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Policy Brief

An Updated Look at Mancozeb: 

Concerns about Farmworker Reproductive and Child 

Developmental Health

Joan Flocks, Jennifer Runkle, Jeannie Economos, Anne 

Dunlop and Jennifer Sass
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An Updated Look at Mancozeb: 

Concerns about Farmworker Reproductive and Child Developmental Health

Executive summary

Mancozeb is a broad-spectrum fungicide commonly used in agriculture worldwide. It is a member of the 

ethylene bisdithiocarbamate (EBDC) group of fungicides, which includes the related active ingredients maneb 

and metiram. Currently, exposure to mancozeb is considered to pose minimal human health risks by most 

regulatory bodies. Despite mounting scientific evidence, the potential adverse reproductive and human health 

developmental effects of mancozeb, especially in sensitive subpopulations (e.g., pregnant women, children, and 

agricultural workers), has not yet been fully considered in the registration and reregistration of this widely used 

fungicide. 

We urge the EPA to systematically evaluate and consider the reproductive, pregnancy, and fetal health impacts 

of the fungicide mancozeb in its upcoming Registration Review. In particular, we urge the Agency to consider 

the full risks – including developmental and reproductive risks - from exposure to mancozeb experienced by 

vulnerable populations, such as all children under the age of 21 and women who work in agricultural operations 

(including in fields, nurseries, and ferneries) and who work with agro-chemicals. The planned scheduled 

opening of the Registration Review docket for mancozeb is the third quarter of FY2015.

We make this request because current EPA policy does not reflect acknowledged data gaps and uncertainty 

about the health impacts of mancozeb. Our systematic review of mancozeb research indicates that the fungicide 

presents a risk of both reproductive and developmental harm to humans. Furthermore, recent cases of infants 

born with severe birth defects to farmworkers possibly exposed to mancozeb in violation of regulations should 

trigger the EPA to act in a precautionary manner by taking action to address the risk concerns from the use of 

mancozeb.

The Problem

Our research with farmworkers of childbearing age demonstrates that: (1) an agricultural work environment 

characterized by multiple stressors, including pesticide exposure, prolonged elevated temperatures, and 

musculoskeletal strain, can influence reproductive health, and (2) the farmworker community is profoundly 

concerned about the health risks associated with agricultural work, the scarcity of guidelines, information or 

regulations protecting the health of pregnant women in the workforce, and (3) the uncertainty regarding the 

impact of these exposures on pregnancy and child health.1,2 One chemical that has been of particular concern in 

the farmworker community is mancozeb.

A systematic review of the relevant scientific literature related to mancozeb is provided in Appendix A. 

Mancozeb is an ethylene bisdithiocarbamate (EBDC) fungicide with the degradate ethylenethiourea (ETU). 

ETU has known teratogenic, mutagenic, and carcinogenic risks. The State of California and the Toxic Release 

Inventory (TRI) lists mancozeb as a carcinogen. The TRI also lists mancozeb as a developmental and 

reproductive toxicant. One study in rodents has reported that mancozeb and ETU are capable of crossing the 

"""""""""""""""""""""""""""""""""""""""""""""""""""""" 1

Flocks J, Kelley M, Economos J, McCauley L. Female farmworkers’ perceptions of pesticide exposure and pregnancy health. J 

Immigr Minor Health 2012:14: 626-632.

2 Flocks J, Thien Mac V, Runkle J, Tovar-Aguilar JA, Economos J, McCauley L. Female farmworkers’ perceptions of heat-related 

illness and pregnancy health. J Agromedicine 2013:18: 350-358. 
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placental barrier and can damage DNA and initiate tumors in fetal cells.3 Mancozeb is also a suspected 

endocrine disruptor and is associated with both hyperthyroidism and hypothyroidism.

3a,b

From 2004-2006, six infants were born with a range of birth defects to farmworkers in southwest Florida. Of 

these, three cases were investigated by the Centers for Disease Control and Prevention in collaboration with 

state health departments in Florida and North Carolina (the two states in which the mothers worked harvesting 

tomatoes for the same agricultural company). In a report on the cases, mancozeb and ETU were implicated 

because in two cases it could be shown that the pregnant mothers had been exposed to the fungicide in violation 

of Restricted Entry Intervals. Also, the specific birth defects of the farmworkers’ infants resembled those 

observed in laboratory studies of rats exposed to mancozeb and ETU.

4 This is concerning since our research

with female farmworkers in Central Florida revealed higher urinary metabolite levels of ETU compared with 

non-detect levels of ETU in other nationally representative women and Mexican Americans reported in the 

National Health and Nutrition Examination Survey (NHANES), suggesting the risk to women and their babies 

may be much more widespread than just these documented cases in Florida.

5

It is not just pregnant women and developing fetuses that are especially sensitive to pesticides, but all workers

under the age of 21. Puberty is recognized as a critical or sensitive window of susceptibility to chemicals, 6

particularly with respect to reproductive and endocrine disrupting effects. For example, exposure of rodents to 

high doses of some phthalates has been shown to delay onset of puberty while pubertal dosing of two rat species 

with the phthalate DEHP has been shown to delay onset of puberty and cause testicular damage7 and exposure 

to various chemicals can delay or accelerate onset of puberty in females.8 Data primarily from laboratory animal 

and cell culture studies have revealed more than 50 pesticides that have known or suspected endocrine 

disrupting properties on the female reproductive system.9,10 These risks of reproductive harm are not fully 

addressed in EPA’s mancozeb registration process.

We recognize that for farmworkers to be adequately protected against occupational hazards, such as pesticides,

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3 Shukla Y, Arora A. Transplacental carcinogenic potential of the carbamate fungicide mancozeb. J Environ Pathol Toxicol Oncol

2001:20: 127–131.

3a Goldner WS, Sandler DP, Yu F, Hoppin JA, Kamel F, and Levan TD. Pesticide use and thyroid disease among women in the 

Agricultural Health Study. Am J Epidemiol 2010: 171 (4): 455-64.

3b Axelstad M, Boberg J, Nellemann C, et al. Exposure to widely used fungicide mancozeb causes thyroid hormone disruption in rat 

dams but no behavioral effects in the offspring. Tox Sci. 2011: 120 (2): 439-46.

4 Calvert GM, Alarcon WA, Chelminski A, et al. Case report: three farmworkers who gave birth to infants with birth defects closely 

grouped in time and place—Florida and North Carolina, 2004–2005. Environ Health Perspect 2007:115: 787–791.

5Runkle JD, Tovar-Aguilar JA, Economos E, Flocks J, Williams B, Muniz J, Semple M, and McCauley L. Pesticide risk perception 

and biomarkers of exposure in Florida female farmworkers. J Occup Environ Med 2013:55: 1286-92.

6 Woodruff TJ, Carlson A, Schwartz JM, Giudice LC. Proceedings of the summit on environmental challenges to reproductive health 

and fertility: executive summary. Fertil Steril 2008:89: 281-300.

7 Noriega NC, Howdeshell KL, Furr J, Lambright CR, Wilson VS, Gray LE Jr. Pubertal administration of DEHP delays puberty, 

suppresses testosterone production, and inhibits reproductive tract development in male Sprague-Dawley and Long-Evans rats. 

Toxicol Sci 2009:111: 163-78. 

8 Woodruff TJ, Carlson A, Schwartz JM, Giudice LC. Proceedings of the summit on environmental challenges to reproductive health

and fertility: executive summary. Fertil Steril 2008:89: 281-300.

9 Bretveld RW, Thomas CMG, Scheepers PTJ, Zielhuis GA, Roeleveld N. Pesticide exposure: the hormonal function of the female 

reproductive system disrupted? Reprod Biol Endocrinol 2006:4: 30.

10 Diamanti-Kandarakis E, Bourguignon JP, Giudice LC, Hauser R, Prins GS, Soto AM, Zoeller RT, Gore AC. Endocrine-disrupting 

chemicals: An endocrine society scientific statement. Endocr Rev 2009:30: 293-342.
4

there must be meaningful regulation that is adequately and uniformly enforced. At present, pesticide regulations 

are often ineffective or unenforced. Furthermore, existing regulations are often not based upon consideration of 

the reproductive health impacts and developmental outcomes, particularly for chronically exposed and 

vulnerable populations such as farmworkers and their children. Accurate exposure and risk characterization of 

mancozeb as a potential reproductive toxicant is increasingly becoming a priority in the scientific community, 

yet regulatory bodies are lagging in their consideration of the potential reproductive health impacts in the 

reassessment of the currently registered uses of mancozeb. EPA must consider the growing evidence of human 

health harm in its upcoming Registration Review docket for mancozeb in 2015.

Regulatory History and Current Policy

According to the 2005 Reregistration Eligibility Decision (RED),11 mancozeb was first registered in the United 

States in 1948 as a broad spectrum fungicide for use in agriculture, turf management, and horticulture. The 

Mancozeb Task Force was formed in 1994 to represent the interests of the mancozeb registrants, which were 

then two companies: Rhom and Haas and E.I. DuPont De Nemours. The 2005 RED states that at that time the 

registrants included Dow AgroSciences, Griffin (now a DuPont subsidiary), and Cerexagri (RED p. 3)

Mancozeb has demonstrated 50 years of fungicidal efficacy for roughly 400 different plant pathogens. It is 

commonly used on a variety of crops including tree fruits, vegetable crops, field crops, ornamental plants, and 

sod farms. In 2005, the EPA reported that approximately 5.6 million pounds of mancozeb were used annually. 

The United States Geological Survey map and chart below show the estimated amount and distribution of 

agricultural use for mancozeb in 2009, the last year with confirmed crop data from the Census of Agriculture, 

and a breakdown of crops on which mancozeb was used. As the map shows, mancozeb use is particularly heavy 

in Florida and North Carolina. 

Source: U.S. Geological Survey, Pesticide Use Maps – Mancozeb,

http://water.usgs.gov/nawqa/pnsp/usage/maps/show_map.php?year=2011&map=MANCOZEB&hilo=L&disp=Mancozeb. (Accessed May 23, 2014).

