Avoidance tests with the oribatid mite Oppia nitens (Acari: Oribatida) in cadmium-spiked natural soils.

Avoidance behavior can be a useful parameter for assessing the ability of organisms to escape from pollutants in their environment. For soil evaluation, a variety of invertebrates is used including the oribatid mite Oppia nitens. Here, we tested the avoidance behavior of O. nitens using a two-chamber test and an escape test with exposures to different cadmium concentrations of up to 800 mg kg-1 dry LUFA 2.2 soil for 2, 4, and 6 days, and up to 7 weeks. With the two-chamber method, the oribatid mites had the choice between clean and polluted soils, whereas they were allowed to escape from a box with polluted soil to clean containers without soil with the escape method. Avoidance of cadmium was observed after 2 days in both tests and the net response of the mites in the two-chamber test increased with increasing cadmium exposure concentrations. Mite responses varied through time, especially with the escape method; with the avoidance behavior becoming more variable and overall non-significant with longer test durations. This is the first study investigating the escape test simultaneously with long-term avoidance of cadmium by O. nitens. This mite species is a promising species for avoidance testing in soil ecotoxicology, but more experiments are needed to evaluate the factors that influence its responses in laboratory tests and the consequences for its distribution in contaminated ecosystems.

Metagenomic analysis of bacterial community structure and diversity of lignocellulolytic bacteria in Vietnamese native goat rumen.

OBJECTIVE: In a previous study, analysis of Illumina sequenced metagenomic DNA data of bacteria in Vietnamese goats' rumen showed a high diversity of putative lignocellulolytic genes. In this study, taxonomy speculation of microbial community and lignocellulolytic bacteria population in the rumen was conducted to elucidate a role of bacterial structure for effective degradation of plant materials. METHODS: The metagenomic data had been subjected into Basic Local Alignment Search Tool (BLASTX) algorithm and the National Center for Biotechnology Information non-redundant sequence database. Here the BLASTX hits were further processed by the Metagenome Analyzer program to statistically analyze the abundance of taxa. RESULTS: Microbial community in the rumen is defined by dominance of Bacteroidetes compared to Firmicutes. The ratio of Firmicutes versus Bacteroidetes was 0.36:1. An abundance of Synergistetes was uniquely identified in the goat microbiome may be formed by host genotype. With regard to bacterial lignocellulose degraders, the ratio of lignocellulolytic genes affiliated with Firmicutes compared to the genes linked to Bacteroidetes was 0.11:1, in which the genes encoding putative hemicellulases, carbohydrate esterases, polysaccharide lyases originated from Bacteroidetes were 14 to 20 times higher than from Firmicutes. Firmicutes seem to possess more cellulose hydrolysis capacity showing a Firmicutes/Bacteroidetes ratio of 0.35:1. Analysis of lignocellulolytic potential degraders shows that four species belonged to Bacteroidetes phylum, while two species belonged to Firmicutes phylum harbouring at least 12 different catalytic domains for all lignocellulose pretreatment, cellulose, as well as hemicellulose saccharification. CONCLUSION: Based on these findings, we speculate that increasing the members of Bacteroidetes to keep a low ratio of Firmicutes versus Bacteroidetes in goat rumen has resulted most likely in an increased lignocellulose digestion.

Coping with living in the soil: the genome of the parthenogenetic springtail Folsomia candida.

BACKGROUND: Folsomia candida is a model in soil biology, belonging to the family of Isotomidae, subclass Collembola. It reproduces parthenogenetically in the presence of Wolbachia, and exhibits remarkable physiological adaptations to stress. To better understand these features and adaptations to life in the soil, we studied its genome in the context of its parthenogenetic lifestyle. RESULTS: We applied Pacific Bioscience sequencing and assembly to generate a reference genome for F. candida of 221.7 Mbp, comprising only 162 scaffolds. The complete genome of its endosymbiont Wolbachia, was also assembled and turned out to be the largest strain identified so far. Substantial gene family expansions and lineage-specific gene clusters were linked to stress response. A large number of genes (809) were acquired by horizontal gene transfer. A substantial fraction of these genes are involved in lignocellulose degradation. Also, the presence of genes involved in antibiotic biosynthesis was confirmed. Intra-genomic rearrangements of collinear gene clusters were observed, of which 11 were organized as palindromes. The Hox gene cluster of F. candida showed major rearrangements compared to arthropod consensus cluster, resulting in a disorganized cluster. CONCLUSIONS: The expansion of stress response gene families suggests that stress defense was important to facilitate colonization of soils. The large number of HGT genes related to lignocellulose degradation could be beneficial to unlock carbohydrate sources in soil, especially those contained in decaying plant and fungal organic matter. Intra- as well as inter-scaffold duplications of gene clusters may be a consequence of its parthenogenetic lifestyle. This high quality genome will be instrumental for evolutionary biologists investigating deep phylogenetic lineages among arthropods and will provide the basis for a more mechanistic understanding in soil ecology and ecotoxicology.

