Large-scale characterisation of hospital wastewater system microbiomes and clinical isolates from infected patients: profiling of multidrug-resistant microbial species.

Hospital-acquired infection (HAI) and infectious agents exhibiting antimicrobial resistance (AMR) are challenges globally. Environmental patient-facing wastewater apparatus including handwashing sinks, showers and toilets are increasingly identified as sources of infectious agents and AMR genes. We performed large-scale metagenomic analysis of the wastewater system in a large teaching hospital in the Republic of Ireland experiencing multidrug-resistant HAI outbreaks. Immediately prior to refurbishment of a medical ward where HAI has been endemic, wastewater pipe sections (n=20) were removed. These comprised toilet u-bends, sink and shower drains and, following DNA extraction, each underwent metagenomic analysis. Diverse taxonomic and resistome profiles were observed, with members of phyla Proteobacteria and Actinobacteria dominating (38.23 ± 5.68% and 15.78 ± 3.53%, respectively). Genomes of five clinical isolates were analysed. These antimicrobial-resistant bacterial isolates were from patients >48 hours post-admission to the ward. Genomic analysis determined that the isolates bore a high number of antimicrobial resistance genes (ARGs). Comparison of resistome profiles of these isolates and wastewater pipe metagenomes revealed a high degree of similarity, with many identical ARGs shared between clinical isolates and the wastewater environment, suggesting probable acquisition post-admission. The highest numbers of ARGs observed were those encoding resistance to significant clinically- and commonly-used antibiotic classes. Average nucleotide identity analysis confirmed presence of highly similar or identical genomes in the clinical isolates and wastewater pipes. These unique large-scale analyses reinforce the need for regular cleaning and decontamination of patient-facing hospital wastewater pipes and effective infection control policies to prevent the transmission of nosocomial infection and emergence of AMR within potential wastewater reservoirs.