outcomes. The Tuberculin Test can be useful in reported to lead to a decreased ability to bind assessing these overall outcomes. In this test, an antigens. This may be related to a decreased organism such as mycobacterium is inoculated amount of antibody production (Desi et al., with an antigen in a sufficient quantity to insure 1986; Lukowicz-Ratajczak and Krechniak, sensitization of all components of the immune 1992). Desi et al. (1986) describe reductions in system. Memory cells are created and distributed humoral response in rabbits dosed with 1/40, in the body and antibodies are primed for 1/20, or 1/10 proportion of the LD50 value for production. When a small amount of this antigen cypermethrin, which they determined to be is placed in the skin of the animal, the immune 3,000 mg/kg. The response was observed within system springs into activity and all facets of this one week of the initial exposure and was dose response contribute to an observable and dependent, with the two higher concentrations measurable swelling and inflammation at the site demonstrating significant reductions in antibody of injection. The resulting size and amount of responses. Lukowicz-Ratajczak and Krechniak swelling, the color of the skin, and duration of (1992) observed similar results when they the reaction are all indicators of the health of the exposed mice to a concentration of deltamethrin immune system. In one experiment for example, of either 6 mg/kg or 15 mg/kg. Furthermore, cypermethrin exposure prior to sensitization plaque-forming cells were also reduced in increased the reaction time and decreased the number, which is another indication of a intensity of the immune response (Tamang et al., diminished humoral response. 1988). This study did not, however, identify what component of the system was diminished. Effects on Thymus Weight As noted previously, perturbations early in Significance of Immune Suppression from development can have graver consequences for Pesticide Exposure lymphocyte populations than later in life. Since Determining the significance of immune the T-lymphocytes are distributed throughout the suppression from exposure to pesticides used in body after leaving the thymus, this organ can be vector-control programs is not a simple exposure removed in adults with minimal effect to the problem. The carefully controlled experiments immune system performance. However, if the where mice or rats are dosed in precise amounts excision or perturbation occurs prior to the provide only a glimpse of the potential scope of migration, significant effects are observable the problem. For example, Banerjee and upon pathogen challenges, thus making the colleagues demonstrated that pesticide exposure effects of pesticides during this stage of great in conjunction with environmental stressors can importance. Santoni et al. (1997) reported that increase the magnitude of observed effects. rats prenatally exposed to cypermethrin during More specifically, diets deficient in protein days 7 to 16 had a reduced thymus weight and contributed to a reduction in immune responsiveness in rats previously exposed to being activated and primed. For instance, these pesticides (Banerjee et al., 1995). Rats that had may be the kinds of effects observed by Rehana been exposed to 50 or 100 ppm DDT for 4 and Rao (1992) and identified as permanent weeks and then maintained on a low protein diet reductions in immune competency. (3%) had a depressed humoral and cell-mediated Coincident with immune suppression in general response when challenged with an antigen. Rats are increased frequencies of infection and cancer consuming more protein-rich diets (12-20%) did (Descotes et al., 1995). For example, not demonstrate this immunosuppressive effect. Kaskhedikar and colleagues (1996) exposed Banerjee et al. (1997) also demonstrated adult mice to graded doses (0.0078, 0.03125, reductions in humoral response associated with 0.125, or 0.5 ml/kg) of malathion in their food stress (immune system performance is known to over the course of five, ten, or fifteen be tightly intertwined with endocrine and consecutive days, and then infected them with nervous system function); mice exposed to 20, 500 viable eggs of a nematode. Twenty-one days 50, or 100 ppm DDT for 4 weeks did not show a later, the intestines were examined for live reduction in the number or response of plaque- nematodes. forming cells when challenged with sheep red The larger the dose of malathion and the longer blood cells (SRBC). When these animals were its duration, the larger the number of surviving handled or held, however, they showed dose- worms (Table II-6), signaling a weakened dependent declines. This raises concerns that immune system response. The researchers did these subtle shifts may lead to adverse health not extend their study to determine possible effects especially to sensitive components of the ramifications of higher parasite loadings on population, such as the very young, developing these mice. However, it is reasonable to suspect individuals, or the elderly. that the stress, lost energy, and physiological Banerjee et al. (1996) also observed that DDE imbalance caused by both the parasites and the and DDD surpass DDT in their ability to damage they may do to the intestinal lining are a suppress the humoral response in rats. DDE detrimental health effect and may affect the accumulates rapidly and readily in the body, and animal’s survival and reproductive success. This is passed freely in breast milk to nursing also represents a model for pathogens and thus offspring. It is not known what exposures during host resistance to other infectious agents may be fetal and postnatal periods of life can do during diminished as well. the critical period when the immune system is

