In situ dosing of monochloramine in a hospital hot water system results in drastic microbial communities changes.

Understanding changes in microbial composition under selective pressures is crucial to assess the emergence of resistant taxa and the survival of drinking water-associated pathogens. This study evaluated the impact of in situ monochloramine disinfection in a hospital hot water system on bacterial (16S rRNA gene amplicon sequencing, 112 samples) and eukaryotic communities (18S rRNA gene amplicon sequencing, 103 samples), and on general microbial measurements (180 samples), including adenosine triphosphate (ATP) and flow cytometry counts. After the onset of treatment, ATP decreased by 1.2- and 3.5-fold, and total cell counts (TCC) dropped by 1- and 2-log at distal and system sites, respectively. During the dosage interruption (27-day), TCC rebounded to pre-treatment levels, but viability percentage decreased, indicating that cells were predominantly damaged. Low-use sites (e.g., showerheads) showed elevated ATP (>15 pg/mL) and TCC (10⁵-10⁶ cells/L). Monochloramine drastically altered bacterial and eukaryotic communities. Alpha-diversity showed increased amplicon sequence variant richness during treatment, driven by new, low-abundant taxa, while Beta-diversity revealed distinct shifts in community composition over time, with tight or looser clusters corresponding to each treatment phase. Post-treatment, temporal and spatial heterogeneity was evident across distal sites, while elevated temperatures, consistent flow, and higher monochloramine concentrations in the hot water system resulted in more uniform communities at system sites. Additionally, the persistence of potential pathogenic strains belonging to Legionella and Mycobacterium genera highlights the value of comprehensive risk assessments. These findings emphasize the need to understand microbial shifts under disinfection stress and their public health implications, offering new insights into how treatment interventions shape microbial ecology and pathogen dynamics.