A Novel DANP-Coupled Hairpin RT-PCR for Rapid Detection of Chikungunya Virus

作者：Chen Huixin; Takei Fumie; Koay Evelyn Siew Chuan; Nakatani Kazuhiko; Chu Justin Jang Hann^*

来源：Journal of Molecular Diagnostics, 2013, 15(2): 227-233.

DOI：10.1016/j.jmoldx.2012.10.004

Chikungunya has re-emerged as an important arboviral infection of global health significance. Because of lack of a vaccine and effective treatment, rapid diagnosis plays an important role in early clinical management of patients. In this study, we have developed a novel molecular diagnostic platform that ensures a rapid and cost-effective one-step RT-PCR assay, with high sensitivity and specificity, for the early detection of the Chikungunya virus (CHIKV). It uses 2,7-diamino-1,8-naphthyridine derivative (DANP)-labeled cytosine-bulge hairpin primers to amplify the nsP2 region of the CHIKV genome, followed by measurement of the fluorescence emitted from DANP-primer complexes after PCRs. The detection Limit of our assay was 0.01 plaque-forming units per reaction of CHIKV. Furthermore, the HP-nsP2 primers were highly specific in detecting CHIKV, without any cross-reactivity with the panel of RNA viruses validated in this study. The feasibility of the DANP-coupled hairpin RT-PCR for clinical diagnosis was evaluated using clinical serum samples from CHIKV-infected patients, and the specificity and sensitivity were 100% (95% CI, 80.0% to 100%) and 95.5% (95% CI, 75.1% to 99.8%), respectively. These findings confirmed its potential as a point-of-care clinical molecular diagnostic assay for CHIKV in acute-phase patient serum samples. (J Mol Diagn 2013, 15: 227-233; http://dx.doi.org/10.1016/j.jmoldx.2012.10.004)

• 出版日期2013-3