The Incidence of the XRCC1 rs25487 and PON1 rs662 Polymorphisms in a Population from Central Brazil: Patterns in an Area with a High Level of Agricultural Activity

In Brazil, high levels of agricultural activity are reflected in the consumption of enormous amounts of pesticides. The production of grain in Brazil has been estimated at 289.8 million tons in the 2022 harvest, an expansion of 14.7% compared with 2021. These advances are likely associated with a progressive increase in the occupational exposure of a population to pesticides. The Paraoxonase 1 gene (PON1) is involved in liver detoxification; the rs662 variant of this gene modifies the activity of the enzyme. The repair of pesticide-induced genetic damage depends on the protein produced by the X-Ray Repair Cross-Complementing Group 1 gene (XRCC). Its function is impaired due to an rs25487 variant. The present study describes the frequencies of the rs662 and rs25487 and their haplotypes in a sample population from Goiás, Brazil. It compares the frequencies with other populations worldwide to verify the variation in the distribution of these SNPs, with 494 unrelated individuals in the state of Goiás. The A allele of the rs25487 variant had a frequency of 26% in the Goiás population, and the modified rs662 G allele had a frequency of 42.8%. Four haplotypes were recorded for the rs25487 (G > A) and rs662 (A > G) markers, with a frequency of 11.9% being recorded for the A–G haplotype (both modified alleles), 30.8% for the G–G haplotype, 14.3% for the A–A haplotype, and 42.8% for the G–A haplotype (both wild-type alleles). We demonstrated the distribution of important SNPs associated with pesticide exposure in an area with a high agricultural activity level, Central Brazil.


Introduction
Brazil is a significant consumer of pesticides due to its intense agricultural activity (Abreu and Alonzo 2014). The National Provisions Company (CONAB 2021) estimates that total grain production in the Brazilian state of Goiás will reach 28.8 million tons in the 2022 harvest, a record for the state, and an increase of 21.1% in comparison with 2021. In addition, the principal factor determining intoxication by pesticides is the use and distribution of these substances. In Goiás, at least 43,000 tons of active pesticide compounds may be applied to the local plantations yearly (IBAMA 2019).
There is detectable misuse of pesticides in Brazilian crops; in addition, the production of soy, corn, cattle, and vegetables is spread throughout the entire state territory, inevitably exposing the population to pesticides. Recent assessments have revealed a high level of pesticides in the city's water supply, whose economy is mainly agriculture based-an example is the city of Rio Verde, situated in the southeast of Goiás state, where there is the presence of nitrite and nitrate in alarming levels, substances derived from fertilizers. Another example is the city of Rubiataba, in the northeastern Goiás state, where from 2018 to 2020, twelve products found in water are related to chronic diseases, such as cancer, including three pesticide formulations (Aldrin + Dieldrin, Chlordane, Dichlorodiphenyltrichloroethane (DDT) and products generated by DDT degradation). Therefore, such examples demonstrated that agriculture in Goiás state spreads unwanted substances in the environment that significantly affect individuals in these areas. Another problem is that many cities in Goiás state do not demonstrate quality tests on the water supplied to the population in recent years (https:// mapad aagua. repor terbr asil. org. br/).
