A novel bidirectional regulation mechanism of mancozeb on the dissemination of antibiotic resistance.

The high abundance of antibiotic resistance genes (ARGs) in the fungicide residual environment, posing a threat to the environment and human health, raises the question of whether and how fungicide promotes the prevalence and dissemination of antibiotic resistance. Here, we reported a novel mechanism underlying bidirectional regulation of a typical heavy-metal-containing fungicide mancozeb on the horizontal transfer of ARGs. Our findings revealed that mancozeb exposure significantly exerted oxidative and osmotic stress on the microbes and facilitated plasmid-mediated ARGs transfer, but its metallic portions (Mn and Zn) were potentially utilized as essential ions by microbes for metalating enzymes to deal with cellular stress and thus reduce the transfer. The results of transcriptome analysis with RT-qPCR confirmed that the expression levels of cellular stress responses and conjugation related genes were drastically altered. It can be concluded mancozeb bidirectionally regulated the ARGs dissemination which may be attributed to the diverse effects on the microbes by its different portions. This novel mechanism provides an updated understanding of neglected fungicide-triggered ARGs dissemination and crucial insight for comprehensive risk assessment of fungicides.

Determination and Dietary Intake Risk Assessment of Pesticide Residues in Fritillariae Thunbergii Bulbs and Cultivated Soils.

BACKGROUND: Pesticide residues in traditional Chinese medicines pose a potential risk to human health. However, little is known about the characteristics of pesticide residues in the fritillariae thunbergii bulbs (FTB). OBJECTIVE: This study aims to establish a method for the determination of pesticide multi-residues in FTB and then measured their residual levels in the FTB collected from nine cultivation regions. METHODS: A modified QuEChERS method coupled with GC/UPLC was used to determine the residues of 24 pesticides in the FTB and soil samples. RESULTS: The recoveries of these pesticides at three concentrations were 72.17-112.48% in the FTB and 70.92-113.74% in the soil with RSD < 11.83%, and the LOD and LOQ ranged from 0.005-10 µg/kg and 0.011-22 µg/kg, respectively. A total of 13 pesticide residues were detected in the FTB samples with the residual levels of 0.0011-509.63 µg/kg, which were all below the referred MRLs in other Chinese herbs or food. Meanwhile, dietary intake risk assessment showed that the risk of pesticide residues in the FTB was acceptable to consumers. CONCLUSIONS: This study developed a method for the determination of pesticide multi-residues in the FTB and cultivated soil samples, and furthermore the dietary intake risk of pesticide residues in the FTB is safe. HIGHLIGHTS: A modified QuEChERS method was established for the determination of 24 pesticides in the FTB with high sensitivity, accuracy, and precision.

Exposure to fungicide difenoconazole reduces the soil bacterial community diversity and the co-occurrence network complexity.

Difenoconazole is a triazole fungicide that is widely used worldwide and has been frequently detected in agricultural soils, but its ecotoxicological effect on soil bacterial community remains unknown. Here, the degradation of difenoconazole and its effect on soil bacterial communities were investigated at three concentrations in five different agricultural soils. Difenoconazole degraded faster in non-sterilized soils than in sterilized soils, suggesting that biodegradation is a major contributor to the dissipation of difenoconazole in soils. Exposure to high concentrations of difenoconazole decreased the soil bacterial community diversity in most soils, and this influence was aggravated with the increasing concentration. The effect of difenoconazole on soil bacterial community diversity was also enhanced with the increasing content of organic matter and total nitrogen in soils. Moreover, difenoconazole exposure also reduced the soil bacterial community network complexity and exhibited a concentration-dependent characteristic. In addition, a core bacterial community (57 operational taxonomic units, OTUs) was identified, and some core OTUs were strongly linked to the degradation of difenoconazole in soils. It is concluded that high concentrations of difenoconazole may have a significant effect on the soil bacterial communities, and co-occurrence networks may improve the ecotoxicological risk assessment of fungicides on soil microbiome.

Tracking resistomes, virulence genes, and bacterial pathogens in long-term manure-amended greenhouse soils.

Organic manure has been implicated as an important source of antibiotic resistance genes (ARGs) in agricultural soils. However, the profiles of biocide resistance genes (BRGs), metal resistance genes (MRGs) and virulence genes (VGs) and their bacterial hosts in manure-amended soils remain largely unknown. Herein, a systematic metagenome-based survey was conducted to comprehensively explore the changes in resistomes, VGs and their bacterial hosts, mobile genetic elements (MGEs), and pathogenic bacteria in manure-amended greenhouse soils. Many manure-borne ARGs, BRGs, MRGs, VGs, and bacterial pathogens could be transferred into soils by applying manures, and their abundance and diversity were markedly positively correlated with greenhouse planting years (manure amendment years). The main ARGs transferred from manures to soils conferred resistance to tetracycline, aminoglycoside, and macrolide-lincosamide-streptogramin. Both statistical analysis and gene arrangements showed a good positive co-occurrence pattern of ARGs/BRGs/MRGs/VGs and MGEs. Furthermore, bacterial hosts of resistomes and VGs were significantly changed in the greenhouse soils in comparison with the field soils. Our findings confirmed the migration and dissemination of resistomes, VGs, and bacterial pathogens, and their accumulation and persistence were correlated with the continuous application of manures.