Table II-6 Parasites Reported in Mice Exposed to Malathion and Subsequently Infected with Known Numbers of Parasites (Kaskhedikar et al., 1996) Duration of Exposure 5 5 10 10 15 15 to Malathion (Days) Dose of Malathion ml/kg body weight WB PWB WB PWB WB PWB 0.0078125 70 14.0 91* 18.2 102* 20.4 0.03125 85* 19.0 108* 21.6 122** 24.4 0.125 102* 20.4 130** 26.0 148** 29.6 0.500 122* 24.4 142** 28.4 160** 32.0

WB= Worm Burden; PWB = percent of worm burden based on the 500 initially infected; * = statistically significant difference; **=highly significant result. DDT and Cancer The U.S. EPA has identified DDT as a probable human carcinogen (ATSDR, 1994a) based on and fat content. These factors account for about lab studies; liver and lung tumors found in mice 30% of breast cancers, leaving the remaining and rats were a result of chronic exposures 70% of the breast cancers unaccounted for. (Kashyap et al., 1977; Terracini et al., 1973; Environmental factors, including synthetic Thorpe and Walker, 1973; Tomatis et al., 1974; chemicals, are thought to be involved, in part, in Rossi et al., 1977). Human epidemiological data the unaccounted breast cancers. Proponents of do not provide clear evidence of the this hypothesis see further support in a report by carcinogenicity of DDT to humans (Houghton the U.S. National Cancer Institute’s and Ritter, 1995). Surveillance, Epidemiology and End Results On the whole, human studies have been Program that described newly diagnosed breast exceedingly rare, being restricted to exposure cancer rates increasing 1% per year between during the manufacture or application of DDT, 1950 and 1979, and 3% between 1980 and 1984. with reports that cancer incidences were not When analyzed by the kind of tumor based on significantly different from control groups. Part the receptor content, estrogen-receptor-negative of the difficulty associated with these studies tumors rose 22-27% between the mid-1970s and relates to the determination of exposure history, mid-1980s, while the number of estrogen- effects of mixtures, and sample sizes. receptor-positive tumors increased an average of Furthermore, the latency period for the 131% (Glass and Hoover, 1990). development of cancer is estimated to be from Pujol and colleagues (Pujol et al., 1994) 10 to 20 years, making direct causal links analyzed 11,195 samples of breast cancer tumors difficult to establish (Dewailly, Ayotte, and collected between 1973 and 1992 and found that Dodin, 1997). This is particularly important if the quantity of estrogen receptors has steadily causal agents such as DDT act in such a way as increased to levels about four times higher over to predispose susceptibility during very early life the 20 year period. They suspect that this change and effects are expressed long after the agent is a reflection of hormonal events that influence disappears from the tissue. breast cancer genesis and growth, and not differences in techniques or screening programs. DDT and Breast Cancer Later age at first pregnancy, early age at There has been considerable recent discussion of menarche, and nulliparity have also been a possible link between DDT and the occurrence associated with estrogen-receptor-positive of breast cancer, although there remains tumors. However, these authors hypothesize that considerable scientific uncertainty and debate the increase in the numbers of estrogen receptor- (Houghton and Ritter, 1995). The case for positive tumors could be associated with concern that DDT is associated with increased induction by estrogen or estrogen-agonists, cancer in humans involves a series of leading to the development of estrogen receptor- observations relating to: 1) known causes of positive cancers (Pujol et al., 1994). breast cancers; 2) changes in frequency of In another study that measured organochlorine cancer in the population; 3) shifts in types of concentration in relation to the estrogen-receptor breast cancers seen; and, 4) patterns of chemical status of the tumor, it was found that estrogen- contaminant levels in tumor tissue. receptor-positive tumors had significantly higher Breast cancer due to mutations of genes only concentrations of DDE than did the controls accounts for about 10% of breast cancers in the (Dewailly et al., 1994). This highlights the need United States (Pollner, 