Mutagenesis, Vol. 18, No. 2, 201-205,
March 2003
© 2003 UK Environmental Mutagen Society/Oxford University Press
Evaluation of genetic damage in workers employed in pesticide production utilizing the Comet assay
Toxicology Unit, Biology Division, Indian Institute of Chemical Technology, Hyderabad 500 007, Andhra Pradesh, India, 1 Deccan College of Medical Sciences and Owaisi Hospital and Research Centre, Hyderabad, Andhra Pradesh, India and 2 Department of Genetics, Osmania University, Hyderabad 500 007, Andhra Pradesh, India
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and methodsResultsDiscussionReferences |
|---|
The use of pesticides has been increasing in recent years, resulting in the need for increased production of pesticides. However, some pesticides may represent a hazard to human health, especially by causing cancer. Genotoxicity tests form an important part of cancer research and risk assessment of potential carcinogens. Therefore, in the current study the potential DNA damage associated with exposure to pesticides of Indian pesticide production workers was assessed using the single cell gel electrophoresis assay or Comet assay. Blood leukocytes of a group of 54 pesticide workers and an equal number of control subjects were examined for genotoxicity in this study. The two groups had similar mean ages and smoking prevalences. The mean comet tail length was used to measure DNA damage. The exposed workers had significantly greater mean comet tail lengths than those of controls (mean ± SD 19.17 ± 2.467 versus 8.938 ± 2.889, P < 0.001). Smokers had significantly larger mean tail lengths than non-smokers (19.75 ± 2.52 versus 18.26 ± 2.13, P = 0.024). Analysis of covariance showed that occupational exposure (P < 0.05) and smoking (P < 0.05) had significant effects on mean tail length, whereas age and gender had no effect on DNA damage. The present study suggests that occupational exposure to pesticides and smoking can cause DNA damage. This investigation confirms the sensitivity of the Comet assay.
|
Introduction |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Pesticides have been a boon, especially to the developing nations in their efforts to eradicate insect-borne endemic diseases, to produce adequate food and to protect forests, plantations and fibers. Nevertheless, many pesticides are a source of potential hazard to the environment and non-target organisms. A large number of epidemiological studies on cancer risk in farmers have produced conflicting results. However, meta-analyses and reviews are available in the scientific literature reporting increased risk of tumors such as leukemia and multiple myeloma (Daniels et al., 1997
; Khuder and Mutgi, 1997; Zahm et al., 1997; Viel et al., 1998). Hence, the undesirable health effects caused by pesticides in humans are of special concern. Among these are their genotoxic effects, including cancer and several other genetic diseases (IARC, 1991). Genetic biomonitoring of populations exposed to potential carcinogens is an early warning system for genetic diseases or cancer. It also allows identification of risk factors at a time when control measures could still be implemented (Kassie et al., 2000). Many approaches and techniques have been developed for monitoring human populations that have been exposed to environmental genotoxins (Hulka et al., 1990). Human biomonitoring can be performed using different genetic markers (Legator and Au, 1994). Cytogenetic markers such as chromosomal aberrations (CA), micronuclei (MN) and sister chromatid exchanges (SCEs) are among the most extensively used markers of genotoxic effects of pesticides.
Several researchers have used cytogenetic assays to evaluate the potential genotoxicity of pesticide exposures in occupationally exposed populations from various countries (Nehez et al., 1981; Dulout et al., 1985; Rupa et al., 1988; Carbonell et al., 1993; Joksic et al., 1997; Davies et al., 1998; Gomez-Arroyo et al., 2000; Zeljezic and Garaj-Vrhovac, 2002). However, there are reports on positive genotoxic effects in populations exposed to pesticides (Paldy et al., 1987; De Ferrari et al., 1991; Bolognesi et al., 1993; Kourakis et al., 1996; Garaj-Vrhovac and Zeljezic, 1999; Antonucci and Colus, 2000) as well as negative findings (Carbonell et al., 1990; Gomez Arroyo et al., 1992; Hoyos et al., 1996; Scarpato et al., 1996; Lucero et al., 2000; Pastor et al., 2001a,b). These conflicting results obtained in the biomonitoring of pesticide-exposed groups could reflect heterogeneity of pesticide exposure (e.g. farm workers, floriculturists, pesticide applicators and pesticide manufacturing workers). The contradictory data may also be due to differences in the pesticides used, in the protective measures employed or in the cytogenetic end-point evaluated. Thus, data from one study in one particular occupational setting cannot be used to draw conclusions on genetic risk in another occupational setting. This justifies this study (and other studies as well), despite the availability in the literature of a large number of studies of this kind (but on different populations, with different exposures).
