Discovery of a novel human picornavirus in a stool sample from a pediatric patient presenting with fever of unknown origin

Abstract

Fever of unknown origin (FUO) is a serious problem in the United States. An unidentified agent was cultured from the stool of an infant who presented with FUO. This virus showed growth in HFDK cells and suckling mice. Using DNase sequence-independent single-primer amplification, we identified several nucleotide sequences with a high homology to Theiler's murine encephalomyelitis virus. Nearly full-length viral genome sequencing and phylogenetic analysis demonstrate that this virus is a member of the Cardiovirus genus of the Picornaviridae family.

FIG. 1.

DNase-SISPA amplification of tissue culture sample containing SAF-V. PCR products were analyzed on a 6.5% acrylamide gel. Lane L contains molecular size markers in base pairs.

FIG. 2.

Schematic representation of SAF-V (A), Theiler virus (B), TMEV (C), and EMCV (D) genomes. The boxes represent the genes that encode proteins. Solid black horizontal lines represent the UTRs. Yellow boxes represent the genes that encode the L* proteins.

FIG. 3.

Alignment of the cardiovirus leader peptides. The zinc finger domain is highlighted in gray, the acidic domain is highlighted in aqua, and the Ser/Thr domain is highlighted in yellow.

FIG. 4.

Alignment of the EF loop structure amino acids in VP2 of cardioviruses SAF-V, TMEV, Theiler-like virus, Vilyuisk virus, EMCV, and mengovirus. Areas highlighted in gray are part of the EF loop structure. The area highlighted in yellow is an addition to the CD loop structure of SAF-V. Amino acids shown in blue are identical to the amino acids in SAF-V.

FIG. 5.

Alignment of the CD loop amino acids in VP1 of cardioviruses SAF-V, TMEV, Theiler-like virus, Vilyuisk virus, EMCV, and mengovirus. Areas highlighted in yellow are part of the CD loop structure. Amino acids shown in red are identical to the amino acids in SAF-V.

FIG. 6.

Evolutionary relationships among Picornaviridae based on full-length genomes of picornaviruses (A) and based on sequences of each genomic region of cardioviruses (B). GenBank accession numbers are given in Materials and Methods. Abbreviations used for virus names: HEchoV 5, 22, and 23, human echoviruses 5, 22, and 23; HRV 1B, human rhinovirus 1B; PEV 1 and 8, porcine enterovirus 1 and 8; HAV, human hepatitis A virus; SHAV, simian hepatitis A virus; AEMV, avian encephalomyelitis virus; LjunganV, Ljungan virus; HPEchoV 2, human parechovirus 2; MengoV, mengovirus; T lV, Theiler-like virus; FMDV, foot-and-mouth disease virus; ERV 1, 2, and 3, equine rhinovirus 1, 2, and 3; AichiV, Aichi virus; PTV, porcine teschovirus.

FIG. 7.

Bootscan analysis of SAF-V based on full-length genomes. For virus names, see the legend of Fig. 6.