CoFe₂O₄-Modified Biochar Is Associated with Microbial Community and Antibiotic Resistance Gene Profiles in Vertical...

The widespread occurrence of antibiotics in aquatic environments has raised concern over the dissemination of antibiotic resistance genes (ARGs). Although vertical flow constructed wetlands (VFCWs) efficiently remove antibiotics from wastewater, the relationships between substrate configuration, microbial communities, and ARG abundance remain poorly understood. Here, five laboratory-scale VFCWs with different substrate configurations, including cobalt ferrite (CoFe₂O₄)-modified biochar and its composite with zeolite, were evaluated using synthetic domestic wastewater amended with oxytetracycline (OTC). CoFe₂O₄ modification increased the specific surface area and pore volume of biochar and altered its adsorption behavior. All VFCWs achieved high OTC removal (>97%), despite differences in OTC accumulation within the substrate. In contrast, ARG and mobile genetic element abundances and bacterial community profiles varied among reactors. Combined tetA and tetX abundance ranged from 2.55 × 10² to 1.90 × 10⁵ copies g^-1, with the CoFe₂O₄-modified biochar-zeolite reactor showing the lowest mean ARG abundance. Exploratory co-occurrence analysis identified associations of Acinetobacter with tetA and Flavobacterium with tetA and tetX (nominal Spearman p < 0.05), although these correlations do not establish ARG host identity or horizontal gene transfer. Overall, the findings indicate that microbial ecological patterns associated with ARG abundance may differ among VFCWs even when antibiotic removal efficiencies are comparable.