RESUMEN

The uptake, translocation, and accumulation of mefentrifluconazole (MFZ), an innovative chiral triazole fungicide, in plants at the enantiomeric level are still unclear. Herein, we investigated the patterns and mechanisms of enantiomeric uptake, bioaccumulation, and translocation through several experiments. Rac-MFZ shows the strongest uptake and bioaccumulation capacity in wheat compared with its enantiomers, while S-(+)-MFZ has the highest translocation potential. Molecular docking provided evidence of the stronger translocation ability of S-(+)-MFZ than R-(-)-MFZ. Split-root experiments showed that MFZ and its enantiomers could undergo long-distance transport within the wheat. Active transport or facilitated and simple diffusion may be involved in the wheat uptake of MFZ. The limited acropetal translocation capability of MFZ may be attributed to the dominant uptake pathway of apoplastic. The concentrations of Rac-MFZ in different subcellular fractions varied greatly. In summary, this study provides novel insights for further understanding the behaviors of MFZ and its enantiomers in plants.

Asunto(s)

Fungicidas Industriales , Triazoles , Triticum , Triticum/metabolismo , Triticum/química , Triazoles/química , Triazoles/metabolismo , Fungicidas Industriales/metabolismo , Fungicidas Industriales/química , Estereoisomerismo , Transporte Biológico , Simulación del Acoplamiento Molecular , Raíces de Plantas/metabolismo , Raíces de Plantas/química

RESUMEN

Genome mining in association with the OSMAC (one strain, many compounds) approach provides a feasible strategy to extend the chemical diversity and novelty of natural products. In this study, we identified the biosynthetic gene cluster (BGC) of restricticin, a promising antifungal agent featuring a reactive primary amine, from the fungus Aspergillus sclerotiorum LZDX-33-4 by genome mining. Combining heterologous expression and the OSMAC strategy resulted in the production of a new hybrid product (1), along with N-acetyl-restricticin (2) and restricticinol (3). The structure of 1 was determined by spectroscopic data, including optical rotation and electronic circular dichroism (ECD) calculations, for configurational assignment. Compound 1 represents a fusion of restricticin and phytotoxic cichorin. The biosynthetic pathway of 1 was proposed, in which the condensation of a primary amine of restricticin with a precursor of cichorine was postulated. Compound 1 at 5 mM concentration inhibited the growth of the shoots and roots of Lolium perenne, Festuca arundinacea, and Lactuca sativa with inhibitory rates of 71.3 and 88.7% for L. perenne, 79.4 and 73.0% for F. arundinacea, and 58.2 and 52.9% for L. sativa. In addition, compound 1 at 25 µg/mL showed moderate antifungal activity against Fusarium fujikuroi and Trichoderma harzianum with inhibition rates of 22.6 and 31.6%, respectively. These results suggest that heterologous expression in conjunction with the OSMAC approach provides a promising strategy to extend the metabolite novelty due to the incorporation of endogenous metabolites from the host strain with exogenous compounds, leading to the production of more complex compounds and the acquisition of new physiological functions.

Asunto(s)

Lactuca , Lolium , Lolium/genética , Lolium/efectos de los fármacos , Lolium/crecimiento & desarrollo , Lolium/metabolismo , Lactuca/efectos de los fármacos , Lactuca/genética , Lactuca/crecimiento & desarrollo , Familia de Multigenes , Festuca/genética , Festuca/metabolismo , Festuca/microbiología , Festuca/efectos de los fármacos , Festuca/crecimiento & desarrollo , Fungicidas Industriales/farmacología , Fungicidas Industriales/química , Fungicidas Industriales/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/química , Vías Biosintéticas , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Raíces de Plantas/efectos de los fármacos , Raíces de Plantas/microbiología , Estructura Molecular , Genoma Fúngico , Ascomicetos/genética , Ascomicetos/efectos de los fármacos , Ascomicetos/metabolismo , Fusarium/efectos de los fármacos , Fusarium/genética , Fusarium/crecimiento & desarrollo

RESUMEN

Fungicides undergo rapid metabolism and are excreted in the urine. There are few methods for screening these ubiquitous compounds, which have a high potential for human exposure. High-resolution mass spectrometry (HRMS) is a suitable technique to assess fungicide exposures; however, there is a lack of spectral libraries for fungicide annotation and in particular for downstream metabolites. We created spectral libraries for 32 fungicides for suspect screening. Fungicide standards were administered to mice, and 24-h urine was analyzed using hydrophilic interaction and reversed-phase chromatography coupled to hybrid quadrupole-orbitrap mass spectrometry. Suspect metabolite MS2 spectra for library creation were selected based on the ratio of exposed-to-control mouse urine. MS2 libraries were applied to urine collected from female university students (n = 73). Several tetraconazole and tebuconazole metabolites were detected in 3% (2/73) of the samples. The creation of comprehensive suspect screening MS2 libraries is a useful tool to detect fungicide exposure for human biomonitoring.

