Epoxiconazole Research Articles

Epoxiconazole disturbed metabolic balance and gut microbiota homeostasis in juvenile zebrafish

Epoxiconazole (EPX) is a broad-spectrum fungicide extensively used in agricultural pest control. Emerging evidence suggests that EPX can adversely affect different endpoints in non-target organisms. Here, the toxicity of EPX was assessed using earlier developmental stage of zebrafish as a model to investigate its effects on metabolism and intestinal microbiota after 21 days of exposure. Our findings indicated that EPX exposure resulted in physiological alterations in juvenile zebrafish, including increase in triglycerides (TG), total cholesterol (TC), low-density lipoprotein (LDL), and glycose (Glu). Nile red staining demonstrated enhanced lipid accumulation in juvenile, accompanied by a marked upregulation in the expression of genes associated with TG synthesis. Moreover, EPX led to alterations in amino acids and carnitines levels in 21 dpf (days post fertilization) zebrafish. We also observed that EPX disrupted intestinal barrier function in juvenile zebrafish, manifested by decreasing mucus secretion and changing in genes related to tight junctions. Moreover, for a more comprehensive analysis of the intestinal microbiota in 21 dpf zebrafish, the intestine tissues were dissected under a microscope for 16S rRNA sequencing analysis. The results revealed that EPX altered the structure and abundance of intestinal microflora in zebrafish, including decreased alpha diversity indices and shifted in bacteria at phylum and genus levels. Notably, the correlation analysis demonstrated strong associations between alterations in various pathogenic bacterial genera and levels of amino acids and carnitines. Overall, these findings confirm that the fungicide EPX promotes metabolic disorders and alterations in the intestinal micro-environment in 21 dpf zebrafish, shedding light on the toxicologic effects of chemicals to aquatic organisms during the development stage.

Absorption, distribution, metabolism, and elimination of epoxiconazole enantiomers in lizards (Eremias argus)

Epoxiconazole (EPX) is a world widely used chiral triazole fungicide in the agriculture field. The excessive application of this triazole may cause damage to lizards. However, limited information is known about the toxicokinetics of EPX on lizards. Our study aimed to investigate the enantioselective absorption, distribution, metabolism, and elimination (ADME) of EPX in lizards following low and high dose exposure (10 and 100 mg kg−1 bodyweitht (bw)). The results demonstrated that (+)-EPX was easier absorbed than (−)-EPX in lizard plasma. Both (+)-EPX and (−)-EPX were detected in the liver, gonad, kidney, skin, brain, and intestine, with (+)-EPX preferentially distributed in these tissues. The elimination of (−)-EPX was faster than that of (+)-EPX in lizard liver and kidney in the high dose groups. Chiral conversion was found between EPX enantiomers in lizard skin. Simultaneously, five metabolites including M2, M4, M10, M18 and M19 were detected in lizard liver and kidney after EPX enantiomers exposure. The relative concentrations of M2, M4, and M10 were higher in the liver and kidney of (−)-EPX groups than those produced from (+)-EPX groups. The metabolic enzymes CYP3A4 and SULT1A1 primarily mediated enantioselective metabolism of EPX. The conclusions drawn from this study significantly enhance our understanding of the enantioselective behaviors of chiral triazole fungicides in reptiles, offering essential guidance for assessing the risks associated with different enantiomers of triazole fungicides.

Mechanistic insights into the parental co-exposure of T-2 toxin and epoxiconazole on the F1 generation of zebrafish (Danio rerio)

Mycotoxins and pesticides frequently coexist in agricultural commodities on a global scale. The potential transgenerational consequences induced by these substances pose a significant threat to human health. However, there is a lack of data concerning the effects of co-contamination by these chemicals in the F1 generation following parental exposure. This investigation delved into the mixture effects of T-2 toxin (T-2) and epoxiconazole (EPO) on the offspring of zebrafish (Danio rerio). The findings revealed that exposure across generations to a combination of T-2 and EPO resulted in toxicity in the larvae of the F1 generation. This was demonstrated by a significant increase in the levels or activities of malondialdehyde (MDA), thyroxine (T4), Caspase3, and cas9, along with a decrease in the levels of cyp19a, ERα, and ERβ. These outcomes suggested that cross-generational exposure to T-2 and EPO in D. rerio disrupted oxidative balance, induced cell apoptosis, and affected the endocrine system. Moreover, these effects were magnified when the F1 generation was continuously exposed to these compounds. Notably, these adverse effects could persist in subsequent generations without additional exposure. This study underscored the potential dangers associated with the simultaneous presence of T-2 and EPO on the development of fish offspring and the resulting environmental hazards to aquatic ecosystems. These findings emphasized the significant health risks posed by cross-generational exposure and highlighted the need for additional legislative measures to address these concerns.

