Analysis of Antibiotic Resistance and Virulence Traits (Genetic and Phenotypic) in Klebsiella pneumoniae Clinical Isolates from Pakistan: Identification of Significant Levels of Carbapenem and Colistin Resistance

Background: The emergence of carbapenem-resistant and hypervirulent hypermucoviscous Klebsiella pneumoniae strains poses a significant public health challenge. We determined the MDR profiles, antibiotic resistance factors, virulence gene complement, and hypermucoviscous features of 200 clinical K. pneumoniae isolates from two major tertiary care hospitals in Islamabad and Rawalpindi, Pakistan. Methods: Susceptibility profiling and phenotypic analysis were performed according to the CLSI guidelines. Genetic determinants of antibiotic resistance and virulence were detected by PCR. Biofilm formation analysis was performed by microtiter plate assay. Results: The isolates displayed a high degree of antibiotic resistance: 36% MDR-CRKP; 38% carbapenem resistance; 55% gentamicin resistance; 53% ciprofloxacin resistance; and 59% aztreonam resistance. In particular, the level of resistance against fosfomycin (22%) and colistin (15%) is consistent with previous reports of increased resistance levels. Combined resistance to carbapenem and colistin was 7%. Genetic factors associated with colistin resistance (mcr-1 and mcr-2 genes) were detected in 12 and 9% of the isolates, respectively. Significant differences in resistance to gentamicin and levofloxacin were observed between the 200 isolates. Many of the isolates harbored genes specifying extended-spectrum and/or carbapenem-resistant beta-lactamases: bla(CTX-M-15) (46%), bla(NDM-1) (39%), and bla(OXA-48) (24%). The prevalence of the hypermucoviscous phenotype was 22% and 13% of the MDR isolates carried the rmpA gene (regulator for mucoid phenotype). Key virulence factor genes detected include those encoding: porins (ompK35 and ompK36; at 56 and 55% prevalence, respectively); adhesins (fimH, mrkD, and ycfM; at 19, 18, and 22% prevalence, respectively); and the polysaccharide regulator, bss, at 16% prevalence. Conclusion: This report highlights carbapenem-resistant K. pneumoniae (CRKP) prevalence, emerging resistance to fosfomycin, and the presence of mcr-1 and mcr-2 in colistin-resistant isolates. Further, the detection of rmpA signifies the prevalence of the hypermucoviscous trait in CRKP clinical isolates from Pakistan.