Asunto(s)

Monitoreo Biológico , Fungicidas Industriales , Fungicidas Industriales/metabolismo , Fungicidas Industriales/orina , Humanos , Femenino , Animales , Ratones , Monitoreo Biológico/métodos , Espectrometría de Masas/métodos , Adulto , Espectrometría de Masas en Tándem/métodos , Adulto Joven

RESUMEN

The metabolic breakdown of propiconazole by fungi was examined, and it was found that the microbial model (Cunninghamella elegans ATCC36112) efficiently degrades the triazole fungicide propiconazole through the action of cytochrome P450. This enzyme primarily facilitates the oxidation and hydrolysis processes involved in phase I metabolism. We observed major metabolites indicating hydroxylation/oxidation of propyl groups of propiconazole. Around 98% of propiconazole underwent degradation within a span of 3 days post-treatment, leading to the accumulation of five metabolites (M1-M5). The experiments started with a preliminary identification of propiconazole and its metabolites using GC-MS. The identified metabolites were then separated and identified by in-depth analysis using preparative UHPLC and MS/MS. The metabolites of propiconazole are M1 (CGA-118245), M2(CGA-118244), M3(CGA-136735), M4(GB-XLIII-42-1), and M5(SYN-542636). To further investigate the role of key enzymes in potential fungi, we treated the culture medium with piperonyl butoxide (PB) and methimazole (MZ), and then examined the kinetic responses of propiconazole and its metabolites. The results indicated a significant reduction in the metabolism rate of propiconazole in the medium treated with PB, while methimazole showed weaker inhibitory effects on the metabolism of propiconazole in the fungus C. elegans.

Asunto(s)

Cunninghamella , Sistema Enzimático del Citocromo P-450 , Fungicidas Industriales , Triazoles , Triazoles/metabolismo , Triazoles/farmacología , Cunninghamella/metabolismo , Fungicidas Industriales/metabolismo , Fungicidas Industriales/farmacología , Sistema Enzimático del Citocromo P-450/metabolismo , Cromatografía de Gases y Espectrometría de Masas , Espectrometría de Masas en Tándem , Oxidación-Reducción , Butóxido de Piperonilo/metabolismo , Butóxido de Piperonilo/farmacología

RESUMEN

Banana Fusarium wilt caused by Fusarium oxysporum f. sp. cubense (Foc TR4) is the most destructive soil-borne fungal disease. Until now, there has been a lack of effective measures to control the disease. It is urgent to explore biocontrol agents to control Foc TR4 and the secretion of mycotoxin. In this study, fluvirucin B6 was screened from Streptomyces solisilvae using an activity-guided method. Fluvirucin B6 exhibited strong antifungal activity against Foc TR4 (0.084 mM of EC50 value) and significantly inhibited mycelial growth and spore germination. Further studies demonstrated that fluvirucin B6 could cause the functional loss of mitochondria, the disorder of metabolism of Foc TR4 cells, and the decrease of enzyme activities in the tricarboxylic acid cycle and electron transport chain, ultimately inhibiting mycotoxin metabolism. In a pot experiment, the application of fluvirucin B6 significantly decreased the incidence of banana Fusarium wilt and the amount of Foc TR4 and controlled fungal toxins in the soil. Additionally, fluvirucin B6 could positively regulate the changes in the structure of the banana rhizosphere microbial community, significantly enriching beneficial microbes associated with disease resistance. In summary, this study identifies fluvirucin B6, which plays versatile roles in managing fungal diseases and mycotoxins.

Asunto(s)

Fungicidas Industriales , Fusarium , Musa , Micotoxinas , Enfermedades de las Plantas , Microbiología del Suelo , Streptomyces , Fusarium/metabolismo , Fusarium/efectos de los fármacos , Enfermedades de las Plantas/microbiología , Enfermedades de las Plantas/prevención & control , Musa/microbiología , Streptomyces/metabolismo , Micotoxinas/metabolismo , Micotoxinas/química , Fungicidas Industriales/farmacología , Fungicidas Industriales/química , Fungicidas Industriales/metabolismo , Bacterias/genética , Bacterias/clasificación , Bacterias/efectos de los fármacos , Bacterias/metabolismo , Bacterias/aislamiento & purificación , Microbiota/efectos de los fármacos

RESUMEN

Fusarium head blight caused by Fusarium graminearum is a devastating disease in wheat that seriously endangers food security and human health. Previous studies have found that the secondary metabolite phenazine-1-carboxamide produced by biocontrol bacteria inhibited F. graminearum by binding to and inhibiting the activity of histone acetyltransferase Gcn5 (FgGcn5). However, the detailed mechanism of this inhibition remains unknown. Our structural and biochemical studies revealed that phenazine-1-carboxamide (PCN) binds to the histone acetyltransferase (HAT) domain of FgGcn5 at its cosubstrate acetyl-CoA binding site, thus competitively inhibiting the histone acetylation function of the enzyme. Alanine substitution of the residues in the binding site shared by PCN and acetyl-CoA not only decreased the histone acetylation level of the enzyme but also dramatically impacted the development, mycotoxin synthesis, and virulence of the strain. Taken together, our study elucidated a competitive inhibition mechanism of Fusarium fungus by PCN and provided a structural template for designing more potent phenazine-based fungicides.

