Molecular typing，drug resistance and virulence analysis of Klebsiella pneumoniae isolated from Type 2 diabetes mellitus complicated with maxillofacial infection

Abstract

Objective To investigate molecular typing and drug resistance patterns of 98 Klebsiella pneumoniae (K.pneumoniae) isolated from type 2 diabetes patients complicated with maxillofacial infection,to research the virulence and resistance mechanisms. Methods The study was a prospective study that adopted the method of continuous sampling from fixed location,from March 2010 to October 2012.The maxillofacial surgery patients diagnosed with type 2 diabetes complicated with maxillofacial infection were chosen in 7 hospitals in Zhengzhou as the research object,and a total of 431 pus sample were collected continuously,in which 98 strains K.pneumoniae were isolated and identified.The Kirby-Bauer disk diffusion test was conducted in 98 strains to determine the resistance to 19 antibacterial agents.K.pneumoniae chromosomal DNA were digested by restriction endonuclease XbaⅠ and analyzed by pulsed-field gel electrophoresis (PFGE).PFGE patterns of K.pneumoniae strains were analyzed using Fingerprinting software.The relationship between the molecular types and resistance phenotype was observed.The extended spectrum β-lactamase-producing K.pneumoniae were screened out by the double disc synergy test (DDST).Polymerase chain reaction (PCR) was used to detect resistant genotypes,serotype and virulence genes.The purified PCR products of resistant genes were cloned and sequenced.Hypermucoviscosity phenotype of all strains were determined by string test. Results Much severer drug-resistance for K.pneumoniae was identified and the result of extended-spectrum beta-lactamase producing rate was 57.1%.Ninety-eight strains were dispatched into 13 groups by PFGE.No dominant bands and specific extended-spectrum beta-lactamase DNA bands were found.The results of PCR showed that among the 56 strains of extended spectrum β-lactamase-producing K.pneumoniae,40 were positive for blaSHV (accounting for 71.4%),28 positive for blaTEM (accounting for 50.5%),21 positive for blaCTX-M (accounting for 37.5%).The sequencing results were as follows: TEM-1,CTX-M-3 and a variety of SHV.Serotype K1,K2,K3,K5,K20,K54 and K57 and 3 kinds of virulence genes were detected,but not in strong toxicity-based.Hypermucoviscosity positive rate was 31.6%(31/98). Conclusion Much severer drug resistance of K.pneumoniae in this study was identified and resistant mechanism was complex,in which strong toxicity serotype and virulence genes exist,which need more attention from clinical.（Chin J Lab Med,2014,37: 136-139） Key words: Diabetes mellitus,type 2; Klebsiella pneumoniae; Drug resistance,bacterial; Serotyping; Virulence