Evolutionary ecology of beta-lactam gene clusters in animals.

Beta-lactam biosynthesis was thought to occur only in fungi and bacteria, but we recently reported the presence of isopenicillin N synthase in a soil-dwelling animal, Folsomia candida. However, it has remained unclear whether this gene is part of a larger beta-lactam biosynthesis pathway and how widespread the occurrence of penicillin biosynthesis is among animals. Here, we analysed the distribution of beta-lactam biosynthesis genes throughout the animal kingdom and identified a beta-lactam gene cluster in the genome of F. candida (Collembola), consisting of isopenicillin N synthase (IPNS), δ-(L-α-aminoadipoyl)-L-cysteinyl-D-valine synthetase (ACVS), and two cephamycin C genes (cmcI and cmcJ) on a genomic scaffold of 0.76 Mb. All genes are transcriptionally active and are inducible by stress (heat shock). A beta-lactam compound was detected in vivo using an ELISA beta-lactam assay. The gene cluster also contains an ABC transporter which is coregulated with IPNS and ACVS after heat shock. Furthermore, we show that different combinations of beta-lactam biosynthesis genes are present in over 60% of springtail families, but they are absent from genome- and transcript libraries of other animals including close relatives of springtails (Protura, Diplura and insects). The presence of beta-lactam genes is strongly correlated with an euedaphic (soil-living) lifestyle. Beta-lactam genes IPNS and ACVS each form a phylogenetic clade in between bacteria and fungi, while cmcI and cmcJ genes cluster within bacteria. This suggests a single horizontal gene transfer event most probably from a bacterial host, followed by differential loss in more recently evolving species.

Priorities for research in soil ecology.

The ecological interactions that occur in and with soil are of consequence in many ecosystems on the planet. These interactions provide numerous essential ecosystem services, and the sustainable management of soils has attracted increasing scientific and public attention. Although soil ecology emerged as an independent field of research many decades ago, and we have gained important insights into the functioning of soils, there still are fundamental aspects that need to be better understood to ensure that the ecosystem services that soils provide are not lost and that soils can be used in a sustainable way. In this perspectives paper, we highlight some of the major knowledge gaps that should be prioritized in soil ecological research. These research priorities were compiled based on an online survey of 32 editors of Pedobiologia - Journal of Soil Ecology. These editors work at universities and research centers in Europe, North America, Asia, and Australia.The questions were categorized into four themes: (1) soil biodiversity and biogeography, (2) interactions and the functioning of ecosystems, (3) global change and soil management, and (4) new directions. The respondents identified priorities that may be achievable in the near future, as well as several that are currently achievable but remain open. While some of the identified barriers to progress were technological in nature, many respondents cited a need for substantial leadership and goodwill among members of the soil ecology research community, including the need for multi-institutional partnerships, and had substantial concerns regarding the loss of taxonomic expertise.

Resilience of Soil Microbial Communities to Metals and Additional Stressors: DNA-Based Approaches for Assessing "Stress-on-Stress" Responses.

Many microbial ecology studies have demonstrated profound changes in community composition caused by environmental pollution, as well as adaptation processes allowing survival of microbes in polluted ecosystems. Soil microbial communities in polluted areas with a long-term history of contamination have been shown to maintain their function by developing metal-tolerance mechanisms. In the present work, we review recent experiments, with specific emphasis on studies that have been conducted in polluted areas with a long-term history of contamination that also applied DNA-based approaches. We evaluate how the "costs" of adaptation to metals affect the responses of metal-tolerant communities to other stress factors ("stress-on-stress"). We discuss recent studies on the stability of microbial communities, in terms of resistance and resilience to additional stressors, focusing on metal pollution as the initial stress, and discuss possible factors influencing the functional and structural stability of microbial communities towards secondary stressors. There is increasing evidence that the history of environmental conditions and disturbance regimes play central roles in responses of microbial communities towards secondary stressors.