Asunto(s)

Proteínas Fúngicas , Fungicidas Industriales , Fusarium , Histona Acetiltransferasas , Fenazinas , Enfermedades de las Plantas , Triticum , Fusarium/metabolismo , Fusarium/efectos de los fármacos , Fusarium/genética , Fenazinas/metabolismo , Fenazinas/farmacología , Fenazinas/química , Proteínas Fúngicas/metabolismo , Proteínas Fúngicas/genética , Proteínas Fúngicas/química , Fungicidas Industriales/farmacología , Fungicidas Industriales/química , Fungicidas Industriales/metabolismo , Enfermedades de las Plantas/microbiología , Histona Acetiltransferasas/metabolismo , Histona Acetiltransferasas/genética , Histona Acetiltransferasas/química , Histona Acetiltransferasas/antagonistas & inhibidores , Triticum/microbiología , Sitios de Unión , Acetilación

RESUMEN

Fenpropidin (FPD), a widely employed chiral fungicide, is frequently detected in diverse environments. In an in vitro rat liver microsomes cultivation (RLMs), the metabolism exhibited the order of R-FPD > S-FPD, with respective half-lives of 10.42 ± 0.11 and 12.06 ± 0.15 min, aligning with kinetic analysis results. CYP3A2 has been demonstrated to be the most significant oxidative enzyme through CYP450 enzyme inhibition experiments. Molecular dynamics simulations unveiled the enantioselective metabolic mechanism, demonstrating that R-FPD forms hydrogen bonds with the CYP3A2 protein, resulting in a higher binding affinity (-6.58 kcal mol-1) than S-FPD. Seven new metabolites were identified by Liquid chromatography time-of-flight high-resolution mass spectrometry, which were mainly generated through oxidation, reduction, hydroxylation, and N-dealkylation reactions. The toxicity of the major metabolites predicted by the TEST procedure was found to be stronger than the predicted toxicity of FPD. Moreover, the enantioselective fate of FPD was studied by examining its degradation in three soils with varying physical and chemical properties under aerobic, anaerobic, and sterile conditions. Enantioselective degradation of FPD occurred in soils without enantiomeric transformation, displaying a preference for R-FPD degradation. R-FPD is a low-risk stereoisomer both in the environment and in mammals. The research presented a systematic and comprehensive method for analyzing the metabolic and degradation system of FPD enantiomers. This approach aids in understanding the behavior of FPD in the environment and provides valuable insights into their potential risks to human health.

Asunto(s)

Fungicidas Industriales , Microsomas Hepáticos , Microsomas Hepáticos/metabolismo , Animales , Ratas , Fungicidas Industriales/metabolismo , Fungicidas Industriales/química , Humanos , Contaminantes del Suelo/metabolismo , Estereoisomerismo , Medición de Riesgo

RESUMEN

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.

Asunto(s)

Compuestos Epoxi , Fungicidas Industriales , Lagartos , Triazoles , Animales , Triazoles/química , Triazoles/toxicidad , Triazoles/metabolismo , Lagartos/metabolismo , Fungicidas Industriales/química , Fungicidas Industriales/metabolismo , Compuestos Epoxi/metabolismo , Compuestos Epoxi/química , Estereoisomerismo , Hígado/metabolismo , Riñón/metabolismo , Masculino , Distribución Tisular