In this context, the exposure to agricultural pesticides can lead to the impairment of a set of essential functions, such as the inhibition of carboxyl ester hydrolases, chymotrypsin, acetylcholinesterase (AchE), butyrylcholinesterase, liver carboxylases, and paraoxonases (Abdollahi et al. 2004;Kampire et al. 2011). Changes in gene expression due to pesticide exposure must be investigated in a concentrationdependent manner, allowing recognition responses that interfere with molecular signaling dynamics to reveal the role of essential genes and, consequently, the related pathways that are more affected (Thomas et al. 2013). Exposure to some pesticides causes, directly or indirectly, damage to the single-or double-stranded DNA by producing Reactive Oxygen Species (ROS) (Ceja Galvez et al. 2021;Franco et al. 2016;Godoy et al. 2014;Rohr et al. 2011;Singh et al. 2011a). Base Excision Repair (BER) is a fundamental pathway for repairing single-stranded DNA breaks, which may help to minimize this type of impact caused by pesticides (Ceja Galvez et al. 2021;de Oliveira et al. 2019). Although recent research has been conducted to recognize such responses in human cells, there are a lack of information about the effects of pesticide mixing, making it difficult to understand the overall impact on humans. Rural workers are not only those who live in cultivated areas but also consumers who are primarily at risk of exposure (Cipullo et al. 2018). Cutting-edge studies using Omics analysis have already explored the possible effects of environmental mixtures found in water on human lineage cells. Demonstrating the importance of recognizing transcriptional responses allows for understanding the most affected pathways and the most relevant forms of cellular response (Fang et al. 2020;Wang et al. 2020;Xia et al. 2020).
The XRCC1 gene (X-Ray Repair Cross-Complementing Group 1), which is located on chromosome 19 (q13.2-13.3), consists of 33 kb, 17 exons, 16 introns, and encodes a protein of 633 amino acids with a molecular weight of 70KDa (Batar et al. 2016, Li et al. 2012a). The XRCC1 gene is an essential component of the Base Excision Repair (BER) pathway, coordinating other components, such as the poly ADP-ribose, DNA polymerase H, and DNA ligase III, which are all involved in the repair of single strands damaged by ROS (de Oliveira et al. 2019;Fujihara, et al. 2016;Mahmoud et al. 2019;Sobiahe et al. 2020;Xu et al. 2019).
An SNP in the XRCC1 gene increases susceptibility to DNA damage (Chang et al. 2009;de Oliveira et al. 2019). The rs25487 G > A SNP of the XRCC1 gene causes a change in codon 399 in the C-terminal domain of the protein, which shifts from arginine to glutamine (Arg399Gln), with the wild-type G allele-producing arginine, and the A allele-producing glutamine (Bukowski and Woźniak 2017;Huang et al. 1 3 2011; Mahmoud et al. 2019). This shift (Arg399Gln) is associated with a decreased level of DNA repair (Batar et al. 2016), the development of skin cancer (Sobiahe et al. 2020) and lung cancer (Wang et al. 2015), and increased susceptibility to viral hepatitis C (Leite et al. 2013;Mahmoud et al. 2019).
The PON1 (Paraoxonase 1) gene is located on chromosome 7 in q21.3. This gene consists of 26.8 kb, nine exons, and eight introns and encodes a protein of 354 amino acids with a molecular weight of 43KDa (Shunmoogam et al. 2018). Paraoxonase 1 is synthesized by the liver and found on high-density lipoproteins (HDL) surfaces. It is an enzyme involved in phase I detoxification and plays a role in protecting the low-density lipoproteins (LDL). Paraoxonase 1 is also involved in the oxidation pathway of the reverse transport of cholesterol, generating anti-atherogenic effects and hydrolyzing and catalyzing organophosphates before they phosphorylate AChE (Durrington et al. 2001;Hofer et al. 2006;Medina-Díaz et al. 2017).
Several polymorphisms of the PON1 gene affect the concentration and activity of the paraoxonase-1 enzyme. These polymorphisms have been associated with an increased risk of developing cardiac and autoimmune diseases and osteonecrosis (Durrington et al. 2001;Hadjigeorgiou et al. 2007;Zayed et al. 2015;Sunay et al. 2015;Sridon et al. 2020;Hernández-Collazo et al. 2021). The PON1 rs662 A > G mutation is responsible for changing amino acid 192 from glutamine to arginine (Gln192Arg). The former paraoxonase isoform (allele A), which contains glutamine, has been shown to have more efficient catalytic properties for the detoxification of the liver and lipoprotein oxidation reactions in comparison with the latter isoform (allele G), which contains arginine (Rossignol et al. 2014;Siller-López et al. 2017;Sridon et al. 2020). The Gln192Arg mutation is associated with the development of atherosclerosis, hypertension, and coronary heart disease (Dardiotis et al. 2019;Han et al. 2015;Siller-López et al. 2017), changes in the action of AChE, and DNA damage resulting from exposure to pesticides (Bukowski and Woźniak 2017;Dardiotis et al. 2019;Leonel Javeres et al. 2020).