The 2005 RED states that mancozeb and other EBDCs have been the subject of two special EPA reviews, 

"""""""""""""""""""""""""""""""""""""""""""""""""""""" 11 EPA 2005. Reregistration Eligibility Decision for mancozeb. Office of Prevention, Pesticides, and Toxic Substances. EPA 738-R04-012
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initiated in 1977 and 1987, because of particular health concerns, including developmental and thyroid effects, 

caused by their common degradate ETU. According to the RED, the final determination of the second special 

review concluded that the dietary risks for EBDCs exceeded the economic benefits for 11 fruit and vegetable 

food/feed crops, resulting in the cancellation of the use of mancozeb and other EBDCs on these specific crops

(RED pp. 2-3). The conclusion of the 2005 RED was to allow for continued use of mancozeb provided that 

certain risk mitigation measures were adopted and labels amended to reflect those measures. Certain uses of 

mancozeb, however, were voluntarily canceled, including its foliar use on cotton, use on residential lawns/turf, 

and use on athletic fields. In the RED, the EPA raised several risk concerns and recognized data gaps for 

developmental, reproductive, and thyroid toxicity, including:

• Risk estimates were not provided for the following scenarios due to lack of worker exposure data: 

applicator using liquid dip for seed-piece treatment, applicator using dusts for commercial or on farm 

seed treatment, and secondary handling for hand planting treated seed pieces. These data will be 

required in the DCI for this RED ( RED p. 44)

• EPA intends to issue a DCI requiring submission of additional environmental fate data for mancozeb 

parent, the mancozeb complex, and the ETU degradate (RED p. 53)

These above studies would likely be very relevant to an evaluation of the exposures and resultant health risks to 

workers, including women and children in agriculture fields. It is not clear whether or not EPA ever received 

these data from the registrants, and, if received, what the data concluded.

In addition to the studies above, the following guideline studies were identified as missing, and required as part 

of a DCI (RED pp. 101-102):

Toxicology:

870.6200 Acute neurotoxicity

Occupational Exposure

875.1100 Dermal exposure monitoring, outdoor (potato seed piece treatment with liquids and

dusts)

875.1300 Inhalation exposure monitoring, outdoor (potato seed piece treatment with liquids

and dusts)

Toxicology

870.3700 Developmental toxicity study in rabbits

870.3800 2 Generation Reproductive Toxicity Study

870.6300 Developmental neurotoxicity study

Special study Comparative thyroid toxicity study in young and adult rats

The RED states that EPA “intends to issue a separate product-specific data call-in (PDCI) outlining specific 

data requirements” (RED p. 103). These above studies are directly and critically relevant to an evaluation of the 

exposures and resultant health risks to women and children – including underage workers - in agriculture fields. 

It is not clear whether or not EPA ever received these data from the registrants, and, if received, what the data 

concluded regarding developmental and reproductive risks to humans from mancozeb.

Since 2005, the EPA has issued final rules on tolerances for residues of mancozeb on various crops including 

almonds, cabbage, lettuce, peppers, and broccoli in 201112 and walnuts and tangerines in 2013.13 In these final 

"""""""""""""""""""""""""""""""""""""""""""""""""""""" 12 Mancozeb; Pesticide Tolerances. 40 CFR Part 180. 2013 ed. 13 Mancozeb; Pesticide Tolerances. 40 CFR Part 180. 2011 ed.
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rules, EPA continued to acknowledge certain gaps for mancozeb and ETU, especially regarding impact on the 

developing thyroid and reproductive systems. 

The 2013 final rule acknowledged that maternal mortality, clinical signs, spontaneous abortion, thyroid effects, 

body weight gain decrements, and decreased pup body weight were observed in rat and rabbit studies with 

mancozeb. It acknowledged fetal malformations, hydrocephaly, and domed head were observed in rat and rabbit 

studies with ETU. 

For mancozeb and ETU, the EPA retained the 10X FQPA safety factor for women of childbearing age and for 

children less than six years old. Yet, despite the data gap and uncertainty, the EPA determined that children 

older than six would be adequately protected if the FQPA safety factor were reduced to 1X, even though these 

are the years of puberty in which hormonal disruption could render children and youth most vulnerable. It is not 

clear how EPA made that determination, given the substantial data gaps and lack of relevant information, along 

with scientific evidence that pregnant women and children under age 21 exposed occupationally to pesticides 

represent vulnerable and inadequately-protected populations.

For vulnerable populations, such as farmworkers, who are more frequently and intensely exposed to 

agrochemicals, the risk associated with data gaps and uncertainties is multiplicative. The EPA has seemingly 

recognized these inherent risks for farmworkers. In the 2005 RED, the EPA stated: “Based on its evaluation of 

mancozeb, the Agency has determined that mancozeb products, unless labeled and used as specified in this 

document, would present risks inconsistent with FIFRA. Accordingly, should a registrant fail to implement any 

of the risk mitigation measures identified in this document, the Agency may take regulatory action to address 

the risk concerns from the use of mancozeb.” It is not clear to us whether or not the registrants fully 

implemented the label requirements to mitigate risk, or if not, whether any Agency regulatory action was taken.

Systematic Review of Research on Mancozeb

The evidence summarized in our review of thirty-six studies suggests that mancozeb presents a risk of both

reproductive and developmental harm to humans. Data from in vitro studies provide strong evidence that 

mancozeb may indirectly disrupt or impair reproduction at the cellular level and should be regarded as a 

reproductive toxicant. Animal studies also provide strong evidence that mancozeb may induce reproductive and 

developmental toxicity in mammals. A summary of the data is below, and is presented in more detail in the 

systematic review in Appendix A.

• In vitro studies provide strong evidence in support of mancozeb as a reproductive toxicant, with over a 

decade of research demonstrating high endocrine disrupting potential, significant cytotoxic and proapoptotic patterns at 10 and 50 ug/ml between 24 and 48 hours, developmental injury in murine preimplementation embryos, oxidative and genotoxic damage associated with chronic exposure, 

premalignant status in somatic cells of mammalian ovarian follicles, and dose-dependent antiandrogenic effects likely involved in fetal disruption of male sexual development. 

• Early in vivo studies failed to find a significant effect on reproduction for female rats exposed to the 

maximum tolerated dose of ETU before and during gestation (80 mg/kg/day); however, significant 

morphological fetal abnormalities, with the fetal brain most commonly affected, were detected at doses 

showing no maternal toxicity or fetal mortality in rat offspring.

• Several studies documented a reduction in reproductive potential in female animals at higher levels of 

exposure, while mixture studies revealed that prenatal exposure to a low dose pesticide mixture 

containing mancozeb may lead to adverse developmental toxicity in F1 generations including 

disruptions in the developing mouse cerebellar cortex, long-term delayed effects at dose levels where 

single pesticides showed no effects, and genital malformations in male rat pups. 
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• Animal studies also confirmed reproductive and developmental toxicity in mammals and suggested

significant changes in physiological, biochemical, and pathological processes involved in normal 

reproduction may lead to infertility in males chronically exposed to mancozeb. 

• Epidemiological studies examining the effect of mancozeb on reproduction were mixed, but results 

suggested that mancozeb may interfere with normal reproductive functioning of women persistently 

exposed (either occupationally or indirectly by spouse’s occupation) and thereby may delay or 

complicate pregnancy. 

• Few population-based studies have examined the association between exposure to mancozeb and 

adverse reproductive or developmental outcomes.

Policy Recommendations

In summary, high confidence ratings from in vitro and animal studies, in combination with moderate confidence 

ratings from human health studies, provide sufficient evidence that mancozeb should be regarded as a presumed

developmental and reproductive hazard in humans. We urge the EPA to systematically consider the 

reproductive, pregnancy, and developmental health impacts of mancozeb. The chronic exposure experienced in

vulnerable populations, such as farmworkers and children, to mancozeb makes this matter urgent. The planned 

scheduled opening of the Registration Review docket for mancozeb is the third quarter of FY2015. 

The EPA has previously acknowledged data gaps in its consideration of mancozeb toxicity. To address this, the 

Agency should clearly identify the data for which it issued a DCI and when it expected to receive that data, a 

check-list of the studies that were submitted to EPA in response to the DCI, a link to the Data Evaluation 

Record (DER), and a short summary of what the data results were. EPA should furthermore indicate if there are 

any outstanding studies that have not been received, or that were received but were found to be unacceptable as 

guideline studies.

In light of the acknowledged data gaps, we urge the agency to reconsider the 1X FQPA safety factor for 

children over six. The inconsistent safety factor pattern makes little sense in the face of continuing uncertainties 

about the health impacts of mancozeb and ETU, especially for vulnerable populations. Currently, girls and boys 

ages 6-13 receive a 1X safety factor, whereas girls and women ages 13-49 receive a 10X safety factor. Yet, 

children under age 21 should not be evaluated as adults. Puberty is recognized as a critical or sensitive window 

of susceptibility to chemicals 14 particularly with respect to reproductive and endocrine disrupting effects. For 

example, exposure of rodents to high doses of some phthalates has been shown to delay onset of puberty while 

pubertal dosing of two rat species with the phthalate DEHP has been shown to delay onset of puberty and cause 

testicular damage15 and exposure to various chemicals can delay or accelerate onset of puberty in females.16

Data primarily from laboratory animal and cell culture studies have shown over 50 pesticides to have known or 

suspected endocrine disrupting properties on the female reproductive system.17,18 Studies have reported that 

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14 Woodruff TJ, Carlson A, Schwartz JM, Giudice LC. Proceedings of the summit on environmental challenges to reproductive health 

and fertility: executive summary. Fertil Steril 2008:89: 281-300.

15 Noriega NC, Howdeshell KL, Furr J, Lambright CR, Wilson VS, Gray LE Jr. Pubertal administration of DEHP delays puberty, 

suppresses testosterone production, and inhibits reproductive tract development in male. Sprague-Dawley and Long-Evans rats. 

Toxicol Sci 2009:111: 163-78.

16 Woodruff TJ, Carlson A, Schwartz JM, Giudice LC. Proceedings of the summit on environmental challenges to reproductive health 

and fertility: executive summary. Fertil Steril 2008:89: 281-300.

17 Bretveld RW, Thomas CMG, Scheepers PTJ, Zielhuis GA, Roeleveld N. Pesticide exposure: the hormonal function of the female 

reproductive system disrupted? Reprod Biol Endocrinol 2006:4: 30.