During the last few years the alkaline single cell gel electrophoresis (SCGE) assay, also known as the Comet assay, has increasingly been used in human biomonitoring studies. This assay is a rapid and sensitive tool to demonstrate the damaging effects of different compounds on DNA at the individual cell level. Cells with damaged DNA display increased migration of DNA fragments from the nucleus, generating a comet shape (Singh et al., 1988; Fairbairn et al., 1995). Despite the fact that the Comet assay has been used in several occupational studies, only a limited number of molecular epidemiological studies have applied this assay to evaluate the genotoxic effect of pesticides in human populations (Lebailly et al., 1998a,b; Moretti et al., 2000; Garaj-Vrhovac and Zeljezic, 2000, 2001; Zeljezic and Garaj-Vrhovac, 2001).
All populations have a degree of risk in relation to pesticide exposures from agricultural and non-agricultural use and through residues in food. However, workers employed in pesticide production and farmers who use pesticides have a higher risk of exposure and hence are more prone to the potential health effects of pesticides. Investigations that have examined the genotoxic effects in pesticide industry workers are scarce (Kiraly et al., 1979; Amr, 1999; Padmavathi et al., 2000; Garaj-Vrhovac and Zeljezic, 1999, 2000, 2001; Zeljezic and Garaj-Vrhovac, 2001, 2002). Pesticide production in India is a year-round activity. Pesticide industry workers log in 8 h/day, 6 days/week. These workers are constantly exposed to a variety of pesticides. Reports on genetic damage in populations exposed to pesticides from India are scanty (Rita et al., 1987; Rupa et al., 1988, 1989a, Rupa et al., b,c, 1991; Padmavathi et al., 2000). The aim of this study was risk characterization of pesticide exposure in workers employed in a pesticide manufacturing unit. The pesticides being produced were organophosphates, carbamates and pyrethroids. Hence, in the current study a group of pesticide production workers from Hyderabad, India, were evaluated for DNA damage in blood leukocytes using the Comet assay.
|
Materials and methods |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
Study population
The study involved 108 subjects divided into two groups. The first group consisted of 54 workers employed in a pesticide manufacturing unit located in Hyderabad, India. The average duration of their employment in pesticide production was 8.57 years (range 3–13 years). During production all subjects were simultaneously exposed to a complex mixture of pesticides (acephate, chloropyriphos, phorate, fenvalerate, cypermethrin, monocrotophos, dimethoate and carbendazim and a wide range of formulations). The control group (54 subjects) was selected from the general population with no history of occupational exposure to pesticides or any particular environmental agent. The selection criteria for study persons was based on a questionnaire. All subjects were asked to complete a face-to-face questionnaire which included standard demographic data (age, gender, etc.) as well as medical (exposure to X-rays, vaccinations, medication, etc.), lifestyle (smoking, coffee, alcohol, diet, etc.) and occupational questions (hours per day working, years of exposure, use of protective measures, etc.). The exposed group was identified and selected by a medical doctor from an initial 160 workforce. Only those subjects who had worked for at least 3 years in the pesticide production unit were considered eligible. It was ensured that the pesticide-exposed workers and the controls did not markedly differ from each other except for occupational exposure. It was also ensured that the exposed subjects and the controls had not been taking any medicines nor had they been exposed to any kind of radiation for 12 months before blood sampling. The subjects who smoked >5 cigarettes/day at least for 1 year were considered smokers, in both the study and control groups. The samples were collected in winter between October and December 2001. Table I
shows the main characteristics of both groups. The study was approved by the local ethical committee. Informed consent was obtained from each individual prior to the beginning of the study.
|
DNA damage analysis using the Comet assay
For the study of DNA damage, blood samples were collected from all the study and control group subjects. A total of 40 µl of blood was taken from a finger prick into a heparinized glass capillary for the Comet assay, which was carried out according toSingh et al.(1988)
, with slight modifications. To avoid possible bias, the blood samples were coded. The samples were transported on ice to the laboratory and were processed within 2 h. Cell viability determined by the trypan blue exclusion technique (Pool Zobel et al., 1994) ranged from 92 to 96% (data not shown). Slides were prepared in duplicate per subject. Fully frosted microscope slides were covered with 140 µl of 0.75% normal melting point agarose (40–42°C). After application of a coverslip the slides were allowed to gel at 4°C for 10 min. An aliquot of 20 µl of whole blood was then added to 0.5% of 110 µl of low melting agarose (37°C). After carefully removing the coverslips a second layer of 110 µl of sample mixture was pipetted onto the pre-coated slides and allowed to solidify at 4°C for 10 min, with the coverslips in place. The coverslips were removed and a third layer of 110 µl of low melting agarose was pipetted onto the slides and allowed to gel at 4°C for 10 min. The slides (without coverslips) were immersed in freshly prepared, cold lysing solution (2.5 M NaCl, 100 mM Na2EDTA, 10 mM Tris–HCl, pH 10, 1% sodium N-lauroyl sarcosinate, 1% Triton X-100 and 10% DMSO, with the DMSO added just before use) and refrigerated overnight. Slides were then placed in alkaline buffer (300 mM NaOH and 1 mM EDTA, pH 13) for 20 min to allow unwinding of the DNA. Electrophoresis was conducted for 25 min at 25 V (0.66 V/cm) adjusted to 300 mA by raising or lowering the buffer level in the tank. Slides were then drained, placed on a tray and washed slowly with three changes for 5 min each of neutralization buffer (0.4 M Tris–HCl, pH 7.5). DNA was precipitated and slides were dehydrated in absolute methanol for 10 min and were left at room temperature to dry. The whole procedure was carried out in dim light to minimize artefactual DNA damage. Slides were stained with 50 µl of ethidium bromide (20 µg/ml) and viewed under a fluorescence microscope. Analysis was performed using a 400x objective with a Leica optiphase microscope equipped with an excitation filter of 515–560 nm and a barrier filter of 590 nm. Slides were randomized and coded to blind the scorer. All slides were scored by one person, to avoid inter-scorer variability.
