Pesticides are used extensively all over the world and in recent years their use has risen. Large amounts of these chemicals are released into the environment, and many of them affect non-target organisms, posing a potential hazard to human health. Pesticide exposure is ubiquitous, not only from agricultural pesticide use and contamination of foods, but also from the extensive use of these products in and around households. Individuals occupationally exposed to pesticides (such as field workers, mixers, loaders, appliers, etc.), who are in direct contact with these chemicals, may provide a good opportunity to study these adverse health consequences.
Exposure to pesticides has been associated with an increase in the incidence of non-Hodgkin’s lymphoma (Hardell and Eriksson, 1999
; Zheng et al., 2001
), multiple myeloma (Khuder and Mutgi, 1997
), soft tissue sarcoma (Kogevinas et al., 1995
), lung sarcoma (Blair et al., 1983
), pancreatic, stomach, liver, bladder and gall bladder cancer (Ji et al., 2001
; Shukla and Arora, 2001
), Parkison’s disease (Gauthier et al., 2001
) and reproductive outcomes (Arbuckle et al., 2001
), among others.
Regarding pesticide exposure, many reports dealing with chromosomal aberrations (Au et al., 1999
; Zeljezic and Garaj-Vrhovac, 2001
), sister chromatid exchange (Shaham et al., 2001
; Zeljezic and Garaj-Vrhovac, 2002
), micronuclei (Falck et al., 1999
; Pastor et al., 2003
) and Comet cells (Zeljezic and Garaj-Vrhovac, 2001
; Grover et al., 2003
) found significant increases in these biomarkers, providing suggestive evidence of genotoxic effects induced by pesticides.
In view of these findings, the detection of populations at risk constitutes a very important topic. Thus, the Micronucleus test (MN) and the Comet assay seem to satisfy many of these criteria, which are used in human biomonitoring studies (Fairbairn et al., 1995
; Moller et al., 2000
; Grover et al., 2003
In order to assess whether prolonged exposure to complex mixtures of pesticides could lead to an increase in cytogenetic damage, vineyard workers and tobacco farmers and workers exposed to pesticides were evaluated using the Micronucleus test and Comet assay in peripheral leukocytes. In order to evaluate whether genetically determined individual variations in xenobiotic metabolizing capacity could modify individual susceptibility to the possible genotoxic effects of pesticides, the subjects were genotyped for several genes.
The study on vineyard workers involved a total number of 173 individuals: 108 were agricultural workers exposed to pesticides and 65 were controls (da Silva et al., 2008
). This study showed a high rate of MN and DNA damage in pesticide-exposed individuals (p < 0.001; Mann-Whitney U
test). In addition, some effects of genetic polymorphisms in PON
in the modulation of MN results were observed in the exposed group, and an association between GSTM1, GSTT1
polymorphisms was suggested. In addition, some effects of genetic polymorphisms in PON
in the modulation of MN in the exposed group were observed, and an association between GSTM1, GSTT1
polymorphisms was suggested.
Considering the importance of repair mechanisms, OGG1 and XRCC1 are examples of important proteins of the BER pathway (Goode et al., 2002
; Au et al., 2004
; Hao et al., 2004
; Muniz et al., 2008
). In another study we evaluated whether the two BER polymorphisms (OGG1 Ser326Cys: rs1052133; and XRCC1 Arg194Trp: rs1799782) or the combined genotypes of these polymorphisms with PON1 Gln192Arg modify individual genotoxic susceptibility to pesticide exposure in vineyard workers, as measured by micronucleus formation and DNA damage induction in peripheral leukocytes. Our study showed that the polymorphisms in the BER pathway could modulate susceptibility to DNA damage caused by pesticide exposure (Rohr et al., 2011
). Significant increases in DI and DF were observed in Cys/- (Cys/Cys or Cys/Ser)-
exposed individuals in relation to OGGI Ser/Ser
individuals (DI: p = 0.032 and DF: p = 0.009).
polymorphism has no effect on the studied biomarkers. Considering the combined influence of the PON1
gene and the OGG1
gene, encoding metabolism and DNA repair functions respectively, we observed higher DNA damage in the exposed group of individuals with a less efficient Cys
allele, regardless of PON1
genotype and significant results in MN formation when combined with XRCC1194
genotype analyses in the exposed group. As this repair pathway is the major cellular defense against oxidative DNA damage, our results support the existing evidence that suggests the involvement of oxidative damage in the pesticide-induced genotoxic effects. Thus, our study reinforces the importance of considering the combined effect of metabolizing and repair variant genotypes on the individual susceptibility to incorporate DNA damage, as these two processes act coordinately in determining the final response to pesticide exposure (Rohr et al., 2011
Located in southern Brazil, the municipality of Santa Cruz do Sul (Rio Grandedo Sul) is an important producer of tobacco leaves (Nicotiana tabacum
). With an annual production of over 687,180 tons, the activity provides employment to over 223,000 farmers (AFUBRA, 2011
). Health risks associated with smoking tobacco and exposure to secondhand smoke are well known. The effects of handling wet tobacco leaves, however are only poorly studied. Green Tobacco Sickness (GTS) is a form of nicotine poisoning that affects workers that have direct contact with tobacco plants during cultivation and harvesting (Mc Bride et al., 1998
). Although GTS has not been associated with mortality or long-term morbidity, it causes significant discomfort and productivity loss among tobacco workers. People involved in preparing and spraying pesticide mixes are the most commonly exposed group of farmers (Bolognesi, 2003
). Many of these compounds are classified as carcinogenic by the International Agency for Research on Cancer (IARC). In this scenario, the evaluation of the effects of occupational exposure to these products becomes an important field of study. In Brazil, tobacco farmers usually apply several pesticides (insecticides, herbicides, fungicides and plant growth regulators) onto tobacco plants to improve crop characteristics and yield.
