A 10-year investigation of the impact of a wastewater treatment plant upgrade on nitrogen-cycling microorganisms and DNA viruses in receiving river

Wastewater treatment plants (WWTPs) are upgraded to improve the quality of river water downstream, but the effects of such upgrades on aquatic microbial and viral communities remain poorly understood. We conducted a 10-year investigation of Tonghui River and Qing River in Beijing that receive effluent from representative WWTPs, during which the WWTP for Tonghui River underwent an upgrade while the other one for Qing River, used as a control, had been upgraded prior to the study. Over this decade, we monitored key water quality parameters and simultaneously analyzed microbial community dynamics via metagenomic sequencing. Metagenomic analysis showed that for nitrogen-cycling bacteria, overall α-diversity (Shannon index) remained steady after the WWTP upgrade, but community composition changed significantly. β-Diversity analysis also revealed a clear shift in microbial community structure in the impacted river after the upgrade compared with the status before it. The partitioning of β-diversity revealed that species nestedness dominated community variation in Tonghui River. Its contribution increased substantially from 68% before the upgrade to 86% after it. These results indicated that community differences were mainly driven by the loss or gain of some taxa while a stable core microbiome was retained, rather than by complete species turnover. Functionally, the median nitrifier-to-denitrifier abundance ratio decreased by 70% following the upgrade, indicating a shift in nitrogen cycling toward enhanced denitrification. Viral community structure remained largely stable over time, with differences mainly attributable to temporary species replacement rather than permanent loss or gain of taxa. Additionally, after the upgrade, the abundance of viral replication and structural genes in the river water increased by approximately 15%~30%, the abundance of host metabolic auxiliary genes decreased by about 20%~40%, and the overall proportion of lysogenic viruses remained unchanged. These changes suggest that better effluent quality may have prompted viruses to shift from assisting host metabolism to active self-replication. Overall, this long-term study provides field evidence that WWTP upgrades alter nitrogen-cycling processes and viral ecology, highlighting the necessity of incorporating viral and microbial markers into post-upgrade water quality assessments and river management. • Impact of a WWTP upgrade on the receiving river’s microbiome was examined. • A 10-year investigation into two urban rivers in Beijing was performed. • The upgrade induced a shift in nitrogen cycling toward enhanced denitrification. • Viral communities exhibited no significant changes following the upgrade.