A total of 50 individual cells were screened per subject (25 cells from each slide). Undamaged cells have an intact nucleus without a tail and damaged cells have the appearance of a comet. The length of DNA migration in the comet tail, which is an estimate of DNA damage, was measured using an ocular micrometer. Quantification of the DNA damage for each cell was calculated as: comet tail length (µm) = (maximum total length) – (head diameter).
Statistical analysis
Analysis was carried out using MINITAB release 11.21 for Windows. Because the distribution of DNA mean tail length did not significantly differ from the normal distribution, untransformed data were used for simple or multifactor analysis of variance (t-test, 
2 test and ANCOVA).
|
Results |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
The effect of occupational exposure to pesticides on the levels of DNA damage in leukocytes of pesticide production workers and control subjects was assessed by the Comet assay. Table I
represents the distribution of subjects with respect to sex, age, smoking and years of exposure. The mean age of the subjects in the control group and in the group exposed to pesticides was similar (33.35 ± 5.47 versus 35.07 ± 5.20, P = 0.097). The gender distribution and smoking habit were also similar (Table I). The mean comet lengths in leukocytes of exposed workers and the control subjects using the Comet assay are summarized in Tables II and III.
|
|
The exposed workers had a significantly greater mean DNA tail length than the controls (P = 0.001). The smokers had a slightly greater mean tail length than the non-smokers (P < 0.024). Although those who were older had a longer mean tail length than those who were younger and men had a longer mean tail length than women, the differences were not significant in pesticide-exposed workers. A slight difference was observed between workers with <10 and those with
10 years exposure (18.28 ± 2.21 versus 20.69 ± 2.15, P = 0.003) (Table II). Age and gender showed significant effects on DNA in control persons only (Table III). Table IV reveals a significant effect of occupational exposure on mean comet tail length in smoking and non-smoking exposed workers when compared with the controls (P = 0.001). Note that age and gender are not taken into account in Table IV. The results of ANCOVA when age was included as a covariate are summarized in Table V. The effects of smoking and occupational pesticide exposure on DNA damage were quite significant (P < 0.05). Age and gender showed no significant effect on mean comet tail length.
|
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
The aim of this study was to evaluate the genotoxic damage in workers employed in pesticide production in India. The investigation was conducted utilizing the Comet assay. The results of this study indicate that occupational exposure to a mixture of pesticides induces a significant increase in the level of DNA damage. In vivo studies that have investigated the genotoxic potential of workers handling pesticides using the Comet assay are few. The assay was used to quantify the level of DNA damage in mononuclear leukocytes of French farmers who were occupationally exposed to a number of pesticides. The findings showed that genetic damage was significantly high (Lebailly et al., 1998a
). In another study by the same authors, the Comet assay was used to assess DNA damage in the farmers. An increase in DNA damage levels was observed after 1 day spraying with pesticide mixtures (Lebailly et al., 1998b). Evaluation of workers involved in the production of a variety of pesticides using the Comet assay revealed an increase in DNA damage in peripheral blood lymphocytes (Garaj-Vrhovac and Zeljezic, 2000). Similarly, a study of possible genetic damage in Croatian workers occupationally exposed to a complex mixture of pesticides showed an increase in the values of the Comet assay parameters (Garaj-Vrhovac and Zeljezic, 2001). Our results are consistent with the above reports. Other studies performed on populations exposed to a variety of pesticides, which examined genotoxicity with different assays, support our findings (Yoder et al., 1973; Dulout et al., 1985; Paldy et al., 1987; Rita et al., 1987; Nehez et al., 1988; Rupa et al., 1989c, 1991; De Ferrari et al., 1991; Kourakis et al., 1992; Bolognesi et al., 1993; Carbonell et al., 1993; Lander and Ronne, 1995; Falck et al., 1999; Munnia et al., 1999; Antonucci and Colus, 2000; Gomez-Arroyo et al., 2000; Shaham et al., 2001). However, results obtained by some researchers using various cytogenetic assays with populations exposed to pesticides elucidated negative results (Crossen and Morgan, 1978; Carbonell et al., 1990; Gomez-Arroyo et al., 1992; Hoyos et al., 1996; Scarpato et al., 1996; Davies et al., 1998; Venegas et al., 1998; D’Arce and Colus, 2000; Lucero et al., 2000; Pastor et al., 2001a,b; Zeljezic and Garaj-Vrhovac, 2001).