The objective of our other study with tobacco farmers was to determine the genotoxic effects in a group of workers occupationally exposed to agrochemicals and nicotine. Peripheral blood samples were collected from 30 agricultural workers (42.10 ± 10.15 years), in different crop times (off-season [I], during pesticide application [II] and leaf harvest [III]), and 30 who were not exposed (42.10 ± 10.15 years). As a biomarker of occupational exposure, we obtained data on DNA damage detected by the Comet assay. The results showed significant increase in the Damage index (I: 14.53 ± 13.98; II: 15.24 ± 12.78; III: 17.59 ± 10.23) and frequency (I: 10.57 ± 7.83; II: 8.76 ± 6.15; III: 13.59 ± 10.23) in tobacco farmers compared to the non-exposed group (DI: 1.55 ± 1.36; DF: 1.34 ± 7.63), for all crop times. No correlation was found between age, PPE use and exposure time in relation to DNA damage observed in the Comet assay. A difference, although not significant, was found between gender and exposure. DNA damage was higher in males than females, though not statistically significant (p > 0.05; ANOVA). Moreover, females presented increased DNA damage during leaf harvest compared to off-season and pesticide application phases (p > 0.05). This investigation suggests DNA damage rises in all tobacco crop stages, though more so during the tobacco leaf harvest phase. More worker protection regulations should be enacted to prevent the health problems of tobacco farmers.
Our study with tobacco farmers demonstrates the presence of genotoxic effects in blood cells and exfoliated buccal cells of agricultural workers exposed to pesticides (p < 0.001). These findings appeared during the high exposure period, when pesticides are more frequently applied, at times daily. During the low exposure period, when the pesticides are almost never used, the values detected in the Comet assay, the Micronucleus test and Superoxide dismutase assay were lower than those observed during the high exposure period, and the values were restored to low levels. However, it is important to emphasize that protective measures are essential to avoid potential risks to human health. In addition, some effects of genetic polymorphisms in the PON1 gene in the modulation of micronucleus results were observed in the exposed group, where the exposed homozygote Gln/Gln individuals showed higher genotoxic effect of these pesticides, as they presented the highest MN frequency. Our findings indicate that agricultural workers have higher levels of DNA damage in somatic cells, suggesting a potential health risk for these workers. Risk assessment of pesticide users is crucial to prevent long-term health hazards from developing into cancer and other degenerative diseases.
Various micronutrients of the diet play an important role in genomic stability. Nutrient imbalances, deficiencies and excessive exposure to environmental mutagens and carcinogens can enhance genetic damage. Deficiency of fo-late and other vitamin B cofactors (B12 and B6) also may cause defective DNA repair. The MTHFR C677T allele presents reduced activity, and therefore promotes functional folate deficiency. Therefore, the aim of this study was to evaluate influences of the intake of the micronutrients B12, B6 and folate and the MTHFR C677T polymorphism on DNA damage in pesticide exposed individuals. We conducted a study that involved 113 individuals: 56 (mean age: 42.89 ± 15.79 years) farmers from Venâncio Aires (State of Rio Grande do Sul, Brazil) and 57 non-exposed individuals (mean age: 43.12 ± 14.83 years). They were sampled during tobacco harvest (July to December of 2008/2009) and pesticide application. Blood was collected and submitted to Comet assay, DNA extraction and PCRRFLP. The nutritional status was evaluated on the basis of the mean results of three recordatory 24-hour inquest (RI). The data were computerized, and the intake of nutrients and micronutrients was estimated using the Food Process program. Our analysis showed comet assay values with a significant increase in damage index (DI) and damage frequency (DF) (p < 0.001) for the exposed group as compared to the control group (Mann-Whitney U-test). In our study the individuals with deficiency of folate and B12 vitamin presented a significant increase in the formation of MN in lymphocytes, supporting the hypothesis that folate adequacy and B12 vitamin may be protective against the mutagenic action of pesticides. Dietary adequacy, for both folate and B12 could be assisting in adequate repair. The analyses of genetic susceptibility didn’t demonstrate significant association of MTHFR C677T polymorphism with DI and DF. In conclusion, our results indicate that the tobacco farmers studied have experienced genotoxic exposure, which is manifest as an increase in repairable DNA damage detected by Comet assay, maybe due to heavy and repeated use of pesticides. This effect was influenced by consumption of folate and B12 as this association is described in literature. In the current post-genomic era, different opportunities and challenges have emerged in the context of nutritional research, therefore nutrigenomics plays a huge role in elucidating the complex relation between nutrition and health and establishing the best individual dietary recommendations.