Coexistence between antibiotic resistance genes and metal resistance genes in manure-fertilized soils

Antibiotics and metals introduced during manure application are believed to be the major drivers of the bloom and spread of antibiotic resistance genes (ARGs) and metal resistance genes (MRGs) in agricultural environments. However, the coexistence of multidrug resistance (MDR) and metal resistance and the effects of manure application on the coexistence are less studied. Here, we profiled the coexistence patterns of 9919 MDR genes (MDRGs) and 25,312 MRGs in paddy fields with manure or inorganic fertilization. The abundances of both MDRGs and MRGs were higher under manure fertilization than inorganic fertilization. Network analysis determined that coexistence between MDRGs and MRGs was much more frequent in manure-fertilized soils than in inorganic-fertilized soils. Manure application drove this coexistence by not only directly introducing resistance genes but also increasing co-selection through antibiotics and metals, and stimulating their potential common hosts, including species of Actinobacterium WWH12, Bacillus, Geobacter, Solirubrobacter, Acidobacteriales, Bacillales, Chloroflexi, Methyloligellaceae and Xanthobacteraceae. Soil antibiotic resistance was significantly predicted by the coexistence of major multidrug efflux genes (mfs, mex, abc, mate and smr) and MRGs. The coexistence between MDRGs and MRGs explained 18% of the increase in soil antibiotic resistance. Such coexistence is nonnegligible and permits the promotion of soil antibiotic resistance, thus posing a potential threat to both agroecosystems and humans.