engleski

Deep Profiling of the Klebsiella pneumoniae OXA-48 Proteome

Klebsiella pneumoniae is a nosocomial pathogen that exhibits multidrug resistance and virulence. Emergence of carbapenem-resistant K. pneumoniae is increasingly reported worldwide, especially strains producing OXA-48 carbapenemases, Ambler class D beta-lactamases encoded by blaOXA-48 gene. These strains cause difficulties to detect in clinical specimens due to many of these isolates falling within the susceptible range for carbapenems. The aim of this study was to identify specific characteristics of K. pneumoniae isolates containing OXA-48 gene. Four clinical isolates were chosen for the analysis, two containing OXA- 48 gene and two strains lacking OXA-48 gene (controls). OXA-48 genes presence was confirmed by a multiplex polymerase chain reaction (PCR). The antimicrobial susceptibility was examined by disc diffusion method and by determination of minimal inhibitory concentrations (MIC) with broth microdilution method according to European Committee on Antimicrobial Susceptibility Testing (EUCAST) standards. Two isolates with present OXA- 48 gene were susceptible to carbapenems (imipenem and meropenem). To distinguish the difference between the analysed strains, a comprehensive proteomics analysis was performed via liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) operating in data‐independent acquisition mode (DIA). DIA technology enabled accurate identification of more than 2000 proteins in all of the strains. Beta- lactamase OXA-48 was detected in OXA-48 positive strains despite low- level resistance. Quantitative analysis revealed an altered expression of membrane proteins in OXA- 48 strains in comparison to controls. Particularly, upregulation of putative porin KPN_04773 and outer membrane lipoprotein SlyB were detected in OXA-48 strains. Furthermore, outer membrane protein OmpF was downregulated in OXA-48 strains. As well, downregulation of peptidase YggG and two proteins with dehydrogenase activity (KPN_01492 and PutA) was also detected in OXA-48 strains. Overall, our results give an insight into the difference in the protein expression between OXA-48 strains and controls and indicate that OXA- 48 gene presence is not the only difference between strains.

Klebsiella pneumoniae ; OXA-48 ; Proteome

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