Establishment of a multiplex PCR for rapid identification of Mycobacterium species
10.3760/cma.j.issn.0254-5101.2019.10.008
- VernacularTitle: 多重PCR快速鉴定分枝杆菌方法的建立
- Author:
Shupeng YIN
1
;
Chenqi YAN
2
;
Zhiguang LIU
2
;
Xiuqin ZHAO
2
;
Xiaoqin LI
1
;
Machao LI
2
;
Haican LIU
2
;
Yongliang LOU
1
;
Kanglin WAN
1
Author Information
1. School of Laboratory Medicine and Life Sciences, Wenzhou Medical University, Wenzhou 325035, China
2. State Key Laboratory of Infectious Diseases Prevention and Control, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
- Publication Type:Journal Article
- Keywords:
Multiplex PCR;
Mycobacterium;
Mycobacterium tuberculosis;
Beijing genotype
- From:
Chinese Journal of Microbiology and Immunology
2019;39(10):771-777
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish and evaluate a multiplex PCR method for rapid identification of Mycobacterium species in order to provide an approach for rapid diagnosis of pathogens causing tuberculosis.
Methods:Four genes including 16S rRNA, Rv0577, RD9 and mtbk_20680 were selected to establish the multiplex PCR method. Specific primers were designed and the reaction system and conditions were optimized. The sensitivity and specificity of the multiplex PCR method were evaluated through identifying Mycobacterium africanum (M.africanum), Mycobacterium bovis (M.bovis), M. bovis BCG, seven common non-tuberculosis Mycobacterium (NTM), seven reference species of common respiratory bacteria and 93 clinical strains of Mycobacterium tuberculosis (MTB) isolated from patients with tuberculosis in Gansu Province of China.
Results:The fragments of 16S rRNA, Rv0577, RD9 and mtbk_20680 genes were 543 bp, 786 bp, 369 bp and 231 bp in length, respectively. MTB strains of the Beijing genotype were positive for all of the four genes, while the non-Beijing genotype strains were negative for mtbk_20680. M. africanum, M. bovis and M. bovis BCG strains were negative for 16S rRNA and Rv0577. NTM strains only carried 16S rRNA and none of four genes were detected in the seven species of respiratory bacteria. Among the 93 clinical MTB strains, 80 (86.02%) belonged to the Beijing genotype and the other 13 (13.98%) were non-Beijing genotype strains. The specificity of the multiplex PCR method was 100%.
Conclusions:The established multiplex PCR method could accurately distinguish Mycobacterium, Mycobacterium tuberculosis complex (MTBC), NTM, MTB, and Beijing and non-Beijing genotype MTB with high sensitivity and specificity.