Rapid Detection and Molecular Typing of Dengue Virus by Using Multiplex-Nested-RT-PCR

Abstract

world. We have evaluated the combination of one-step RT-PCR and multiplex nested PCR assays for detecting dengue viruses from clinical samples. Twelve patients were screened for the dengue virus, using a pair of primers that conserve for several Flavivirus. The results showed that in 12 suspect patients, 100% were positive for Flavivirus and there are some genotypic variation among them, that indicated by several RT-PCR products higher than 511 bp, the expected product for RT-PCR. Further assay was performed to clarify the presence and serotypes of dengue virus using multiplex nested PCR. Serotyping results indicated that 83,3% of samples can be confirmed for dengue virus. Among the dengue virus positive 16,7 % are dengue-2, 16.7 % are dengue-3, and the most common 50% are dengue-4, whereas dengue-1 were not found among the patients. The combination of RT-PCR and multiplex nested PCR assay can be used for rapid analysis dengue samples in early phase which is potentially useful for clinical, epidemiology and also evolutionary studies. Key words: Flavivirus, dengue virus, serotype, RT-PCR, multiplex nested PCR