Changes in soil oribatid communities associated with conversion from conventional to organic agriculture.

We investigated the effects of switching from conventional management to organic management on the abundance and community composition of soil-living oribatid mites in clover fields in an experimental agricultural station at Al-Fayoum, Egypt. The site had two adjacent fields with identical vegetation cover but different management. Fifteen random soil samples were collected monthly from each of three plots per field, from October to March. We characterized the soils with respect to various physicochemical variables as well as fungal community composition, and estimated mite densities through core sampling. Organic fields had a significantly more abundant oribatid community than did conventional fields. Also the abundance of soil fungi was greater in the organically managed field. Organic management promoted common oribatid mite species with a wide ecological amplitude that already had a high abundance where such common species are more responsive to changes in agricultural management. However, some species of mite responded indifferent or negative to the switch from conventional to organic management. Overall, the differences between the two ecological systems were mainly quantitative. Species diversities of both mite and fungal communities did not differ much between the two management systems. Diversity (H0) and equitability (E) of soil oribatid communities were higher in conventional plots than in the organic plots during the first 2 months but indistinguishable thereafter. Our study confirmed that organic management stimulates soilorganic matter build-up, with positive effects on both fungal and oribatid mite abundance and possible long-term effects on soil function.

Combined Transcriptomics Analysis for Classification of Adverse Effects As a Potential End Point in Effect Based Screening.

Environmental risk assessment relies on the use of bioassays to assess the environmental impact of chemicals. Gene expression is gaining acceptance as a valuable mechanistic end point in bioassays and effect-based screening. Data analysis and its results, however, are complex and often not directly applicable in risk assessment. Classifier analysis is a promising method to turn complex gene expression analysis results into answers suitable for risk assessment. We have assembled a large gene expression data set assembled from multiple studies and experiments in the springtail Folsomia candida, with the aim of selecting a set of genes that can be trained to classify general toxic stress. By performing differential expression analysis prior to classifier training, we were able to select a set of 135 genes which was enriched in stress related processes. Classifier models from this set were used to classify two test sets comprised of chemical spiked, polluted, and clean soils and compared to another, more traditional classifier feature selection. The gene set presented here outperformed the more traditionally selected gene set. This gene set has the potential to be used as a biomarker to test for adverse effects caused by chemicals in springtails to provide end points in environmental risk assessment.

Genome-wide genetic diversity of rove beetle populations along a metal pollution gradient.

To what extent chemical contamination affects genetic diversity of wild populations remains an open question in ecotoxicology. Here we used a genome-wide approach (615 nuclear RADseq loci containing 3017 SNPs) and a mtDNA fragment (ATP6) to analyze the effect of long-term exposure to elevated concentrations of metals (Cd, Pb, Zn) on genetic diversity in rove beetle (Staphylinus erythropterus) populations living along a pollution gradient in Poland. In total, 96 individuals collected from six sites at increasing distance from the source of pollution were analyzed. We found weak differentiation between populations suggesting extensive gene flow. The highest genetic diversity was observed in a population inhabiting the polluted site with the highest metal availability. This may suggest increased mutation rates, possibly in relation to elevated oxidative stress levels. The polluted site could also act as an ecological sink receiving numerous migrants from neighboring populations. Despite higher genetic diversity at the most polluted site, there was no correlation between the genetic diversity and metal pollution or other soil properties. We did not find a clear genomic signature of local adaptation to metal pollution. Like in some other cases of metal tolerance in soil invertebrates, high mobility may counteract possible effects of local selective forces associated with soil pollution.

Knowledge, attitude, and practice of Indonesian farmers regarding the use of personal protective equipment against pesticide exposure.