1993), leaving 90% of to view cancers not as a single endpoint but as a the cases for non-genetic causes. class of outcomes involving different cells and Epidemiological studies have tried to identify tissues, promoters, and developmental pathways. other known risk factors for breast cancer and When estrogen receptor-positive breast cancer have identified a wide range of associations: tumors become associated with elevated early menarche, nulliparity, late age of first concentrations of chemicals such as DDT or birth, onset of menopause, oral contraception, DDE, it is reasonable to suggest a causal link estrogen replacement therapy, ionizing radiation, (Pujol et al., 1994). Furthermore, this linkage extends to endocrine-disrupting chemicals since Summary hormonal agonists or antagonists have the Concerns over DDT’s carcinogenicity, potential to mimic the behaviour of natural bioaccumulation, persistence, hazards to wildlife hormones that may be involved in pathways that and other chronic effects have led to its ban in lead to cancer. DDT is a likely candidate for 49 countries and restriction in 29 more. The consideration as a causal agent for breast cancer weight of scientific evidence about its negative since one isomer is known to be an estrogen health and environmental effects continues to agonist (o,p’-DDT) and can therefore act as a mount. Many of these effects, particularly cancer promoter. At present, there are no data to endocrine disruption, were poorly studied when support these concerns. the WHO Study Group on Vector Control for When comparing women who developed breast Malaria and other mosquito-borne diseases made cancer with controls, Wolfe and associates found its recommendations in 1993 to continue the use higher concentrations of DDT and DDE in of DDT. At that time, there were insufficient women who had developed breast cancer, toxicological data to suggest that indoor though only DDE concentrations were spraying might be harmful to humans. This is no statistically significant. More recently, Hunter et longer the case, as this section has shown and al. (1997) report that with more careful the following one further illustrates. screening of participants and an increase in the In its 1993 decision, the Study Group did not sample size, no association was detected with take the well-documented effects of DDT use on DDE and breast cancer incidence. Such studies wildlife and conservation into consideration. involving associations of breast cancer and Today those effects are better understood and serum pesticide levels rely on measurement in more far-reaching than previously known. Given adulthood, but since breast cancer is a disease that many biological processes are conserved with a long initial period of development, the across species, DDT’s known and suspected role of a chemical such as DDT could be effects on wildlife and laboratory animals should obscured. Plasma concentrations can vary, be considered as probable effects on humans. especially when women are breast feeding. Moreover, the possible role of these chemicals Concerns for the subtle endocrine disrupting in diseases like breast cancer has not been and/or developmental effects are not limited to assessed for developmental effects of exposures DDT and its metabolites. Recent research in utero, lactational exposures during early highlights the fact that biologically active development, and exposures later in life. These chemicals can have unanticipated effects, some may well promote such diseases, but when of which can lead to an irreversible cascade of associations are sought, current concentrations events that alters the future functioning of are not representative of exposure history. This tissues and organs. The few synthetic is not meant to argue that DDT does or does not pyrethroids studied demonstrate such effects to cause cancer, only that the designs of neural development, neural performance, and experiments published to date do not target or expressed behaviour, as well as forms of consider developmental consequences in their immune suppression. Although these pesticides assessments of association. If synthetic may lack the persistence and ability to chemicals that mimic, block, or otherwise bioaccumulate seen in DDT, chronic, low dose influence hormones play a role in the occurrence exposures may lead to subtle developmental of breast cancer in the same “trans-generational” effects. A more rigorous research program is manner as they influence other health endpoints, necessary to identify such effects and to then studies of adult concentrations can provide determine their significance. only limited insight. To truly establish whether there is a link between DDT (and other synthetic chemicals) and the occurrence of breast cancer, it is essential to determine if breast cancer victims were exposed to such chemicals in the womb or during early childhood. III. EXPOSURE AND ITS (Torres-Arreola et al., 1998). IMPLICATIONS The concentrations of DDT in breast milk in Table I-5 illustrate the variability of exposure, Levels