Molecular characterization of multi-drug resistant (MDR) gram-negative bacterial pathogens from environments, patients and staff in a teaching hospital in Ghana.

Abstract

Multidrug resistant Gram-negative bacteria (MDR GNB) are implicated in serious infections both of community and nosocomial origin and may be disseminated in the hospital in the absence of efficient infection prevention and control (IPC) practices. The prevalence and risk factors for rectal colonization of MDR GNB among patients, the carriage of MDR GNB on healthcare workers’ (HCWs’) hands and the contamination patients’ environments with MDR GNB were investigated in a teaching hospital in Ghana. In this prospective study, conducted between April 2021 to July 2021, the phenotypic profiles of the MDR GNB isolates were determined using the VITEK 2 system. Risk factors for colonization with MDR GNB were assessed using univariate and multivariate analysis of associated data. The resistome, virulome, mobilome and genetic relatedness of MDR extended-spectrum β-lactamase (ESBL)-producing Escherichia coli and ESBL-producing or carbapenem resistant Klebsiella pneumoniae isolates from patients and their environment were also determined using whole genome sequencing performed on the Nextseq 550 (2 x 150 bp) and bioinformatics analysis. A total of 585 samples were collected from patients, HCWs’ hands and the hospital environment within the study period. The prevalence of MDR GNB rectal colonization among patients was 50.62% on admission and 44.44% after 48 hours. MDR GNB, frequently E. coli and K. pneumoniae were isolated from 6 (5.26%) and 24 (11.54%) of HCW’s hand swabs and environmental swabs, respectively. Previous hospitalization (p-value = 0.021, OR,95% CI= 7.170 (1.345-38.214) was significantly associated with colonization by MDR GNB after 48 hours of admission while age (21-30 years) (p-value =0.022, OR, 95% CI =0.103(0.015-0.716) was significantly identified as a protective factor associated with a reduced risk of rectal MDR GNB colonization. Rectal carriage and acquisition of ESBL-producing E. coli among patients was 13.65% and 11.32% respectively. blaTEM-1B and blaCTX-M-15 were commonly associated with IncFIB plasmid replicons and co-occurred with aminoglycoside, macrolide, and sulfamethoxazole/trimethoprim resistance. Multiple virulence genes, predominantly, terC were detected in the ESBL E. coli isolates. Sequence types (STs) were diverse and included one novel ST (ST13846) present in two isolates. Phylogenetic analysis grouped the ESBL E. coli isolates into four main clusters. High genetic relatedness was observed between two carriage isolates of ST940 and between a carriage isolate and an environmental isolate of ST648. Isolates with different STs, collected at different times and locations, also showed genetic similarities. Of the ten selected MDR K. pneumoniae isolates, the β-lactamase gene, blaCTX-M-15 was observed in six isolates. Mutations were found in both ompK36 and ompK37 in all isolates (both carriage isolates and isolates from hospital environments). Genes encoding resistance to fluoroquinolone (qnrB), aminoglycosides (aadA1, aadA2, aac(3)-IIa, aac(6')-Ib-cr,aph(3'')-Ib , aph(6)-Id) sulphamethoxazole/trimethoprim (sul1, sul2, dfrA14, dfrA15) were also detected. The K. pneumoniae isolates belonged to seventeen different STs with ST39 most commonly observed and common to both carriage isolates and isolates from hospital environments. A myriad of virulence genes, including irp1, irp2, iutA, gndA, ompA, fes, fep, mrkD and fimH were detected in both carriage and isolates from the hospital environment. IncFIB was the most abundant plasmid replicon occurring in nine (four carriage isolates and five isolates from hospital environments).

ESBL-producing K. pneumoniae isolates appeared to be introduced into the hospital from the community. The high colonization of MDR GNB in patients, the carriage of MDR GNB on HCW’s hands, the contamination of hospital environments and the circulation of ESBL-producing E. coli and K. pneumoniae isolates with diverse genomic characteristics, highlights the need for patient screening, and stringent infection prevention and control practices to prevent the spread of MDR GNB in hospitals. The observed clonal relatedness among isolates from patients and the hospital environment, as well as between different patients, suggests a possible transmission within and between sources, hence infection prevention and control practices need to be enhanced to prevent the dissemination and transmission of these resistant strains in the hospital. This study further highlights the usefulness of whole genome sequencing as an effective tool in AMR surveillance.