Since the clastogenicity may be due to cumulative effects of all or some of the pesticides, it is not possible to attribute damage to any particular agent. The results of this study, as well as the results of research performed on subjects occupationally exposed to pesticides using a variety of genotoxic assays, suggest that mixtures of pesticides in long-term occupational exposure could act as clastogens on the DNA of somatic cells. The detected DNA damage could be due to cytotoxic and/or genotoxic effects. The genetic damage demonstrated in the current study and evaluated as an increase in comet tail length could possibly originate from DNA single-strand breaks, repair of DNA double-strand breaks, DNA adduct formation or DNA–DNA and DNA–protein crosslinks. Occupational exposure to xenobiotics may result in their covalent binding to DNA, which may lead to chromosome alterations and could be an initial event in the process of chemical carcinogenesis (Fairbairn et al., 1995; Shah et al., 1997). Exposure to known genotoxic compounds could induce DNA damage not only directly but also through other mechanisms, such as oxidative stress or inflammatory processes (Lebailly et al., 1998b). It is well known that increased genotoxicity in individuals occupationally exposed to pesticides is related to cancer risk and genetic illness. The individual genetic variability in the enzymes which metabolize agricultural chemicals may also be involved in this process. When they are not efficient in detoxification, the metabolic sub-products accumulate, contributing to the turmorigenic process (Hodgson et al., 1991).
The genetic damage seen in the current investigation may be due to insufficient protective measures taken by the exposed workers. Similarly, positive genotoxicity has been reported in studies where the pesticide-exposed workers had used very few protective measures (Gomez-Arroyo et al., 2000; Padmavathi et al., 2000). In contrast, some cytogenetic studies of pesticide sprayers showed that using proper safety procedures significantly reduced the genetic risk (Venegas et al., 1998; Lucero et al., 2000). Therefore, using more or fewer safety measures is also a factor that may explain the positive findings of this study. However, no data on environmental measurements in the work area are available.
This study has demonstrated significantly greater DNA migration in smokers of the control group as well as in the pesticide industry workers. Likewise, a significantly higher level of DNA damage was found in smokers after a period of pesticide production with the Comet assay (Zeljezic and Garaj-Vrhovac, 2001). A significantly increased comet tail length was observed in pesticide manufacturing workers with 10 years exposure. A similar increase in genotoxic effects in workers exposed to pesticides with increase in duration of exposure has been reported (Carbonell et al., 1993; Padmavathi et al., 2000). Our investigation has revealed an insignificant effect of age on DNA damage in the study subjects. These results are in agreement with those ofLebailly et al.(1998a). Our study did not show a significant difference between genders. Similarly, a significant difference in gender was not detected in a study of occupational exposure to pesticides (Zeljezic and Garaj-Vrhovac, 2001).
In the present investigation the blood samples were collected in a single season (October–December) for the study of genetic damage in pesticide industry workers. However, in some studies variations in genetic markers were evaluated at different times in pesticide exposed workers and contrasting results were observed (Carbonell et al., 1995; Laurent et al., 1996).
Biomonitoring studies of populations exposed to pesticides are rather specific because different populations have different lifestyles, nutritional habits, climatic and environmental conditions and are exposed to different mixtures of pesticides. This could explain why some studies have found an increase in genetic damage in populations exposed to pesticides whilst in other studies the results were negative. Although the exposure level can be considered significant and information is available on the genotoxicity of several of the pesticides used, there is a lack of information concerning genotoxic effects of complex mixtures. Another explanation for the genotoxic damage observed could be the lack of protective measures taken by the workers. Therefore, there is a need to educate those who work with pesticides about the potential hazards of occupational exposure and the importance of using protective measures. Since DNA damage is an important step in events leading from carcinogen exposure to cancer, our study represents an important contribution to the correct evaluation of the potential health risks associated with agrochemical exposure.
