Tracking mitigation and expression patterns of the antibiotic resistome during full-scale livestock wastewater treatment: Comparison between summer and winter conditions.

Livestock wastewater is a critical reservoir of antibiotic resistance genes (ARGs) that poses significant public health risks. This study comprehensively evaluated the seasonal dynamics and associated risks of ARGs in a full-scale livestock wastewater treatment plant using an integrated metagenomic and metatranscriptomic approach. The results showed that untreated livestock wastewater harbored high abundance (4.69 ± 1.93 copies/cell, GPC) and high diversity (838 subtypes) ARGs comparable to and sometimes higher than those found in human and pig guts. These ARGs primarily encode resistance to common veterinary antibiotics including Tetracycline (7.4 %), Macrolide-Lincosamide-Streptogramin (10.4 %), and Aminoglycoside (13.2 %). Notably, 39 % of livestock wastewater ARGs (based on relative abundance) were further identified as high-risk (i.e., mobilizable, pathogen-carriable and clinically relevant) ARGs. Moreover, these livestock-associated ARGs showed marked seasonal variation in removal efficacy: summer and autumn treatments reduced ARG abundance by 48-55 % in biological units (anaerobic digestion and activated sludge), while winter witnessed a 40 % increase in ARGs, driven by certain plasmid-associated ARGs notably enriched during anaerobic digestion but inefficiently removed in the subsequent activated sludge process due to slow microbial community turnover. Transcriptional profiling revealed 66 % to 93 % of ARGs were actively expressed, with disproportionately high activity of high-risk ARGs (e.g., erm(B) and cfr) in winter. The findings emphasize spreading potential of antibiotic resistome through land application of winter livestock wastewater treatment and suggest that temperature control of treatment facilities or membrane filtration might be a promising strategy to remove high-risk ARGs and their hosts. This study provides actionable insights for optimizing livestock wastewater treatment to mitigate spread risks of antimicrobial resistance.