RESUMEN

The enantioselective environmental behavior of difenoconazole, a widely utilized triazole fungicide commonly detected in agricultural soils, has yet to be comprehensively explored within the earthworm-soil system. To address this research gap, we investigated the bioaccumulation and elimination kinetics, degradation pathways, biotransformation mechanisms, spatial distribution, and toxicity of chiral difenoconazole. The four stereoisomers of difenoconazole were baseline separated and analyzed using SFC-MS/MS. Pronounced enantioselectivity was observed during the uptake phase, with earthworms exhibiting a preference for (2R,4R)-difenoconazole and (2R,4S)-difenoconazole. A total of five transformation products (TPs) were detected and identified using UHPLC-QTOF/MS in the earthworm-soil system. Four of the TPs were detected in both earthworm and soil, and one TP was produced only in eaerthwroms. Hydrolysis and hydroxylation were the primary transformation pathways of difenoconazole in both earthworms and soil. Furthermore, a chiral TP, 3-chloro, 4-hydroxy difenoconazole, was generated with significant enantioselectivity, and molecular docking results indicate the greater catalytic bioactivity of (2R,4R)- and (2R,4S)-difenoconazole, leading to the preferential formation of their corresponding hydroxylated TPs. Furthermore, Mass Spectrometry Imaging (MSI) was applied for the first time to explore the spatial distribution of difenoconazole and the TPs in earthworms, and the "secretory zone" was found to be the dominant region to uptake and biodegrade difenoconazole. ECOSAR predictions highlighted the potentially hazardous impact of most difenoconazole TPs on aquatic ecosystems. These findings are important for understanding the environmental fate of difenoconazole, evaluating environmental risks, and offering valuable insights for guiding scientific bioremediation efforts.

Asunto(s)

Biotransformación , Dioxolanos , Fungicidas Industriales , Oligoquetos , Contaminantes del Suelo , Triazoles , Oligoquetos/metabolismo , Triazoles/metabolismo , Triazoles/química , Fungicidas Industriales/metabolismo , Fungicidas Industriales/química , Animales , Dioxolanos/metabolismo , Dioxolanos/química , Contaminantes del Suelo/metabolismo , Estereoisomerismo , Suelo/química , Espectrometría de Masas en Tándem , Biodegradación Ambiental

RESUMEN

Chlorothalonil (2,4,5,6-tetrachloroisophthalonitrile; TPN) is an environmentally persistent fungicide that sees heavy use in the USA and is highly toxic to aquatic species and birds, as well as a probable human carcinogen. The chlorothalonil dehalogenase from Pseudomonas sp. CTN-3 (Chd, UniProtKB C9EBR5) degrades TPN to its less toxic 4-OH-TPN analog making it an exciting candidate for the development of a bioremediation process for TPN; however, little is currently known about its catalytic mechanism. Therefore, an active site residue histidine-114 (His114) which forms a hydrogen bond with the Zn(II)-bound water/hydroxide and has been suggested to be the active site acid/base, was substituted by an Ala residue. Surprisingly, ChdH114A exhibited catalytic activity with a kcat value of 1.07 s-1, ~ 5% of wild-type (WT) Chd, and a KM of 32 µM. Thus, His114 is catalytically important but not essential. The electronic and structural aspects of the WT Chd and ChdH114A active sites were examined using UV-Vis and EPR spectroscopy on the catalytically competent Co(II)-substituted enzyme as well as all-atomistic molecular dynamics (MD) simulations. Combination of these data suggest His114 can quickly and reversibly move nearly 2 Å between one conformation that facilitates catalysis and another that enables product egress and active site recharge. In light of experimental and computational data on ChdH114A, Asn216 appears to play a role in substrate binding and preorganization of the transition-state while Asp116 likely facilitates the deprotonation of the Zn(II)-bound water in the absence of His114. Based on these data, an updated proposed catalytic mechanism for Chd is presented.

Asunto(s)

Histidina , Nitrilos , Pseudomonas , Pseudomonas/enzimología , Pseudomonas/metabolismo , Nitrilos/metabolismo , Nitrilos/química , Histidina/química , Histidina/metabolismo , Hidrólisis , Biocatálisis , Dominio Catalítico , Fungicidas Industriales/química , Fungicidas Industriales/metabolismo , Halogenación , Hidrolasas/metabolismo , Hidrolasas/química

RESUMEN

The study evaluated Ceremonia 25 EC®, a plant protection product (PPP) containing difenoconazole, in tomato crops, to identify potential risks associated with PPPs, and in addition to this compound, known metabolites from difenoconazole degradation and co-formulants present in the PPP were monitored. An ultra high performance liquid chromatography coupled to quadrupole-Orbitrap mass analyser (UHPLC-Q-Orbitrap-MS) method was validated with a working range of 2 µg/kg (limit of quantification, LOQ) to 200 µg/kg. Difenoconazole degradation followed a biphasic double first-order in parallel (DFOP) kinetic model in laboratory and greenhouse trials, with high accuracy (R2 > 0.9965). CGA-205374, difenoconazole-alcohol, and hydroxy-difenoconazole metabolites were tentatively identified and semi-quantified in laboratory trials by UHPLC-Q-Orbitrap-MS from day 2 to day 30. No metabolites were found in greenhouse trials. Additionally, 13 volatile co-formulants were tentatively identified by gas chromatography (GC) coupled to Q-Orbitrap-MS, detectable up to the 7th day after PPP application. This study provides a comprehensive understanding of difenoconazole dissipation in tomatoes, identification of metabolites, and detection of co-formulants associated with the applied PPP.