of DDT in Humans long-distance transport, and long half-life. There are reports that low-birth-weight babies and It is reported that worldwide levels of DDT and premature babies had higher levels of DDE in its degradation products have been slowly their blood compared to normal-weight and full- declining over the past 25 years as countries term babies (O’Leary et al., 1970). Also, higher have banned its use. In humans and animals, levels of DDT and its metabolites were found in DDT levels have declined from a global average the breast milk of women who had the of 12 ppm to below 7 ppm (IEM on POPs, premature babies (Berkowitz et al., 1996). Annex II). These levels vary widely however, Evidence has also shown that elevated depending on the location and various concentrations of DDE are associated with characteristics of sub-populations. For instance, reduced lactation by human mothers (Gladen the concentrations of p,p’-DDE in breast milk in and Rogan, 1995). women in Zimbabwe are 25 times higher than American women (Table I-5). Even within Even after dramatic reductions in DDT use regional populations concentrations vary world-wide, breast feeding women in the most dramatically. Sampling of breast milk in remote locales today are unintentionally feeding Veracruz, Mexico in 1994 and 1995 showed their infants concentrations of a known concentrations ranging from 0.99 to 26.9 mg per endocrine disruptor at a critical stage of their kg of fat. Infants drinking this milk are ingesting offspring’s development (Tables III-1 & III-2). from 5.5 to 150.6 mg/kg of body weight per day DDT levels in the milk of Inuit women in the (Waliszewski et al., 1996). (The Total Daily Arctic are five-fold higher than those of women Intake [TDI] guideline for DDT is 20 mg/kg of living in southern Canada, reflecting their body weight.) Indeed, research in Mexico and greater consumption of traditional foods which elsewhere has revealed measured concentrations are high on the food chain. A 5-kg Inuit infant of DDE in humans that exceed health consuming 750 ml of milk per day with 3% fat authorities’ guidelines for acceptable exposure would take in approximately 5 ppt per day.

Table III-1: Concentrations of DDE in Human Milk Fat in non-Aboriginal and Aboriginal Populations [adapted from Jensen, J., K. Adare, and R. Shearer (eds.) Canadian Arctic Contaminants Assessment Report. (Ottawa, Ontario, Canada: Department of Indian Affairs and Northern Development, 1997)] Location Year Mean concentration (ppt lipid) Southern Canada 1992 222 Southern Quebec 1989/1990 340 Lower North Shore Quebec 1991 823 Nunavik, Northern Quebec (Inuit) 1989/1990 1212 Greenland (Inuit) 1993 3844 Table III-2: DDE Concentrations in Newborn Cord Blood Samples in Canada (sampled in 1993-1995) [adapted from Jensen, J., K. Adare, and R. Shearer (eds.) Canadian Arctic Contaminants Assessment Report. (Ottawa, Ontario, Canada: Department of Indian Affairs and Northern Development, 1997)] Population p,p’-DDE Concentration p,p’-DDT Concentration (ppm) (ppm) Dene/Metis NWT* 0.32 0.02 Non-Aboriginals NWT 0.63 0.04 Southern Quebec 0.66 0.03 Nunavik, Northern Quebec (Inuit) 1.28 0.06 NWT (Inuit) 0.47 0.03 *Northwest Territories

DDT and Reduction in Lactation agonism by the estrogen mimicking isomer, o,p’-DDT may be the reason for the shortening Endocrine disruption may have an indirect effect of duration, primarily because estrogen is known on the developing infant by reducing the amount to reduce milk volume. Humans are not the only of breast milk it can obtain. It is known that species that breast feed their young – all concentrations of DDE in the body of a lactating mammals do – and the biological mechanisms mother can shorten the duration of lactation for bioaccumulation of DDT and passing it (Gladen and Rogan, 1995; Rogan et al., 1987). along to infants are the same. This relationship was first noted in women from the general population in North Carolina. Women with concentrations of DDE in breast Levels of DDT and Effects in Other milk ranging from 0.31 to 2 ppm had a median species duration of lactation of 26 weeks while those with higher concentrations showed a decrease in Eggshell thinning duration as DDE increased. Those with 5 to 23 ppm had median values ranging from 9 to 10 DDE has been identified as the principal weeks (Rogan et al., 1987). Gladen and Rogan pollutant causing eggshell thinning and (1995) extended this study to Tlahualido, reproductive failure in predatory birds. Since the Mexico where further evidence suggests that banning of DDT in the early 1970s, many of the DDE shortens the duration of lactation. In this bird populations facing extinction in the 1960s study, the duration of lactation ranged from 7.5 and 1970s have recovered. However, there are months for women with 0 to 2.5 ppm to a instances where DDE levels in eggs and prey median duration of 3.0 months for those with species around the world are still high enough to 12.5 ppm or greater. They proposed that cause reproductive failure in recent times (Table III-3).

