Development of a triplex real-time RT-PCR assay to detect Zika, Chikungunya and Mayaro viruses
10.3760/cma.j.issn.1003-9279.2019.06.015
- VernacularTitle: 寨卡病毒、基孔肯雅病毒和马雅罗病毒三重实时荧光定量RT-PCR检测方法的建立
- Author:
Lijin LAI
1
;
Aqian LI
2
;
Quanfu ZHANG
2
;
Lina SUN
2
;
Chuan LI
2
;
Wei WU
2
;
Qin WANG
2
;
Mifang LIANG
2
;
Dexin LI
2
;
Yan WEI
1
;
Jiandong LI
2
Author Information
1. Key Laboratory of Environmental Pollution Monitoring and Disease Control, Ministry of Education, Department of Occupational and Environmental Health, School of Public Health, Guizhou Medical University, Guiyang 550025, China
2. National Institute for Viral Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China
- Publication Type:Journal Article
- Keywords:
Zika virus;
Chikungunya virus;
Mayaro virus;
Triplex Real-time RT-PCR
- From:
Chinese Journal of Experimental and Clinical Virology
2019;33(6):632-636
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To establish a method for the simultaneous identification of Zika, Chikungunya and Mayaro viruses.
Methods:The complete genome sequences of Zika, Chikungunya and Mayaro virus were retrieved from Global Shared Database for comparative analysis, estimate its conservative region and determine the target gene location, specific primers and probes were designed, then a triplex real-time RT-PCR assay was developed. The specificity, sensitivity and repeatability of the assay were assessed by viral nucleic acid of Zika virus, Chikungunya virus a, in vitro transcriptional RNA of Mayaro virus, normal human serum and related virus simulation sample.
Results:The result showed that the established method could detect Zika virus, Chikungunya virus, as well as simulated Mayaro virus samples, the limit of detection (LOD) of Zika and Chikungunya virus was 16.22 Copy/PCR and 12.02 Copy/PCR, respectively, the LOD for simulated Mayaro virus RNA was 2.82 Copy/PCR, no significant difference was detected between the triplex and monoplex assays. No cross reaction was found in the detection of dengue virus, Hantavirus, severe fever with thrombocytopenia syndrome (SFTS) virus, yellow fever virus and influenza virus, and 100 healthy adults blood samples, the specificity of the method was 100%. The repeatability result showed that the standard deviation of all three detections were blow 0.5 and the coefficient of variation was less than 2% by selecting viral nucleic acids or transcribed RNA with high, medium and low concentration gradients.
Conclusions:A triplex real-time RT-PCR assay for detection of Zika, Chikungunya and Mayaro virus has been established with an acceptable specificity, sensitivity and repeatability.
