Metagenomic Profiling of Pathogens in Simulated Reclaimed Water Distribution Systems Operated at Elevated Temperature Reveals Distinct Response of Mycobacteria to Filtration and Disinfection Conditions

Abstract

Distribution of reclaimed water presents distinct challenges for pathogen control relative to the distribution of conventional potable water. The elevated temperatures common in arid regions can exacerbate such challenges by accelerating disinfectant decay and increasing microbial growth rates. Here, we carried out a controlled, continuous-flow, head-to-head study of six simulated reclaimed water distribution systems (RWDSs) in order to assess whether treatments identified to be effective for microbiological control at a lower temperature (22 °C) were also effective at 30 °C. Shotgun metagenomic sequencing was carried out to comprehensively profile fecal pathogens (FPs) (e.g., Escherichia and Klebsiella spp.), non-FPs (non-FPs) (e.g., Legionella and Mycobacterium spp.), and antibiotic-resistance genes (ARGs). Consistent with prior studies, generally, all FP, non-FP, and ARG markers were lowest when the influent water was biologically active carbon (BAC) filtered and disinfected. However, in contrast to observations at the lower temperature, where BAC filtration combined with chlorine disinfection effectively controlled Mycobacterium spp., they were markedly elevated in BAC-filtered and chlorine- or chloramine-disinfected conditions. This study highlights the need for further research to improve the understanding of conditions that select for different mycobacterial species and identify strategies to effectively control them in RWDSs under elevated temperature conditions.