Carbendazim (CBZ), a broad-spectrum pesticide frequently detected in fruits and vegetables, could trigger potential toxic risks to mammals. To facilitate the assessment of health risks, this study aimed to characterize the cytochrome P450 (CYPs)-mediated metabolism profiles of CBZ by a combined experimental and computational study. Our results demonstrated that CYPs-mediated region-selective hydroxylation was a major metabolism pathway for CBZ in liver microsomes from various species including rat, mouse, minipig, dog, rabbit, guinea pig, monkey, cow and human, and the metabolite was biosynthesized and well-characterized as 6-OH-CBZ. CYP1A displayed a predominant role in the region-selective hydroxylation of CBZ that could attenuate its toxicity through converting it into a less toxic metabolite. Meanwhile, five other common pesticides including chlorpyrifos-methyl, prochloraz, chlorfenapyr, chlorpyrifos, and chlorothalonil could significantly inhibit the region-selective hydroxylation of CBZ, and consequently remarkably increased CBZ exposure in vivo. Furthermore, computational study clarified the important contribution of the key amino acid residues Ser122, and Asp313 in CYP1A1, as well as Asp320 in CYP1A2 to the hydroxylation of CBZ through hydrogen bonds. These results would provide some useful information for the metabolic profiles of CBZ by mammalian CYPs, and shed new insights into CYP1A-mediated metabolic detoxification of CBZ and its health risk assessment.

Fungicides, usually refer to the chemical agents that can effectively control or kill the pathogenic microorganisms. Here, we revealed the effects of three different fungicides, imazalil (IMZ), chlorothalonil (CTL) and carbendazim (CBZ), which are typical broad-spectrum fungicides that are detected at high levels in the natural environment, on heterogeneous human epithelial colorectal cells (Caco-2 cells). All three fungicides had the potential to induce different degrees of toxicity, cause apoptosis, reactive oxygen species (ROS) and even change the cell cycle in the cells. The half maximal inhibitory concentration (IC50) of CTL is the lowest among these three fungicides, suggesting that it may have the highest exposure risk, followed by IMZ, and CBZ. The results of the real-time PCR, Western blotting, and mitochondrial membrane potential (MMP) assays and the activities of key enzymes suggested that CTL induced apoptosis in Caco-2 cells via a mitochondrial-dependent pathway, as indicated by the upregulation of the expression of the apoptotic p53 and bax genes, the increase of the apoptosis marker cytochrome-c, the decrease of mRNA level of bcl-2 gene, and the decrease in the MMP. Exposure to two other fungicides also upregulated the transcriptional level of bax and the expression of cytochrome-c, but the mRNA level of bcl-2 was increased (IMZ) or unchanged (CBZ), suggesting that other pathways may be involved in the induction of cellular apoptosis by these two fungicides. In addition, all three of the fungicides could induce oxidative stress in Caco-2 cells. Our data showed that the three different kinds of fungicides all caused toxic effects in Caco-2 cells through various pathways.

The ecological and economic contributions of pollinator bees to agricultural production have been threatened by the inappropriate and excessive use of pesticides. These pesticides are often applied in areas with ecological peculiarities (e.g., the Neotropical savannah-like region termed as Cerrado) that were not considered during the product development. Here, we conducted field experiments with melon (i.e., Cucumis melo L.) plants cultivated under Brazilian Cerrado conditions and evaluated the impacts of botanical (i.e., neem-based insecticide) and synthetic (i.e., the pyrethroid insecticide deltamethrin and the fungicides thiophanate-methyl and chlorothalonil) pesticides on the flower visitation rates of naturally occurring pollinator bees. Our results revealed that both honey bees (i.e., Apis mellifera L.) and non-Apis bees visited melon flowers and the intensity of bee visitation was moderately correlated with yield parameters (e.g., number of marketable fruits and fruit yield). Pesticide treatments differentially affected bee species. For instance, Plebeia sp. bees were not affected by any pesticide treatment, whereas both A. mellifera and Halictus sp. bees showed reduced visitation intensity after the application of deltamethrin or neem-based insecticides. Fungicide treatment alone did not influence the bee's visitation intensity. Deltamethrin-treated melon fields produced significantly lighter marketable fruits, and the melon yield was significantly lower in melon fields treated with the neem-based insecticide. Thus, our findings with such pollinator bees reinforce the idea that field applications of botanical pesticides may represent as risky as the applications of synthetic compounds, indicating that these alternative products should be submitted to risk assessments comparable to those required for synthetic products.

