Biodegradation of Antibiotics and Pharmaceuticals in Activated Sludge: Insights from OECD 301F Tests and 16S rRNA Amplicon Sequencing

Wastewater treatment plants (WWTPs) are recognized as hotspots for the release of pharmaceuticals and antibiotics into aquatic environments. In the current study, we assessed the biodegradation potential of 26 pharmaceuticals (PhACs) using the OECD 301F manometric respirometry test with activated sludge from a WWTP in Mascara (Algeria) by assessing oxygen consumption and characterized the associated microbial community using 16S rRNA amplicon sequencing combined with culture-based isolation. Biodegradability, expressed as theoretical oxygen demand (ThOD) percent, exhibited marked variability among the compounds. After 28 days, spiramycin (93.4%) and doxycycline (66.7%) met the criteria for ready biodegradability, whereas several antibiotics (e.g., gentamicin, sulfamethoxazole, and clarithromycin) and pharmaceuticals (e.g., fluvastatin, esomeprazole, and tramadol) exhibited low or negligible biodegradation (<30%). Amplicon-based community analysis indicated that the dominate phyla in the activated sludge were Proteobacteria and Firmicutes, with prevalent genera including Pseudomonas, Enterococcus, Bacillus, Citrobacter, Klebsiella, and Enterobacter. Multivariate analysis identified six clusters of microbial communities associated with different enrichment conditions. In parallel, 39 bacterial strains were isolated from biodegradation assays and taxonomically affiliated with Pseudomonas, Enterococcus, Bacillus, Klebsiella, and Citrobacter, supporting their potential involvement in the PhAC transformation processes. Overall, the results demonstrated a compound-specific biodegradation potential in activated sludge and highlighted the association between microbial community composition and biodegradation performance. While oxygen consumption-based assays provide valuable screening information, further studies integrating chemical analyses and functional omics are required to confirm specific biodegradation pathways and the roles of individual taxa.