18 Diamanti-Kandarakis E, Bourguignon JP, Giudice LC, Hauser R, Prins GS, Soto AM, Zoeller RT, Gore AC. Endocrine-disrupting 

chemicals: An endocrine society scientific statement. Endocr Rev 2009:30: 293-342.
8

dioxin causes abnormal breast development in pre-pubertal girls, the plastics-component Bisphenol A (BPA) 

has shown to cause precocious puberty in girls, the pesticide endosulfan has been reported to affect pubertal 

boys by slowing down the timing of reproductive maturation, and the fungicides vinclozolin and iprodione have 

been shown in pubertal male rats to impair normal puberty hormone levels.19,20,21,22 EPA has not adequately 

addressed the potential risks for reproductive harm from childhood exposures to mancozeb. Until this is done, 

the full 10X FQPA factor should be retained for all children, male and female, under the age of 21.

EPA should also consider the occupational health of vulnerable populations, such as farmworkers in the case of 

mancozeb, as part of its risk assessments. Farmworkers experience multiple stressors in addition to pesticides, 

including poverty, lack of sanitation, heat stress from outdoor field work, lack of access to health care, poor 

housing conditions, risk of pesticide drift into farmworker housing, lack of stability of home life, separation 

from family, rape and sexual assault risks, and other significant issues. Because of economic instability, it is not 

uncommon for farmworker women to continue working while pregnant, sometimes well into the third trimester 

and our research has indicated that there are no special workplace accommodations made for pregnant 

farmworkers, who risk direct contact to pesticide residues or drift.23 Furthermore, some low-income 

farmworkers do not have access to childcare, and farmworkers have been known to bring their children to the 

workplace with them. Farmworkers are paid by a piece rate system and the resulting production demands often 

means they do not have or take the time to practice occupational safety behavior such as frequent handwashing, 

wearing adequate protective clothing, or replacing defective equipment.24,25 Sometimes workers are not provided 

with proper PPE and/or decontamination supplies by their employer and must provide it themselves.26 Despite 

federal regulations, farmworkers often lack access to basic information about pesticides applied at their 

worksites. Widespread lack of compliance and inadequate enforcement of current Worker Protection Standards 

means that many farmworkers receive no or minimal pesticide health and safety training or that information 

about pesticides applied is not being posted correctly.27 Language barriers can impede the understanding of 

workers of the risks posed by the pesticides around which they are working, including those who mix, handle, 

and/or apply the pesticides. Pesticide labels are generally in English only. Even in the best scenarios in which 

"""""""""""""""""""""""""""""""""""""""""""""""""""""" 19 Cowin PA, Gold E, Aleksova J, O'Bryan MK, Foster PM, Scott HS, Risbridger GP. Vinclozolin exposure in utero induces 

postpubertal prostatitis and reduces sperm production via a reversible hormone-regulated mechanism. Endocrinology 2010: 151:783-

92. 

20 Blystone CR, Lambright CS, Cardon MC, Furr J, Rider CV, Hartig PC, Wilson VS, Gray LE Jr. Cumulative and antagonistic effects

of a mixture of the antiandrogens vinclozolin and iprodione in the pubertal male rat. Toxicol Sci 2009:111:179-88.

21 Roy JR, Chakraborty S, Chakraborty TR. Estrogen-like endocrine disrupting chemicals affecting puberty in humans--a review. Med 

Sci Monit 2009:15:RA137-45. 

22 Schoeters G, Den Hond E, Dhooge W, van Larebeke N, Leijs M. Endocrine disruptors and abnormalities of pubertal development. 

Basic Clin Pharmacol Toxicol 2008:102:168-75.

23 Runkle JD, Tovar-Aguilar JA, Economos E, Flocks J, Williams B, Muniz J, Semple M, and McCauley L. Pesticide risk perception 

and biomarkers of exposure in Florida female farmworkers. J Occup Environ Med 2013:55: 1286-92.

24 Mayer B, Flocks J, and Monaghan P. The role of employers and supervisors in promoting pesticide safety behavior among Florida 

farmworkers. Am J Ind Med 2010:53:814-24.

25 Runkle JD, Tovar-Aguilar JA, Economos E, Flocks J, Williams B, Muniz J, Semple M, and McCauley L. Pesticide risk perception 

and biomarkers of exposure in Florida female farmworkers. J Occup Environ Med 2013:55: 1286-92.

26 Flocks J, Kelley M, Economos J, McCauley L. Female farmworkers’ perceptions of pesticide exposure and pregnancy health. J 

Immigr Minor Health 2012:14: 626-32.

27 Flocks J, Monaghan P, Albrecht S, Bahena A. Florida farmworkers’ perceptions and lay knowledge of occupational pesticides. J 

Commun Health 2007:32:181-94.
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an employer conscientiously complies with all existing regulations, current WPS training requirements do not 

require information about reproductive health or risks posed by take-home exposures, although this is being 

considered under the proposed regulations currently open for public comment. 

In 2009, the Committee on Improving Risk Analysis Approaches Used by the U.S. EPA, part of the National 

Research Council (NRC), emphasized the need for cumulative risk assessments, as defined by EPA, “that 

include combined risks posed by aggregate exposure to multiple agents or stressors; aggregate exposure 

includes all routes, pathways, and sources of exposure to a given agent or stressor. Chemical, biologic, 

radiologic, physical, and psychologic stressors are considered in this definition.” 28 The NRC report points out 

that, “although cumulative risk assessment has been used in various contexts, there has been little consideration 

of nonchemical stressors, vulnerability, and background risk factors” and recommends EPA seriously consider 

adopting the recommendations of a NRC report on phthalates29 that “addresses issues related to the framework 

within which dose-response assessment can be conducted in the context of simultaneous exposures to multiple 

stressors.”The NRC report also emphasizes that “variability in human susceptibility has not received sufficient 

or consistent attention in many EPA health risk assessments... greater attention to susceptibility in practice is 

needed, particularly for specific population groups that may have greater susceptibility because of their age, 

ethnicity, or socioeconomic status.” 30 EPA should implement the recommendations of these NRC reports with 

mancozeb and other pesticide assessments.

Finally, more detail is needed for EPA’s assessment of endocrine effects of mancozeb. EPA should describe 

how the agency will move forward on its analysis of endocrine disrupting effects and specifically address how it 

considers non-linear dose responses and adverse health effects occurring at extremely low doses (e.g. parts per 

trillion) for which there is a growing body of scientific evidence. For example, pesticides, at extremely low 

levels, can interfere with the proper functioning of hormone systems including estrogen, androgens, thyroid, and 

the pituitary/hypothalamic axis. When exposures, even at very low levels, coincide with critical windows of 

development, they may be more potent than higher dose exposures, or the low dose exposure may otherwise 

exert a non-traditional dose-response curve.The consequences of exposure to an endocrine disruptor may also 

be delayed with additive or synergistic effects from a myriad of chemical exposures. Data primarily from 

laboratory animal and cell culture studies have shown that more than 50 pesticides have known or suspected 

endocrine disrupting properties on the female reproductive system.31,32

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28 National Research Council. Science and Decisions: Advancing Risk Assessment. Washington, DC: The National Academies Press, 

2009, 478 pages.

29 National Research Council. Phthalates and Cumulative Risk Assessment: The Task Ahead. Washington, DC: The National 

Academies Press, 2008, 208 pages.

30 National Research Council. Science and Decisions: Advancing Risk Assessment. Washington, DC: The National Academies Press, 

2009, 478 pages.

31 Bretveld RW, Thomas CMG, Scheepers PTJ, Zielhuis GA, Roeleveld N. Pesticide exposure: the hormonal function of the female 

reproductive system disrupted? Reprod Biol Endocrinol 2006:4: 30.

32 Diamanti-Kandarakis E, Bourguignon JP, Giudice LC, Hauser R, Prins GS, Soto AM, Zoeller RT, Gore AC. Endocrine-disrupting 

chemicals: An endocrine society scientific statement. Endocr Rev 2009:30: 293-342.
10

Appendix A: Systematic Review Findings

In Vitro Studies. Seven in vitro studies published between 2000 and 2012 met our inclusion criteria (Figure 1, 

Table 1). We found seven in vitro studies in which the reproductive toxicity of mancozeb was examined. The 

media for most of these studies was mammalian cells. All of these studies provide evidence that mancozeb may 

indirectly disrupt or impair reproduction at the cellular level. 

Lin et al.1 showed mancozeb induced cytotoxicity and apoptotic patterns of cell death at doses of 10 and 50 

ug/ml between 24 and 48 hours of exposure, while ETU produced apoptotic induction at much higher 

concentrations (1000 ug/ml). A later study examining adrenocortical steriodogenic cells of rainbow trout 

demonstrated significant impairment of cortisol secretion at low-doses (50 ug/ml) and showed mancozeb as the 

highest ranked endocrine disruptor based on LC50/EC50 values.2 Mancozeb exposure did not result in increased 

embryotoxicity or a higher incidence in developmental abnormality in chicken embryos, but when mixed with 

copper sulfate showed significant embryomortality compared to individual doses.3 Another mixture study 

revealed that single exposure to mancozeb induced developmental injury in murine pre-implemenation embryos 

and a fungicide mixture (including mancozeb) reduced blastocyst development and increased apoptosis.4

Calviello et al.5 studied the DNA damaging and proapoptotic effects of mancozeb in RAT-1 fibroblasts and 

peripheral blood mononucleated cells (PBMC) isolated from Wistar rats at one and four hours post-exposure. 

Following one hour of exposure to 500 ug/ml of mancozeb, fibroblast cells produced dose-dependent induction 

of single strand breaks and elevated levels of DNA oxidation and reactive oxygen studies. After four hours, 

mancozeb induced dose-dependent apoptosis in fibroblasts and PBMC cells. 

Finally, two recent studies demonstrate mancozeb should be regarded as a reproductive and developmental 

toxicant. Paro et al. examined the toxic effects of mancozeb exposure on granulosa cells from mice and women 

undergoing assisted reproductive procedures.6 Granulosa cells from mice exposed to mancozeb showed timeand dose-dependent changes in morphology, an acquired ability to migrate, decreased levels of p53, but no 

changes in apoptosis. Human granulosa cells showed dose-dependent morphology changes and reduced p53 

expression post-exposure to mancozeb. This study demonstrates that mancozeb damages the somatic cells of 

mammalian ovarian follicles by inducing a premalignant-like status in both mice and human granulosa cells. In 

2013, Kjeldsen et al.7 demonstrated the in vitro effects of mancozeb on the androgen receptor (AR) function in 

human breast carcinoma MVLN cells and hamster ovary CHO-K1 cells. Results showed concentrationdependent effects on the agonist-induced AR transactivity, with mancozeb showing the highest inhibitory 

effect, suggesting a link between mancozeb as an AR disruptor and disorders of male sexual health. Because 

mancozeb has been shown to cross-placental barriers in mice, it is likely that mancozeb can disrupt fetal 

reproductive development in males. 