Table III-3: Eggshell Thinning in Predatory Birds Species/Location Concentrations Endpoints Critical Reference in eggs concentrations Bald DDE: 4-20 ppm 10% of population had Egg: 5 ppm DDE Anthony et Eagles/Columbia thinner eggshell Egg: 15-20 ppm al., 1993 River Estuary (1980- Only 30% of the nests DDE 1987), USA had fledging young reproductive failure Peregrine Falcons/ DDE: 4.5 (0.8-28) 15% thinner eggshell Eggshell critical Johnstone Rankin Inlet, NWT ppm w.w. 28% of the sample thickness: 17% of et al., 1996 (1991-1994), had eggshell thinner average pre-DDT Canada than critical level shell thickness Species/Location Concentrations Endpoints Critical Reference in eggs concentrations Peregrine Falcons/ DDE: 7.6 (1.8- 16% thinner eggshells Court et al., Keewatin District/ 29.3) ppm w.w. eggshells with 29% 1990 Rankin Inlet (1981- thinner than pre-DDT 1986), Canada average did not hatch Peregrine Falcons DDE: 2-5 ppm 11% thinner eggshell Henny et al., (1991), Russia w.w. 1994 African Goshawk ΣDDT: 18-326 18% and 22% thinner Critical level: 130 Hartley and (1988-1991), ppm d.w. eggshell 35% to 45% ppm d.w. Douthwaite, Zimbabwe population decline 1994

DDT continues to be found in certain wildlife in the Northwest Territories still suffer from and human tissues throughout the world at eggshell thinning. And, based on residue levels, concentrations able to cause population-level it is probable that reproduction in many other effects. The bald eagle population in the populations of fish-eating birds and birds of prey Columbia River Estuary and Peregrine falcons are also affected.

Table III-4: Action and Advisory Levels for DDT and Metabolites

Agency DDT and metabolites criteria Great Lakes Water Quality Agreement Objectives (whole fish) 1 ppm wet weight Health Canada, Tolerable Daily Intake (TDI) 20 ppb/day FAO/WHO, Tolerable Daily Intake 20 ppb/day WHO, Drinking water guideline 1 ppb WHO DDT Guideline, milk (in fat) 1 ppm US Food and Drug Administration Action Level for fish (wet weight) 5 ppm Health Canada, maximum allowable concentration Fish 5 ppm Eggs and fresh vegetables 0.5 ppm Dairy products, meat and meat by-products 1 ppm Drinking water 1 ppm Michigan Department of Public Health, fish consumption advisories 5 ppm US EPA Minimal Risk Level (MRL) 0.5 ppt/day US EPA recommended action level: Most fruit and vegetables 0.1- 0.5 ppm Eggs 0.5 ppm Grain 0.5 ppm Milk 0.05 ppm Meat 5 ppm

Routes of Exposure absorbed by inhalation, direct contact with skin Overview of pathways (dermal exposure), or oral ingestion For DDT, oral exposure through the ingestion of during pesticide preparation (mixing) or contaminated foods is considered to be the most application (IEM on POPs, Annex II). Once important exposure route (ATSDR, 1993). sprayed, DDT does not disappear or degrade However, these substances may also be into harmless byproducts. A stable and persistent substance, it can easily move on soil or dust particles and through waterways to end up in “mass balance” model to explore this issue aquatic and terrestrial ecosystems both near and further. It provides an accounting of the fate of far. DDT then moves up the food web and DDT and other pesticides used for indoor house because of its lipophilic nature, bioaccumulates spraying (Feltmate et al., 1998). The model uses at high concentrations in the fats of fish, birds, the concept of fugacity - the tendency of and animals, including humans. A certain level chemicals to move from one or more of DDT due to historical uses continues to cycle “compartments” of the environment to others - through ecosystems. It is appropriate to inquire to estimate how the pesticide moves, over what if vector-control uses such as indoor spraying period of time, and where the pesticide will end add to these levels. up. The key parameters used in the model are:

Levels From House Spraying – A model  the basic physical and chemical and assessment of the fate and exposure of properties of the pesticide; DDT  the physical characteristics of the room DDT is sprayed on the walls of homes and and its contents; other buildings as a control measure mainly against Anopheline mosquitoes for control of  the affinities of the pesticide for different malaria, and sand flies (Phlebotomus) for control components of the environment, for of leishmaniasis. The use of DDT for indoor instance with DDT, its affinity for fat house spraying has generally been assumed to be versus air or water; a minor source of exposure to residents. This may have been the situation when DDT was in  the rates of phase transfer, e.g., widespread and voluminous use in agriculture in degradation and vapourization; and, the 1970s and before, and also a prevalent contaminant in the food web. For instance, the  the behaviour of the people in the room, 1993 review for WHO of anti-malaria tools e.g., inhalation and food consumption considered pre-1977 data showing comparable rates. DDT residues in the fat of residents whose houses were regularly sprayed with DDT and in The objective of the model is to provide a the general population (Mouchet, 1994). quantitative picture of the fate of DDT or other However, recent data indicate that many of the pesticides which are sprayed indoors. Of highest concentrations of DDT residues in particular interest are