In this context, recognizing human variation by detecting the distribution of essential genotypes and comparing them with worldwide populations of 1000 genomes has become relevant (Abecasis et al. 2012, de Melo e Silva et al. 2018. In recent years, after sequencing the human genome, describing ethnic characteristics in a population based on genetic signatures is an essential guideline for future research, especially when genetic information is scarce, as in Brazil. Furthermore, it is essential in the characterization of genotype-phenotype relationships, and, ultimately, this study helps predict groups at risk of adverse effects due to environmental contamination. As the economy of Goiás state is dominated by its agricultural sector, occupational exposure to pesticides may be widespread, leading to the frequent intoxication of rural workers (IBAMA 2019;de Araújo Nascimento et al. 2020;Tavares et al.2020). This emphasizes the need to determine the distribution of the rs25487 and rs662 polymorphisms, given their association with susceptibility to intoxication and DNA damage. Understanding the susceptibility of the Goiás population to pesticide exposure will be essential to develop adequate preventive measures by government agencies. In this context, the present study describes the frequencies of the rs25487 and rs662 SNPs and their haplotypes in a sample of the population of Goiás. It compares the frequencies with other groups from Brazil and around the world to understand the distribution pattern in this Brazilian state.

Study Population
The study population included 494 unrelated individuals, all rural workers in the southeast and southwest regions of the Goiás state ( Fig. 1), aged between 30 and 65 years.
The population of Goiás, according to the 2010 census, was 6,154,996 inhabitants, being the twelfth most populous federation unit in the country, concentrating approximately 3.1% of the Brazilian population (IBGE 2010). Goiás economy is based manly in agricultural and livestock. Therefore, the sample group consisted of farmers occupationally exposed to a complex mixture of pesticides and manipulated herbicides, insecticides, and fungicides, for at least five years. They do not present neither autoimmune diseases, nor cancer. All the participants were informed of the objective of the study and the procedures used to collect data. Once each participant had filled out a questionnaire (Supplementary material 1) with their details and signed the Informed Consent Term. A 10-ml sample of peripheral blood was collected. The study population from Goiás state was designated "SAMPLE (GO)" for the comparative analyses. It was obtained approval for this study from the Research Ethics Committee of the Federal University of Goiás (reference number 2.648.494). All research procedures were according to the principles of the regulatory guidelines and standards described in Resolution No. 466/12 of the National Health Council, which approves the regulatory guidelines and standards for research involving human beings in Brazil.

DNA Extraction and SNP Genotyping by Real-Time Quantitative PCR
The genomic DNA was isolated from the blood samples using the ReliaPrep™ Blood gDNA Miniprep System kit (Promega, U.S.A.), following the manufacturer's protocol. The DNA was diluted to a final concentration of 20 ng/μL for quantification using a NanoVue Plus™ spectrometer, following the manufacturer's protocol.
The DNA samples were used for genotyping the rs25487 SNP of the XRCC1 gene and the rs662 SNP of the PON1 gene, using the 5′-GGG TTG GCG TGT GAG GCC TTA CCT C[A/G]GGG AGG GCA GCC GCC GAC GCA TGC G-3′ and 5′-ACT TTT CCC CCT AGTTG'TGC ATT CCGT-3′ regions, respectively, to detect the alleles. In addition, 1 μl of the DNA was used for real-time PCR amplification with the TaqMan SNP Genotyping Assays system (Thermo Fisher Scientific, Inc., Waltham, MA, U.S.A.).
