Clonal diversity and genetic profiling of antibiotic resistance among multidrug/carbapenem-resistant Klebsiella pneumoniae isolates from a tertiary care hospital in Saudi Arabia

Abstract

Background: The nexus between resistance determinants, plasmid type, and clonality appears to play a crucial role in the dissemination and survival of carbapenem-resistant Klebsiella pneumoniae (CRKP). The incidence of infections involving CRKP in Saudi Arabia is increasing and there is a need for detailed molecular profiling of this pathogen for CRKP surveillance and control. Methods: The resistance determinants of 71 non-redundant CRKP isolates were investigated by polymerase chain reaction (PCR) and sequencing. Plasmid typing was performed using PCR-based replicon typing and the clonality of isolates was determined by multilocus sequence typing. Capsular polysaccharide synthesis genes and other virulence factors were examined using multiplex PCR. Diversity was calculated using DIVEIN, clonal relationship was determined using eBURST, and phylogenetic analysis was performed using SplitsTree4. Results: A polyclonal OXA-48 gene alone was the most common carbapenemase detected in 48/71 (67.6%) isolates followed by NDM-1 alone in 9/71 (12.7%) isolates. Coproduction of OXA-48 and NDM-1 was observed in 6/71 (8.5%) isolates. Both carbapenemase genes could be transferred into an Escherichia coli recipient. CTX-M-15 was the most abundant extended-spectrum β-lactamase gene detected in 47/71 (66.2%) isolates, whereas clone-specific CTX-M-14 (ST-199 and -709) was found in 15/71 (21%) isolates. Sixty-seven of 71 isolates were positive for one or more plasmid replicons. The replicons detected were: IncFII; IncFIIK; IncFIA; IncFIB; L/M; IncI1; and IncN. FIIK and L/M were predominant, with 69 and 67% positivity, respectively. All isolates were negative for the magA (K1), rmpA, and K2 genes and presented a non-hypermucoviscous phenotype. Conclusion: A polyclonal CRKP reservoir of sequence types (STs)-37, - 199, and - 152 was observed and ST-152 appeared to be a "frequent carrier" of the NDM-1 gene. ST-199, a singleton not previously reported, showed a sequence diversity suggestive of positive selection. A significant association was evident between resistance determinants and the clonal types of K. pneumoniae: all ST-152 isolates were positive for NDM-1 but negative for OXA-48; ST-199 isolates were positive for OXA-48 but negative for NDM-1; and ST-709 and -199 isolates were positive for CTX-M-14. The incidence of certain clonal types in large numbers predicts an outbreak-like situation and warrants stringent surveillance and infection control.