Profiling Antibiotic Resistance Genes and Microbial Community in Greenhouse Soil by Using SmartChip qPCR and Miseq Platform

Abstract

Since Alexander Fleming’s discovery of penicillin in 1928, antibiotics have contributed greatly to prolonging human life and improving the level of public health. However, rapid development and overuse of antibiotics led to side effects such as the emergence of antibiotic-resistant bacteria (ARB). ARB and antibiotic resistance genes (ARGs) were widely distributed in the environment, especially in the agriculture field using livestock manure compost and agricultural antibiotics. Antibiotics overused in livestock led to excrement after incomplete digestion and cannot be completely removed in composting process and sprayed on the soil. Residual antibiotics in compost contribute to the accumulation and dissemination of ARB and ARGs in soil by selective pressure. The main objective of this study was to profile ARGs and microbial community structure in greenhouse soil using livestock manure compost in contrast with control soil and to select core ARGs and indicator species that can serve as basic data for ARGs contamination. Soil sampling was performed at a total of 13 greenhouses in the Jeolla-do area, and control soil was sampled at a total of 4 mountain soil. The sampled soil used a soil component analysis through soil component analyzer, and after extracting gDNA from soil samples, SmartChip qPCR for ARGs analysis was performed, and Miseq analysis for microbial community analysis was performed. Raw data were converted to fit statistical analysis and various visualization of PCA, NMDS, and heatmap were performed, and as a result, it was found that the greenhouse soil and control soil differed statistically significantly (p < 0.05) in ARGs distribution and microbial community with groups such as sample type, soil type, cultivation year and crop. In addition, a total of 19 (16 ARGs and 3 MGEs) core ARGs and 5 indicator species were selected. There is a significant difference between greenhouse soil and control soil in aspects of ARGs and microbial community, and core ARGs selected in the heatmap, and indicator species can be used as a standard in greenhouse soils.