For this, 5 μl of TaqMan™ Master MIX, 0.25 μL of the TaqMan™ SNP Genotyping Probe, and 3.75 μL of milli-Q H2O were added to the diluted DNA. The Step Plus Real-Time PCR system thermocycler (Applied Biosystems Inc., Foster City, CA, U.S.A.) for processing. The plate was exposed to a temperature of 95 °C for 10 min for initial denaturation, 15 s at 95 °C for DNA strand denaturation, 1 min at 60 °C for primer annealing, with 45 repetition cycles, and a final 5 min at 72 °C for extension and the emission and detection of the fluorescence. Following the manufacturer's protocol (Thermo Fisher Scientific Inc., Waltham, MA, USA), the reactions were run in duplicate, with a positive control for each genotype and negative control, which ensured the absence of contamination.

Populations Included in the Study for Comparison
The data collected in the present study were compared with those from other studies in Brazil and other parts of the world (Fig. 2). For this comparison, studies of the rs25487 and rs662 polymorphisms-from the period between 2007 and 2021-were surveyed on the Web of Science and Google Scholar platforms using the search terms: "rs25487" OR "rs662" AND "pesticide" OR "pesticides." It is noteworthy that the papers identified in this search contained data appropriate for comparison with the parameters recorded in the present study; in particular, the frequencies of the alleles and genotypes of the rs662 (PON1) and rs25487 (XRCC ) SNPs.
Data on the rs25487 and rs662 polymorphisms were also compiled from 26 populations (2504 individuals) representing five ethnic groups sampled by the 1000 Genomes Project. These data were obtained in.vcf files, downloaded from the project's online database (www. 1000g enomes. org). These data were used for comparison with the sequences of the Goiás study population.

Statistical Analysis
The genotype and allele frequencies of the rs662 (PON1) and rs25487 (XRCC1) polymorphisms were calculated using GENEPOP 4, while Arlequin 3.5.2.2 was used to calculate the haplotype frequencies and Hardy-Weinberg equilibrium. Fisher's Exact Test was run in the RStudio software to compare the allele and haplotype frequencies distribution between populations.

Results
Overall, the 494 unrelated rural workers sampled in the present study from the state of Goiás had a mean age of 48.18±15.56 years, 129 (26%) were female, and 365 (74%) were male. For comparative purposes, studies of rs25487 and rs662 were compiled from 56 other populations, including 26 from the 1000 Genomes Project database (Table 1) and 16 other from rs25487 and rs662 studies (Tables 2 and 3).
In the Goiás study population, the mutant A allele of the rs25487 polymorphism has a frequency of 26% (Table 4), while the frequency of the mutant G allele of the rs662 polymorphism was 42.8% (Table 4). In the case of the rs25487 polymorphism, the highest frequency (49.0%) of the mutant A allele was recorded in Indian farmers (SAS) exposed to pesticides (EXP [IND]). For the rs662 polymorphism, the highest frequency of the mutant G allele was 80.8% in Luhya in Webuye, Kenya (LWK), representing the African ethnicity in the 1000 Genomes project (Table 4). The frequencies of the rs662 polymorphism of the Goiás study deviated significantly (p = 0.03) from the values expected under the Hardy-Weinberg equilibrium, while those of the rs25487 polymorphism were in equilibrium. All 26 populations of the 1000 Genomes database were in equilibrium for both polymorphisms. In the Goiás study population, the mutant (A-G) haplotype had a frequency of 11.9%, while the wild-type G-A haplotype was the most common, with a frequency of 42.8% (Table 5).
