RT-PCR Detection of Coxsackievirus B۳: A Viral Myocarditis

Backgrounds and Aims: Coxsakievirus B۳ (CVB۳), one of the six Coxsakievirus B serotypes, is a member of the Enterovirus genus within the Picornaviridae family. CVB۳ is an important pathogen of viral myocarditis, which accounts for more than ۵۰% of viral myocarditis cases. The genome of CVB۳, like that of other Entroviruses, is a single-stranded, sense, polyadenylated RNA molecule with ۷۴۰۰ nucleotides in length and a single open reading frame (ORF), flanked by ۵΄ and ۳΄ non-translated regions. The capsids of coxcakieviruses are composed of the four structural proteins: viral protein-۱ (VP۱), VP۲, VP۳, and VP۴. In the present study, a new set of primers were designed based on the VP۱ for RT-PCR detection of CVB۳. Materials and Methods: Total RNA was extracted from CVB۳-infected HeLa cell line and cDNA was synthesized using random primers. Then, PCR was carried out by specific primers and the PCR product analysis was performed using ۱% agarose gel electrophoresis. Moreover, the sensitivity and specificity of this method were determined using serial dilution of CVB۳ cDNA and three genuses of entroviruses, respectively. Results: RT-PCR assay revealed a ۲۳۴ bp specific amplified fragment. The sensitivity of this test was determined ۵.۷۲ fg/µl cDNA. On the other hand, the specificity was successful in comparison with coxsackievirus A۱۶, Echovirus ۳۶ and Rhinovirus. Conclusions: The RT-PCR is a highly sensitive and rapid technique for detecting CVB۳ infection. Moreover, this method can be used as an easy diagnostic test in regard with CVB۳ detection in the clinical laboratories.

کلیدواژه ها:

Myocarditis ، RNA extraction ، RT-PCR