Overall, in vitro studies provide strong evidence that mancozeb is a reproductive or endocrine disrupting 

xenobiotic. 

Animal Studies. Fifteen experimental studies in mice, rats, rabbits, and pigeons published between 1973 and 

2012 met our inclusion criteria (Figure 1, Table 2). In all fifteen studies, mancozeb was either administered 

orally in food or delivered directly into the stomach (gavage) for lengths of time ranging from 30 days to 1 or 

more years. While each study included in the review measured individual exposure to mancozeb, several studies 

simultaneously examined exposure to other chemical agents including: maneb, probineb, endosulfan, and 

phosphamidon. Four studies assessed independent exposure to mancozeb and then as a mixture with copper 

oxycloride (one study), paraquat (one study), and procymidone, epoxicanazole, tebuconazole, and prochloraz 

(two studies). Duration of exposure and dose ranges in female studies (twelve studies) were categorized as: (1) 

15 to 30 before gestational days (lowest dose: 500 mg/kg/day and highest dose: 800 mg/kg/day); (2) 1 to 30 
11

gestational days (lowest dose: 5 mg/kg/day and highest dose: 1,300 mg/kg/day); and (3) postnatal days (lowest 

dose: 6.25 mg/kg/day and highest dose: 500 mg/kg/day). For studies examining only males (n=3), length of 

experiment time ranged from 5 to 360 days, with daily doses ranging from 10 to 1,500 mg/kg. Details are 

below.

Reproductive & developmental endpoints 

Females. Two early experimental studies dating back to the 1970s examined the teratogenic effects of 

mancozeb. Khera investigated prenatal survival and teratogenicity in rats and rabbits.8 Results indicated no 

significant effect on reproduction for female rats exposed to the maximum tolerated dose of ETU before and 

during gestation (80 mg/kg/day). However, significant morphological fetal abnormalities, with the fetal brain 

most commonly affected, were detected at doses showing no maternal toxicity or fetal mortality in rat offspring.

While increased resorption sites and reduced brain weight at 80 mg/kg/day were noted for female rabbits, rabbit 

offspring showed no sensitivity to mancozeb at any dose level and no teratogenic effects on developing brains. 

Larsson et al.9 reported profound internal and skeletal malformations in rat offspring. For the highest level of 

mancozeb exposure (1330 mg/kg/day), the frequency of internal malformations was markedly higher than that 

of external malformations. The low-dose group (380 mg/kg/day) resulted in significant reductions in fetal 

weight for exposed mice. Following a twenty-year gap, a collection of animal studies examined the effect of 

mancozeb exposure on reproductive and pregnancy health in female rats with mixed results. While an earlier 

study10 showed no change in the number of estrous cycles post-mancozeb exposure, several studies documented 

a reduction in the number of estrous cycles, duration of proestrus, estrus, and metestus, healthy follicles and 

number of corpora lutea with greater than 600 mg/kg/day mancozeb treatment (700 mg/kg/day and 800 

mg/kg/day).11-13 A significant increase in thyroid weight was also observed in all rats treated with mancozeb,

except for the group treated with 500 mg/kg/d.11,12 High prenatal doses of mancozeb (500 mg/kg) resulted in 

reduced maturity and fertilizability of oocytes in females of the F1 generation.14 These studies mark the first 

report on the effects of mancozeb on reproductive potential in animals.

Whole animal studies examining prenatal exposure to mancozeb report significant adverse effects. Axelstad et 

al.15 observed toxic effects in rat dams treated with high doses of mancozeb (150 mg/kg/d) including severe 

weight loss and temporary paralysis in hind limbs. Results also showed a dose-dependent decline in T4 levels 

for all dams exposed to mancozeb (50 mg/kg, 100 mg/kg, and 150 mg/kg) on gestation day fifteen. Four 

mixture studies included in the review revealed that prenatal exposure to a low dose pesticide mixture 

containing mancozeb may lead to adverse developmental toxicity including disruptions in the developing mouse 

cerebellar cortex16, long-term delayed effects at dose levels where single pesticides showed no effects17, and 

genital malformations in male rat pups.18

Males. Experimental studies showed significant declines in male reproductive functioning including, 

spermatoxic effects in mice, and biochemical and structural alterations in testes of rats and mice following 

chronic exposure. Khan & Sinha19 observed decreased sperm counts and significantly higher sperm head 

abnormalities in all three groups exposed to mancozeb (3, 6, or 1,000 mg/kg/day). Abnormal spermatic head 

size occurred most frequently in mice treated with mancozeb. A long-term experimental study20 in male rats 

resulted in higher mortality-related acute effects in all groups fed mancozeb. During months 5 through 12, male 

rats demonstrated prolonged reproductive effects at higher doses of mancozeb (1,000 and 1,500 mg/kg/d) 

including a small increase in testes weight and reduction in epididymis size. Animals in the high dose group 

(1,500 mg/kg/d) demonstrated temporal changes in pathology for seminiferous tubules, with pronounced 

damage in epithelial cells of epididymis resulting in reduced sperm count. Ksheerasagar & Kaliwal21 also found 

time-dependent declines in weight of testes and other organs (liver, kidneys, and spleen), as well as, significant 

increases in thyroid and thymus weight of treated mice. 
12

Animal studies provide strong evidence for acute toxicity in mammals and suggest significant changes in 

physiological, biochemical, and pathological processes involved in normal reproduction may lead to infertility 

in males chronically exposed to mancozeb. 

Human Health Studies. Fourteen epidemiologic studies published between 2001 and 2007 met our inclusion 

criteria (Figure 1, Table 3.) The majority of studies included used a case-control (six studies) and crosssectional design (four studies), with the exception of one case study and three cohort studies (one prospective 

study and two retrospective studies). We observed substantial differences in mancozeb exposure assessment. 

Most studies assessed exposure indirectly, using pesticide use questionnaires or database registries (eleven 

studies). Three studies examining Italian vineyard workers obtained individual measures of recent mancozeb 

exposure by measuring urinary metabolite levels of ETU.22-24

Pre-conception. Studies examining the fertility effects of mancozeb or other members of the ethylene 

bisdithiocarbamate (EBDC) group of fungicides, which includes the related active ingredients maneb and 

metiram, were mixed. Farr et al.25 found that women who reported using mancozeb or maneb were four times 

more likely to experience longer menstrual cycles and two times more likely to have a missed a menstrual cycle

compared to unexposed women living on farms in Iowa or North Carolina. In a case study of three women 

working in similar agricultural industries around the same time, two of the women were exposed to mancozeb 

during the first trimester, and all three women gave birth to infants with severe malformations. 

Mancozeb is considered an endocrine disruptor. While Rull et al. observed no apparent effect on neural tube 

defects when mancozeb was applied within 1,000 meters of maternal residences, authors found an increased 

odds ratio of 1.5 and 2.0 of having a baby with a neural tube defect or anencephaly, respectively, among 

pregnant women living within 1,000 meters of where two or more endocrine disrupting chemicals were 

applied.26

Some studies report on the fertility impacts of fungicide exposures generally, but do not identify which 

fungicides the subjects were exposed to. A trend in longer time to pregnancy was observed in female flower 

production workers exposed to fungicides year-round.27 In an early study, Arbuckle et al. observed elevated 

risk of late-term spontaneous abortions among farm couples exposed to fungicides and double the risks of 

spontaneous abortions in older women. 28 A later study demonstrated no difference in birth rate for families of 

fungicide applicators compared to applicators using herbicides and other agrochemicals (2.85 vs 2.80). Garry et 

al.29 examined spouses of pesticide applicators in the Red River Valley and found an association between 

personal use of pesticides and an increased risk of fetal loss. This study also observed an increased frequency of 

miscarriages for summer, fall, and winter combined among spouses of pesticide applicators who worked with 

fungicides.

A modest risk of fetal death during the first and second trimesters, especially the 3rd and 4th month of pregnancy, 

was found in women exposed to thiocarbamates during pregnancy.30 In this study, mancozeb was assigned to 

the carbamate/thiocarbamate pesticide class. One study showed an increased risk of birth defects following 

parental exposure to fungicides during pregnancy, including an increased risk of anencephaly in children of 

mothers working in agriculture during the three months before conception and whose fathers also applied 

pesticides at work.

31

These results provide some evidence that mancozeb is a reproductive toxicant. Mancozeb exposure may 

interfere with normal reproductive functioning of women persistently exposed (either occupationally or 

indirectly by spouse’s occupation) and thereby may delay or complicate pregnancy. Very few population-based 

studies have examined the association between exposure to mancozeb and adverse reproductive or 

developmental outcomes.
13

Figure 1. Flow diagram of study selection process for systematic review. 

Overview of Systematic Review Protocol

(Adapted from National Toxicology Program Office of Health Assessment and Translation approach for 

systematic review and evidence integration from literature-based health assessments) 

1. Draft protocol strategy

a. Discuss literature search strategy

b. Determine criteria for selection of studies relevant to review question

c. Discuss data abstraction components (risk of bias assessment and confidence in evidence)

2. Search strategy

a. Decide on search criteria

b. Search literature

c. Read abstracts and select studies to include in review; obtain full paper

3. Data abstraction

a. Develop and test data abstraction form

b. Extract data & assess quality of study using OHAT criteria (BPA example)

c. Assess confidence in studies

4. Translate confidence ratings into level of evidence for health effect
14

Table 1. In vitro studies (seven studies) examining exposure to Mancozeb and reproductive and developmental outcomes. 

Source Type of 

cell/tissue

Compound (Daily Dose) Incubation Outcomes Confidence*

Lin & Garry, 20001 MCF-7 breast 

cancer cell line

Herbicides:

2,4-D LV4 (0.1-10 ug/ml)

2,4-D Amine (1-10 ug/ml)

2,4-Dichlorophenoxyacetic acid, isooctyl 

ester (1-10 ug/ml)

2,4-Dichlorophenoxyacetic acid 

Roundup (1-10 ug/ml)

Glyphosate (0.228-2.28 ug/ml)

Fungicides:

Triphenyltin acetate 

Mancozeb (reagent grade) (10-50 ug/ml)

Manzate (commercial grade, 75% 

mancozeb) 

Tilt (reagent grade) 

Tilt (commercial grade, 42% 

propiconazole)

Test chemicals 

added within 2 to 4 

hours of incubation 

and cultured for 72 

hours in media

Three fungicides (triphenyltin, 

mancozeb,and Tilt) induced cytotoxicity 

over the concentration range tested (50–

70% decrease in cell number at the 

highest concentration tested)

Triphenyltin (4.1 ug/ml) and mancozeb 

(at 10 and 50 μg/ml) showed an apoptotic 

pattern of cell death between 24 and 

48hrs.