Asunto(s)

Dioxolanos , Fungicidas Industriales , Solanum lycopersicum , Triazoles , Solanum lycopersicum/metabolismo , Solanum lycopersicum/química , Dioxolanos/metabolismo , Triazoles/metabolismo , Triazoles/análisis , Triazoles/química , Fungicidas Industriales/metabolismo , Fungicidas Industriales/análisis , Cromatografía Líquida de Alta Presión , Espectrometría de Masas/métodos , Contaminación de Alimentos/análisis , Residuos de Plaguicidas/análisis , Residuos de Plaguicidas/metabolismo

RESUMEN

DNA adduction in the model yeast Saccharomyces cerevisiae was investigated after exposure to the fungicide penconazole and the reference genotoxic compound benzo(a)pyrene, for validating yeasts as a tool for molecular toxicity studies, particularly of environmental pollution. The effect of the toxicants on the yeast's growth kinetics was determined as an indicator of cytotoxicity. Fermentative cultures of S. cerevisiae were exposed to 2 ppm of Penconazole during different phases of growth; while 0.2 and 2 ppm of benzo(a)pyrene were applied to the culture medium before inoculation and on exponential cultures. Exponential respiratory cultures were also exposed to 0.2 ppm of B(a)P for comparison of both metabolisms. Penconazole induced DNA adducts formation in the exponential phase test; DNA adducts showed a peak of 54.93 adducts/109 nucleotides. Benzo(a)pyrene induced the formation of DNA adducts in all the tests carried out; the highest amount of 46.7 adducts/109 nucleotides was obtained in the fermentative cultures after the exponential phase exposure to 0.2 ppm; whereas in the respiratory cultures, 14.6 adducts/109 nucleotides were detected. No cytotoxicity was obtained in any experiment. Our study showed that yeast could be used to analyse DNA adducts as biomarkers of exposure to environmental toxicants.

Asunto(s)

Benzo(a)pireno , Aductos de ADN , Contaminantes Ambientales , Saccharomyces cerevisiae , Aductos de ADN/metabolismo , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Benzo(a)pireno/toxicidad , Benzo(a)pireno/metabolismo , Contaminantes Ambientales/toxicidad , Contaminantes Ambientales/metabolismo , Mutágenos/toxicidad , Mutágenos/metabolismo , ADN de Hongos/genética , Fungicidas Industriales/toxicidad , Fungicidas Industriales/metabolismo

RESUMEN

Biochar and organic compost are widely used in agricultural soil remediation as soil immobilization agents. However, the effects of biochar and compost on microbial community assembly processes in polluted soil under freezingthawing need to be further clarified. Therefore, a freezethaw cycle experiment was conducted with glyphosate (herbicide), imidacloprid (insecticide) and pyraclostrobin (fungicide) polluted to understand the effect of biochar and compost on microbial community assembly and metabolic behavior. We found that biochar and compost could significantly promote the degradation of glyphosate, imidacloprid and pyraclostrobin in freezethaw soil decrease the half-life of the three pesticides. The addition of immobilization agents improved soil bacterial and fungal communities and promoted the transformation from homogeneous dispersal to homogeneous selection. For soil metabolism, the combined addition of biochar and compost alleviated the pollution of glyphosate, imidacloprid and imidacloprid to soil through up-regulation of metabolites (DEMs) in amino acid metabolism pathway and down-regulation of DEMs in fatty acid metabolism pathway. The structural equation modeling (SEM) results showed that soil pH and DOC were the main driving factors affecting microbial community assembly and metabolites. In summary, the combined addition of biochar and compost reduced the adverse effects of pesticides residues.

Asunto(s)

Carbón Orgánico , Compostaje , Glicina , Glifosato , Herbicidas , Neonicotinoides , Nitrocompuestos , Microbiología del Suelo , Contaminantes del Suelo , Estrobilurinas , Neonicotinoides/metabolismo , Neonicotinoides/toxicidad , Nitrocompuestos/metabolismo , Nitrocompuestos/toxicidad , Estrobilurinas/metabolismo , Estrobilurinas/toxicidad , Contaminantes del Suelo/metabolismo , Contaminantes del Suelo/toxicidad , Carbón Orgánico/química , Glicina/análogos & derivados , Glicina/metabolismo , Glicina/toxicidad , Herbicidas/metabolismo , Herbicidas/toxicidad , Carbamatos/metabolismo , Carbamatos/toxicidad , Microbiota/efectos de los fármacos , Fungicidas Industriales/toxicidad , Fungicidas Industriales/metabolismo , Pirazoles/metabolismo , Pirazoles/toxicidad , Insecticidas/metabolismo , Insecticidas/toxicidad , Biodegradación Ambiental , Suelo/química , Bacterias/metabolismo , Bacterias/efectos de los fármacos

