Genomic Determinants of Cardiovirulence in Coxsackievirus B3 Clinical Isolates: Localization to the 5′ Nontranslated Region

Abstract

Coxsackievirus B3 (CVB3) infections can cause myocarditis in humans and are implicated in the pathogenesis of dilated cardiomyopathy. The natural genetic determinants of cardiovirulence for CVB3 have not been identified, although using strains engineered in the laboratory, cardiovirulence determinants have been identified in the CVB3 5′ nontranslated region (5′NTR) and capsid. The myocarditic phenotypes of two CVB3 clinical isolates were determined using an established murine model of inflammatory heart disease. The 5′NTRs and capsid proteins of the noncardiovirulent CVB3/CO strain and cardiovirulent CVB3/AS strain were examined to determine their influence on the cardiovirulence phenotype. Six intratypic chimeric viruses were constructed in which 5′NTR and capsid sequences of the infectious cDNA copy of the cardiovirulent CVB3/20 genome were replaced by homologous sequences from CVB3/CO or CVB3/AS. Chimeric strains were tested for cardiovirulence by inoculation of C3H/HeJ mice. Sections of hearts removed at 10 days postinoculation were examined for evidence of myocarditis by light microscopy and assayed for the presence of virus. Replacement of the CVB3/20 capsid coding region by that from the homologous region of CVB3/CO resulted in no change in the cardiovirulent CVB3/20 phenotype, with virus recoverable from the heart at 10 days postinoculation. However, recombinant virus containing the CVB3/CO 5′NTR alone or the 5′NTR and capsid sequences together were not myocarditic, and infectious virus was not recovered from the myocardium. Chimeric viruses containing the CVB3/AS 5′NTR alone, capsid sequence alone, or both together preserved the myocarditic phenotype. These data support the 5′NTR as the primary site in the determination of the natural cardiovirulence phenotype of CVB3.