Metagenomic Characterization of Biofilm and Suspended Microbial Communities in a Hybrid Algal Turf Scrubber‐Based Wastewater Treatment System

This study investigates a hybrid wastewater treatment system combining a biofilm-based Algal Turf Scrubber (ATS) with a membrane-coupled High Rate Algal Pond (ATS-MHRAP) for shrimp aquaculture effluents. Shotgun metagenomic sequencing was used to compare microbial composition, functional pathways, and antibiotic resistance genes (ARGs) across attached biofilm (ATS1) and suspended biomass (ATS2, HRAP1) under three nutrient loading stages. Biofilm samples (ATS1) exhibited higher microbial richness and evenness, with Shannon index values up to 9.25, compared to 6.93 in suspended cultures. Functional pathways enriched in ATS1 included nitrogen cycling, amino acid metabolism, and terpenoid biosynthesis, with elevated expression of amoA, nirK, and nirS genes under moderate loading. These traits coincided with higher removal efficiency of COD (up to 88.6%), phosphate (82.1%), and total nitrogen (73.4%). ARGs were more diverse in ATS1, with up to 11 resistance classes detected, including β-lactam and sulfonamide genes co-occurring with intI1, indicating possible horizontal gene transfer. The ATS-MHRAP system offers a robust and biologically enriched platform for nature-based aquaculture wastewater treatment. Our findings reveal microbial and functional differentiation between attached and suspended communities, with implications for optimizing dissolved oxygen, nutrient ratios, and retention time.