|
Acknowledgments |
|---|
We are especially grateful to Dr Y.R.Ahuja (Head, Genetics Unit, Mahavir Hospital) for his encouragement and guidance throughout this study. The authors express their sincere thanks to Dr K.V.Raghavan (Director, Indian Institute of Chemical Technology, Hyderabad) for his encouragement during the study.
|
Notes |
|---|
3 To whom correspondence should be addressed. Tel: +91 40 7193135; Fax: +91 40 7160387; 7160757; Email: grover{at}iict.ap.nic.in
|
References |
|---|
|
TopAbstractIntroductionMaterials and methodsResultsDiscussionReferences |
|---|
-
Amr,M.M. (1999) Pesticide monitoring and its health problems in Egypt, a Third World country. Toxicol. Lett., 107, 1–13.[CrossRef][Web of Science][Medline]
Antonucci,G.A. and Colus,I.M. (2000) Chromosomal aberrations analysis in a Brazilian population exposed to pesticides. Teratog. Carcinog. Mutagen., 20, 265–272.[CrossRef][Web of Science][Medline]
Bolognesi,C., Parrini,M., Bonassi,S., Ianello,G. and Salanitto,A. (1993) Cytogenetic analysis of a human population occupationally exposed to pesticides. Mutat. Res., 285, 239–249.[Web of Science][Medline]
Carbonell,E., Puig,M., Xamena,N., Creus,A. and Marcos,R. (1990) Sister chromatid exchanges in lymphocytes of agricultural workers exposed to pesticides. Mutagenesis, 5, 403–405.
Carbonell,E., Xamena,N., Creus,A. and Marcos,R. (1993) Cytogenetic biomonitoring in a Spanish group of agricultural workers exposed to pesticides. Mutagenesis, 8, 511–517.
Carbonell,E., Valbuena,A., Xamena,N., Creus,A. and Marcos,R. (1995) Temporary variations in chromosomal aberrations in a group of agricultural workers exposed to pesticides. Mutat. Res., 344, 127–134.[CrossRef][Web of Science][Medline]
Crossen,P.E. and Morgan,W.F. (1978) Cytogenetic studies of pesticide and herbicide sprayers. N. Z. Med. J., 88, 192–195.[Web of Science][Medline]
D’Arce,L.P.G. and Colus,I.M. (2000) Cytogenetic and molecular biomonitoring of agricultural workers exposed to pesticides in Brazil. Teratog. Carcinog. Mutagen., 20, 161–170.[CrossRef][Web of Science][Medline]
Daniels,J.L., Olshan,A.F. and Savitz,D.A. (1997) Pesticides and childhood cancers. Environ. Health Perspect., 105, 1068–1077.[Web of Science][Medline]
Davies,H.W., Kennedy,S.M., Teschke,K. and Quintana,P.J.E. (1998) Cytogenetic analysis of South Asian berry pickers in British Columbia using the micronucleus assay in peripheral lymphocytes. Mutat. Res., 416, 101–113.[Web of Science][Medline]
De Ferrari,M., Artuso,M., Bonassi,S., Bonatti,S., Cavalieri,Z., Pescatore,D., Marchini,E., Pisano,V. and Abbondandolo,A. (1991) Cytogenetic biomonitoring of an Italian population exposed to pesticides: chromosome aberrations and sister chromatid exchange analysis in peripheral blood lymphocytes. Mutat. Res., 260, 105–113.[CrossRef][Web of Science][Medline]
Dulout,F.N., Pastori,M.C., Olivero,O.A., Gonzalez Cid,M., Loria,D., Matos,E., Sobel,N., de Bujan,E.C. and Albiano,N. (1985) Sister chromatid exchanges and chromosomal aberrations in a population exposed to pesticides. Mutat. Res., 143, 237–244.[CrossRef][Web of Science][Medline]
Fairbairn,D.W., Olive,P.L. and O’Neill,K.L. (1995) The comet assay: a comprehensive review. Mutat. Res., 339, 37–59.[Web of Science][Medline]
Falck,G.C.M., Hirvonen,A., Scarpato,R., Saarikoski,S.T., Migliore,L. and Norppa,H. (1999) Micronuclei in blood lymphocytes and genetic polymorphism for GSTM1, GSTT1 and NAT2 in pesticide exposed greenhouse workers. Mutat. Res., 441, 225–237.[Web of Science][Medline]
Garaj-Vrhovac,V. and Zeljezic,D. (1999) Chromosomal aberrations and frequency of micronuclei in workers employed in pesticide production. Biologia (Bratisl.), 54, 707–712.
