Detection of human parechoviruses from clinical stool samples in Aichi, Japan.

Between April 1999 and March 2008, a total of 4,976 stool specimens collected from patients with suspected viral infection through infectious agent surveillance in Aichi, Japan, were tested for the presence of human parechoviruses (HPeVs). We detected HPeVs in 110 samples by either cell culture, reverse transcriptase PCR (RT-PCR), or both. Serotyping either by neutralization test or by nucleotide sequence determination and phylogenetic analysis of the VP1 region and 5' untranslated region (5'UTR) regions revealed that 63 were HPeV type 1 (HPeV-1), followed by 44 HPeV-3 strains, 2 HPeV-4 strains, and 1 HPeV-6 strain. The high nucleotide and amino acid sequence identities of the Japanese HPeV-3 isolates in 2006 to the strains previously reported from Canada and Netherlands confirmed the worldwide prevalence of HPeV-3 infection. Ninety-seven percent of the HPeV-positive patients were younger than 3 years, and 86.2% younger than 12 months. The clinical diagnoses of HPeV-positive patients were gastroenteritis, respiratory illness, febrile illness, exanthema, "hand, foot, and mouth disease," aseptic meningitis, and herpangina. Among 49 HPeV-positive patients with gastroenteritis, 35 were positive with HPeV-1 and 12 with HPeV-3, and out of 25 with respiratory illness, 11 were positive with HPeV-1 and 14 with HPeV-3. HPeV-3 seemed to be an important etiological agent of respiratory infection of children. While HPeV-1 was detected predominantly during fall and winter, the majority of the HPeV-3 cases were detected during summer and fall. A different pattern of clinical manifestations as well as seasonality suggested that there are different mechanisms of pathogenesis between HPeV-1 and HPeV-3 infections.

Isolation and characterization of a new species of kobuvirus associated with cattle.

A cytopathic agent was isolated using Vero cells from the culture medium of HeLa cells that had been used for more than 30 years in our laboratory. This agent, termed U-1 strain, was serially passed in Vero cells with distinct CPE. Particles of U-1 strain negatively stained with phosphotungstic acid exhibited a distinct surface that resembled Aichi virus. The RNA genome of U-1 strain comprises 8374 nt, with a genome organization analogous to that of picornaviruses. Possible cleavage sites of the large ORF, which encoded a leader protein prior to the capsid protein region, were assigned following amino acid alignment with Aichi virus. The virus sequence had 33 and 75 % amino acid identity with the Aichi virus VP1 and 3D regions, respectively, but no more than 23 and 36 % with those of the prototype strains of other PICORNAVIRIDAE: The dendrogram based on the P1, P2 and P3 proteins indicated that U-1 strain is genetically included in the genus Kobuvirus but is distinct from Aichi virus. Of 72 cattle sera, 43 (59.7 %) were positive for neutralizing antibody against U-1 strain at a titre of 1 : 8 or more. However, sera from 190 humans, 242 monkeys, 139 pigs, 5 horses, 22 dogs and 9 cats did not neutralize U-1 strain at a 1 : 4 dilution. RNA corresponding to U-1 strain was detected in 12 (16.7 %) of 72 faecal samples from cattle by RT-PCR. These results indicated that U-1 strain, suspected to be a contaminant from calf sera, is a new species of the genus Kobuvirus, now termed bovine kobuvirus.