ETU induced apoptosis at a much higher 

concentration (1000 ug/ml).

+++

Bisson et al. 20022 Adrenocortical 

cells of rainbow 

trout

Atrazine (0.0005, 0.05, 5, 500 uM)

Diazinon (0.05, 0.5, 5, 50, and 500 uM)

Endosulfan (0.05, 0.5, 5, 50, and 500 

uM)

Mancozeb (0.5, 5, 50, 500 and 5,000 

uM) 

60 minutes Mancozeb exposed cells showed 

significant effects on cortisol secretion 

and on viability. 

• Significant impairment of 

cortisol secretion and no 

significant reduction of viability

at a concentration of 500 uM. 

• Significant reduction of cortisol 

secretion (no reduction in

viability reported) at 50 uM of 

mancozeb.

• Mancozeb had the highest 

LC50/EC50 ratio*.

*The ratio LC50/ EC50 for mancozeb 

was 16—a concentration about 16-fold 

higher was required to kill 50% of cells 

than the concentration required to impair 

50% of cortisol secretion.

++++
15

Fejes et al. 20023 288 chicken 

embryos

80% mancozeb containing formulation 

(Dithane M-45)

Copper-sulfate (0.1% applied 

concentration)

19 days Mancozeb did not increase 

embroytoxicity or result in an increased 

incidence in developmental 

abnormalities.

Increased embryomortality was observed 

in embryos exposed to both mancozeb 

and copper-sulfate. 

++

Greenlee et al. 20044 Mice embryos Several agrochemicals were tested at

low-doses in the study.

Mancozeb (.003 ug/mL)

96 hours Mancozeb induced developmental injury 

in pre-implementation embryos

Fungicide mixture (including mancozeb) 

reduced development to blastocyst and 

increased apoptosis 

+++

Calviello et al. 20065 Wistar rats:

1) RAT-1 

fibroblasts

2) Peripheral 

blood 

mononucleated 

cells (PBMC) 

Mancozeb and Zineb (concentrations 

ranging from 125 to 500 ng/ml)

1 hour and 4 hours At 1 hour of mancozeb exposure 

(500ng/mL):

• Cells produced dose-dependent 

induction of single strand breaks 

in RAT-1

• Increased levels of markers of 

DNA oxidation and reactive 

oxygen species in RAT-1

• PBMC less responsive than 

RAT-1 to oxidative insult

After 4 hours of exposure, results showed

dose-dependent:

• Oxidative effect and prooxidant 

action of Mancozeb

• Apoptosis in PBMC and RAT-1 

cells

++++

Paro et al. 20126 Mouse granulosa 

cells and human 

granulosa cells 

from women 

undergoing 

assisted 

reproductive 

therapy

Mancozeb (increasing concentrations 

(0.001-1 ug/ml)

1, 24, and 36 hours Mouse granulosa cells exposed to 

mancozeb showed time- and dosedependent changes in morphology, 

acquired ability to migrate, decreased 

level of p53, and no change in apoptosis

Human granulosa cells showed dosedependent changes in morphology and 

reduction in p53 expression postmancozeb exposure. 

++++

Kjeldsen et al. 20137 human breast 

carcinoma 

MVLN cells and

14 pesticides were tested

Mancozeb concentrations tested singly: 

48 hours When tested as a single compound, 

mancozeb exposure showed 

concentration-dependent antiandrogenic 

++++
16

hamster ovary 

CHO-K1 cells

LOEC (M): 1 x 10-5

MOEC (M): 1 x 10-5

% of Solvent control (0.02% DMSO): 45

effects on agonist-induced AR 

transactivity. 

At the lowest tested dose inducing the 

maximum effect (MOEC), mancozeb 

showed the highest inhibitory effect in 

the form of an agonist-reduced response. *

Confidence#ratings#in#body#of#evidence#ratings:#High%confidence (++++)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#highly#likely#to#be

reflected#by#the#apparent#relationship;#Moderate#confidence#(+++)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#may#be reflected#in#the#

apparent#relationship;#Low%confidence (++)in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#likely#to#be different#than#the#apparent#

relationship;#and#Very%low confidence (+)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#highly#likely#to#be different#than#the#apparent#

relationship.##

Table 2. In vivo studies (fifteen studies) examining exposure to Mancozeb and reproductive and developmental outcomes.

Source Experimental 

Animal

n Compound (route) Daily Dose Duration Outcomes Rating*

Khera 19768 Nulliparous adult 

Wistar female rats 

& New Zealand 

White rabbits

209 rats

33 rabbits

Mancozeb (oral) 0, 5, 10, 20, 

40, or 80 

mg/kg/day

Rats: 

Group I: 21-42 

days BG and 

GD 1-15; 

Group II: 6-15 

GD and 

Group III: GD 

7-20

Rabbits: 

30 GDs

Prenatal survival in rats. 

Numbers of corpora lutea, live fetuses, 

and fetal death at all doses were similar 

to controls in all the three experiments. 

Doses of 80 mg/kg in Group 2 and

40 mg/kg in Groups 1 and 3 reduced

the mean fetal weight as compared to

matching control groups.

ETU at the maximum tolerated doses 

administered before and during 

gestation had no significant

effect on any reproductive parameter.

Teratogenicity in rats. 

ETU was teratogenic in rats at doses that 

produced no apparent maternal toxicity 

or fetal lethality. 

Significant morphological fetal 

abnormalities were observed (including 

jaw, tongue, eyelids, short or absent tail 

malformations); fetal brain appeared to 

be the most commonly affected organ.

++++
17

Prenatal survival in rabbits. 

Trend observed for (i) increased 

frequency of resorption sites (including 

dead fetuses), and (ii) reduced brain 

weigh at 80 mg/kg. 

Teratogenicity in rabbits. 

No skeletal abnormalities observed in 

animals treated with mancozeb. Rabbit 

fetal brain was insensitive to teratogenic 

effects at any dose level.

Larsson et al. 19769 Three-month-old 

NMRI mice and 

virgin female 

Sprague-Dawley 

rats

56 mice

56 rats

Maneb, Mancozeb, 

and Propineb (oral)

Mice received 

400, 770, or 

1420 mg/kg

Maneb

380, 730, or 

1330 mg/kg 

Mancozeb

400, 760, or 

2300 mg/kg 

Propineb

18 GDs Mouse: 

No adverse maternal effects or fetal 

effects were observed for the different 

dosages of maneb or mancozeb. 

Fetal weight was not reduced in the 

experimental group compared to control

groups except in the fetuses from

mothers treated with the lowest dose of 

mancozeb on day 9.

Rat: 

Gross malformations occurred in all 

surviving fetuses after treatment with 

the medium and high dosages of maneb 

and in 25% of those exposed to the 

highest dose of mancozeb. 

Superficial hemorrhages were found in 

the 2 highest mancozeb dosage groups. 

Frequency of internal malformations

was extremely high compared with that 

of external malformations for highest 

dose of mancozeb.

+++

Khan & Sinha, 

199619

males of Swiss 

albino mice

56 Endosulfan, 

Phosphamidon, and 

Mancozeb (oral)

Vitamin C (injection)

Pesticides: 

3, 6 or 1000 

mg/kg/day

Vitamin C: 

35 days Significant decreases in mean sperm 

counts in each of the three pesticidetreated groups compared to controls.

All three categories of sperm head 

++++
18

10 (lower 

dose), 20 

(middle dose) 

and 40 mg/kg 

body wt/day 

(higher dose)

abnormalities increased significantly 

(compared to control) in the groups 

treated with each of the three pesticides.

Sperm with abnormal head size were 

most common in the mancozeb-treated 

group.

Kacker et al. 199720 Male albino rats 200 Mancozeb in peanut 

oil (oral)

500, 1,000, 

and 1,500 

mg/kg/day

30, 60, 90, 

180, and 360 

days

Mancozeb at all doses produced signs of 

poisoning (diarrhea, dyspnea, salivation, 

nasal bleeding, hind limb paralysis). 

Mortality was more pronounced during 

days 0-90 as compared to days 90-360.

Higher doses (1,000 and 1,500 mg/kg/d) 

over a period of 180 and 360 days 

produced a slight increase in relative 

weight of testes and a decrease in 

epididymis.

Testes of rats dosed with mancozeb 

(1,500 mg/kg/d) produced time 

dependent pathological changes in 

seminiferous tubules and showed 

damaged epithelial cells in the tubules 

of epididymis with declines in sperm 

count. 

Increased activity of LDH, ALP and 

decreased activity of ACP and SDH in 

testes and epididymis.

++++

Roperto & Galati 

199832

nonmigratory 

pigeons

10 mixture of 30% 

mancozeb and 15% 

copper oxychloride

-- Longterm 

exposure over 

years

Pigeons living in control environment: 

No apparent optical and ultrastructural 

lesions were found in the trachea or 

lungs via histopathology, scanning EM, 

or x-ray. The metallic components of 

the fungicidal mixture (Cu and Zn) were 

not detected in the lungs via 

spectrometer. 

Pigeons living in the exposed 

environment: 

Severity of the histopathological lesions 

+++
19

was dependent on age. 

Intimal proliferation of the intraparabronchial arteries and scattered foci 

of fibrosis were also evident as was 

deposition of Zn and Cu particles 

throughout lungs. 

Castro et al. 199910 Wister rats 120 Mancozeb (mixed 

into diet)

0, 2000 or 

3.000 ppm

Group A: GD 

1-6, 

Group B: GD 

6-15, 

Mancozeb exposure did not interfere 

with estrous cycle in pregnant females, 

maternal weight gain, nor percentage of 

birth viability of pups.

Pups of mancozeb exposed dams 

experienced delays in physical 

development.

Prenatal mancozeb exposure also 

resulted in developmental motor 

alterations in pups. 

