Surveillance of Antibiotic Resistance in Clinical and Hospital Water Isolates of K. pneumoniae

Abstract

Klebsiella pneumoniae (K. pneumoniae, a prominent nosocomial pathogen, has emerged as a leading cause of neonatal septicemia worldwide, determining substantial morbidity and mortality. K. pneumoniae belongs to the ESKAPE pathogen group, comprising six critically significant microorganisms that have developed resistance to antibiotics. This opportunistic bacterium has developed extensive antibiotic resistance via the acquisition of genes that encode enzymes like ESBLs and carbapenemases. In the last decade, K. pneumoniae carbapenemase (KPC) has played a role in the increase in carbapenem-resistant Enterobacteriaceae (CRE) cases. To elucidate the molecular basis of resistance, virulence factors, and genetic context of key resistance genes in multidrug-resistant K. pneumoniae, we conducted short-read whole-genome sequencing of eleven isolates from adult patients, neonates, and water samples. The draft genomes displayed size variation (5.48 to 6.68 Mbp), indicating genomic plasticity within this species. All sequenced isolates harbored genes conferring resistance to various antibiotic classes, including aminoglycosides, quinolones, sulfonamides, tetracycline, and trimethoprim. The gene qacEdelta1, conferring resistance to biocides, was detected in most of the strains. Diverse sequence types and capsular types were identified among the isolates. Notably, key antibiotic resistance genes were flanked by various mobile genetic elements (MGEs), highlighting the role of transposons, integrons, and insertion sequences in shaping resistance gene transmission. Crucial components involved in the transposition of resistance include IS26, Tn3, IS903B, ISEcp1, and ISKpn19. We also observed spontaneous mutations in genes that indirectly contribute to antibiotic resistance such as efflux pumps, outer membrane proteins etc. Furthermore, the loss or deficiency of outer membrane porins OmpK35 and OmpK36, coupled with ESBL production, potentially played a significant role in carbapenem resistance in our sequenced isolates. Phylogenetic analysis suggested an evolutionary relationship between our isolates and strains from China, India, and the USA, implying a shared evolutionary history and potential dissemination of similar genes. This study provides critical insights into the mechanisms of carbapenem resistance, including the gain of multiple resistance genes through MGEs and mutations in other carbapenem resistance-associated genes, in K. pneumoniae. Our analysis underscores the utility of whole-genome sequencing for monitoring antibiotic resistance patterns and prescribing treatment decisions based on the spread of acquired II resistance genes. In addition, co-infection of K. pneumoniae and Mycobacterium tuberculosis poses a significant health threat because of delayed diagnosis and insufficient treatment. Because of the rise of multidrug-resistant strains, the development of prophylactic and immunotherapeutic vaccines is imperative. In this study, a reverse vaccinology approach is utilized to assess immunogenic epitopes associated with K. pneumoniae OmpA, and M. tuberculosis Rv1698 and Rv1973, to develop a chimeric vaccine. The multi-epitopic vaccine that was designed underwent an evaluation to assess its antigenicity, allergenicity, and physicochemical characteristics. Molecular docking and simulations demonstrated the immunogenicity and stability of the complex. The designed multi-epitopic vaccine demonstrated strong immunogenicity and appears to be a promising proactive solution against these pathogens.