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

Chen, Jiun-Han
Siu, L. K.
Fung, Chang-Phone
Lin, Jung-Chung
Yeh, Kuo-Ming
Chen, Te-Li
Tsai, Yu-Kuo
Chang, Feng-Yee

¹Department of Medical Laboratory Science and Biotechnology, Yuanpei University, Hsinchu, Taiwan, ROC
²Division of Infectious Diseases, National Health Research Institutes, Miaoli, Taiwan, ROC
³Section of Infectious Diseases, Department of Medicine, Taipei Veterans General Hospital, Taipei, Taiwan, ROC
⁴Division of Infectious Diseases and Tropical Medicine, Department of Internal Medicine, Tri-Service General Hospital, Taipei, Taiwan, ROC

Corresponding author. Tel: +886 2 8792 7257; Fax: +886 2 8792 7258; E-mail: [email protected]

† These authors contributed equally.

Received September 24, 2009

Accepted February 4, 2010

Journal of Antimicrobial Chemotherapy 65(5):p 986-990, May 2010. | DOI: 10.1093/jac/dkq056

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 (ΔOmpK36) 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 ΔOmpK36 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 (∼4 × 10⁴ cfu) of the parental strain and ΔOmpK36 led to significantly fewer viable bacteria in the liver 24 h post-inoculation in ΔOmpK36-inoculated mice. In the mice LD₅₀ (the bacterial dose that caused 50% death) assay, the parental strain was ∼100-fold more lethal (∼10³ cfu) than the ΔOmpK36 mutant (∼10⁵ 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.