+++

Mahadevaswami et 

al. 200013

Female Wistar

virgin rats

36 Mancozeb (oral) 500, 600, 700, 

and 800 

mg/kg/day

15 BG days Treatment with 700 and 800 mg/kg/day 

mancozeb showed a decrease in ovarian 

hypertrophy. 

No significant change in the number of 

estrous cycles and duration of each 

phase of the estrous cycle with 500 

mg/kg/day mancozeb. 

Decreased number of estrous cycles, 

duration of proestrus, estrus, and 

metestrus with significant increase in

the duration of the diestrus phase with > 

600 mg/kg/day mancozeb treatment. 

Decreased number of healthy follicles 

with increase in the number of atretic 

follicles at higher doses of mancozeb. 

No significant changes in the body and 

organ weight with any dose level of 

mancozeb.

++++
20

Treatment with 600, 700, and 800 

mg/kg/day mancozeb showed a 

significant decrease in the levels of 

protein, glycogen, total lipid, 

phospholipid, and neutral lipid in the 

liver, uterus, and ovary, with the 

exception of liver total lipid and uterine 

glycogen.

Mancozeb treatment reduced the

number of healthy follicles with a 

concomitant increase in the number of 

atretic follicles.

Baligar & Kaliwal 

200112

Wister rats (virgin) 40 Mancozeb (oral) 500, 600, 700 

and 800 

mg/kg/d

30 days Significant decrease in the number of 

estrous cycle, duration of proestrus, 

estrus and metestrus with concomitant 

increase in duration of diestrus in all 

mancozeb treated groups.

Significant decrease in the number of 

healthy follicles and increase in atretic 

follicles in all mancozeb treated groups.

Reduced number of corpora lutea and 

size of ovary in high doses of mancozeb 

treated rats.

Significant increase in thyroid weight in 

all mancozeb treated rats except in 500 

mg/kg/d dosage group.

+++

Ksheerasagar &

Kaliwal 200321

Male Swiss

albino mice

Mancozeb (oral) 800 

mg/kg/day

5, 10, 20 and 

30 days

Weight of testes decreased significantly 

with increasing durations of pesticide 

treatment in mice.

Increasing durations of mancozeb 

resulted in a significant decrease in the

weight of liver, kidneys and spleen.

Significant increase in the thyroid and

thymus weight in duration dependent 

++++
21

manner.

Baligar & Kaliwal 

200411

Female virgin 

albino rats

70 Mancozeb 

(75% wettable 

powder) dissolved in 

olive oil (oral)

700 mg/kg/d 5, 10, 20 or 30 

BG days

Mancozeb treatment for 

1) 5 days- significant increase in the 

duration of diestrus and decrease 

in state II and total number of 

healthy follicles.

2) 10 days- significant increase in 

the number of estrous cycles and 

duration of estrous, with increase 

in diestrus; decrease in stages I, 

II, and IV and in total number of 

healthy follicles. 

3) 20 and 30 days-number of estrous 

cycles and duration of proestrus, 

estrus, and metestrus were 

significantly decreased, with 

increase in duration of diestrus; 

increase in atretic follicles and 

thyroid weight

+++

Miranda-Contreras 

et al. 200516

Pregnant NMRI 

mice

160 Paraquat [PQ] and 

Mancozeb [MZ] 

(injection)

Group I: 

saline, 

Group II: PQ 

(10 mg/kg), 

Group III: MZ 

(30 mg/kg) or 

Group IV: the 

combination 

of PQ and MZ 

(10 mg/kg PQ 

+ 30 mg/kg 

MZ)

Pregnant mice: 

GD 1-30

Dams: PD 12 

and PD 20

Body Weight

Second postnatal week: Significant 

reductions in body weight were found in 

mice exposed to PQ + MZ

After 1 month: 

• marked decline (27%) in body 

weight as compared with the 

control group (permanent 

effects of prenatal exposure to 

PQ + MZ on the physical 

growth of the animals). 

• PQ-exposed group showed 

significant declines 21% and 

17% for the MZ-exposed 

animals 

Pups showed significant reductions in 

excitatory amino acid neurotransmitters 

during cerebellar cortex development of 

prenatally exposed mice to chronic 

doses of MZ and PQ + MZ.

++++
22

In MZ-exposed important variations in 

the developmental variation in Gly 

levels compared to control were 

registered: a 36% reduction at P3, a 

150% increment at P15 and a 43% fall 

at P30.

MZ exposed groups showed general 

trend of reduction in inhibitory 

neurotransmitters during cerebellar 

cortex development.

Locomotor activity

Mice prenatally exposed to MZ were 

more active by 3.1 fold compared to 

controls; at P30 locomotor activity was 

significantly lower than controls for all 

three experimental groups. 

Prenatal exposure to either PQ or MZ or 

the combination of both may lead to 

alterations to the chronology and 

magnitude of synaptic transmission in 

developing mouse cerebellar cortex.

Rossi et al. 200614 Swiss CD1 female 

mice

25 Mancozeb in sesame 

oil (oral)

50 and 500 

mg/kg

GD 2 to PD 20 No significant difference in mean 

number of oocytes from F1 generation 

treated with either low (50 mg/kg) or 

high (500 mg/kg) doses of mancozeb 

compared to controls. 

Production of MII-arrested oocytes in

F1 generation was slightly, although 

significantly, reduced in mice exposed 

to high doses (500 mg/kg) of mancozeb; exposure to low doses (50mg/kg) 

was ineffective.

82% of the oocytes derived from control 

or low dose treatment groups formed 

male and female pronuclei 

+++
23

71% of oocytes derived from the high 

dose treatment 

Axelstad et al. 

201115

Wister rats 64 (litters) Mancozeb in corn oil 

(oral)

0, 50, 100, or 

150 mg/kg/d

GD 7 to PD16 Doses of 150 mg/kg bw/day and above 

caused toxic effects in the dosed dams 

(e.g., severe weight loss and transient 

paralysis of the hind limbs) 

T4 levels showed a dose-dependent and 

significant decline in dams from all 

three dose groups on gestational day 15

(offspring T4 levels, thyroid weights 

and histology were unaffected on 

postnatal day 16).

++++

Hass et al. 201218 Time-mated 

nulliparous, young 

adult female Wistar 

rats

198 procymidone, 

epoxiconazole, 

tebuconazole, 

mancozeb, and 

prochloraz (gavage)

procymidone: 

12.5 and 50 

mg/kg/day;

epoxiconazol: 

3.75 and 15 

mg/kg/day;

tebuconazole: 

12.5 and 50 

mg/kg/day;

mancozeb: 

6.25 and 25 

mg/kg/day; 

and 

prochloraz: 

8.75 and 35 

mg/kg/day

GD 7-21, PN 

1-16

No significant effect on gestation length, 

maternal or pup body weight gain for 

rats exposed to mancozeb.

Gestation length was significantly 

increased in the two highest mixture 

groups compared to controls.

Nipple retention in male offspring was 

significantly increased in the groups 

exposed to the mixture and in the groups 

exposed to the high dose of mancozeb, 

tebuconazole and both doses of 

procymidone.

One male in each of the groups dosed 

with mancozeb showed malformation of 

the seminal vesicle.

All pesticides except for mancozeb 

exhibited AR antagonism in vitro.

Combined developmental exposure to 

endocrine disrupting pesticides at dose 

levels below NOAELs for the single 

pesticides caused adverse effects on 

male sexual development and gestation 

length.

++++ 

Jacobsen et al. Time-mated 198 procymidone, Procymidone: Dams were One animal from each of the mancozeb ++++
24

201217 nulliparous, young 

adult female Wistar 

rats

epoxiconazole, 

tebuconazole, 

mancozeb, and 

prochloraz (gavage)

12.5 and 50 

mg/kg/day;

epoxiconazole

: 3.75 and 15 

mg/kg/day;

tebuconazole: 

12.5 and 50 

mg/kg/day;

mancozeb: 

6.25 and 25 

mg/kg/day; 

prochloraz: 

8.75 and 35 

mg/kg/day 

mixture: 

0, 14.6 (8.3% 

of NOAEL), 

29.2 (17% of 

NOAEL) or 

43.8 (25% of 

NOAEL) 

mg/kg/day of 

the mixture of 

the 5 

pesticides

dosed daily by 

gavage, from 

GD 7 to pup 

day (PD) 16

groups and one animal from the low 

dose mixture group had more than 

double thyroid weight com- pared to 

controls.

Significant increase in the number of 

animals with follicles dominated by 

columnar to cuboidal epithelium was 

seen in mancozeb exposed animals 

compared to the control group (observed 

in 4 of 9 animals from the high dose 

mancozeb group compared to 1 of 17 

control animals).

Significantly higher activity levels were 

observed in females from the high dose 

mancozeb group. 

**Note: Lowest dose of each pesticide 

was similar to the dose included in the 

highest mixture dose and then highest 

dose of the single pesticides was 4 times 

higher, corresponding to 25% of 

NOAEL 

BG=Before#Gestation,#GD=Gestation#day,#PD=Postnatal#day,#mg/kg/d=#mg/kg#body#weight/day,#NF=not#found *

Confidence#ratings#in#body#of#evidence#ratings:#High%confidence (++++)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#highly#likely#to#be

reflected#by#the#apparent#relationship;#Moderate#confidence#(+++)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#may#be reflected#in#the#

apparent#relationship;#Low%confidence (++)in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#likely#to#be different#than#the#apparent#

relationship;#and#Very%low confidence (+)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#highly#likely#to#be different#than#the#apparent#

relationship.##
25

Table 3. Epidemiologic studies (fourteen studies) examining exposure to Mancozeb and reproductive and developmental outcomes. 

¶

Design—Co: Cohort; RCo: Retrospective Cohort; CaCo: Case-Control; CrSe: Cross-sectional; and CaS: Case Series/Case report.

Source Design¶ Population Mancozeb Assessment Outcome Confidence*

Arbuckle et al., 

200128

RCo Farm couples in 

the Ontario Family 

Health Study 

(N=2,110 women)

Pooled pesticide exposure 

information from the farm 

operator, husband, and wife to 

construct a history of monthly 

agricultural and residential 

pesticide use. 

Four pesticide classes: 

1) Herbicide, 2) Insecticide, 

3) Fungicide and 

4) Miscellaneous. 

