RT-PCR Detection of Coxsackievirus B3: A Viral Myocarditis

Backgrounds and Aims: Coxsakievirus B3 (CVB3), one of the six Coxsakievirus B serotypes, is a member of the Enterovirus genus within the Picornaviridae family. CVB3 is an important pathogen of viral myocarditis, which accounts for more than 50% of viral myocarditis cases. The genome of CVB3, like that of other Entroviruses, is a single-stranded, sense, polyadenylated RNA molecule with 7400 nucleotides in length and a single open reading frame (ORF), flanked by 5΄ and 3΄ non-translated regions. The capsids of coxcakieviruses are composed of the four structural proteins: viral protein-1 (VP1), VP2, VP3, and VP4. In the present study, a new set of primers were designed based on the VP1 for RT-PCR detection of CVB3.
Total RNA was extracted from CVB3-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 1% agarose gel electrophoresis. Moreover, the sensitivity and specificity of this method were determined using serial dilution of CVB3 cDNA and three genuses of entroviruses, respectively.
RT-PCR assay revealed a 234 bp specific amplified fragment. The sensitivity of this test was determined 5.72 fg/µl cDNA. On the other hand, the specificity was successful in comparison with coxsackievirus A16, Echovirus 36 and Rhinovirus.
The RT-PCR is a highly sensitive and rapid technique for detecting CVB3 infection. Moreover, this method can be used as an easy diagnostic test in regard with CVB3 detection in the clinical laboratories.