The highest frequency (20.1%) of the mutant (A-G) haplotype was recorded in the Japanese (JPT) population of the EAS group, in which the frequencies ranged from 11 to 20%. In contrast, in the SAS group, the frequencies of the mutant haplotype ranged between 8 and 13% (Fig. 3). The G-G haplotype was relatively common

Table 2
Populations from other studies that genotyped the rs25487 polymorphism of the XRCC1 gene included in the present study to compare the allele frequencies    in the African (AFR) populations, with frequencies between 40 and 74%, which were higher than those of the wild (G-A) haplotype, although the mutant (A-G) haplotype was the least common haplotype in this group. Individuals from India exposed to organophosphates Kaur and Kaur et al. (2020)

0.0012
Health individuals from Egypt Zayed et al. (2015) 0.6664 Individuals from China with lung cancer Wang et al. (2015) 1.0000 Individuals from Egypt, with symptoms, exposed to organophosphates Zayed et al. (2015) 0.1048 Health individuals from South Korea Kim et al. (2014) 1.0000 Individuals from Egypt, without symptoms, exposed to organophosphates Zayed et al. (2015) 0.4774 Biochemical Genetics (2023) 61:1675-1703 The rs25487 allele frequencies recorded in the Goiás study population were significantly different (p < 0.05) from those of seven populations (Table 6: Health individuals from the Paraná State Population, BR (dos Reis et al. 2013); Individuals from India exposed to organophosphates [Kaur and Kaur et al. 2020], Esan in Nigeria; Mende in Sierra Leone; Yoruba in Ibadan, Nigeria; Colombian in Medellin, Colombia; and Tuscan from Italy). The last five ones are from the 1000 genomes population. In the case of the rs662 polymorphism, the allele frequencies of the Goiás population were significantly different (p < 0.05) from almost half the other populations ( Table 6).
The haplotype frequencies (rs25487+rs662) of the Goiás study population were compared with those of the 1000 Genomes project using Fisher's exact test (Fig. 4). This analysis indicated significant differences between the haplotype frequencies of the study population and those of all the African (AFR) and East Asian (EAS) populations. In contrast, none of the American (AMR) or South Asian (SAS) presented any significant variation.

Discussion
The identification of genetic risk and individual susceptibility biomarkers, such as the rs25487 and rs662 SNPs, can be valuable for the monitoring of individuals exposed to carcinogenic and genotoxic compounds (Kvitko et al. 2012;Lima Sombra et al. 2011;Manel et al. 2003;Leite et al. 2013;Alves et al. 2019). Few previous studies in the state of Goiás, Central Brazil have examined the relationship between the rs25487 and rs662 polymorphisms and the outcomes of pesticide exposure. Understanding the distribution of deleterious variants is of great importance when it comes to the individualization of the genetic characteristics of a group of individuals, still representing a total population. Recognizing such markers can demonstrate the relevance of maintaining public policies that reinforce the importance of monitoring susceptibilities of a specific population.
In this context, residents of a Brazilian state, as Goiás, Central Brazil, need more attention concerning biomarkers with relevance in situations of pesticide exposure. They live in areas repeatedly affected by different forms of agriculture, not only on farms, but also on properties around cities. From childhood, pesticides can also affect humans if they live in rural areas (Vidi et al. 2017;Arcury et al. 2021). However, pesticides are not the only concern, there are several other forms of exposure that have been investigated, especially in a child (Quintana et al. 2019), and genetic markers, as evaluated in this study, are of great relevance in how an individual can respond to agrochemicals exposure during the entire life.
The A allele of the rs25487 of the XRCC1 gene was recorded in 26% of the individuals sampled from Goiás state. The A/A genotype has been associated with skin cancer in individuals with sunburn (Nelson et al. 2002), squamous cell skin cancer (Kang et al. 2007), and Hodgkin's disease (El-Zein et al. 2009). It is also related to an inadequate response to esophageal cancer and an increased risk of side effects to the treatment of head and neck cancer (Gong et al. 2021), in addition to an increased risk of radiation-induced lymphopenia in patients who have been treated for lung cancer (Xie et al. 2021).
A significantly higher frequency (p = 0.001) of the A allele of the rs25587 was recorded in the EXP (IND) population (49%) in comparison with the Goiás study population. Kaur and Kaur (2020) associated high frequencies of the altered genotypes of the OGG1 rs1052133 (Ser326Cys, 1245CA), XRCC1 rs1799782 (Arg194Trp, 26304CT), and XRCC1 rs25487 (Arg399Gln, 2815GA) genes with the DNA damage observed in agricultural workers exposed to organophosphates in the Punjab, India. These alterations can be linked to pesticides, which can be genotoxic and cytotoxic and cause reproductive hormone disorders. These toxic compounds can induce DNA damage and cytotoxic effects in humans and animals through oxidative pathways (Singh et al. 2011a;Kim et al. 2010;Miranda-Contreras et al. 2013).