**Note: Mancozeb was not 

directly mentioned or 

measured in the study

Preconception exposure to thiocarbamates (OR = 1.8; 95% CI, 

1.1–3.0), glyphosate (OR = 1.7; 95% CI, 1.0–2.9), fungicides (OR 

= 1.4; 95% CI, 0.9–2.1), and the miscellaneous class of pesticides 

(OR = 1.5; 95% CI, 1.0–2.4) were associated with elevated risks 

of late spontaneous abortions (12-19 weeks).

Exposure to both fungicides and herbicides before conception 

doubled the risk relative of spontaneous abortions compared to 

woman exposed only to fungicides (OR = 2.0; 95% CI, 1.1–3.5). 

Exposure to fungicides doubled the risk of having a spontaneous 

abortion compared to those not exposed (OR = 2.4; 95% CI, 1.0–

5.9) in older women (age 35+). No increased risk was observed in 

younger women.

+++

Bell et al., 200130 CaCo Cases=319 

neonatal deaths 

within 24 hours of 

birth and controls= 

611 normal live 

births in 10 

California counties 

in 1984. 

County maps to locate the 

township, range, and section 

for each maternal address 

(maternal TRS) was 

determined by using the public 

land survey system. Pesticide 

exposure was determined by 

linking the maternal TRS to 

the township, range, and 

section of each pesticide 

application located in the 

California Pesticide Use 

Report database.

Five categories were selected 

for analysis: phosphates, 

carbamates, pyrethroids, 

halogenated hydro-carbons, 

and endocrine disruptors. 

Mancozeb was assigned to the 

Carbamates/thiocarbamates 

class.

Pesticide classes were not strongly associated with fetal death,

regardless of trimester of exposure. 

Weakly elevated risks were observed for applications of 

halogenated hydrocarbons, carbamates/thiocarbamates, estrogenic 

pesticides, and carbamate acetylcholinesterase inhibitors during 

the second trimester, with HRs of 1.3 (95 percent confidence 

interval (CI): 1.0, 1.8), 1.3 (95 percent CI: 1.0, 1.8), 1.4 (95 

percent CI: 0.8, 2.5), and 1.3 (95 percent CI: 1.0,1.8), respectively.

Modestly increased risk of fetal death was observed when 

exposure to halogenated hydrocarbons occurred during the fourth 

and fifth months of gestation, with HRs of 1.4 (95 percent CI: 1.0, 

2.0) for both months. Results were similar for exposure to 

carbamates and carbamate inhibitors during the third and fourth 

months of gestation.

Slightly elevated HRs were observed for exposure to carbamates 

during the first and second trimesters of gestation, the monthly 

analysis showed that risk increased only for exposure during the 

third and fourth months of gestation.

++

Colosio et al., 

200222

CaCo 

(pilot)

Italian vineyard 

workers (N=13) 

First urine spot sample 

collected in the morning preUrinary ETU was significantly increased at the end of shift (2.5, 0.5–95.2 mg/g creatinine) in workers compared with baseline 

++
26

and controls 

(N=13)

mancozeb application. 

Dermal exposure assessment 

captured morning in which 

mancozeb mixture was 

prepared and applied by 

workers (pads were placed on 

clothes and skin)

* assessment of the exposure 

was performed in the absence 

of previous applications of 

mancozeb and/or other 

pesticides

levels. 

End-shift urinary ETU levels were higher in operators using open 

tractors (n=7) than in those using closed tractors (n=5) (16.2 vs. 

2.4 mg/g creatinine), but not statistically significant. 

End-shift urinary ETU was positively correlated with dermal 

exposure to mancozeb determined both over the clothes and on the 

skin.

Garry et al., 

200229

CrSe 1,070 licensed 

pesticide 

applicators 

(males), 851 were 

married or had a 

marriage-like 

relationship, 

residing in five 

counties in Red 

River Valley, 

Minnesota, USA

Pesticide use 

Each certified pesticide 

applicator was initially interviewed by phone to obtain 

current and past pesticide use 

with specific attention to 

product name and the number 

of days per year applied. 

6 months follow-up 

questionnaire to document 

common pesticide use by 

pesticide class, acreage 

treated, type of crop, and use 

of personal protective gear. 

Overlap between the two 

questionnaires was intentional 

to validate use of pesticides by 

class (herbicides, insecticides, 

fumigants, and fungicides).

Male Pesticide Use, Fetal Loss, and Miscarriages

*Sample: 540 farm families with children, where the applicator 

was the biologic father or whose spouse had been pregnant in the 

relationship, participated and gave detailed pesticide use data.

Among couples who had children or who had been pregnant, one 

in four women had a miscarriage. 

Spouses of pesticide applicators who reported use of herbicides, 

insecticides, and fungicides had more miscarriages than any other 

pesticide application group.

Specific Fungicide Use

The unadjusted odds ratio for specific use of organotin (OR = 

1.55; CI 1.01–2.37) and/or use of EBDC (ethylene 

bisdithiocarbamate) fungicides (OR = 1.77; CI 1.11–2.83) such as 

mancozeb demonstrates a significantly increased frequency of 

fetal loss compared to nonuse of fungicides by applicators.

Maternal Pesticide Exposure

*Sample: Only 36 of 379 women respondents applied or mixed 

pesticides. 

Personal use of pesticides, (e.g., mixing, loading and pesticide 

applications) by the female spouse, was a significant risk factor for 

fetal loss (fetal loss/pregnancy OR = 1.81; CI 1.04–3.12). 

30% had a fetal loss compared to 21% of those women who did 

not mix or apply pesticides. 

The number of fetal losses per pregnancy is significantly elevated 

+++
27

in those who did mix or apply pesticides (15% vs. 8.9%).

Miscarriage and Season

# of first trimester miscarriages in spring was significantly 

elevated compared to all other seasons. 

Spouses of applicators who applied fungicides and other pesticides 

showed no peak in the miscarriage frequency in spring. 

Frequency of first-trimester miscarriages was significantly higher 

in summer, fall, and winter combined for the spouses of 

applicators of fungicides compared to applicator families whose 

male spouse did not apply fungicides.

Significant shift in the sex ratio of children born to the spouse of 

appliers who applied fungicides.

• 25% reduction in male births in the children of 

applicators who applied herbicides, insecticides, and 

fungicides or among those applicators who applied

mancozeb and/or triphenyltin fungicides was significant.

Garry et al., 

200233

CrSe Families of 

licensed pesticide 

applicators in Red 

River Valley, 

Minnesota, USA

(N=536 couples)

Phone survey on current and 

past pesticide use in 

agriculture with 6 month 

follow-up interview to validate 

pesticide class. 

Class of pesticide

Herbicides 

 Chlorophenoxy 

 Oxyphenoxy 

 Sulfonylurea 

 Carbanilate 

 Bromophenol 

 Benzothiazole 

 Nitroaniline

Insecticides 

 Organophosphate 

 Synthetic pyrethroids 

 Carbamate

Fungicides

 Organotin

 Ethylene bisdithiocarbamate

 Triazole 

In families whose male partner did not apply fungicides, increased 

frequency of males born compared to men who applied fungicide 

products. 

Increased frequency of male children born with birth defects than 

female children (M/F sex ratio = 1.8) when no fungicides were

applied. 

If fungicides were applied by the male partner, far fewer male 

children with birth defects were born (M/F sex ratio = 0.57, p = 

0.02). 

No difference observed in birth rate for families of fungicide 

applicators compared to applicators of herbicides and other 

products (2.85 vs. 2.80 children per family).

++
28

Fumigants

 Phosphide

 Other

Sallmen et al., 

200334

Co Finnish 

greenhouse 

worker: 

employers 

(N=419) and 

employees

(N=14,733)

Exposure to pesticides was 

assessed on the basis of 

questionnaire information and 

data gathered from the 

employers 

Reduced fecundability in exposed greenhouse workers not wearing 

proper personal protection equipment (FDR 0.67 95%CI: 0.33-

1.35).

0.92 (85%CI: 0.45-1.88) and 0.77 (95%CI 0.46-1.29) for high 

exposure, moderate, and low exposure groups compared to 

unexposed greenhouse workers.

Fertility was comparable between exposed workers who used PPE 

and unexposed greenhouse workers

++

Farr et al., 200425 CrSe Women living on 

farms in Iowa and 

North Carolina 

(N=3,103)

pesticide exposure questions 

assessed lifetime use of any 

pesticides and hormonally 

active pesticides

women were asked, in their 

lifetime, had they ever mixed 

or applied any of 50 different 

pesticides

Carbamate pesticides use associated with increased odds of long 

menstrual cycles (OR = 2.1) and decreased odds of irregular cycles 

(OR = 0.38).

Women who used mancozeb or maneb had four times the odds of 

experiencing long cycles and two times the odds of missed periods 

as women who had never used pesticides. 

No other outcomes were associated with the use of mancozeb or 

maneb.

A subanalysis excluding female licensed applicators and spouses 

who had not likely used hormonally active pesticides in the last 

year results showed elevated odds of long cycles, missed periods, 

and intermenstrual bleeding among women exposed to lindane, 

atrazine, or mancozeb or maneb compared to unexposed women.

*Models adjusted for age, body mass index, and current smoking 

status.

++

Corsini et al., 

200524

CaCo 13 Vineyard 

workers and 13 

matched healthy 

controls in 

Northern Italy

Workers: urine and blood 

samples were collected before 

the beginning of the spraying 

season, prior to any exposure 

to pesticides, and at the end of 

the work shift of the last day 

of Mancozeb application.

Controls: urine and blood 

samples were collected only 

Urinary ETU metabolite results:

Significant increase in urine ETU concentration among applicators 

in post-exposure compared to baseline values and controls 

samples.

No difference observed in baseline levels of urine ETU in workers 

or controls.

Blood cell counts:

Significant increase in CD19+ cells and decrease in CD25+ cells 

+++
29

once in the morning at the 

same time of the year of 

workers.

were detected in follow-up blood samples compared to controls. 

Significant reduction in eosinophil and serum IgE in workers postexposure.

Significant increase in the proliferative response to phorbol 

myristate acetate plus ionomycin (PMA + ionomycin) was 

observed in the post-exposure group compared to controls and 

baseline.

Significant decrease observed in LPS-induced TNF-a release postexposure in workers.

Idrovo et al., 

200527

CrSe Female flower 

production 

workers in Bogota, 

Colombia 

(N=2,085).

Exposure matrix constructed 

based on work in flowers and 

time in agriculture; exposure 

was categorized as low, 

medium, and high. 

