Establishment and application of fluorescence PCR method for genotyping of Mycoplasma pneumoniae based on specimen nucleic acid
10.3760/cma.j.cn112309-20240627-00229
- VernacularTitle:基于标本核酸的肺炎支原体基因分型荧光PCR方法的建立和应用
- Author:
Yiwei ZHANG
1
;
Liyong LIU
;
Lihua HE
;
Fanliang MENG
;
Ruixue GU
;
Jie GONG
;
Shaoli LI
;
Fei ZHAO
Author Information
1. 中国疾病预防控制中心传染病预防控制所，传染病溯源预警与智能决策全国重点实验室，北京 102206
- Keywords:
Mycoplasma pneumoniae;
MGB (minor groove binder) probe;
Genotyping;
Real-time PCR
- From:
Chinese Journal of Microbiology and Immunology
2024;44(8):653-659
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish a nucleic acid detection and genotyping method for Mycoplasma pneumoniae ( Mp) based on nucleic acid in clinical samples. Methods:Through genomic comparison, the specific target sequences of Genotype 1 and Genotype 2 Mp strains were selected to design synthetic primers and probes, and a PCR detection and classification method for Mp dual fluorescent probe was established, and the specificity, accuracy, detection limit and repeatability of the method were evaluated. The established fluorescence PCR method was used to detect the nucleic acid of clinical specimens and compared with the reported fluorescent PCR methods. Results:The nucleic acid of 18 pathogens, including other species of Mycoplasma and common respiratory bacteria and viruses, which were closely related to the Mp species, were detected, and the results showed that there was no cross-reactivity. The accuracy of detection and typing of 90 Mp nucleic acid was 100%. The detection limits of Genotype 1 and Genotype 2 Mp samples were 1.0 copy/μl, and the experimental coefficient of variation of repeatability within groups and between groups was less than 2.5%. In the detection of 88 nucleic acid of clinical specimens, the Kappa value was 0.675 and the P value was 0.267 compared with the reported real-time PCR method, showing a high degree of agreement. Conclusions:The method for detecting and genotyping Mp in this study has high sensitivity, specificity, and accuracy, which can be applied to the monitoring and prevention and control of Mp in the disease control system of provinces and cities at all levels in China. This method promotes the improvement of the Mp prevention and control system in China, strengthens the surveillance ability, and is of great significance for the early warning and prediction of Mp.