RESUMEN

Hexaconazole (HEX) is an azole fungicide widely used in agricultural practices across various countries and numerous studies have reported the toxic effects of HEX, such as endocrine disruption, immunotoxicity, neurotoxicity and carcinogenicity. Despite its widespread agricultural use and toxic effects, the metabolism of HEX is not completely understood, and information on urinary elimination of HEX or its metabolites is limited. Therefore, in the present study, we aimed to identify HEX metabolites in rat and human liver microsomes followed by their in vivo confirmation using a urinary excretion study in rats to identify potential candidate for exposure biomarkers for human biomonitoring studies. From the in vitro assay, a total of 12 metabolites were observed, where the single oxidation metabolites (M5 and M6) were the most abundant metabolites in both rat and human liver microsomes. The triple oxidation followed by dehydration metabolite, M8 (which could also be hexaconazole acid or hydroxy keto-hexaconazole), and the double oxidation metabolite (M9) were the major metabolites found in rat urine and were detectable in rat urine longer than the parent. These metabolites increased with decreasing concentrations of HEX in the rat urine samples. Therefore, metabolites M8, M9 and M5 could be pursued further as potential biomarkers for assessing and monitoring human exposure to HEX.

Asunto(s)

Biomarcadores , Fungicidas Industriales , Microsomas Hepáticos , Triazoles , Animales , Triazoles/metabolismo , Triazoles/orina , Ratas , Microsomas Hepáticos/metabolismo , Humanos , Fungicidas Industriales/orina , Fungicidas Industriales/metabolismo , Biomarcadores/orina , Biomarcadores/metabolismo , Masculino , Ratas Sprague-Dawley , Monitoreo Biológico

RESUMEN

Tebuconazole is a chiral triazole fungicide used globally in agriculture as a racemic mixture, but its enantiomers exhibit significant enantioselective dissimilarities in bioactivity and environmental behaviors. The steric hindrance caused by the tert-butyl group makes it a great challenge to synthesize tebuconazole enantiomers. Here, we designed a simple chemoenzymatic approach for the asymmetric synthesis of (R)-tebuconazole, which includes the biocatalytic resolution of racemic epoxy-precursor (2-tert-butyl-2-[2-(4-chlorophenyl)ethyl] oxirane, rac-1a) by Escherichia coli/Rpeh whole cells expressed epoxide hydrolase from Rhodotorula paludigensis (RpEH), followed by a one-step chemocatalytic synthesis of (R)-tebuconazole. It was observed that (S)-1a was preferentially hydrolyzed by E. coli/Rpeh, whereas (R)-1a was retained with a specific activity of 103.8 U/g wet cells and a moderate enantiomeric ratio (E value) of 13.4, which was remarkably improved to 43.8 after optimizing the reaction conditions. Additionally, a gram-scale resolution of 200 mM rac-1a was performed using 150 mg/mL E. coli/Rpeh wet cells, resulting in the retention of (R)-1a in a 97.0% ees, a 42.5% yields, and a 40.5 g/L/d space-time yield. Subsequently, the synthesis of highly optical purity (R)-tebuconazole (>99% ee) was easily achieved through the chemocatalytic ring-opening of the epoxy-precursor (R)-1a with 1,2,4-triazole. To elucidate insight into the enantioselectivity, molecular docking simulations revealed that the unique L-shaped substrate-binding pocket of RpEH plays a crucial role in the enantioselective recognition of bulky 2,2-disubstituted oxirane 1a.

Asunto(s)

Biocatálisis , Epóxido Hidrolasas , Proteínas Fúngicas , Fungicidas Industriales , Rhodotorula , Triazoles , Rhodotorula/enzimología , Rhodotorula/química , Rhodotorula/metabolismo , Triazoles/química , Triazoles/metabolismo , Fungicidas Industriales/química , Fungicidas Industriales/metabolismo , Fungicidas Industriales/síntesis química , Epóxido Hidrolasas/metabolismo , Epóxido Hidrolasas/química , Estereoisomerismo , Proteínas Fúngicas/química , Proteínas Fúngicas/metabolismo , Simulación del Acoplamiento Molecular , Escherichia coli/enzimología , Escherichia coli/metabolismo