The use of synthetic pesticides in tropical countries has increased over the years, following the intensification of agriculture. However, awareness among farmers of the importance of protecting themselves from hazards associated with pesticide application is still lacking, especially in Indonesia. This paper reports results of an inventory on knowledge and attitudes regarding pesticide use by melon farmers of a village in Central Java, Indonesia. The importance of using personal protective equipment such as hats, masks, goggles, boots, and gloves on agricultural land is known and well understood by the farmers. However, in practice, only 3.8 % were wearing glasses and 1.9 % were using boots. In fact, the masks used only consisted of a part of their shirt tied around the mouth. The farmers were not wearing long pants and shirts with long sleeves and used the same clothes for more than 1 day without washing. Almost no farmers used personal protective equipment that was standard, in good condition, and complete. Based on the results of statistical analysis, no significant relationship was found between knowledge and attitude on the required practices on the one hand and the use of personal protective equipment in practice on the other hand. This shows that improved knowledge and attitudes are not enough to change the behavior of farmers to work in a healthy and safe way. The gap between knowledge and practice needs to be bridged by a more interactive and participatory training model. It is therefore of paramount importance to develop a special toolkit for pesticide risk reduction which is developed in a participatory manner involving the farmers as the main actors through a series of focus group discussions and field simulations.

Transcriptome assembly and microarray construction for Enchytraeus crypticus, a model oligochaete to assess stress response mechanisms derived from soil conditions.

BACKGROUND: The soil worm Enchytraeus crypticus (Oligochaeta) is an ecotoxicology model species that, until now, was without genome or transcriptome sequence information. The present research aims at studying the transcriptome of Enchytraeus crypticus, sampled from multiple test conditions, and the construction of a high-density microarray for functional genomic studies. RESULTS: Over 1.5 million cDNA sequence reads were obtained representing 645 million nucleotides. After assembly, 27,296 contigs and 87,686 singletons were obtained, from which 44% and 25% are annotated as protein-coding genes, respectively, sharing homology with other animal proteomes. Concerning assembly quality, 84% of the contig sequences contain an open reading frame with a start codon while E. crypticus homologs were identified for 92% of the core eukaryotic genes. Moreover, 65% and 77% of the singletons and contigs without known homologs, respectively, were shown to be transcribed in an independent microarray experiment. An Agilent 180 K microarray platform was designed and validated by hybridizing cDNA from 4 day zinc- exposed E. crypticus to the concentration corresponding to 50% reduction in reproduction after three weeks (EC50). Overall, 70% of all probes signaled expression above background levels (mean signal + 1x standard deviation). More specifically, the probes derived from contigs showed a wider range of average intensities when compared to probes derived from singletons. In total, 522 significantly differentially regulated transcripts were identified upon zinc exposure. Several significantly regulated genes exerted predicted functions (e.g. zinc efflux, zinc transport) associated with zinc stress. Unexpectedly, the microarray data suggest that zinc exposure alters retro transposon activity in the E. crypticus genome. CONCLUSION: An initial investigation of the E. crypticus transcriptome including an associated microarray platform for future studies proves to be a valuable resource to investigate functional genomics mechanisms of toxicity in soil environments and to annotate a potentially large number of lineage specific genes that are responsive to environmental stress conditions.

A functional isopenicillin N synthase in an animal genome.

Horizontal transfer of genes is widespread among prokaryotes, but is less common between microorganisms and animals. Here, we present evidence for the presence of a gene encoding functional isopenicillin N synthase, an enzyme in the ß-lactam antibiotics biosynthesis pathway, in the genome of the soil-living collembolan species, Folsomia candida (FcIPNS). At present, this gene is only known from bacteria and fungi, as is the capacity to produce ß-lactam antibiotics. The FcIPNS gene was located on two genomic contigs, was physically linked to a predicted insect ATP-binding cassette transporter gene, and contained three introns each flanked by eukaryotic splicing recognition sites (GT/AG). Homology searches revealed no similarity between these introns and the FcIPNS regions of bacteria or fungi. All amino acids conserved across bacteria and fungi were also conserved in F. candida. Recombinant FcIPNS was able to convert its substrate amino δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine into isopenicillin N, providing strong evidence that FcIPNS is functional. Phylogenetic analysis clustered FcIPNS outside the bacterial IPNS clade, and also outside the fungal IPNS clade, suggesting an ancient gene transfer followed by divergence in the F. candida genome. In conclusion, the data suggest that the soil-living collembolan F. candida has assimilated the capacity for antibacterial activity by horizontal gene transfer, which may be an important adaptive trait in the microbe-dominated soil ecosystem.

Interactions between accumulated copper, bacterial community structure and histamine levels in crayfish meat during storage.

