Detection of Carbapenem-Resistant Bacteria in Skilled Nursing Facility Wastewater

Wastewater surveillance at healthcare facilities for carbapenem-resistant (CR) bacteria harboring carbapenemase genes could result in earlier detection of circulating or emerging antimicrobial resistance (AR) threats. However, knowledge gaps exist regarding the CR bacterial community and carbapenemase genes present in facility wastewater and plumbing, which need to be addressed. Wastewater effluent samples from three skilled nursing facilities (SNFs) (Georgia, USA) were collected weekly over six months each. Selective chromogenic media were examined for their performance in screening the samples for targeted CR bacteria. Culture-dependent and culture-independent methods (i.e., PCR) were applied to enriched wastewater samples (n=73) to screen isolates and pooled samples for the most common carbapenemase genes circulating in the United States ( bla _KPC , bla _NDM , bla _VIM , bla _OXA-48-like , and bla _IMP ), and method sensitivity was compared. Whole genome sequencing (WGS) was used to identify all AR genes present in isolates. Bacteria (n=209 isolates) were identified using MALDI-TOF and included >20 different genera. Clinically relevant ( Acinetobacter baumannii complex, Enterobacter cloacae complex, Enterococcus faecium, Stenotrophomonas spp.) and environmental/opportunistic ( Comamonas spp., Pandoraea spp ., Pseudomonas putida group) CR bacteria were identified in each of the three SNFs. In Facility A and C, only bla _KPC was detected in 9 (26.5%) of 34 and 14 (11.4%) of 123 recovered isolates, respectively. There were no PCR-positive results in Facility B. WGS confirmed the presence or absence of all genes screened for by PCR (100% concordance). These data provide a proof of concept and insights into CR bacteria present in healthcare facility wastewater using culture, PCR, and sequencing methods.