Contribution of outer membrane protein K36 to antimicrobial resistance and virulence in Klebsiella pneumoniae.

OBJECTIVES Loss of outer membrane protein (Omp) is commonly encountered in multidrug-resistant Klebsiella pneumoniae. However, little is known about the association between Omp loss and virulence. In the present study, this association was investigated in K. pneumoniae. METHODS An OmpK36-deficient mutant (DeltaOmpK36) was derived from a virulent clinical isolate by targeted gene insertion. Antimicrobial susceptibility was tested by microbroth dilution and disc diffusion. Virulence was assessed by serum resistance, phagocytosis, clearance of viable bacteria in the liver and lethality in mice following inoculation with bacteria. RESULTS Susceptibility tests showed that DeltaOmpK36 contributed to the resistance to cefazolin and cefoxitin but not to resistance to late-generation cephalosporins. In vitro assays demonstrated that loss of OmpK36 decreased the resistance to neutrophil phagocytosis and increased the resistance to serum killing during the first hour of the assay, but did not influence the growth rate when compared with the parental strain. Intraperitoneal injection of similar doses ( approximately 4 x 10(4) cfu) of the parental strain and DeltaOmpK36 led to significantly fewer viable bacteria in the liver 24 h post-inoculation in DeltaOmpK36-inoculated mice. In the mice LD(50) (the bacterial dose that caused 50% death) assay, the parental strain was approximately 100-fold more lethal ( approximately 10(3) cfu) than the DeltaOmpK36 mutant ( approximately 10(5) cfu). CONCLUSIONS Loss of OmpK36 in K. pneumoniae resulted in increased antimicrobial resistance, increased susceptibility to neutrophil phagocytosis, increased resistance to serum killing and reduced virulence.