Selective attenuation of wastewater resistomes and residual ARG-MGE co-occurrence in semi-arid lagoon-treated effluents.
Abstract
Antimicrobial resistance genes (ARGs), mobile genetic elements (MGEs), and clinically relevant bacterial taxa are environmental-health concerns in wastewater-impacted ecosystems. However, the capacity of lagoon systems to attenuate these AMR-related signals before effluent discharge or reuse remains insufficiently characterized in semi-arid regions. Shotgun metagenomics was applied to 34 influent and treated-effluent samples collected approximately weekly during January-March 2025 from two Moroccan lagoon systems: Settat (urban, n = 18) and Ouled Said (rural, n = 16). TPM-normalized taxonomic, resistome, and mobilome profiles were analyzed using diversity metrics, PERMANOVA, differential-abundance testing, and correlation networks. Lagoon treatment produced site-dependent microbial restructuring, with a marked shift in Ouled Said (Aitchison PERMANOVA, R2 = 0.376, p = 0.002) but a weaker, non-significant shift in Settat (R2 = 0.125, p = 0.054). ESKAPE-associated bacteria declined from approximately 38% to 32% of CARD-annotated taxa but remained detectable in effluents. The resistome was broadly attenuated, but last-resort antibiotic resistance genes (LR-ARGs) responded unevenly: most mcr variants decreased by 82.4-89.9%, with significant reductions for several variants, whereas blaVIM-1 and tet(X) showed limited or non-significant reductions. The mobilome declined, although transposases and insertion sequences remained prominent. Network analysis indicated denser residual statistical ARG-MGE-taxa co-occurrence after treatment, without implying increased genetic mobility or HGT. Overall, the systems acted as selective, context-dependent barriers, reducing AMR-related burdens while retaining clinically relevant residual signals. The contrasting site responses indicate that treatment performance should not be assumed to be uniform across lagoon systems and support targeted effluent surveillance for safer discharge and reuse.