The presence of in situ sulphamethoxazole degraders and their interactions with other microbes in activated sludge as revealed by DNA stable isotope probing and molecular ecological network analysis

Abstract

Wastewater treatment plants (WWTPs) are the main hotspots for the release of antibiotics, including the widely used sulphonamides. Microbes play important roles in eliminating sulphonamides in WWTPs, and knowledge about these degraders and their interactions within the microbial community is crucial for operating and optimising WWTPs. In the present study, stable isotope probing (SIP) coupled with high-throughput sequencing as culture-independent approach revealed four operational taxonomic units (OTUs) involved in sulphamethoxazole (SMX) degradation in activated sludge. Except for the OTU affiliated with Gammaproteobacteria, the others have not been previously reported to possess the ability to metabolise SMX. The isolated SMX degrader by culture-dependent method did not participate in SMX biodegradation in situ according to the SIP analysis, and showed weak correlations with other members in the activated sludge. The complex interactions between in situ active SMX degraders and non-degrading microbes might explain our failure to isolate these degraders. In addition, sul1 genes associated with SMX resistance were also labelled with 13C, suggesting that they might benefit from SMX degradation and/or originate from the active SMX degraders. These findings broaden our understanding of the diversity of SMX-degrading microbes and their associated characteristics in WWTPs. Keywords: Antibiotic resistance genes, Activated sludge, Sulphamethoxazole-degrading bacteria, Molecular ecological network analysis, Stable isotope probing, Sulphonamide biodegradation