Micro-droplet Digital Polymerase Chain Reaction and Real-Time Quantitative Polymerase Chain Reaction Technologies Provide Highly Sensitive and Accurate Detection of Zika Virus
The establishment of highly sensitive diagnostic methods is critical in the early diagnosis and control of Zika virus (ZIKV) and in preventing serious neurological complications of ZIKV infection. In this study, we established micro-droplet digital polymerase chain reaction (ddPCR) and real-time quantitative PCR (RT-qPCR) protocols for the detection of ZIKV based on the amplification of the NS5 gene. For the ZIKV standard plasmid, the RT-qPCR results showed that the cycle threshold (Ct) value was linear from 101 to 108 copy/μL, with a standard curve R2 of 0.999 and amplification efficiency of 92.203%; however, a concentration as low as 1 copy/μL could not be detected. In comparison with RT-qPCR, the ddPCR method resulted in a linear range of 101–104 copy/μL and was able to detect concentrations as low as 1 copy/μL. Thus, for detecting ZIKV from clinical samples, RT-qPCR is a better choice for high-concentration samples (above 101 copy/μL), while ddPCR has excellent accuracy and sensitivity for low-concentration samples. These results indicate that the ddPCR method should be of considerable use in the early diagnosis, laboratory study, and monitoring of ZIKV.
KeywordsZika virus Nucleic acid detection Micro-droplet digital polymerase chain reaction (ddPCR) Real-time fluorescence quantitative polymerase chain reaction (RT-qPCR)
This study was supported by the National Natural Science Foundation of China (Nos. 31470271 and 81730110) and Guangzhou Science and Technology Program key projects (No. 201803040006). We thank Changwen Ke, De Wu, and Jiufeng Sun from the Guangdong Center for Disease Control and Prevention for providing the Asian ZIKV Z16006 strain. The authors would like to thank anonymous referees for their valuable suggestions, which have improved the paper immensely.
ZQ and YH designed the experiments. ZW, LZ, JL, XL, HF, and SF carried out the experiments. JY, XH and WL analyzed the data. WX, and QW wrote the paper. BZ and WZ checked and finalized the manuscript. All authors read and approved the final manuscript.
Compliance with Ethics Standards
Conflict of interest
The authors declare that they have no conflict of interest.
Animal and Human Rights Statement
The study was approved by the Ethics Committees of Southern Medical University. All participants provided written informed consent.
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