BACKGROUND: Pollution in aquaculture areas may negatively impact edible species and threaten seafood quality and safety. The aim of this study was to determine the interaction between copper and bacteria in the aquatic habitat and their impact upon crustaceans. Marbled crayfish was chosen as a model of aquatic crustaceans and the influence of metal contamination on bacterial community structure in water used to culture crayfish and in crayfish themselves was investigated. Histamine, an allergen commonly formed by certain groups of bacteria in crustacean edible tissue during storage, was also determined. RESULTS: Copper exposure increased its concentration in crayfish meat by 17.4%, but the copper concentration remained within acceptable food safety limits. Elevated copper levels affected the bacterial community both in the water used to cultivate crayfish and in the marbled crayfish themselves. Cluster analysis of 16S rRNA-gene based microbial community fingerprints revealed that copper impacted the bacterial community in the water and in the crayfish meat. However, copper exposure reduced the formation of histamine in crayfish meat during storage by 66.3%. CONCLUSION: Copper from the habitat appears to reduce histamine accumulation in crayfish meat during storage by affecting the bacterial community structure of the cultivation water and most likely also in the intestine of the crayfish. From a food safety point of view, copper treatment during the aqua culturing of crustaceans has a positive impact on the postharvest stage.

Pesticide residues in sediments and aquatic species in Lake Nokoué and Cotonou Lagoon in the Republic of Bénin.

Lake Nokoué and Cotonou Lagoon are the most important and most productive continental freshwaters in Bénin, with an estimated fish production of over 2 tonnes per hectare in Lake Nokoué. Organochlorine pesticides are used in agriculture and to repel tsetse flies, malaria mosquitoes and other diseases raised. Sediment, fish, shrimp and oyster species were collected in Lake Nokoué and Cotonou Lagoon for pesticide residues analysis. The main pesticides identified in sediment were pp'-DDT and its metabolites pp-DDE and pp'-DDD, with residue levels between the detection limit and 24.4 µg/kg dry weight. Fish species commonly consumed such as Elops lacerta, Podamasys jubelini, Gobbienellus occidentalis, Ethmalosa fimbriata, Mugil cephalus and Hemichromis fasciatus were contaminated with residues of seven to nine pesticides, including pp-DDE, op'-DDD, pp'-DDD, op'-DDT, pp'-DDT, α-endosulfan, aldrin, dieldrin and γ-hexachlorocyclohexane. The levels ranged from detection limit to 289 ng/g lipid. The same pesticides were also detected in other aquatic species, such as shrimp and oysters. A summed risk assessment, comparing pesticide intake levels through fish consumption with tolerable daily intake levels proposed by the World Health Organization, showed in all cases a low risk for human health.

Do fungivores trigger the transfer of protective metabolites from host plants to arbuscular mycorrhizal hyphae?

A key objective in ecology is to understand how cooperative strategies evolve and are maintained in species networks. Here, we focus on the tri-trophic relationship between arbuscular mycorrhizal (AM) fungi, host plants, and fungivores to ask if host plants are able to protect their mutualistic mycorrhizal partners from being grazed. Specifically, we test whether secondary metabolites are transferred from hosts to fungal partners to increase their defense against fungivores. We grew Plantago lanceolata hosts with and without mycorrhizal inoculum, and in the presence or absence of fungivorous springtails. We then measured fungivore effects on host biomass and mycorrhizal abundance (using quantitative PCR) in roots and soil. We used high-performance liquid chromatography to measure host metabolites in roots, shoots, and hyphae, focusing on catalpol, aucubin, and verbascoside. Our most striking result was that the metabolite catalpol was consistently found in AM fungal hyphae in host plants exposed to fungivores. When fungivores were absent, catalpol was undetectable in hyphae. Our results highlight the potential for plant-mediated protection of the mycorrhizal hyphal network.

Male-male competition leads to less abundant but more attractive sperm.

Males employ complex strategies to optimize their reproductive success when faced with male-male competition; for instance, they can adjust the ejaculate characteristics. In copulating species, a male may also strategically adjust his ejaculate expenditure according to female quality. Quantifying the relative contribution of ejaculate plasticity in male reproductive success is often difficult, especially when females exert postcopulatory cryptic choice. One way to quantify the functional significance of ejaculate plasticity is offered by mating systems in which the reproductive partners do not meet each other during insemination. In the collembolan Orchesella cincta, males deposit their ejaculates (spermatophores) irrespective of the presence of females. We tested whether Orchesella males adjust spermatophore number when exposed to the presence of another male and whether changes in spermatophore production influence female choice. We found that Orchesella males display plasticity in spermatophore allocation. Males decreased the spermatophore number when exposed to a rival male. Moreover, females preferentially took up spermatophores of males that were exposed to a competitor. The reduction in spermatophore number suggests, besides an adaptive response to the risk of ejaculate removal by rival males, an optimization strategy owing to the costs of more attractive spermatophores.

