Effects of Bellamya Stocking Density on Water Quality and Bacterial Community Responses in Aquaculture Effluent

Aquaculture effluent commonly contains suspended solids, inorganic nitrogen, reactive phosphate and algal biomass, creating a need for low-input ecological treatment approaches. This study assessed endpoint water-quality and bacterial-community responses to Bellamya stocking density in aquaculture effluent. A no-snail control (CON) and four stocking-density treatments (LD, MD, MHD and HD) were established with three independent tank replicates per treatment. Suspended solids (SS), NH₄⁺-N, NO₂^--N, NO₃^--N, PO₄^3--P and chlorophyll a (Chl-a) were quantified, and bacterial communities were characterized by 16S rRNA gene sequencing of the V3-V4 region. At the 60-day endpoint, all Bellamya-stocked treatments had lower NH₄⁺-N and NO₃^--N concentrations than the control, and MHD and HD also had lower PO₄^3--P and Chl-a. SS showed a nonsignificant downward tendency, whereas NO₂^--N showed a nonsignificant upward tendency. Bray-Curtis NMDS visualized treatment-associated separation, and PERMANOVA detected significant differences among treatments (R² = 0.62, p = 0.001; stress = 0.0822). However, PERMDISP was also significant (F_4,10 = 4.99, p = 0.001), indicating heterogeneous within-treatment dispersion and requiring cautious interpretation of the PERMANOVA result. Representative genera showed distinct treatment-associated abundance patterns, and 46 of 90 genus-environment associations remained significant after Benjamini-Hochberg correction. Predicted KEGG Level 3 pathways varied numerically among treatments, but none remained significant after false-discovery-rate correction. These findings indicate that Bellamya stocking may provide a low-input ecological component of aquaculture-effluent management, while the microbiome results should be interpreted as community-level associations and predicted functional trends rather than evidence of microbial causality or pathway activation.