Residual enrofloxacin in cattle manure increased persistence and dissemination risk of antibiotic resistance genes during anaerobic digestion

Anaerobic digestion is a common approach to dispose and recycle livestock manures, and the agricultural application of anaerobic digestives represents an important pathway of spreading antibiotic resistance genes (ARGs) from livestock manures to soils. Enrofloxacin is a clinically important fluoroquinolone antibiotic with high residual concentrations in livestock manure, and propagation of fluoroquinolone resistance genes poses a huge risk to public health. Compared with other antibiotics, enrofloxacin is relatively durable in anaerobic digestion system. However, its effect on the persistence of ARGs during anaerobic digestion and its mechanism are not clear. In this study, we investigated effects of 0, 4, and 8 mg/L enrofloxacin on the abundance, persis-tence, and transferring risk of five plasmid-mediated fluroquinolone ARGs and five typic clinically important non-fluoroquinolone ARGs during cattle manure digestion. The responses of integrons and microbial commu-nities to enrofloxacin were assessed to uncover the underlying mechanisms. All the ten detected ARGs were highly persistent in anaerobic digestion, among them seven ARGs increased over 8.2 times after digestion. Network analysis revealed that the potential hosts of ARGs were critical functional taxa during anaerobic digestion, which can explain the high persistence of ARGs. Residual enrofloxacin significantly increased the abundance of aac(6 ')-ib-cr, sul1, intI1, and intI2 throughout the digestion, but had no impact on the other ARGs, demonstrating its role in facilitating horizontal gene transfer of the plasmid-mediated aac(6 ')-ib-cr. The influence of enrofloxacin on microbial communities disappeared at the end of digestion, but the ARG profiles remained distinctive between the enrofloxacin treatments and the control, suggesting the high persistence of enrofloxacin induced ARGs. Our results suggested the high persistence of ARGs in anaerobic digestion system, and highlighted the role of residual enrofloxacin in livestock manure in increasing dissemination risk of fluroquinolone resistance genes.

Automated summary

Anaerobic digestion is a common method for disposing and recycling livestock manures. However, this process can contribute to the spread of antibiotic resistance genes (ARGs) from manures to soils. In this study, the researchers investigated the effects of enrofloxacin, a commonly used antibiotic, on the persistence and transfer of ARGs during cattle manure digestion. They found that enrofloxacin significantly increased the abundance of certain ARGs and played a role in facilitating the transfer of plasmid-mediated resistance genes. The results indicate the high persistence of ARGs in anaerobic digestion systems and highlight the importance of residual antibiotics in increasing the dissemination of resistance genes.