The rs25487 (Agr399Gln) SNP is related to a reduction in the capacity of the organism to repair DNA due to the down-regulation of the interaction of the XRCC1 enzyme with the poly-ADP-ribose polymerase, which can cause mutagenic damage (Mahmoud et al. 2019;Floris et al. 2020;Sobiahe et al. 2020). Interestingly, the Biochemical Genetics (2023) 61:  wild-type rs25487 G allele, which produces arginine, has been associated with an increased risk of late fibrosis of the subcutaneous and deep tissues in people treated with radiotherapy (Andreassen 2005).
Some patients with the wild G/G rs25487 genotype may experience severe neutropenia during the treatment of ovarian cancer with chemotherapy (Khrunin et al. 2010). This genotype may also be associated with increased hematological toxicity following radiotherapy in breast cancer patients (Petty et al. 2007) and increased skin cell toxicity (Gossage and Madhusudan 2007). Alsbeih et al. (2010) suggested that the A allele may confer a degree of resistance to high doses of radiation, which would be advantageous in normal tissue during radiotherapy treatment, given the apparent increased risk of skin reactions in individuals with the G allele following radiotherapy. The toxicity induced by radiotherapy may provoke several effects, including specific responses in normal tissues, primarily in the mucosa and blood, leading to apoptosis, with the loss of the capacity for reproduction and cell replacement (Gossage and Madhusudan 2007). According to current research evidence, the G and A alleles of the rs25487 polymorphism may be associated with different outcomes depending on the exposure scenario, such as radiotherapy or pesticide exposure. Because chronic pesticide exposure has been linked to the pathogenesis of several diseases and the presence of less effective detoxification and repair enzymes, this study emphasizes the importance of assessing the presence of these polymorphic genes in the Goiás population.
The frequency of the G allele in the Goiás study population was 74%, while that of the A allele was 26%. The distribution of the G allele of the rs662 polymorphism in the Pakistani and Thai populations exposed to pesticides [EXP (PAK) and EXP (THA)] was significantly different from that of the Goiás population. Some authors (Leonel Javeres et al. 2020;Sridon et al. 2020) related the incidence of the G allele of the rs662 polymorphism in these populations to an increase in DNA damage when exposed to pesticides. By contrast, the frequencies recorded in the Cameroonian, Colombian, and Egyptian (EXP-WS and EXP-NS) populations exposed to pesticides did not vary significantly compared to the Goiás study population. It is nevertheless interesting to note that the authors of those studies found a significant relationship between the G allele of rs662 and increased DNA damage (Leonel Javeres et al. 2020;Siller-López et al. 2017;Zayed et al. 2015).
The paraoxonase-1 enzyme plays an essential role in the inactivation of organophosphate pesticides (Rossignol et al. 2014), converting the pesticides into more polar metabolites (Lieber et al. 2004;Li et al. 2012b) that are also more electrophilic, genotoxic, and highly carcinogenic. Subsequently are biotransformed into more water-soluble compounds (Tanaka 1999;Liu 2006;Moura 2005;Singh et al. 2011a). The G/G rs662 genotype codifies a less active isoform of the protein in terms of its capacity to oxidize lipids and inactivate pesticides. This isoform has also been associated with increased DNA damage and decreased activity in the acetylcholinesterase enzymes of rural workers exposed to pesticides in some different studies (Hernández et al. 2013;Leonel Javeres et al. 2020;Singh et al. 2011a, b, c;Han et al. 2015;Sridon et al. 2020;Zayed et al. 2015). This genotype is also associated with the development of autism (D'Amelio et al. 2005;Rossignol et al. 2014;Siller-López et al. 2017;Sridon et al. 2020), depression, and bipolar disorder (Yildiz et al. 2017).