Epoxiconazole degradation in water samples: a comparative study of Fenton, photo-Fenton, solar photo-Fenton, and solar photolysis processes.

Epoxiconazole (EPO) is classified as a persistent organic pollutant due to its ability to persist in the environment for prolonged periods. Its degradation is pivotal in mitigating its environmental impact. This investigation focuses on assessing the degradation of EPO using various methodologies, namely Fenton, photo-Fenton, solar photo-Fenton, and solar photolysis, conducted in both Milli-Q water and groundwater. These experiments encompassed evaluations at both the standard pH typically used in photo-Fenton reactions and the natural pH levels inherent to the respective aqueous environments. Additionally, EPO degradation products were analyzed after a 60-min reaction. Notably, in systems utilizing groundwater, the inclusion of additional iron was unnecessary, as the naturally occurring iron content in the groundwater facilitated the intended processes. Specifically, in Milli-Q water, solar photo-Fenton demonstrated an EPO degradation efficiency of 97%. Furthermore, the substitution of Milli-Q water with groundwater in Fenton-like processes did not significantly affect the efficacy of EPO degradation. These findings underscore the potential of solar photo-Fenton as an economically viable and environmentally sustainable strategy for EPO degradation.

Epoxiconazole altered hepatic metabolism in adult zebrafish based on transcriptomic analysis

Epoxiconazole (EPX) is a triazole fungicide, which has been widely used in pest control of cereal crops. However, its extensive use has led to concerning levels of residue in water bodies, posing substantial risks to aquatic life. In this study, we characterized the toxicological effects of EPX on 6-month-old male and female zebrafish at 70 and 700 μg/L, respectively. The results revealed that EPX exposure markedly increased both body length and weight in zebrafish of both sexes, consequently elevating their condition factor. Besides, EPX exposure resulted in notable alterations in hepatic histopathology. These changes included loosened hepatocyte structure, ballooning degeneration, nucleolysis, and disappearance of cell line, with male zebrafish exhibiting more severe damage. High concentration of EPX also significantly increased hepatic lipid accumulation in male zebrafish, as well as increased hepatic triglyceride (TG) levels. Correspondingly, there was a notable alteration in the transcription of genes including cyp51, hmgcr, and PPAR-γ, which associated with cholesterol and lipid metabolism. Interestingly, with the hepatic transcriptomic analysis, high concentration of EPX produced 195 upregulated and 107 downregulated differential expression genes. Both KEGG and GO analyses identified significant enrichment of these genes in lipid and amino acid metabolism pathways. Notably, some key genes involved in the steroid synthesis pathway were marked upregulated. In addition, molecular docking study confirmed that EPX could bind CYP51 protein well (△G = −7.7 kcal/mol). Taken together, these findings demonstrated the multiple toxic effects of EPX on adult zebrafish.

Border cell population size and oxidative stress in the root apex of Triticum aestivum seedlings exposed to fungicides.

Fungicides reduce the risk of mycopathologies and reduce the content of mycotoxins in commercial grain. The effect of fungicides on the structural and functional status of the root system of grain crops has not been studied enough. In this regard, we studied the phytocytotoxic effects tebuconazole (TEB) and epoxiconazole (EPO) and azoxystrobin (AZO) in the roots of Triticum aestivum seedlings in hydroponic culture. In the presence of EPO and AZO (but not TEB) inhibition of the root growth was accompanied by a dose-dependent increase in the content of malondialdehyde, carbonylated proteins, and proline in roots. TEB was characterized by a dose-dependent decrease in the total amount of border cells (BCs) and the protein content in root extracellular trap (RET). For EPO and AZO, the dose curves of changes in the total number of BCs were bell-shaped. AZO did not affect the protein content in RET. The protein content in RET significantly decreased by 3 times for an EPO concentration of 1µg/mL. The obtained results reveal that the BC-RET system is one of the functional targets of fungicides in the root system of wheat seedlings. Studied fungicides induce oxidative stress and structural and functional alterations in the BC-RET system that can affect their toxicity to the root system of crops.

