Genomic epidemiology and longitudinal sampling of ward wastewater environments and patients reveals complexity of the transmission dynamics of blaKPC-carbapenemase-producing Enterobacterales in a hospital setting

Background Healthcare-associated wastewater reservoirs and asymptomatic gastrointestinal patient colonisation by carbapenemase-producing Enterobacterales (CPE) can be important in nosocomial CPE dissemination and infection. We characterised these niches and within-niche diversity in a blaKPC-associated CPE (KPC-E) endemic healthcare setting, to better understand transmission potential. Methods We systematically sampled wastewater sites and patients across three units (six wards) over 6-12 months in 2016 in a KPC-E endemic hospital. We used Illumina sequencing to characterise up to five isolates per sample. Recombination-adjusted phylogenies were used to define genetically related strains; assembly and mapping-based typing approaches were used to characterise antimicrobial resistance gene and insertion sequence profiles, and Tn4401 types/target site sequences. The wider accessory genome was evaluated in a subset of the largest clusters, and those crossing niches. Findings Wastewater site KPC-E-positivity was substantial (101/349 sites [28.9%] positive, 319/4,488 [7.1%] sampling events positive); 183/4,425 (4.1%) of patients were CPE culture-positive over the same timeframe. 13 species and 109 KPC-E strains were observed across niches, and 24% of wastewater and 26% of patient KPC-E-positive samples harboured ≥1 strain. Most diversity was explained by the individual niche, suggesting highly localised factors are important in selection and spread. Tn4401+target site sequence (TSS) diversity was greater in wastewater sites (p<0.001), suggesting these might favour Tn4401-associated transposition/evolution and dissemination. Shower/bath and sluice/mop-associated sites were more likely to be KPC-E-positive (Adjusted Odds Ratio [95% CI]: 2.69 [1.44-5.01], p=0.0019 and 2.60 [1.04-6.52], p=0.0410, respectively). Different strains had different transmission and blaKPC dissemination dynamics. Interpretation There may be substantial KPC-E colonisation of wastewater sites and patients in KPC-E-endemic healthcare settings. Niche-specific factors, and different strains with different transmission dynamics influence carbapenemase gene dissemination. New transmission models incorporating complex, multi-level dynamics are needed to better quantify CPE dissemination to inform interventions and reduce transmission. Funding This study was supported by the National Institute for Health Research, UK.