Genome-resolved metagenomics of a neotropical wastewater treatment plant reveals community-wide metal and biocide-resistance genes and the diversity of phosphate-accumulating and denitrifying bacteria

Wastewater treatment plants (WWTPs) rely on complex microbial communities to remove nutrients and contaminants, yet tropical systems remain poorly characterized. This study applied genome-resolved metagenomics to investigate the taxonomic composition, functional potential, and adaptive traits of activated sludge from a domestic neotropical WWTP. From six sludge samples sampled across two different seasons in Costa Rica (rainy and dry), we reconstructed 177 non-redundant metagenome-assembled genomes (MAGs), of which 62 % were high-quality (90 % completeness and less 5 % contamination). Biocide and metal resistance genes (BMRGs) were widespread, particularly in Actinomycetia and Gammaproteobacteria, reflecting adaptation to selective environmental pressures. We identified 64 potential phosphate-accumulating organisms (PAOs), 78 denitrifying bacteria (DBs), and 32 denitrifying PAOs (DPAOs), with some MAGs showing negative correlations with effluent nitrogen and phosphorus concentrations, indicating roles in nutrient removal. While complete denitrification pathways were observed, nitrification genes were only partially recovered, underscoring both the limitations of metagenome-assembled genomes and the underrepresentation of some genetic variants in databases. Additionally, a high-quality MAG affiliated with Candidatus (Ca.) Accumulibacter clade IIF exhibited genomic divergence from known species, suggesting the presence of a potential novel lineage of this relevant genus. These findings demonstrate that tropical activated sludge harbors both well-known and novel taxa with key metabolic capacities, along with diverse adaptive traits to environmental stressors. Collectively, this study expands current knowledge of microbial diversity and functional potential in tropical WWTPs and provides a framework for future research aimed at improving wastewater treatment in underexplored regions.