the extent of uptake by humans are in areas where indoor house residents and extent of migration to the outdoor spraying with DDT is a common vector control environment. measure although it cannot be ascertained whether there is unauthorized agricultural use as The mathematical model yields an estimate of well (Bouwman et al., 1991). Conversely, the applied pesticide that remains on the walls; concentrations are declining where DDT use has is transferred to air, food, and other been discontinued. “compartments” of the house via different routes; is transferred to the outside environment In addition, there persists an assumption that the via different routes; is taken up by residents via use of DDT indoors contributes only tiny different routes; is degraded, etc. In this case, the amounts of DDT to the environment (Mouchet, model only addresses a single adult male 1994). As DDT use has been deliberately and inhabitant. The very different behaviour, effectively scaled back in agriculture, the consumption patterns, and inhalation rates of potential for indoor house spaying to contribute children, especially their frequent hand to mouth more to residents’ body burden of DDT, and to activity which exposes them to a great deal more environmental contamination, increases. contaminants via ingestion and dermal exposure, were not modelled but could conceivably yield quite different results. The potential transfer of WWF commissioned the development of a DDT to infants via breast milk has not been modelled either. months) after the initial application. In the indoor environment, with limited light and The mass balance model estimated the fate of biological activity, degradation of DDT would DDT 180 days following a single application of be especially slow. Subsequent applications will 670 grams applied at the rate of 2 grams per cause some build-up, but within a year or so a square meter. Because of uncertainties in certain steady state situation will develop in which there model parameters, for example the room is a fairly constant average amount of between ventilation rate, the quantities are given as 400-500 grams remaining on the wall surfaces, ranges rather than as single values. with the range being from 100-800 grams and the rate of application and the rate of loss from Physical removal and transfer of the room approximately equal. With a room of DDT to outdoors. 360 square meters, this corresponds to 1.1 to 1.4 Between 400 to 550 grams (60 to 82%) of the gram per square meter. While still almost half of total DDT applied is physically removed from the DDT applied, it is a much lower amount than the walls and transferred outdoors. The model the 2 g/m2 strived for as an active dose. This assumed that, because of its crystalline form, would explain why standard efficacy tests to DDT would flake off the walls and onto measure mosquito mortality show reduced surfaces, and would ultimately be mopped or contact insecticidal performance with time. swept outdoors. Alternatively, the DDT could be removed from the walls by washing and Human uptake. transferred to the outdoor environment via The one adult male resident is estimated to take washwater, as surveys conducted by WWF in up DDT from the indoor application in the order Mexico indicate. of 1 microgram per hour or 20 micrograms per day by inhalation. This represents a very small Absorption into food. fraction of the DDT applied. Although DDT is likely to be absorbed from air and dust concentrations in the air are calculated to be into food, especially fatty foods such as butter much higher than those in the outside air, and milk which have a high affinity for DDT. inhalation is a relatively unimportant route of The concentrations achieved in the food may be human exposure. On the other hand, uptake quite large - in the range of parts per million - through consumption of food into which DDT but the total mass of DDT in the food will be has deposited or migrated and dermal contact are small compared to the mass in the room, i.e. less significant routes of exposure. The latter would than 1 gram. be especially relevant for those who clean the walls and floors, and for infants and children Evaporation. who are in regular contact with contaminated surfaces. Direct evaporation from the wall is calculated to be minimal but does occur constantly and is The total uptake over a 6 month period is based on the chemical’s vapour pressure. estimated to be in the range of 0.1 to 0.3 grams. However, since concentrations indoors are Since approximately 50% would be excreted, calculated to be three orders of magnitude higher there is a net retention of 0.05 to 0.15 grams than in outdoor air, it is certain that there is total DDT in the fat. The model estimates that, transfer of DDT to the outdoors in the gaseous with continuous exposure, i.e., every 6 months, phase. concentrations of DDT in the fat would increase over time in the range of 3-9 ppm of fat per year for an adult male. However, this would not continue indefinitely since, after approximately five years, the concentration would start to level Remaining on wall surface. off at 10 to 30 µg/g fat. This reflects a saturation Between 120 to 270 grams (18 to 40%) remain point in the body and is within the range actually on the wall and other surfaces 