Bioavailability evaluation of epoxiconazole and difenoconazole in rice and the influence of dissolved organic matter in reducing uptake and translocation

The aim of this study was to assess the bioavailability of epoxiconazole (EPO) and difenoconazole (DIF) in rice plants by evaluating their uptake, translocation, and accumulation. The results showed that the concentration of DIF in the roots was approximately three times higher than EPO, and both accumulated mainly in the roots. In addition, EPO continued to be transported from stems to leaves, causing a rise in its concentration in leaves. Contrastingly, only a minimal amount of DIF was transported to the leaves. This phenomenon is mainly governed by their differing octanol-water partition coefficient. The effects of dissolved organic carbon (DOC) on the accumulation of EPO and DIF in the roots were similar to those of the freely dissolved concentration measured by OECAMs. The concentrations of EPO and DIF in the roots and OECAMs consistently decreased with increasing DOC levels. Furthermore, a significant linear relationship was observed between the EPO and DIF concentrations in root and OECAMs. We also confirmed the accuracy and usefulness of the OECAMs method in predicting the bioavailability of EPO and DIF in rice roots. Therefore, OECAMs show good potential for use as a passive sampler to evaluate the bioavailability of EPO and DIF.

Analytical studies on some pesticides with antifungal effects: Simultaneous determination by HPLC, investigation of interactions with DNA and DNA damages

A simple, and fast method was developed for the simultaneous determination of five fungicides, namely thiram (THR), epoxiconazole (EPO), hexaconazole (HEX), tebuconazole (TEB), and diethofencarb (DIE), in different matrices by HPLC-UV. Parameters influencing the peak shape and resolution, such as the composition of mobile phase, pH and concentration of buffer solution, and column temperature, were examined and optimized. The proposed method was validated in terms of linearity, sensitivity, precision, and accuracy. Forced degradation studies were carried out for all analytes to demonstrate the specificity of the method and to evaluate the stability of analytes under different conditions. DNA interaction and DNA damage studies were conducted by HPLC and comet assay, respectively. All fungicides were found to bind DNA, except for DIE. While the binding coefficients for EPO, HEX, and TEB were of the order of 104, THR was found to interact more strongly with DNA with a binding coefficient of higher than 106. DIE did not induce DNA damage at any concentration tested. On the other hand, TEB, HEX, and EPO induced DNA damage up to 30 µg/mL. THR showed cytotoxic effects at 20 and 30 µg/mL and caused significant DNA damage at lower concentrations.

ConA-Loaded PEGylated Graphene Oxide for Targeted Nanopesticide Carriers against Magnaporthe Oryzae

Rice blast is a destructive rice disease that is rapidly spread by the drift of Magnaporthe oryzae spores. Conidial attachment to the rice leaf surface is the most crucial step in the rapid spread of rice blast. Therefore, blocking the chance of contact between spores and the rice leaf surface is extremely vital for controlling rice blast. In this study, based on the strong affinity between the lectin protein-ConA and M. oryzae spores, we successfully constructed ConA-loaded PEGylated graphene oxide (GO-PEG-ConA) as targeted nanopesticide carriers to load epoxiconazole (EPX) against M. oryzae. The ConA loading capacity of the GO-PEG-EPX-ConA nanocarrier was 14.7% by the sodium dodecyl sulfate–polyacrylamide gel electrophoresis assay. The GO-PEG-ConA nanocarrier could obviously inhibit the germination of M. oryzae spores by 41.7% and showed 2.4-fold higher inhibitory activity than the inhibition ratio (17.4%) of GO-PEG nanocarriers, which was mainly due to the higher affinity between the GO-PEG-ConA nanocarrier and M. oryzae spores. Moreover, the greenhouse experiment indicated that the GO-PEG-EPX-ConA nanopesticide could significantly reduce rice blast lesions in rice seedlings, fundamentally settling the transmission of rice blast. Therefore, GO-PEG-ConA could be a targeted nanopesticide carrier to realize precise release of pesticides, which would offer an important reference value for improving pesticide utilization efficiency.

Oral Exposure to Epoxiconazole Disturbed the Gut Micro-Environment and Metabolic Profiling in Male Mice.