Chlorothalonil is a broad spectrum fungicide with high annual production and environmental contamination. Despite its high consumption, studies regarding the potential ecological risks of chlorothalonil, especially its metabolites, to aquatic organisms are still limited. In this study, we selected the zebrafish (Danio rerio) as the in vivo model and first identified the metabolite (4-hydroxychlorothalonil) of chlorothalonil in zebrafish by tandem quadrupole/orthogonal-acceleration time-of-flight (Q-TOF). Then, the in vivo and in vitro models were applied to comprehensively estimate the embryo toxicity and potential endocrine effect of chlorothalonil and 4-hydroxychlorothalonil. The data from zebrafish embryo toxicity showed that the lowest observed effect concentrations of both chlorothalonil and 4-hydroxychlorothalonil were 50 μg/L after 96 h of exposure. The mortality rate of the 4-hydroxychlorothalonil was 2.6-fold higher than that of the parent compound at the concentration of 50 μg/L. Dual-luciferase reporter gene assays indicated that both chlorothalonil and 4-hydroxychlorothalonil exerted estrogen receptor α (ERα) agonist activity with REC20 values of 2.4 × 10−8 M and 3.6 × 10−8 M, respectively. However, only 4-hydroxychlorothalonil exhibited both thyroid receptor β (TRβ) agonistic and antagonistic activities. Lastly, we employed molecular docking to predict the binding affinity of chlorothalonil and 4-hydroxychlorothalonil with ERα or TRβ. The results revealed that the potential endocrine effect of chlorothalonil and 4-hydroxychlorothaloni might be attributed to the different binding affinities with the receptors. In conclusion, our studies revealed that 4-hydroxychlorothalonil exhibited potent endocrine-disrupting effects compared to its parent compound, chlorothalonil. The results provided here remind us that the assessment of the potential ecological and health risks of the metabolites of fungicides in addition to their parent compounds should arouse great concerns.

Understanding which factors affect the process of leaf litter decomposition is crucial if we are to predict changes in the functioning of stream ecosystems as a result of human activities. One major activity with known consequences on streams is agriculture, which is of particular concern in tropical regions, where forests are being rapidly replaced by crops. While pesticides are potential drivers of reduced decomposition rates observed in agricultural tropical streams, their specific effects on the performance of decomposers and detritivores are mostly unknown. We used a microcosm experiment to examine the individual and joint effects of an insecticide (chlorpyrifos) and a fungicide (chlorothalonil) on survival and growth of detritivores (Anchytarsus, Hyalella and Lepidostoma), aquatic hyphomycetes (AH) sporulation rate, taxon richness, assemblage structure, and leaf litter decomposition rates. Our results revealed detrimental effects on detritivore survival (which were mostly due to the insecticide and strongest for Hyalella), changes in AH assemblage structure, and reduced sporulation rate, taxon richness and microbial decomposition (mostly in response to the fungicide). Total decomposition was reduced especially when the pesticides were combined, suggesting that they operated differently and their effects were additive. Importantly, effects on decomposition were greater for single-species detritivore treatments than for the 3-species mixture, indicating that detritivore species loss may exacerbate the consequences of pesticides of stream ecosystem functioning.

In many countries, including Morocco, groundwater contamination with pesticides such as globally banned organochlorides (e.g., dichlorodiphenyltrichloroethane (DDT)) and some accredited organophosphates and pyrethroids poses ecological and human health risks. To assess these risks, we herein monitored pesticides in Saïss plain groundwater (Morocco) during the summer of 2017 and the winter of 2018 using polar organic chemical integrative samplers. The two types of passive samplers were deployed in 22 traditional wells for 14–20 days and subjected to solid-phase extraction. The extracts were analyzed by gas chromatography-mass spectrometry and liquid chromatography-tandem mass spectrometry using a multiresidue method, and 27 pesticides were detected in total. In the summer campaign, 22 pesticides with individual concentrations ranging from <limit of quantitation (LOQ) to 243.1 ng L⁻¹ were identified, whereas 17 compounds with concentrations ranging from <LOQ to 53.8 ng L⁻¹ were detected in the winter campaign. In the summer period, the maximum individual concentrations of chlorothalonil, DDT, and α-hexachlorocyclohexane (α-HCH) equaled 111.7, 36.1, and 22.3 ng L⁻¹, respectively, with the respective values for the winter period equaling 18.14, 16.62, and 22.2 ng L⁻¹. Health risk assessment indicated that the carcinogenic α-HCH, β-HCH, DDT, and dichlorodiphenyldichloroethylene present in groundwater may also contaminate drinking water and thus pose a threat to human health, particularly to that of infants and children. Further analysis revealed that the Saïss aquifer presents a high ecological risk. Thus, the monitoring of pesticides in groundwater by passive sampling was effective and could be combined with human health and ecological risk assessment to develop ways of reducing human and environmental exposure to pesticides.