Genetic variation in parthenogenetic collembolans is associated with differences in fitness and cadmium-induced transcriptome responses.

Ecotoxicological tests may be biased by the use of laboratory strains that usually contain very limited genetic diversity. It is therefore essential to study how genetic variation influences stress tolerance relevant for toxicity outcomes. To that end we studied sensitivity to cadmium in two distinct genotypes of the parthogenetic soil ecotoxicological model organism Folsomia candida. Clonal lines of both genotypes (TO1 and TO2) showed divergent fitness responses to cadmium exposure; TO2 reproduction was 20% less affected by cadmium. Statistical analyses revealed significant differences between the cadmium-affected transcriptomes: i) the number of genes affected by cadmium in TO2 was only minor (~22%) compared to TO1; ii) 97 genes showed a genotype × cadmium interaction and their response to cadmium showed globally larger fold changes in TO1 when compared to TO2; iii) the interaction genes showed a concerted manner of expression in TO1, while a less coordinated pattern was observed in TO2. We conclude that (1) there is genetic variation in parthenogenetic populations of F. candida, and (2) this variation affects life-history and molecular end points relative to cadmium toxicity. This sheds new light on the sources of biological variability in test results, even when the test organisms are thought to be genetically homogeneous because of their parthenogenetic reproduction.

Transcriptional responses indicate attenuated oxidative stress in the springtail Folsomia candida exposed to mixtures of cadmium and phenanthrene.

Since the 'omics revolution', the assessment of toxic chemical mixtures has incorporated approaches where phenotypic endpoints are connected to a mechanistic understanding of toxicity. In this study we determined the effect of binary mixtures of cadmium and phenanthrene on the reproduction of Folsomia candida and investigated the cellular mechanisms underlying this response. Mixture toxicity modeling showed an antagonistic deviation from concentration addition for reproduction effects of the mixtures. Subsequent transcriptional response analysis was done using five mixtures at the modeled 50 % effect level for reproduction. The transcription profiles of 86 high throughput RT-qPCR assays were studied by means of partial least squares regression analysis. The first and second principal components (PCs) were correlated with global responses to cadmium and phenanthrene, while correlations with the mixture treatments were found in the higher PCs. Specifically associated with the mixture treatments were a biotransformation phase II gene, four mitochondrial related genes and a gene involved in the biosynthesis of antioxidant selenoproteins. Membrane integrity related gene inductions were correlated with the single phenanthrene treatment but not with the mixtures. Immune and inflammatory response assays did not correlate with any of the mixtures. These results suggest moderated oxidative stress, a higher mitochondrial maintenance and less compromised membrane function in the mixture exposed samples compared to the separate cadmium or phenanthrene exposures. The antagonism found for inhibition of reproduction may partially originate from these differences. Mechanistic studies on mixture toxicity can ultimately aid risk assessment by defining relevant toxicity pathways in organisms exposed to real-world mixture exposures present in the field.

Health risks associated with pesticide residues in sediments, fish, and plants from the Ouémé Valley in the Republic of Bénin.

To determine possible human and environmental health risks, organochlorine pesticide residues were determined in vegetables grown in floodplains along the Ouémé River near Lowé in Bénin. Testing of vegetables found 13 pesticides with ΣDDT, α-endosulfan, Σdrin, and lindane being most important. The same pesticides were also detected in plants eaten by bovine cattle, sheep, and herbivorous fish. Human pesticide intake by vegetable consumption was compared with tolerable daily intake (TDI) values reported by the World Health Organization. Pesticide intake by fish consumption was estimated from residue levels in whole fish collected from the Ouémé River in 2004 and reported earlier. Fish consumption does not pose a risk for human health, but consuming vegetables that contain pesticide residues may lead to exceedance of TDI values. Based on these findings, concerns are warranted, and more work is needed to understand the full exposure profile for the local population.