RESUMEN

Plant pathogens have frequently shown multidrug resistance (MDR) in the field, often linked to efflux and sometimes metabolism of fungicides. To investigate the potential role of metabolic resistance in B. cinerea strains showing MDR, the azoxystrobin-sensitive strain B05.10 and -resistant strain Bc242 were treated with azoxystrobin. The degradation half-life of azoxystrobin in Bc242 (9.63 days) was shorter than that in B05.10 (28.88 days). Azoxystrobin acid, identified as a metabolite, exhibited significantly lower inhibition rates on colony and conidia (9.34 and 11.98%, respectively) than azoxystrobin. Bc242 exhibited higher expression levels of 34 cytochrome P450s (P450s) and 11 carboxylesterase genes (CarEs) compared to B05.10 according to RNA-seq analysis. The expression of P450 genes Bcin_02g01260 and Bcin_12g06380, along with the CarEs Bcin_12g06360 in Saccharomyces cerevisiae, resulted in reduced sensitivity to various fungicides, including azoxystrobin, kresoxim-methyl, pyraclostrobin, trifloxystrobin, iprodione, and carbendazim. Thus, the mechanism of B. cinerea MDR is linked to metabolism mediated by the CarE and P450 genes.

Asunto(s)

Botrytis , Carboxilesterasa , Sistema Enzimático del Citocromo P-450 , Farmacorresistencia Fúngica , Proteínas Fúngicas , Fungicidas Industriales , Pirimidinas , Estrobilurinas , Fungicidas Industriales/farmacología , Fungicidas Industriales/metabolismo , Estrobilurinas/farmacología , Estrobilurinas/metabolismo , Estrobilurinas/química , Pirimidinas/farmacología , Pirimidinas/metabolismo , Sistema Enzimático del Citocromo P-450/metabolismo , Sistema Enzimático del Citocromo P-450/genética , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Botrytis/genética , Botrytis/efectos de los fármacos , Carboxilesterasa/metabolismo , Carboxilesterasa/genética , Farmacorresistencia Fúngica/genética , Enfermedades de las Plantas/microbiología , Metacrilatos/farmacología , Metacrilatos/metabolismo

RESUMEN

Biological control is a more sustainable and environmentally friendly alternative to chemical fungicides for controlling Fusarium spp. infestations. In this work, Bacillus siamensis Sh420 isolated from wheat rhizosphere showed a high antifungal activity against Fusarium graminearum as a secure substitute for fungicides. Sh420 was identified as B. siamensis using phenotypic evaluation and 16S rDNA gene sequence analysis. An in vitro antagonistic study showed that Sh420's lipopeptide (LP) extract exhibited strong antifungal properties and effectively combated F. graminearum. Meanwhile, lipopeptides have the ability to decrease ergosterol content, which has an impact on the overall structure and stability of the plasma membrane. The PCR-based screening revealed the presence of antifungal LP biosynthetic genes in this strain's genomic DNA. In the crude LP extract of Sh420, we were able to discover several LPs such as bacillomycin, iturins, fengycin, and surfactins using ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry. Microscopic investigations (fluorescent/transmission electron microscopy) revealed deformities and alterations in the morphology of the phytopathogen upon interaction with LPs. Sh420 LPs have been shown in grape tests to be effective against F. graminearum infection and to stimulate antioxidant activity in fruits by avoiding rust and gray lesions. The overall findings of this study highlight the potential of Sh420 lipopeptides as an effective biological control agent against F. graminearum infestations.IMPORTANCEThis study addresses the potential of lipopeptide (LP) extracts obtained from the strain identified as Bacillus siamensis Sh420. This Sh420 isolate acts as a crucial player in providing a sustainable and environmentally friendly alternative to chemical fungicides for suppressing Fusarium graminearum phytopathogen. Moreover, these LPs can reduce ergosterol content in the phytopathogen influencing the overall structure and stability of its plasma membrane. PCR screening provided confirmation regarding the existence of genes responsible for biosynthesizing antifungal LPs in the genomic DNA of Sh420. Several antibiotic lipopeptide compounds were identified from this bacterial crude extract using ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry. Microscopic investigations revealed deformities and alterations in the morphology of F. graminearum upon interaction with LPs. Furthermore, studies on fruit demonstrated the efficacy of Sh420 LPs in mitigating F. graminearum infection and stimulating antioxidant activity in fruits, preventing rust and gray lesions.

Asunto(s)

Bacillus , Fungicidas Industriales , Fusarium , Antifúngicos/química , Fusarium/genética , Fungicidas Industriales/metabolismo , Fungicidas Industriales/farmacología , Antioxidantes/farmacología , Antioxidantes/metabolismo , Lipopolisacáridos/metabolismo , Lipopéptidos/farmacología , ADN/metabolismo , Ergosterol , Enfermedades de las Plantas/prevención & control , Enfermedades de las Plantas/microbiología

