Optimization of Pulse-Field Gel Electrophoresis for Subtyping of Klebsiella pneumoniae

Hui Han,Haijian Zhou,Baoliang Xu,Zhi Lu,Haishan Li,Kongxin Hu,Yuan Gao

doi:10.3390/ijerph10072720

Abstract

A total of 110 strains of Klebsiella pneumoniae were used to optimize pulsed-field gel electrophoresis (PFGE) for subtyping of K. pneumoniae. For optimization of electrophoresis parameters (EPs) of XbaI-PFGE, 11 isolates were analyzed with XbaI digestion using three EPs. The EP of a switch time of 6 to 36 s for 18.5 h gave clearest patterns and was declared the optimal EP for XbaI PFGE of K. pneumoniae. By software analysis and pilot study, AvrII was chosen as another PFGE enzyme. Both XbaI- and AvrII-PFGE gave D-values higher than 0.99 for 69 K. pneumoniae isolated from different sources. Our results also showed good typeability, reproducibility of both XbaI- and AvrII-PFGE for K. pneumoniae subtyping. Furthermore, the established PFGE method also had good discriminatory power to distinguish outbreak K. pneumoniae strains and a high degree of consistency with multilocus sequence typing method. A rapid PFGE protocol was established here, which could be used for genotyping and other researches of K. pneumoniae.

Highlights

Klebsiella pneumoniae is an opportunistic pathogen that usually causes hospital- and communityacquired bacterial infections in humans
An electrophoresis parameters (EPs) could be recommended by the CHEF Mapper equipment, based on the sizes of restriction fragments
The EP recommended for XbaI digestion of K. pneumoniae was a switch time of 6 to 20 s for 18 h (EP-a)

Summary

Introduction

Klebsiella pneumoniae is an opportunistic pathogen that usually causes hospital- and communityacquired bacterial infections in humans. Invasive K. pneumoniae has been reported to be an emerging infectious disease causing pyogenic liver abscesses and complications, such as meningitis or endophthalmitis [1,2,3]. The emergence and rapid spread of drug-resistant K. pneumoniae isolates is becoming a serious antibiotic management problem and causing great concern worldwide [4,5]. Standardized PFGE protocols had been established for pathogens such as Salmonella spp., Shigella spp., Escherichia coli O157:H7, Vibrio cholerae and Vibrio parahaemolyticus [11,12,13]. The use of standardized PFGE protocols allows for rapid comparison of DNA fingerprints between different laboratories to enhance disease surveillance

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