Development of a multiplex PCR system and its application in detection of bla(SHV), bla(TEM), bla(CTX-M-1), bla(CTX-M-9) and bla(OXA-1) group genes in clinical Klebsiella pneumoniae and Escherichia coli strains

作者：Ogutu, James O.; Zhang, Qingmeng; Huang, Ying; Yan, Huo; Su, Lijie; Gao, Bo; Zhang, Wenli; Zhao, Jizi; Cai, Wenhui; Li, Wenjing; Zhao, Hong; Chen, Yang; Song, Wuqi; Chen, Xiaobei; Fu, Yingmei^*; Zhang, Fengmin

来源：Journal of Antibiotics, 2015, 68(12): 725-733.

DOI：10.1038/ja.2015.68

• SPECTRUM-BETA-LACTAMASE
• QUINOLONE RESISTANCE DETERMINANTS
• CTX-M
• PREVALENCE
• ENTEROBACTERIACEAE
• SURVEILLANCE
• NATIONWIDE
• DOMINANCE
• VARIANTS
• SHV

Resistance to beta-lactam antibiotics through beta-lactamase production by Enterobacteriaceae continues to burden the health-care sector worldwide. Traditional methods for detection of beta-lactamases are time-consuming and labor-intensive and newer methods with varying capabilities continue to be developed. The objective of this study was to develop a multiplex PCR (M-PCR) system for the detection of bla(SHV), bla(TEM), bla(CTX-M-1), bla(CTX-M-9) and bla(OXA-1) group genes and to apply it in clinical Klebsiella pneumoniae and Escherichia coli strains. To do this, we used group-specific PCR primers in singleplex reactions followed by optimization into multiplex reactions. Specificity and sensitivity of the M-PCR were then evaluated using 58 reference strains before its application to detect bla group genes in 203 clinical Enterobacteriaceae strains. PCR amplicons were sequenced to determine the beta-lactamase subtypes. The M-PCR system exhibited 100% specificity and sensitivity. In all, 83.7% of K. pneumoniae and 89.8% of E. coli clinical strains harbored bla group genes with 46.9%, 40.1%, 15.0%, 21.1% and 6.1% of K. pneumoniae having bla(SHV), bla(TEM), bla(CTX-M-1), bla(CTX-M-9) and bla(OXA-1) group genes, respectively, whereas 12.2%, 77.6%, 22.4%, 36.7% and 8.2% of E. coli had bla(SHV), bla(TEM), bla(CTX-M-1), bla(CTX-M-9) and bla(OXA-1) group genes, respectively. Bla(SHV-1), bla(SHV-11), bla(SHV-27), bla(SHV-33), bla(SHV-144), bla(TEM-1), bla(TEM-135), bla(OXA-1), bla(CTX-M-3), bla(CTX-M-9), bla(CTX-M-14), bla(CTX-M-15), bla(CTX-M-27), bla(CTX-M-55), bla(CTX-M-65) and bla(CTX-M-104) were detected. In conclusion, the M-PCR system was efficient and versatile with an advantage of simultaneously detecting all the targeted bla group genes. Hence, it is a potential candidate for developing M-PCR kits for the screening of these genes for clinical or epidemiological purposes.

• 出版日期2015-12
• 单位哈尔滨医科大学; 大连医科大学