Garaj-Vrhovac,V. and Zeljezic,D. (2000) Evaluation of DNA damage in workers occupationally exposed to pesticides using single-cell gel electrophoresis (SCGE) assay. Pesticide genotoxicity revealed by comet assay. Mutat. Res., 469, 279–285.[Web of Science][Medline]
Garaj-Vrhovac,V. and Zeljezic,D. (2001) Cytogenetic monitoring of a Croation population occupationally exposed to a complex mixture of pesticides. Toxicology, 165, 153–162.[CrossRef][Web of Science][Medline]
Gomez-Arroyo,S., Noriega-Aldana,N., Osorio,A., Galicia,F., Ling,S. and Villalobos-Pietrini,R. (1992) Sister-chromatid exchange analysis in a rural population of Mexico exposed to pesticides. Mutat. Res., 281,173–179.[CrossRef][Web of Science][Medline]
Gomez-Arroyo,S., Diaz-Sanchez,Y., Meneses-Perez,M.A., Villalobos-Pietrini,R. and DeLeon-Rodriguez,J.D. (2000) Cytogenetic biomonitoring in a Mexican floriculture worker group exposed to pesticides. Mutat. Res., 466, 117–124.[Web of Science][Medline]
Hodgson,E., Silver,I.S., Butler,L.E., Lawton,M.P. and Levi,P.E. (1991) Metabolism. In Hayes,W.J. and Laws,E.R. (eds), Handbook of Pesticides Toxicology, Vol. 1. Academic Press, San Diego, CA, pp. 106–167.
Hoyos,L.S., Carvajal,S., Solano,L., Rodriguez, J, Orozco,L., Lopez,Y. and Au,W.W. (1996) Cytogenetic monitoring of farmers exposed to pesticides in Columbia. Environ. Health Perspect., 104 (suppl. 3), 535–538.
Hulka,B.S., Wilcosky,T.C. and Griffith,J.D. (1990) Biological Markers in Epidemiology. Oxford University Press, New York, NY.
IARC (1991) International Agency for Research on Cancer Monographs, no. 53, Occupational Exposures in Insecticide Application and Some Pesticides. IARC, Lyon.
Joksic,G., Vidakovic,A. and Spasojevic-Tisma,V. (1997) Cytogenetic monitoring of pesticide sprayers. Environ. Res., 75, 113–118.[Medline]
Kassie,F., Parzefall,W. and Knasmuller,S. (2000) Single cell gel electrophoresis assay: a new technique for human biomonitoring studies. Mutat. Res., 463, 13–31.[CrossRef][Web of Science][Medline]
Khuder,S.A. and Mutgi,A.B. (1997) Meta-analyses of multiple myeloma and farming. Am. J. Ind. Med., 32, 510–516.[CrossRef][Web of Science][Medline]
Kiraly,J., Szentesi,I., Ruzicska,M. and Czeizel,E. (1979) Chromosome studies in workers producing organophosphate insecticides. Arch. Environ. Contam. Toxicol., 8, 309–319.[CrossRef][Web of Science][Medline]
Kourakis,A., Mouratidou,M., Kokkinos,G., Barbouti,A., Kotsis,A., Mourelatos,D. and Dozi-Vassiliades,J. (1992) Frequencies of chromosomal aberrations in pesticide sprayers working in plastic green houses. Mutat. Res., 279, 145–148.[CrossRef][Web of Science][Medline]
Kourakis,A., Mouratidou,M., Barbouti,A. and Dimikiotou,M. (1996) Cytogenetic effects of occupational exposure in peripheral blood lymphocytes of pesticide sprayers. Carcinogenesis, 17, 99–101.
Lander,F. and Ronne,M. (1995) Frequency of sister chromatid exchange and hematological effects in pesticide-exposed greenhouse sprayers. Scand. J. Work Environ. Health, 21, 283–288.[Web of Science][Medline]
Laurent,C., Jadot,P. and Chabut,C. (1996) Unexpected decrease in cytogenetic biomarkers frequencies observed after increased exposure to organophosphorus pesticides in a production plant. Int. Arch. Occup. Environ. Health, 68, 399–404.[Web of Science][Medline]
Lebailly,P., Vigreux,C., Lechevrel,C., Ledemeney,D., Godard,T., Sichel,F., LeTalaer,Y., Henry-Amar,M. and Gauduchon,P. (1998a) DNA damage in mononuclear leukocytes of farmers measured using the alkaline comet assay: discussion of critical parameters and evaluation of seasonal variations in relation to pesticide exposure. Cancer Epidemiol. Biomarkers Prev., 7, 917–927.[Abstract]
Lebailly,P., Vigreux,C., Lechevrel,C., Ledemeney,D., Godard,T., Sichel,F., LeTalaer,J.Y., Henry-Amar,M. and Gauduchon,P. (1998b) DNA damage in mononuclear leucocytes of farmers measured using the alkaline comet assay: modifications of DNA damage levels after a one day field spraying period with selected pesticides. Cancer Epidemiol. Biomarkers Prev., 7, 929–940.[Abstract]
Legator,M.S. and Au,W.W. (1994) Application of integrated genetic monitoring: the optimal approach for detecting environmental carcinogens. Environ. Health Perspect., 102, 125–132.