Enterobacteriaceae predominate in the endophytic microbiome and contribute to the resistome of strawberry.

Antibiotic resistance genes (ARGs) harbored by plant microbiomes have been implicated as a potential risk to public health via food chain, especially directly edible fruits and vegetables. Here, we investigated the microbiome and antibiotic resistome in soil-strawberry ecosystem using shotgun metagenomic sequencing. The results showed that the enterobacterial population dominated the endophytes of strawberry fruits. Moreover, 85 subtypes of ARGs, including several clinically important ARGs, were detected in the strawberry fruit metagenomes. Additionally, host tracking analysis in combination with antibiotic-resistant bacterial isolate screening suggested that fruit-borne ARGs were mainly carried by members of the Enterobacteriaceae family. Unexpectedly, most of fruit-borne isolates were found to be resistant to several clinically important antimicrobials, e.g., erythromycin and cephalexin. Our findings provide broad insights into endophytic antibiotic resistomes of direct edible strawberry fruits and their potential hosts, and highlight the potential exposure risks of plant microbiomes to the human food chain.

Fungicides enhanced the abundance of antibiotic resistance genes in greenhouse soil.

Long-term substantial application of fungicides in greenhouse cultivation led to residual pollution in soils and then altered soil microbial community. However, it is unclear whether residual fungicides could affect the diversity and abundance of antibiotic resistance genes (ARGs) in greenhouse soils. Here, the dissipation of fungicides and its impact on the abundance of ARGs were determined using shotgun metagenomic sequencing in the greenhouse and mountain soils under laboratory conditions. Our results showed the greenhouse soils harbored more diverse and abundant ARGs than the mountain soils. The application of carbendazim, azoxystrobin, and chlorothalonil could increase the abundance of total ARGs in the greenhouse soils, especially for those dominant ARG subtypes including sul2, sul1, aadA, tet(L), tetA(G), and tetX2. The abundant ARGs were significantly correlated with mobile genetic elements (MGEs, e.g. intI1and R485) in the greenhouse soils but no significant relationship in the mountain soils. Meanwhile, the co-occurrence patterns of ARGs and MGEs, e.g., sul2 and R485, sul1 and transposase, were further verified via the genetic arrangement of genes on the metagenome-assembled contigs in the greenhouse soils. Additionally, host tracking analysis indicated that ARGs were mainly carried by enterobacteria in the greenhouse soils but actinomyces in the mountain soils. These findings confirmed that some fungicides might serve as the co-selectors of ARGs and elevated their abundance via MGEs-mediated horizontal gene transfer in the greenhouse soils.

Deposition, dissipation, and minimum effective dosage of the fungicide carbendazim in the pepper-field ecosystem.

BACKGROUND: In order to reduce application dosage of carbendazim (CBD), the effects of different droplet sizes and application rate on initial deposition, retention rate, and field control efficacy of CBD in the pepper plant (Capsicum annuum L.) were studied under laboratory and field conditions. RESULTS: Based on eight types of nozzles at six concentrations of 646.8, 582.2, 517.5, 414.0, 310.5, and 258.7 g a.i./hm2 , the initial deposition amount of CBD in the pepper plant showed a first increased and then decreased trend with the increasing application dosages. The initial deposition amount of CBD in the pepper leaves and stems was highest when using the nozzle ST110-03 at 200 µm volume median diameter (VMD) and 350 L/hm2 application rate. HPLC-MS/MS analysis revealed that CBD was first converted to 2-aminobenzimidazole, which then transformed into benzimidazole and 2-hydroxybenzimidazole by deamination and hydroxylation, and ultimately mineralized to carbon dioxide and water via the cleavage of the imidazole ring in pepper plant. The dissipation rate of CBD was fruits > roots/stems > leaves > soils. After 7 days of application, the field efficacy of CBD at six concentrations on pepper anthracnose (Colletotrichum spp.) were 94.1%, 91.3%, 82.3%, 76.5%, 47.0%, and 41.2%, respectively. CONCLUSION: The results revealed the deposition and dissipation characteristics of CBD, and the minimum effective dosage on pepper anthracnose decreased to 80% of the minimum recommended dose in the open field pepper ecosystem. The study will contribute to the CBD reduction by improving its utilization rate in the pepper-field ecosystem. © 2019 Society of Chemical Industry.