Metagenomic analysis reveals the fate of antibiotic resistance genes in two-stage and one-stage anaerobic digestion of waste activated sludge

Waste activated sludge (WAS) from wastewater treatment plants is an important reservoir of antibiotic resistance genes (ARGs). The fate of ARGs in this process was not revealed previously. The present study applied metagenomic approach to examine the occurrence and fate of ARGs in thermophilic alkaline fermentation followed by mesophilic anaerobic digestion (TM), by comparison with mesophilic alkaline fermentation followed by mesophilic anaerobic digestion (MM) and one-stage mesophilic anaerobic digestion (M) process. The removal efficiency of two-stage anaerobic digestion (AD) to total ARGs is higher than that of one-stage AD. The hydrogen and methane production stages of two-stage AD processes have dissimilar impact on the fate of ARGs. Macrolide, lincosamide, and streptogramin (MLS) resistance genes were enriched, especially in the hydrogen production reactors of TM and MM processes. Statistical analysis of metagenomic profiles analysis suggested that bacA may be the differential ARG subtype of two-stage AD process. ARG-like sequences encoding antibiotic efflux pump, antibiotic inactivation and antibiotic target alteration mechanisms were identified as the dominant ARGs resistance mechanisms in all samples. Procrustes analysis showed that microbial community composition structured the resistome. Co-occurrence patterns between ARGs and microbial phylogeny revealed that 26 bacterial species might be potential hosts of 94 ARG subtypes.