Lucero,L., Pastor,S., Suarez,S., Durban,R., Gomez,C., Parron,T., Creus,A. and Marcos,R. (2000) Cytogenetic biomonitoring of Spanish greenhouse workers exposed to pesticides: micronuclei analysis in peripheral blood lymphocytes and buccal epithelial cells. Mutat. Res., 464, 255–262.[Web of Science][Medline]
Moretti,M., Villarini,M., Sforzolini,G.S. and Pasquini,R. (2000) Pesticide-induced primary DNA damage in peripheral blood leukocytes of farm workers evaluated by the computerized ‘comet’ assay. Biomarkers, 5, 192–204.[CrossRef]
Munnia,A., Puntoni,R., Merlo,F., Parodi,S. and Peluso,M. (1999) Exposure to agrochemicals and DNA adducts in Western Liguria, Italy. Environ. Mol. Mutagen., 34, 52–56.[CrossRef][Web of Science][Medline]
Nehez,M., Berencsi,G., Paldy,A., Selypes,A., Czeizel,A., Szentesi,I., Csanko,J., Levay,K., Maurer,J. and Nagy,E. (1981) Data on the chromosome examinations of workers exposed to pesticides. Regul. Toxicol. Pharmacol., 1, 116–122.[CrossRef]
Nehez,M., Boros,P., Ferke,A., Mohos,J., Palotas,M., Vetro,G., Zimanyi,M. and Desi,I. (1988) Cytogenetic examination of people working with agrochemicals in the southern region of Hungary. Regul. Toxicol. Pharmacol., 8, 37–44.[CrossRef][Web of Science][Medline]
Padmavathi,P., Prabhavathi,A.P. and Reddy,P.P. (2000) Frequencies of SCEs in peripheral blood lymphocytes of pesticides workers. Bull. Environ. Contam. Toxicol., 64, 155–160.[CrossRef][Web of Science][Medline]
Paldy,A., Puskas,N., Vincze,K. and Hadhazi,M. (1987) Cytogenetic studies on rural populations exposed to pesticides. Mutat. Res., 187, 127–132.[CrossRef][Web of Science][Medline]
Pastor,S., Gutierrez,S., Creus,A., Cebulska-Wasilewska,A. and Marcos,R. (2001a) Micronuclei in peripheral blood lymphocytes and buccal epithelial cells of Polish farmers exposed to pesticides. Mutat. Res., 495, 147–156.[Web of Science][Medline]
Pastor,S., Gutierrez,S., Creus,A., Xamena,N., Piperakis,S. and Marcos,R. (2001b) Cytogenetic analysis of Greek farmers using the micronucleus assay in peripheral lymphocytes and buccal cells. Mutagenesis, 16, 539–545.
Pool Zobel,B.L., Lotzmann,N., Knoll,M., Kuchenmeister,F., Lambertz,R., Leucht,U., Schroder,H.G. and Schmezer,P. (1994) Detection of genotoxic effects in human gastric and nasal mucosa cells isolated from biopsy samples. Environ. Mol. Mutagen., 24, 23–45.[Web of Science][Medline]
Rita,P., Reddy,P.P. and Reddy,S.V. (1987) Monitoring of workers occupationally exposed to pesticides in grape gardens of Andhra Pradesh. Environ. Res., 44, 1–5.[Medline]
Rupa,D.S., Rita,P., Reddy,P.P. and Reddi,O.S. (1988) Screening of chromosomal aberrations and sister-chromatid exchanges in peripheral lymphocytes of vegetable garden workers. Hum. Toxicol., 7, 333–336.[Web of Science][Medline]
Rupa,D.S., Reddy,P.P. and Reddi,O.S. (1989a) Frequencies of chromosomal aberrations in smokers exposed to pesticide in cotton fields. Mutat. Res., 222, 37–41.[CrossRef][Web of Science][Medline]
Rupa,D.S., Reddy,P.P. and Reddi,O.S. (1989b) Analysis of sister-chromatid exchanges, cell kinetics and mitotic index in lymphocytes of smoking pesticides sprayers. Mutat. Res., 223, 253–258.[CrossRef][Web of Science][Medline]
Rupa,D.S., Reddy,P.P. and Reddi,O.S. (1989c) Chromosomal aberrations in peripheral lymphocytes of cotton field workers exposed to pesticides. Environ. Res., 49, 1–6.[Medline]
Rupa,D.S., Reddy,P.P. and Reddi,O.S. (1991) Clastogenic effect of pesticides in peripheral lymphocytes of cotton-field workers. Mutat. Res., 261, 177–180.[CrossRef][Web of Science][Medline]
Scarpato,R., Migliore,L., Angotzi,G., Fedi,A., Miligi,N. and Loprieno,N. (1996) Cytogenetic monitoring of a group of Italian floriculturists: no evidence of DNA damage related to pesticide exposure. Mutat. Res., 367, 73–82.[CrossRef][Web of Science][Medline]