180 days (6 found in human fat in regions of the world where DDT is used or highly concentrated. applications of DDT also serve to compound impacts on human health. There are few experimental or monitoring data against which to validate the results of this mass While the human data on the health effects of balance model, although actual data on residue low-dose exposures and endocrine disruption levels on walls and surfaces, of DDT effects are sparse at this point, there is a large concentrations in air, food, and residents’ fat body of evidence regarding exposures, residues, should not be difficult to obtain. The model and health effects in wildlife. Given that many would also benefit from refinement of the input essential biochemical processes are common to data, including food storage and consumption all species, it would seem prudent to regard patterns, cleaning behaviour, and specific effects on such processes in other species, information about children’s food consumption including humans, as potentially significant. and indoor behaviours.

Summary IV. RECOMMENDATIONS Using the mass balance model as a screening FOR RESEARCH tool indicates, overall, that much of the pesticide sprayed on walls and furniture during indoor Formulating disease-vector-control strategies spraying operations ends up outdoors. In requires a complex assessment and balancing of addition, a small but significant amount is a broad range of factors including effectiveness, transferred via food to residents, which can cost, sustainability, nature of the disease threat contribute substantially to their body burden of (morbidity vs. mortality) and such DDT. environmental considerations as impacts on non- target species and hazards to workers handling chemicals and humans living in treated Synthetic Pyrethroids environments. Despite some questions raised Synthetic pyrethroids are being substituted for about their cost, synthetic pyrethroids in spraying DDT in houses and they are the only particular appear to be growing in popularity as chemicals available for impregnating bednets. an alternative to DDT, especially since they are The limited research conducted to date on not as persistent and bioaccumulative as DDT. synthetic pyrethroids is insufficient to fully assess exposure to them. The weight of scientific evidence regarding the connection between wildlife health and human health is growing. Adverse health impacts Interpretation of Human Exposure observed in wildlife and laboratory animals from Data concentrations of DDT and other POPs are Many human populations depend on fish and indicators of the potential human situation other wildlife for a large portion of their diet. because biological processes of the endocrine, Thus, they may accumulate high levels of immune, nervous, and reproductive systems are persistent pesticide residues, including DDT, common to all animals. WHO’s scientific from these sources. Inuit in northern Canada are experts appear to have focused largely on what one group whose traditional diet and mothers’ might be called traditional health endpoints - milk have become dangerously contaminated. cancer and acute toxicity. There has been little, The strict pesticide residue regulations on if any, attention to the new science on imported foods by the United States, Europe, transgenerational impacts of DDT and other and some Asian countries reflect health pesticides. Since WHO’s last significant review authorities’ concerns about human health of DDT in a public health context occurred in impacts. Where malaria control programs spray 1993, and most of the scientific literature on the houses repeatedly, DDT residues in impacts of these hormone-disrupting chemicals householders and applicators are particularly on reproductive, neural, immune, and high (Bouwman et al., 1991). Illegal agricultural behavioural outcomes post-dates this review, this is not surprising. There is now a compelling, science-based case for the re-examination of previously reported to be the No Observed DDT and other recommended chemical Adverse Effects Level (NOAEL) for bisphenol alternatives. A (vom Saal, 1997). Other researchers working with PCBs (polychlorinated biphenols) found A survey of the currently published literature significant neurotoxic effects in the offspring of involving vector control pesticides highlights the exposed female rats, effects that showed up at need for much more research in order to better low doses but were not evident at high doses understand the range of impacts on both humans (Holene et al., 1995). Dutch researchers looking and wildlife. In particular, assessment of the at PCBs, dioxins, and furans report that hazards associated with the ‘newer’ chemicals “relatively subtle adverse effects on which are replacing DDT are needed. It is neurobehavioural development and thyroid crucial for research to consider a broad range of hormone alterations have been observed in endpoints including subtle effects on the infants and children exposed to background immune system and nervous system, levels” (Brouwer et al., 1995). reproductive outcomes, as well as behavioural impacts. While this paper cannot provide a detailed listing of specific research objectives, Testing for Transgenerational given the broad potential for endocrine Effects