Epoxiconazole (EPX), a triazole fungicide, is widely used in agriculture to control pests and diseases. High residual and occupational exposure to EPX increases health risks, and evidence of potential harm to mammals remains to be added. In the present study, 6-week-old male mice were exposed to 10 and 50 mg/kg bw EPX for 28 days. The results showed that EPX significantly increased the liver weights. EPX also decreased the mucus secretion of the colon and altered intestinal barrier function in mice including a reduced expression of some genes (Muc2, meprinβ, tjp1). Moreover, EPX altered the composition and abundance of gut microbiota in the colon of mice. The alpha diversity indices (Shannon, Simpson) in the gut microbiota increased after exposure to EPX for 28 days. Interestingly, EPX increased the ratio of Firmicutes to Bacteroides and the abundance of other harmful bacteria including Helicobacter and Alistipes. Based on the untargeted metabolomic analysis, it was found that EPX altered the metabolic profiles of the liver in mice. KEGG analysis of differential metabolites revealed that EPX disrupted the pathway related to glycolipid metabolism, and the mRNA levels of related genes were also confirmed. In addition, the correlation analysis showed that the most altered harmful bacteria were associated with some significantly altered metabolites. The findings highlight that EPX exposure changed the micro-environment and lipid metabolism disturbance. These results also suggest that the potential toxicity of triazole fungicides to mammals cannot be ignored.

A note on the impact of CYP51 alterations and their combination in the wheat pathogen Zymoseptoria tritici on sensitivity to the azole fungicides epoxiconazole and metconazole

Septoria tritici blotch caused by the fungal pathogen Zymoseptoria tritici is the most economically damaging disease of winter wheat in Ireland. As azoles have been intensively used, Z. tritici has developed different means of resisting their toxic effects with multiple alterations now reported in individual strains. Using previously characterised Irish collections of Z. tritici, the relationships between these different alterations have been examined using conditional inference trees and random forest. The results from this study highlight the importance of specific alterations I381V and S524T, with both contributing most to the reductions in epoxiconazole (EPZ) and metconazole (MTZ) sensitivity. As the azole class of fungicides is an extensive and diverse group, it is possible these alterations do not impact other azoles in a similar manner. Further analysis of these and contemporary collections to additional azoles, including mefentrifluconazole, which continues to provide good field control of Z. tritici, is warranted.

Stereoselective accumulation and biotransformation of chiral fungicide epoxiconazole and oxidative stress, detoxification, and endogenous metabolic disturbance in earthworm (Eisenia foetida)

>80 % of applied pesticides in agriculture will enter the soil and be exposed to soil animals. Little is known about the stereoselective metabolic effects of epoxiconazole (EPO) on soil animals. In this study, EPO-mediated stereoselective enrichment, biotransformation, oxidative stress, detoxification, and global metabolic profiles in earthworms were investigated by exposure to EPO and its enantiomers at 1 mg/kg and 10 mg/kg doses. Preferential enrichment of (−)-EPO was observed, and the five transformation products (TPs) exhibited the chemically specific stereoselective accumulation with inconsistent configurations. Biochemical markers related to reactive oxygen species (ROS) and detoxification (·OH− content, SOD, CAT, GST, and CYP450 enzymes) showed a significant stereoselective activation overall at the low-level exposure (p-value <0.05). Based on untargeted metabolomic analysis, the steroid biosynthesis and ROS-related biotransformation, glutathione metabolism, TCA cycle, amino acid metabolism, purine and pyrimidine metabolism of earthworms were significantly interfered with by EPO and its enantiomer exposure. More pronounced stereoselectivity was observed at the level of the global metabolic profile, while comparable levels of metabolic perturbations were identified at the individual metabolite level. This study provides novel insights into the stereoselective effects of the chiral fungicide EPO, and valuable evidence for soil environmental risk assessments.

Adsorption behavior of azole fungicides on polystyrene and polyethylene microplastics

