Interplay of antibiotic resistance genes in anaerobic digestion: Focus on full-scale plants treating livestock waste and sewage sludge

This study aimed to bridge a research gap — understanding the dynamics of antibiotic resistance genes (ARGs) and their microbial associations operating under different microbial communities and regional conditions — by studying six full-scale anaerobic digestion (AD) plants treating livestock waste (LW) and sewage sludge (SS). Based on residual gene fractions (RGFs) calculated from absolute abundance, distinct behaviors of ARGs were observed depending on the substrate type. In plants treating LW, increased RGF values (i.e., > 1.0) were consistently found for qnr D, bla _TEM, bla _SHV, and bla _CTX, while the majority of other ARGs exhibited decreased abundances. In contrast, in SS-treated plants, tet T and sul 3 showed marked enrichment. In addition, tet C, tet D, tet E, tet M, tet O, tet Q, tet W, tet X, and qnr D in all plants exhibited high RGFs based on relative abundance, indicating potential horizontal gene transfer during AD. Correlation analysis revealed strong associations (Spearman's correlation coefficient r > 0.9, p -value < 0.01) between specific bacterial genera (e.g., Sphingobacterium , Butyrivibrio , Raoultibacter) and tetracycline or sulfonamide resistance genes. Notably, Methanobrevibacter , a hydrogenotrophic methanogen, showed significant correlation with tet W, highlighting the potential role of archaea in the persistence of ARGs. These findings underscore the need for comprehensive investigations of full-scale AD plants operated under various conditions to develop more effective strategies for mitigating ARGs and reducing their potential release into the environment.