*Exposure to pesticides is 

year-round and fungicides 

(especially dithiocarbamates 

such as propineb and 

mancozeb) are more 

commonly used for 1 out of 5 

industries. An estimated 5.7% 

of industries used mancozeb.

A trend in longer time to pregnancy (TTP) was observed in 

women engaged in any type of waged work. 

Positive dose-response relationship between duration of work in 

flower production and TTP.

++

Nordby et al., 

200535

RCo National registers 

in Norway, 

identifying female

(n=105 403) and 

male farmers (n= 

131,243) and their 

children (n=300, 

805) born in 1925-

1971.

Data on farm production and 

fungal forecasts (humid and 

temperate weather conditions) 

in 1973–1990, obtained from

agricultural censuses and 

meteorological measurement 

stations, respectively, served 

as the mancozeb exposure 

indicators.

Neural tube defects (131 cases, prevalence 12.8/10 000 births) was 

moderately associated with potato cultivation (PR 1.6, 95% CI 

1.1–2.3) and paternal work of >500 hours/year (PR 1.6, 95% CI 

1.1–2.5).

319 thyroid cancer cases were identified: 

 1) 141 in female farmers (incidence 10.2/100 000 person years),

 2) 79 in male farmers (incidence 3.2/100 000 person-years),

 3) 99 in offspring (female and male incidence 3.4 and 0.6/100 

000 person-years). 

Mancozeb exposure was not associated with thyroid cancer.

+++

Lascana et al., 

200631

CaCo

(paired)

151 cases of 

anencephaly of > 

Questionnaire obtaining 

information on: 

Children of mothers who worked in agriculture in the ARP (acute 

risk period), that is periconceptional period, had a greater risk of 

+++
30

20 wks’ gestation 

151 controls born 

in three Mexican 

states

1) lifetime occupational 

activities (start/end dates of 

job); 

2) time involved in 

agricultural work, application 

of pesticides, and/or other 

activities with potential 

exposure to these substances 

(manufacture, formulation, 

sale, or distribution of 

pesticides; work in fruit or 

vegetable warehouses; pest 

control work or gardening);

3) direct handling of pesticides 

and time periods involved.

.

Work in agriculture was 

categorized as nonagricultural, 

agricultural, and applicators. 

anencephaly (OR = 4.57, 95% CI 1.05 to 19.96). 

Risk of fathers having a child with anencephaly was greater in

those who applied pesticides irrespective of whether it was done in 

the periconceptonal or the non-acute risk period (OR = 2.50, 

95%CI 0.73 to 8.64; and OR= 2.03, 95% CI 0.58 to 7.08, 

respectively).

Pesticides most used by the parents of the cases in decreasing 

order of frequency were: permethrin (pyrethroid), methamidophos 

(organophosphate), methyl parathion (organophosphate), atrazine 

(triazine), 2,4-dichlorophenoxyacetic acid (chlorinated phenoxy), 

chlorpyrifos (organophosphate), mancozeb (dithiocarbamate), 

picloram (pyridine), dimethoate (organophosphate), and 

carbofuran (carbamate); pesticides most used by the parents of the 

controls in decreasing order of frequency were: methyl parathion 

(organophosphate), methamidophos (organophosphate), 2,4-

dichloro- phenoxyacetic acid (chlorinated phenoxy), chlorpyrifos 

(organophosphate), and monocrotophos (organophosphate).

Rull et al., 200626 CaCo Infants with neural 

tube defects 

(NTDs) (n=928) 

and normal formed 

controls (n=1263) 

delivered in 

California between 

1987 and 1991

Potential exposures to specific 

restricted-use agricultural 

pesticides were evaluated by 

using a geographic metric 

based on linking pesticide-use 

reports from the California 

Department of Pesticide 

Regulation with land-use 

survey maps of crops from the 

Public Land Survey System. 

A physiochemical category for 

Dithiocarbamate and pesticide 

category for Mancozeb were 

included in the analysis. 

Mancozeb was classified as an 

endocrine disruptor.

Effect estimates for mancozeb applied within 1,000 meters of 

maternal residences on neural tube defects, California, 1987-1991

Cases: 6/731

Controls: 11/940

Conventional logistic regression:

1) Single-pesticide models:

OR: 0.7 95%CI: 0.2, 1.9

2) Multiple-pesticide model:

OR: 0.5 95%CI: 0.1, 2.3

Hierarchical logistic regression:

1) Multiple-pesticide model:

OR: 0.5 95%CI: 0.2, 1.4

Exposure to any endocrine disruptors applied within 1,000 m of 

maternal residences resulted in an increased odds of 1.5 (95%CI: 

1.2, 2.0) for any NTDs compared to no exposure of any endocrine 

disruptors. 

The effect of exposure to 2+ endocrine disruptors applied within 

1,000 meters of maternal residences resulted in 2 times the odds of 

having a baby born with anencephaly compared to babies in the no 

++
31

exposure group (95%CI: 1.3, 3.0). 

Calvert et al., 

200736

CaS Three mothers and 

their infants born 

with congenital 

abnormalities 

within 8 weeks of 

one another and 

whose mother 

worked for the 

same tomato 

grower

Exposure information was 

obtained from North Carolina 

Department of Agricultural 

and Consumer Services 

(NCDACS) and the Florida 

Department of Agriculture and 

Consumer Services 

(FLDACS), which obtained 

pesticide application and 

worker assignment records 

from the grower.

Maternal Exposure History 

Case 1: The period for limb development is 24–36 days after 

fertilization. During this period, this child’s mother worked in 

violation of the REI for up to 4 days involving exposure to several 

pesticides, including mancozeb. 

Case 2: During gestational days 14–57, this child’s mother worked 

in violation of the REI for up to 8 days. On seven of these days, 

the pesticides applied to the fields where the mother worked 

included methamidophos. In addition, on gestational days 7 and 

10, the mother worked in fields when an REI was possibly in 

effect (mancozeb on both days, and abamectin and 

methylpyrrolidone on day 7). The mother has three other living 

children, none of whom are known to have birth defects. This 

mother also had one previous stillbirth but without obvious birth 

defects.

Case 3: During gestational days 14–59, the mother worked in 

violation of REIs for up to 10 days. On eight of these days, the 

REI for methamidophos was in effect on some of the fields where 

the mother worked. Abamectin and methylpyrrolidone were 

applied to some of the fields on two other days, but the mother 

may have worked in those fields before the applications were 

made. The mother had two previous pregnancies. One pregnancy 3 

years earlier involved a malformed fetus and ended in miscarriage. 

The mother could not recall her employment or whether she had 

any toxic exposures during that pregnancy. The other previous 

pregnancy resulted in a normal child.

Birth Outcomes

Case 1: This infant was born with tetraamelia (absence of all four 

limbs). 

Case 2: This infant was born with mild Pierre Robin syndrome 

(micrognathia, high arched palate, and mild persistent palatine 

rugae). 

Case 3: This infant had multiple severe malformations including 

cleft lip and palate, imperforate anus, solitary kidney, vertebral 

anomalies, dysplastic low-set ears, and ambiguous genitalia.

Cases 1 and 2 were potentially exposed to mancozeb during the 

+
32

maximal sensitivity period. 

There is evidence to suggest that each mother was exposed to 

pesticides during the maximal sensitivity period for the organ 

system/structure that was affected.

During the period of organogenesis (approximately 14–59 days 

after fertilization) when birth defects are most likely to occur, all 

three mothers unknowingly worked in tomato fields that were 

under an REI because the fields were recently treated with 

pesticides.

Colosio et al., 

200723

CaCo Ninety-three 

subjects entered 

the study: 48 

vineyard workers 

intermittently 

exposed to 

mancozeb (14 

from Pavia and 34 

from Trento) and 

45 control subjects 

(11 from Pavia and 

34 from Trento) 

not exposed to 

known sources of 

pesticides in 

Northern Italy. 

Recent exposure to mancozeb 

was assessed by measuring 

urinary ETU. 

Immunotoxic effects of 

mancozeb were studied using 

several blood cellular and 

serum parameters: complete 

and differential blood cell 

counts, lymphocyte 

subpopulations,

aglycoproteins, erythrocyte 

sedimentation rate (ESR), 

complement 3 and 4 fractions 

(C3, C4), antinuclear (ANA), 

antismooth muscle (SMA) and 

antimitochondrial (AMA) 

autoantibodies, and serum 

immunoglobulins (IgG1, 

IgG4, IgM, IgA, and total and 

specific IgE to most common 

allergens). 

Three time points: 

1) T0 baseline—before start of 

seasonal use of mancozeb; 

2) T30-- exposure to 

mancozeb for 30 days; and 

3) T45-- 40-45 days. Only 

time point 1 (T0) and 3 (T45) 

were obtained for controls. 

Urinary ETU

T0: A low but detectable amount of ETU observed in vineyard 

workers and controls at baseline. 

T30: Significant increase of urinary ETU in agricultural workers 

following exposure confirmed that absorption of mancozeb took 

place during application; 

Postexposure urinary ETU exceeded the reference value of 5 mg/

g creatinine.

Blood

T0: Higher prevalence of cold or flu symptoms, lower percentage 

of monocytes, higher absolute count of T lymphocytes, CD4 and 

natural killer cells, and lower plasma levels of IgA and IgM in 

workers. These differences did not continue at T30 or T45. 

At T0, the agricultural workers showed lower plasma levels of IgA 

and IgM in comparison with controls. 

Within agricultural workers a significant increase of serum 

concentrations of IgA and IgM was observed after exposure. 

No biologically relevant differences were observed between 

groups (farmworker vs control) or at different times in other serum 

parameters. 

No statistically significant differences were observed in 

farmworker compared to control groups in the total or specific IgE 

levels.

+++
33

¶Design—Co:#Cohort; RCo:#Retrospective#Cohort; CaCo:#CaseYControl;#CrSe:#CrossYsectional;#and#CaS:#Case#Series/Case#report.

*Confidence#ratings#in#body#of#evidence#ratings:#High%confidence (++++)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#highly#likely#to#be

reflected#by#the#apparent#relationship;#Moderate#confidence#(+++)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#may#be reflected#in#the#

apparent#relationship;#Low%confidence (++)in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#likely#to#be different#than#the#apparent#

relationship;#and#Very%low confidence (+)#in#the#association#between#exposure#to#mancozeb#and#the#outcome.#The#true#effect#is#highly#likely#to#be different#than#the#apparent#

relationship.##
34

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