Agricultural plastic films and triazole fungicides are widely used in agricultural production process. Exposure to natural environment, agricultural plastic films will degrade into micron plastic particles, which will adsorb pesticide molecules and may affect their toxicity, biological activity and persistence. The long-term coexistence of microplastics (MPs) and triazole fungicides will bring potential harms to the agricultural ecological environment. Therefore, two kinds of triazole fungicides flusilazole (FLU) and epoxiconazole (EPO) were selected as cases and the adsorption behaviors of them on polystyrene and polyethylene were investigated. A series of factors which could affect the adsorption behavior were evaluated. Specifically, the particle size of MPs could affect its adsorption capacity, and the smaller the particle size, the stronger the adsorption capacity. Moreover, with the increase of pH value from 6.0 to 9.0, the adsorption capacity of MPs to target compounds gradually increased. The effect of ionic strength was evaluated by NaCl, and 0.05% of NaCl was beneficial to the adsorption process, while the continuous increase of NaCl concentration inhibited the adsorption. Oxalic acid and humic acid decreased the adsorption capacity of flusilazole on PE by 15.99–32.00% and PS by 35.02–48.67%, respectively. In addition, compared with the single pesticide system, the adsorption capacity of MPs for flusilazole and epoxiconazole in the binary pesticides system decreased by 36.13–37.93% and 44.36–51.35%, respectively, indicating that competitive adsorption occurred between the two pesticides. Meanwhile, the adsorption process was evaluated by adsorption kinetics and adsorption isotherms and were consistent with pseudo-second-order kinetic model and Freundlich isotherm model, respectively. Finally, several characterization analyses were conducted to investigated the adsorption mechanism, and hydrogen, halogen bonding and hydrophobic interaction proved to play an important role. The study on the adsorption behavior and mechanism of pesticide on MPs was the basis of assessing the risk of joint exposure.

Isolation, characterization and application of the epoxiconazole-degrading strain Pseudomonas sp. F1 in a soil-vegetable system

Epoxiconazole (EPX) has a long half-life in soil and causes various toxicological effects in both the ecosystem and mammals. In this study, eight strains of bacteria capable of degrading EPX were isolated from pesticide-contaminated soil, with strain F1 showing the best effect. This strain was identified as Pseudomonas sp. by 16S rRNA gene sequencing and physiological-biochemical analyses. Our results indicated that strain F1 has a high capacity to degrade EPX, removing 92.1% of EPX within 6 days. The temperature and pH were the two most important environmental factors affecting EPX degradation, followed by substrate concentration and inoculum dose. In addition, strain F1 has a high capacity to promote EPX degradation in soils, with a lower t1/2 value (2.64 d) in F1-inoculated soil compared to the control (t1/2 = 96.3 d) without strain F1. The strain could efficiently colonize rhizosphere soil and enhance degradation of EPX, leading to a significant decrease in the accumulation and translocation of EPX in vegetables, thereby alleviating the effects of EPX-induced stress on plants. Moreover, we observed that strain F1-gfp was able to colonize the roots, stems and leaves of Brassica rapa var. chinensis. Such colonization may play a role in the efficient degradation of EPX within plants. To our knowledge, this is the first study to demonstrate biodegradation of EPX in a soil-vegetable system using an EPX-degrading bacterium. This study indicates that strain F1 is a promising candidate for simultaneous bioremediation of soil contaminated with EPX and safe food production.

Epoxiconazole alters the histology and transcriptome of mouse liver in a transgenerational pattern

BackgroundThe use of phytosanitary products is always associated with their safety concern on the environment and on health. The associated adverse effects are very broad and substance-dependent. Among these substances, epoxiconazole (EPOX) is one of the most widely used fungicides, especially in beet crops. Although its use is questionable or even prohibited in the European Union, it is still widely used and its consequences (transgenerational effects) on future generations is unknown. ObjectivesWe aimed to investigate the hepatic effects of epoxiconazole in the descendants of perinatally exposed low-dose C57Bl/6J mice, focusing on liver histological and transcriptomic analyses. MethodsFrom day 0 of gestation up to day 21 postnatal, only pregnant F0 C57BL6/J mice were exposed to EPOX (1.75 μg/kg bw/day). F1 males and females were mated to obtain the F2 generation and similarly, F2 mice were crossed to obtain F3. Histological and transcriptomic analyses of the liver were performed. Gene set enrichment analysis was realized to determine an a priori defined set of genes with significantly altered mRNA expression. Plasma parameters were also measured. Results and conclusions: Perinatal exposure to EPOX induces transgenerational effects with phenotypic, histological and transcriptomic changes in the liver. These changes are highly dependent on the sex and generation of the animal. All these modifications lead to an alteration of the hepatic metabolism resulting in a difference in the size of the hepatocytes. Beyond these specific mechanisms, EPOX also seems to have a more general impact on hepatic metabolism via the circadian rhythm.

