Occurrence and transfer characteristics of bla genes among Escherichia coli in anaerobic digestion systems treating swine waste.

Livestock waste is a known reservoir of Escherichia coli (E. coli) carrying clinically important CTX-M-type extended-spectrum β-lactamase genes (bla), however, the occurrence and transfer characteristics of bla genes during anaerobic digestion (AD) remain unclear. Herein, four full-scale and two parallel lab-scale AD systems treating swine waste under ambient and mesophilic conditions were investigated by both molecular- and culture-based methods to reveal the occurrence and transfer behaviors of bla genes during AD. Real-time TaqMan polymerase chain reaction revealed 1.3 × 10-6.8 × 10 and 3.0 × 10-7.0 × 10 copies/mL of bla groups 1 and 9 in all feeding substrates. While AD reduced the absolute abundance of groups 1 and 9 by 0.63-2.24 and 0.08-1.30 log (P < 0.05), 5.0 × 10-4.1 × 10 and 1.1 × 10-3.5 × 10 copies/mL of groups 1 and 9 remained in the anaerobic effluent, respectively. In total, 141 bla-carrying E. coli isolates resistant to cefotaxime were obtained from the AD reactors. Whole-genome sequencing showed that bla mainly carried by E. coli ST155 was the most frequently detected group 9 subtype in the feeding substrate; whereas bla associated with the dominant clones E. coli ST6802 and ST155 became the major subtype in AD effluent. Furthermore, bla was flanked by ΔIS26 upstream and ΔIS903B downstream. The ΔIS26-bla-ΔIS903B element was mainly located on the IncHI2 plasmid in E. coli ST48 and ST6802 and also the IncFIB plasmid in ST155 in anaerobic effluent. Conjugation assays showed that the plasmids harboring bla could be successfully transferred at a frequency of 10-10 cells per recipient cell. This study revealed that bla genes remained in both the full-scale and lab-scale AD effluents of swine waste. Thus, additional efforts should be implemented to block the discharge and spread of antibiotic resistance genes to the environment.