Molecular determinants for virulence in coxsackievirus B1 infection

Abstract

Studies demonstrated that a strain derived from an infectious clone of coxsackievirus B1 (CVB1N) (N. Iizuka, H. Yonekawa, and A. Nomoto, J. Virol. 65:4867-4873, 1991) was 3 to 4 log 10 less virulent than the myotropic Tucson strain of CVB1 (CVB1T) following intraperitoneal inoculation of newborn mice. Replacement of nucleotides (nt) 69 to 804 from the 5 untranslated region (5 UTR) and 1A coding region of CVB1N or nt 117 to 161 from the 5 UTR with the corresponding part from CVB1T restored greater than 90% of the virulence. Sequencing of the 5 UTR of CVB1T demonstrated areas with a greater similarity to particular echoviruses than to CVB1N, suggesting that recombination events might have occurred, perhaps influencing the virulence phenotype.

Molecular determinants of disease in coxsackievirus B1 murine infection

“…The cDNA engineered behind the T7 promoter was in vitro transcribed after linearization with Xba I. The resultant RNA was transfected into HeLa cells to produce infectious virus, which was purified partially by pelleting through a 30% sucrose cushion as described previously [Rinehart et al, 1997].…”

“…The viral variant CVB1Nm was plaque‐purified twice and semi‐purified as described above. The viral RNA was isolated and subjected to RT‐PCR as described previously [Rinehart et al, 1997]. Products obtained from nucleotides 1 to 3,448, 1,571 to 5,135, and 4,916 to polyA were cloned into pGEM‐T vector (Promega, Madison, WI).…”

“…In addition, it has been suggested that the 5′ UTR stem loop II (SLII) structure [Lee et al, 1997; Dunn et al, 2000] plays a major role in myocarditis [Dunn et al, 2003; Stadnick et al, 2004]. Furthermore, SLII has been found to determine virulence in CVB1‐inoculated newborn mice [Rinehart et al, 1997]. Studies have demonstrated that single mutation at VP2‐165 (in the “puff” loop) [Knowlton et al, 1996; Stadnick et al, 2004] or at VP1‐155 [Cameron‐Wilson et al, 1998] are important for the myocarditic phenotype.…”

Nucleotide Sequence of the 5′Nontranslated and Virion Polypeptides Regions of Coxsackievirus B6

“…Since several reports suggested the occurrence of recombination among enteroviruses (3,6,7,38,39,49), we were interested in searching the sequence traits of recombination between CVB6 and other CVBs. We found that 19 amino acid residues at the carboxyterminus of VP4 were the same between CVB6 and CVB4.…”

“…Previous studies also showed that the cardiovirulent phenotype of CVB3 was determined by a single site in the 5' nontranslated region (5' NTR) which do not contribute to serotypes (33,46). Therefore, the comparison of CVB sequences is important not only for determining serotypes but for virulence variation (38). So far, entire nucleotide sequences of CVB1, CVB3, CVB4 and CVB5 genomes have been determined (13,15,18,22,49).…”