A strategy for controlling Cercospora leaf spot, caused by Cercospora beticola, by combining induced host resistance and chemical pathogen control and its implications for sugar beet yield

Cercospora beticola populations resistant to methyl benzimidazole carbamate (MBC), quinone outside inhibitor (QoI), and demethylation inhibitor (DMI) fungicides have been reported in Egypt. This pathogen causes a difficult to control variant of Cercospora leaf spot (CLS) on sugar beet. We conducted field experiments over two years to investigate treatment efficacies on MBC- and QoI-resistant, but partially DMI-sensitive, C. beticola from a research farm in Sakha, Egypt. DMI fungicides showed moderate efficacy with half the maximal effective concentration (EC50). Efficacy was 67.2–69.1% and 63.4–63.6% for epoxiconazole (EPO) and propiconazole (PRO), respectively. When each fungicide was mixed with salicylic acid (SA) at EC50 (EPO + SA) and EC50 (PRO +SA), efficacy was 77.5–79.1% and 77.0–78.2%, respectively. The QoI fungicide, azoxystrobin, showed lower efficacy (46.4–49.5%) when applied alone, but enhanced efficacy when combined with SA at EC50/EC50 (77.0–78.2%). Carbendazim alone was only 47.5–45.1% effective, but when mixed with SA, had a 67.1% co-toxicity factor in vitro, indicating antagonism between components. An Artemisia cina aqueous extract (WEA) combined with SA (WEA/SA) showed 76.0–78.2% efficacy, compared with 69.0–71.0% and 49.0–52.7% for WEA and SA, respectively. Single treatments were classified into five sensitivity classes from sensitive (<1) to resistant (>100) based on their EC50 values. Significant increases in insensitivity were observed from 2015/16 to 2017/18. All treatments increased total yield, sucrose percentage, and recoverable white sugar compared with the untreated control. A combination of control strategies could allow for effective disease and fungicide resistance management.

Effects of biochar on the fate of conazole fungicides in soils and their bioavailability to earthworms and plants.

The study showed novel findings about changes in the fate and bioavailability of conazole fungicides (CFs) after biochar (BC) addition to soil. Two contrasting soils (low- and high-sorbing of CF; L soils, H soils) were amended by three BCs (low-, moderate-, and high-sorbing of CF; L-BC, M-BC, H-BC) at 0.2% and 2% doses. Epoxiconazole (EPC) and tebuconazole (TBC) were then added to the soil-BC mixtures, and their degradation, bioaccumulation in earthworms (Eisenia andrei), and bioconcentration in lettuce (Lactuca sativa) were studied for three months. Also, stir bar sorptive extraction (SBSE) was performed to determine CF (bio)accessibility. The EPC and TBC degradation in the soil-BC mixtures followed usually the first-order decay kinetics. The BC addition prevalently decreased the pesticides degradation in the L soil mixtures but often increased it in the H soil mixtures. In general, EPC degraded less than TBC. BC type and dose roles in the pesticides degradation were unclear. The BC addition significantly reduced pesticide uptake to the earthworms in the L soil mixtures (by 37-96%) and in the H soil mixtures (by 6-89%) with 2% BC. The BC addition reduced pesticide uptake to the lettuce roots and leaves significantly-up to two orders of magnitude, and this reduction was strong in H soil mixtures at 2% of BC. The BC addition reduced the CF (bio)accessibility measured by SBSE in all L soil mixtures and some H soil mixtures with 2% BC. Although not significant, it also seems that the pesticide bioaccumulation, bioconcentration, and (bio)accessibility were decreasing according to the BC type (L-BC > M-BC > H-BC). The pesticide concentrations in the earthworms and lettuce correlated significantly to the SBSE results, which indicates this technique as a possible predictor of biotic uptake. Our results showed that the interactions were hard to predict in the complex soil-BC-pesticide system.

