Cu and Zn exert a greater influence on antibiotic resistance and its transfer than doxycycline in agricultural soils.

Livestock manure is a main source of heavy metals, antibiotics and antibiotic resistance genes (ARGs) in agricultural soils. The co-existence of heavy metals and ARGs needs to be systematically studied, since manure application is greatly encouraged. In this study, we examined soils for alterations in antibiotic resistance where doxycycline, Cu, and Zn were added equivalent to those found in typical pig manure applications. The results indicated that high levels of Cu inhibited soil respiration and urease for the first 10 days. Metagenomic analysis demonstrated that Cu and Zn additions caused profound alterations in bacterial community, metal resistance genes (MRGs) and mobile genetic elements. Among the differential ARGs, efflux pump genes took a significantly high ratio compared with control for the first 5 days, emphasizing their important roles in the profile of antibiotic resistance. Moreover, the number of differential MRGs was < 30 for doxycycline treatment, but 66-87 for Cu and Zn treatments. The number of differential integrative and conjugative elements was 3 for doxycycline treatment, and 6-13 for Cu and Zn treatments. Overall, high Cu and Zn levels caused a greater influence than did doxycycline on bacterial communities and transfer of antibiotic resistance in soil.