Development and evaluation of the detection method of microfluidic real-time RT-PCR assays for Zika, Dengue, Yellow fever and Chikungunya viruses
10.3760/cma.j.cn112866-20191216-00201
- VernacularTitle:寨卡病毒、登革病毒、黄热病毒、基孔肯雅病毒微流控荧光定量RT-PCR检测方法的建立及评价
- Author:
Dongyang YU
1
;
Aqian LI
;
Yang LIU
;
Naizhe LI
;
Yu ZHU
;
Chuan LI
;
Jiandong LI
;
Shiwen WANG
;
Dexin LI
;
Mifang LIANG
;
Yan LIU
Author Information
1. 安徽医科大学基础医学院微生物学教研室，合肥 230032
- Keywords:
Microfluidic qRT-PCR;
Zika virus;
Dengue virus;
Yellow fever virus;
Chikungunya virus
- From:
Chinese Journal of Experimental and Clinical Virology
2020;34(2):186-190
- CountryChina
- Language:Chinese
-
Abstract:
Objective:To develop a rapid nucleic acid detection method for Zika virus (ZIKV), Dengue virus (DENV), Yellow fever virus (YFV), Chikungunya virus (CHIKV) based on microfluidic fluorescence quantitative RT-PCR technologies, in order to achieve rapid diagnosis of these four viral infections.Methods:Four sets of specific primers and probes were designed targeting the NS1 gene of ZIKV, the NS5 gene of DENV, and YFV, the E1 gene of CHIKV, respectively. The sensitivity was evaluated using in vitro transcribed RNA of ZIKV, DENV, YFV and CHIKV, and the specificity were evaluated using other viral nucleic acid. ZIKV, YFV and CHIKV detection method were verified using simulated positive samples, and DENV detection method was verified using clinical patient samples, the result of which were also compared with the quantitative RT-PCR detection method . Results:The limit of detection (LOD) of ZIKV, DENV, YFV, and CHIKV microfluidic qRT-PCR method were 14.57 copies/μl, 94.27 copies/μl, 8.25 copies/μl, and 223.19 copies/μl, respectively, and the four detection method showed no cross-reactivity with other viral nucleic acids. The prepared ZIKV, YFV and CHIKV simulated positive samples were 100% detected, and the variation coefficient of Ct value measured at each concentration were all around 2%; the 20 clinical patient specimens of DENV infection were 100% detected, which is consistent with the result of fluorescent quantitative RT-PCR detection.Conclusions:The ZIKV, DENV, YFV, and CHIKV microfluidic quantitative RT-PCR detection method showed good sensitivity, specificity, and stability. The detection could be completed within 25 minutes, which could be used for laboratory detection and early diagnosis.