Shah,R.G., Lagueux,J., Kapur,S., Levallois,P., Ayotte,P., Tremblay,M., Zee,J. and Poirier,G.G. (1997) Determination of genotoxicity of the metabolites of pesticides guthion, sencor, lorox, reglone, daconil and admire by 32P postlabeling. Mol. Cell. Biochem., 169, 177–184.[CrossRef][Web of Science][Medline]
Shaham,J., Kaufman,Z., Gurvich,R. and Levi,Z. (2001) Frequency of sister-chromatid exchange among greenhouse farmers exposed to pesticides. Mutat. Res., 491, 71–80.[Web of Science][Medline]
Singh,N.P., McCoy,M.T., Tice,R.R. and Schneider,E.L. (1988) A simple technique for quantitation of low levels of DNA damage in individual cells. Exp. Cell Res., 175, 184–191.[CrossRef][Web of Science][Medline]
Venegas,W., Zapata,I., Carbonell,E. and Marcos,R. (1998) Micronuclei analysis in lymphocytes of pesticide sprayer from Concepcion Chile. Teratog. Carcinog. Mutagen., 18, 123–129.[CrossRef][Web of Science][Medline]
Viel,J.F., Challier,B., Pitard,A. and Pobel,D. (1998) Brain cancer mortality among French farmers: the vineyard pesticide hypothesis. Arch. Environ. Health, 53, 65–70.[Web of Science][Medline]
Yoder,J., Watson,M. and Benson,W.W. (1973) Lymphocyte chromosome analysis of agricultural workers during extensive occupational exposure to pesticides. Mutat. Res., 21, 335–340.[CrossRef][Web of Science][Medline]
Zahm,S.H., Ward,M.H. and Blair,A. (1997) Pesticides and cancer. Occup. Med., 12, 269–289.[Medline]
Zeljezic,D. and Garaj-Vrhovac,V. (2001) Chromosomal aberration and single cell gel electrophoresis (comet) assay in the longitudinal risk assessment of occupational exposure to pesticides. Mutagenesis, 16, 359–363.
Zeljezic,D. and Garaj-Vrhovac,V. (2002) Sister chromatid exchange and proliferative rate index in the longitudinal risk assessment of occupational exposure to pesticides. Chemosphere, 46, 295–303.[Medline]
Received on April 29, 2002; revised on October 7, 2002; accepted on November 25, 2002.
CiteULike 
Connotea 
Del.icio.us What's this?
This article has been cited by other articles:
|
|
 |
|
  J. da Silva, C. R. Moraes, V. D. Heuser, V. M. Andrade, F. R. Silva, K. Kvitko, V. Emmel, P. Rohr, D. L. Bordin, A. C. Andreazza, et al. Evaluation of genetic damage in a Brazilian population occupationally exposed to pesticides and its correlation with polymorphisms in metabolizing genes Mutagenesis, September 1, 2008; 23(5): 415 - 422. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Chandrasekhar, P. V. Rekhadevi, N. Sailaja, M. F. Rahman, J. P. Reddy, M. Mahboob, and P. Grover Evaluation of genetic damage in operating room personnel exposed to anaesthetic gases Mutagenesis, July 1, 2006; 21(4): 249 - 254. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Bull, K. Fletcher, A.R. Boobis, and J.M. Battershill Evidence for genotoxicity of pesticides in pesticide applicators: a review Mutagenesis, March 1, 2006; 21(2): 93 - 103. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. A. Bhalli, Q.M. Khan, M.A. Haq, A.M. Khalid, and A. Nasim Cytogenetic analysis of Pakistani individuals occupationally exposed to pesticides in a pesticide production industry Mutagenesis, March 1, 2006; 21(2): 143 - 148. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Hoffmann, J. Hogel, and G. Speit The effect of smoking on DNA effects in the comet assay: a meta-analysis Mutagenesis, November 1, 2005; 20(6): 455 - 466. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. D. Meeker, N. P. Singh, L. Ryan, S. M. Duty, D. B. Barr, R. F. Herrick, D. H. Bennett, and R. Hauser Urinary levels of insecticide metabolites and DNA damage in human sperm Hum. Reprod., November 1, 2004; 19(11): 2573 - 2580. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||