RESUMEN

Triadimefon, a type of triazole systemic fungicide, has been extensively used to control various fungal diseases. However, triadimefon could lead to severe environmental pollution, and even threatens human health. To eliminate triadimefon residues, a triadimefon-degrading bacterial strain TY18 was isolated from a long-term polluted site and was identified as Enterobacter hormaechei. Strain TY18 could grow well in a carbon salt medium with triadimefon as the sole nitrogen source, and could efficiently degrade triadimefon. Under triadimefon stress, a total of 430 differentially expressed genes (DEGs), including 197 up-regulated and 233 down-regulated DEGs, were identified in strain TY18 using transcriptome sequencing (RNA-Seq). Functional classification and enrichment analysis revealed that these DEGs were mainly related to amino acid transport and metabolism, carbohydrate transport and metabolism, small molecule and pyrimidine metabolism. Interestingly, the DEGs encoding monooxygenase and hydrolase activity acting on carbon-nitrogen were highly up-regulated, might be mainly responsible for the metabolism in triadimefon. Our findings in this work suggest that strain E. hormaechei TY18 could efficiently degrade triadimefon for the first time. They provide a great potential to manage triadimefon biodegradation in the environment successfully.

Asunto(s)

Biodegradación Ambiental , Enterobacter , Fungicidas Industriales , Perfilación de la Expresión Génica , Triazoles , Enterobacter/genética , Enterobacter/metabolismo , Enterobacter/aislamiento & purificación , Fungicidas Industriales/farmacología , Fungicidas Industriales/metabolismo , Triazoles/farmacología , Transcriptoma

RESUMEN

Fenhexamid are fungicides that act against plant pathogens by inhibiting sterol biosynthesis. Nonetheless, it can trigger endocrine disruption and promote breast cancer cell growth. In a recent study, we investigated the mechanism underlying the lipid accumulation induced by fenhexamid hydroxyanilide fungicides in 3 T3-L1 adipocytes. To examine the estrogen receptor alpha (ERα)-agonistic effect, ER transactivation assay using the ERα-HeLa-9903 cell line was applied, and fenhexamid-induced ERα agonist effect was confirmed. Further confirmation that ERα-dependent lipid accumulation occurred was provided by treating 3 T3-L1 adipocytes with Methyl-piperidino-pyrazole hydrate (MPP), an ERα-selective antagonist. Fenhexamid mimicked the actions of ERα agonists and impacted lipid metabolism, and its mechanism involves upregulation of the expression of transcription factors that facilitate adipogenesis and lipogenesis. Additionally, it stimulated the expression of peroxisome proliferator-activated receptor (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), fatty acid synthase (FAS), and sterol regulatory element-binding protein 1 (SREBP1) and significantly elevated the expression of fatty acid-binding protein 4 (FABP4). In contrast, in combination with an ERα-selective antagonist, fenhexamid suppressed the expression of adipogenic/lipogenic transcription factors. These results suggest that fenhexamid affects the endocrine system and leads to lipid accumulation by interfering with processes influenced by ERα activation.

Asunto(s)

Amidas , Receptor alfa de Estrógeno , Fungicidas Industriales , Ratones , Animales , Receptor alfa de Estrógeno/genética , Receptor alfa de Estrógeno/metabolismo , Fungicidas Industriales/toxicidad , Fungicidas Industriales/metabolismo , Adipocitos/metabolismo , Adipogénesis , Metabolismo de los Lípidos , Proteína alfa Potenciadora de Unión a CCAAT/metabolismo , Factores de Transcripción/metabolismo , Factores de Transcripción/farmacología , Lípidos , Células 3T3-L1 , PPAR gamma/metabolismo

RESUMEN

Pathogenic fungi pose a significant threat to crop yields and human healthy, and the subsequent fungicide resistance has greatly aggravated these agricultural and medical challenges. Hence, the development of new fungicides with higher efficiency and greater environmental friendliness is urgently required. In this study, luvangetin, isolated and identified from the root of Zanthoxylum avicennae, exhibited wide-spectrum antifungal activity in vivo and in vitro. Integrated omics and in vitro and in vivo transcriptional analyses revealed that luvangetin inhibited GAL4-like Zn(II)2Cys6 transcriptional factor-mediated transcription, particularly the FvFUM21-mediated FUM cluster gene expression, and decreased the biosynthesis of fumonisins inFusarium verticillioides. Moreover, luvangetin binds to the double-stranded DNA helix in vitro in the groove mode. We isolated and identified luvangetin, a natural metabolite from a traditional Chinese edible medicinal plant and uncovered its multipathogen resistance mechanism. This study is the first to reveal the mechanism underlying the antifungal activity of luvangetin and provides a promising direction for the future use of plant-derived natural products to prevent and control plant and animal pathogenic fungi.

Asunto(s)

Fumonisinas , Fungicidas Industriales , Fusarium , Zanthoxylum , Animales , Humanos , Fungicidas Industriales/farmacología , Fungicidas Industriales/metabolismo , Antifúngicos/farmacología , Antifúngicos/metabolismo , Zanthoxylum/metabolismo , Fumonisinas/metabolismo