disruption related effects, it is clear that research should focus on low-dose testing, testing for Recent statements from government officials transgenerational effects, and the assessment of and concerned scientists have clearly indicated synthetic pyrethroid exposure to children and the that current regulatory approaches to toxic developing fetus from bednets. chemicals fail to address the special health needs of infants and children. Scientists now acknowledge that it is not sufficient to assess Low-Dose Testing effects only on infants and children, but it is Traditional government-mandated toxicology necessary to go even further back in time – to testing, as noted numerous times in this paper, focus more systematically on the role of focuses on administering high doses of chemical contaminants on the developing chemicals, usually to adult animals. But a new embryo and fetus. Reacting to reports of adverse paradigm of concern is emerging, specifically health effects in wildlife and laboratory animals, with regard to hazards associated with the researchers are increasingly targeting in humans exposure of fetuses and embryos to extremely the very critical 266 days from conception to low doses of chemicals that disrupt the hormonal birth, and exposure during that time to outside systems of the body. Organisms can be chemicals that bypass protective blood and chronically exposed to such doses in the placental barriers. The remarkable sensitivity of environment. the fetus and embryo to extremely low doses of chemicals (both industrial chemicals and Traditional screening and testing programs for pesticides) means their exposure to assessing hazards from pesticides and other extraordinarily low amounts of toxic chemicals toxic chemicals are not designed to capture the can dramatically influence their future full range of undesirable effects of chemicals, development and well-being. particularly effects from exposures to very low doses in the womb or post-natally. The need for attention to low doses is signaled in a number of Assessment of Synthetic Pyrethroid laboratory studies. For example, researchers Exposure to Children and the (Nagel et al., 1997) have found that a 2 ppb (parts per billion) dose of the plasticizer Developing Fetus from Bednets bisphenol A administered to pregnant mice led As the house-spray modeling indicated, there are to a significant increase in their male offspring’s critical questions which should be answered prostate weights; this dose is 25,000 times lower empirically regarding the resident and than the 50 ppm (parts per million) dose that was environmental exposure from indoor spraying and the use of bednets. While all alternatives to DDT ought to be examined for their endocrine precautionary principle includes a substitution disruption potential at low doses, it is especially clause enacted in 1991. This includes avoiding important to test the synthetic pyrethroids chemical products for which less hazardous because pregnant women and children around substitutes are available. The law also states that the world will be exposed to them inside their a scientifically-based suspicion of risk shall homes and under bednets on a continuing basis. constitute sufficient grounds for the government These hazards may prove to be quite low. Even to take measures against a chemical (Hileman, if they are sizable, these hazards may be worth 1998). incurring if the alternative is a high risk of death or severe disease resulting from contact with disease-bearing vectors. But these should be fully-informed decisions, and where lacking, the appropriate data should be developed expeditiously.

In Closing – the Precautionary Principle The release and use of toxic substances, the exploitation of resources, and physical alterations of the environment have had substantial unintended consequences affecting human health and the environment. Growing evidence of high rates of learning deficiencies, asthma, cancer, birth defects, and species extinction; along with global climate change, stratospheric ozone depletion, and worldwide contamination with toxic substances, has moved some countries to adopt policies based on the precautionary principle. According to the precautionary principle, when substantial scientific evidence suggests good reason to believe that an activity, technology, or substance may be harmful, action should be taken to prevent harm. In other words, if an activity raises threats of harm to the environment or human health, precautionary measures should be taken even if some cause and effect relationships are not fully established scientifically (Wingspread Statement on the Precautionary Principle, 1998). The precautionary principle, as a general approach to environmental policy, is not entirely new. It already forms the basis of at least a dozen treaties and laws, including the 1987 Montreal Protocol on Substances That Deplete the Ozone Layer, the 1990 Massachusetts Toxics Use Reduction Act, the 1992 United Nations Framework Convention on Climate Change, and the 1994 Maastricht Treaty of the European Union. Sweden’s interpretation of the The precautionary principle places much less emphasis on risk assessment and cost-benefit analysis than does current practice. 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