Epoxiconazole profoundly alters rat brain and properties of neural stem cells

Epoxiconazole (EPX), a widely used fungicide for domestic, medical, and industrial applications, could cause neurodegenerative diseases. However, the underling mechanism of neurotoxicity is not well understood. This study aimed to investigate the possible toxic outcomes of Epoxiconzole, a triazole fungicide, on the brain of adult rats in vivo, and in vitro on neural stem cells derived from the subventricular zone of newborn Wistar rats. Our results revealed that oral exposure to EPX at these concentrations (8, 24, 40, 56 mg/kg bw representing respectively NOEL (no observed effect level), NOEL × 3, NOEL × 5, and NOEL × 7) for 28 days caused a considerable generation of oxidative stress in adult rat brain tissue. Furthermore, a significant augmentation in lipid peroxidation and protein oxidation has been found. Moreover, it induced an elevation of DNA fragmentation as assessed by the Comet assay. Indeed, EPX administration impaired activities of antioxidant enzymes and inhibited AChE activity. Concomitantly, this pesticide produced histological alterations in the brain of adult rats. Regarding the embryonic neural stem cells, we demonstrated that the treatment by EPX reduced the viability of cells with an IC50 of 10 μM. It also provoked the reduction of cell proliferation, and EPX triggered arrest in G1/S phase. The neurosphere formation and self-renewal capacity was reduced and associated with decreased differentiation. Moreover, EPX induced cytoskeleton disruption as evidenced by immunocytochemical analysis. Our findings also showed that EPX induced apoptosis as evidenced by a loss of mitochondrial transmembrane potential (ΔΨm) and an activation of caspase-3. In addition, EPX promoted ROS production in neural stem cells. Interestingly, the pretreatment of neural stem cells with the N-acetylcysteine (ROS scavenger) attenuated EPX-induced cell death, disruption of neural stem cells properties, ROS generation and apoptosis. Thus, the use of this hazardous material should be restricted and carefully regulated.

Combining Culture-Dependent and Independent Approaches for the Optimization of Epoxiconazole and Fludioxonil-Degrading Bacterial Consortia.

Epoxiconazole (EPO) and fludioxonil (FLU) are two widely used fluorinated pesticides known to be highly persistent and with high ecotoxicological potential, turning them into pollutants of concern. This work aimed to optimize two degrading bacterial consortia, previously obtained from an agricultural soil through enrichment with EPO and FLU, by characterizing the contribution of their corresponding bacterial isolates to the biodegradation of these pesticides using both culture-dependent and independent methodologies. Results showed that a co-culture of the strains Hydrogenophaga eletricum 5AE and Methylobacillus sp. 8AE was the most efficient in biodegrading EPO, being able to defluorinate ca. 80% of this pesticide in 28 days. This catabolic performance is likely the result of a commensalistic cooperation, in which H. eletricum may be the defluorinating strain and Methylobacillus sp. may assume an accessory, yet pivotal, catabolic role. Furthermore, 16S rRNA metabarcoding analysis revealed that these strains represent a minority in their original consortium, showing that the biodegradation of EPO can be driven by less abundant phylotypes in the community. On the other hand, none of the tested combinations of bacterial strains showed potential to biodegrade FLU, indicating that the key degrading strains were not successfully isolated from the original enrichment culture. Overall, this work shows, for the first time, the direct involvement of two bacterial species, namely H. eletricum and Methylobacillus sp., in the biodegradation of EPO, while also offering insight on how they might cooperate to accomplish this process. Moreover, the importance of adequate culture-dependent approaches in the engineering of microbial consortia for bioremediation purposes is also emphasized.

Accumulation of epoxiconazole from soil via oleic acid-embedded cellulose acetate membranes and bioavailability evaluation in earthworms (Eisenia fetida)

A passive sampler in the soil environment is a relatively novel technique and has had quite limited applications, especially for pesticides. Oleic acid-embedded cellulose acetate membranes (OECAMs) were developed to evaluate the bioavailability of epoxiconazole (EPO) to earthworms (Eisenia fetida). The uptake of EPO by OECAMs (R2 = 0.975) and earthworms (R2 = 0.938) was compared and found to follow a two-compartment kinetic model. EPO sampling by OECAMs reached equilibrium (94%) within 2 d. OECAM could be used to determine the concentration of EPO in soil porewater. Furthermore, a significant linear relationship (R2 = 0.990) was observed between the EPO concentrations in earthworms and the OECAMs. The EPO concentrations in the porewater and OECAMs were lower in soils with a higher organic matter (OM) content. The EPO concentrations in the porewater, earthworms, and OECAMs decreased by 64.4, 49.0, and 56.1%, respectively, in the presence of 0.5% biochar, compared with the control. Furthermore, the use of OECAMs versus earthworms for soil testing also allows you to avoid factors that increase variance in organisms, such as avoidance behaviors or feeding. Therefore, OECAMs show good potential for use as a passive sampler to evaluate the bioavailability of EPO.