Elasmobranchs are particularly prone to accumulating contaminants due to their life history patterns and relatively high trophic position. However, several compounds, especially contaminants of emerging concern, have still not been well studied in this group. Here, we aimed to determine the occurrence and concentrations of several inorganic and organic contaminants in different tissues of the Brazilian guitarfish Pseudobatos horkelii. This species is a critically endangered species, endemic from the Southwest Atlantic which uses southern Brazilian waters as a nursery habitat. Polycyclic aromatic hydrocarbons (PAHs), emerging pesticides, pharmaceutical and personal care products (PPCPs) and trace metals were determined in five biological tissues in order to assess the accumulation and organotropism of these compounds. Except for chlorothalonil and triclosan, all compounds were detected in, at least, one tissue, mostly in liver samples. All compounds differed among tissues, with liver presenting the higher concentrations of several contaminants, followed by muscle and gills. PAHs and PPCPs were the most detected analytes and presented the highest concentrations among tissues. Diclofenac levels were determined, for the first time in elasmobranchs, and were relatively high, when compared to other fishes. Finally, relatively high concentrations of PAHs, dichlofluanid and octocrylene in muscle might be suggestive of chronic exposure, presenting also human health implications. Regarding trace metals, contrary to most elasmobranch studies, Hg levels were low in all tissues, whereas Cd and Pb here higher in liver, and gills and blood samples, respectively. Our results indicate that P. horkelii is exposed to several organic and inorganic which might affect this species in a long-term scale. Concerning the determination of emerging contaminants, it is likely that other elasmobranchs are also exposed to these compounds and special attention should be given to this issue in order to predict future effects on this group.

Pesticide exposure is regarded as a contributing factor to the high gross loss rates of managed colonies of Apis mellifera. Pesticides enter the hive through contaminated nectar and pollen carried by returning forager honey bees or placed in the hive by beekeepers when managing hive pests. We used an in vitro rearing method to characterize the effects of seven pesticides on developing brood subjected dietary exposure at worse-case environmental concentrations detected in wax and pollen. The pesticides tested included acaricides (amitraz, coumaphos, fluvalinate), insecticides (chlorpyrifos, imidacloprid), one fungicide (chlorothalonil), and one herbicide (glyphosate). The larvae were exposed chronically for six days of mimicking exposure during the entire larval feeding period, which is the worst possible scenario of larval exposure. Survival, duration of immature development, the weight of newly emerged adult, morphologies of the antenna and the hypopharyngeal gland, and gene expression were recorded. Survival of bees exposed to amitraz, coumaphos, fluvalinate, chlorpyrifos, and chlorothalonil was the most sensitive endpoint despite observed changes in many developmental and physiological parameters across the seven pesticides. Our findings suggest that pesticide exposure during larvae development may affect the survival and health of immature honey bees, thus contributing to overall colony stress or loss. Additionally, pesticide exposure altered gene expression of detoxification enzymes. However, the tested exposure scenario is unlikely to be representative of real-world conditions but emphasizes the importance of proper hive management to minimize pesticide contamination of the hive environment or simulates a future scenario of increased contamination.

Brassinosteroids (BRs), a group of steroid phytohormones, are involved in multiple aspects of plant growth, development and stress responses. Despite recent studies on BRs-promoted pesticide metabolism in plants, the underlying mechanisms remain poorly understood. Here, we showed that 24-epibrassinolide (EBR) significantly enhanced the expression of RESPIRATORY BURST OXIDASE HOMOLOG1 (RBOH1) and H2O2 accumulation in the apoplast of chlorothalonil (CHT, a broad spectrum nonsystemic fungicide)-treated tomato plants. Silencing of RBOH1 significantly decreased the efficiency of EBR-induced CHT metabolism. Moreover, the EBR-induced upregulation in the transcripts of glutaredoxin gene GRXS16 was suppressed in RBOH1-silenced plants. Further studies indicated that silencing of GRXS16 compromised EBR-induced increases in glutathione content, activity of glutathione S-transferase (GST) and transcript of GST1, leading to an increase in CHT residue. By contrast, overexpression of tomato GRXS16 enhanced the basal levels of glutathione content and GST activity that eventually decreased CHT residues in transgenic plants. Our results reveal that BR-mediated induction of a modest oxidative burst is essential for the acceleration of glutathione-dependent pesticide metabolism via redox modulators, such as GRXS16. These findings shed new light on the mechanisms of BR-induced pesticide metabolism and thus have important implication in reducing pesticide residues in agricultural products.