Overall, the G/G genotype appears to represent a significant risk factor for individuals that are exposed to pesticides due to the variation in the PON1 enzyme. However, the A allele of the rs662, which produces glutamine, has also been associated with an increased risk of hypertension and cardiovascular disease in rural workers exposed to pesticides (Siller-López et al. 2017). In a study of South African farmers (Lee et al. 2003), the A allele was identified as a predictor of chronic poisoning, and the genotypes with this allele had a higher incidence of chronic symptoms. Ji et al. (2012) also found an association between the A allele and increased DNA damage in the sperm in a Chinese study of 1657 men who had infertility. These associations may reflect that the A allele produces isoforms with more intense activity (Rossignol et al. 2014;Siller-López et al. 2017;Sridon et al. 2020). The more active isoform generates more genotoxic and carcinogenic metabolites, which may not be entirely biotransformed in the second stage of liver detoxification. Ramos et al. (2016), for example, found that the alleles that cause the over-expression of CYP2E1 lead to an increase in enzyme activity by forming more genotoxic metabolites and increasing DNA damage, as indicated by the polymorphisms of the CPY2E1 gene in alcoholics from Goiás state. However, Sunay et al. (2015) obtained contrasting results from the analysis of a Turkish population exposed to pesticides regarding the influence of the rs662 polymorphism on the enzymatic activity of the PON1. In that study, the G/G allele was responsible for increased PON1 activity in both the exposed and control groups. The A-G mutant haplotype has low frequencies in most other populations, reaching a maximum of 20% in the Japanese population (EAS), while that of the Goiás study population was 11.9%.
The high G-G haplotype frequencies recorded in AFR and EAS populations highlight the intrinsic variability of these loci. A high proportion of the individuals in the African and East Asian populations had haplotypes with at least one mutant allele (i.e., A-G, G-G, and A-A), with mean frequencies of 78% (AFR) and 75% (EAS). These haplotypes were recorded in 57% of the individuals sampled in the Goiás study population. The high frequencies of these haplotypes in the AFR and EAS populations make them more vulnerable to intoxication by pesticides (Kvitko et al. 2012;Lima Sombra et al. 2011;Leite et al. 2013).
The Goiás state population may be at relatively high risk of intoxication by pesticides, given the large proportion (57%) of haplotypes with at least one mutant allele (A-G, G-G, or A-A). It is noteworthy that Goiás is a central agricultural region. Tavares et al. 2020 demonstrated in the state of Goiás from 2000 to 2013, an enlarged agricultural productivity of sugarcane, corn, and soy, accompanied by an increase in the number of poisoning victims, suicide attempts using pesticides, neoplasms, and congenital malformations. Araújo Nascimento et al. (2020) have already observed a direct relationship between the increase in the cultivated area in Goiás state and the frequency of intoxication by pesticides. However, further studies monitoring the daily exposure of rural workers, along with several individual tests, such as acetylcholinesterase or intoxication biomarkers and genotoxic tests, could explain the real risk to this population.

Conclusion
Our findings highlight the importance of further research in Brazilian populations, especially those occupationally exposed to pesticides, to determine any specific pattern in genetic biomarkers. Several populations have been observed to have high frequencies of haplotypes with at least one mutated allele, which is cause for concern. However, any rs25487 polymorphic alleles (A or G) can be associated with the worst pesticide-related exposure outcome. Only 11% of the sample from Central Brazil contains genotypes with both mutated alleles, indicating the need for additional research identifying indicators of genetic susceptibility to pesticide exposure related to DNA damage. Furthermore, rs662 (G) polymorphisms may interact, as well as whether there is a negative additive effect in pesticide-exposed individuals. However, further studies will be needed to determine to what extent the mutant alleles of the rs25487 (A) and rs662 (G) polymorphisms may interact and whether there is an additive adverse effect in individuals exposed to pesticides.