Relationships among porcine and human P[6] rotaviruses: Evidence that the different human P[6] lineages have originated from multiple interspecies transmission events

Abstract

Porcine rotavirus strains (PoRVs) bearing human-like VP4 P[6] gene alleles were identified. Genetic characterization with either PCR genotyping or sequence analysis allowed to determine the VP7 specificity of the PoRVs as G3, G4, G5 and G9, and the VP6 as genogroup I, that is predictive of a subgroup I specificity. Sequence analysis of the VP8* trypsin-cleavage product of VP4 allowed PoRVs to be characterized further into genetic lineages within the P[6] genotype. Unexpectedly, the strains displayed significantly higher similarity (up to 94.6% and 92.5% at aa and nt level, respectively) to human M37-like P[6] strains (lineage I), serologically classifiable as P2A, or to the atypical Hungarian P[6] human strains (HRVs), designated as lineage V (up to 97.0% aa and 96.1% nt), than to the porcine P[6] strain Gottfried, lineage II (<85.1% aa and 82.2 nt), which is serologically classified as P2B. Interestingly, no P[6] PoRV resembling the original prototype porcine strain, Gottfried, was detected, while Japanase P[6] PoRV clustered with the atypical Japanase G1 human strain AU19. By analysis of the 10th and 11th genome segments, all the strains revealed a NSP4B genogroup (Wa-like) and a NSP5/6 gene of porcine origin. These findings strongly suggest interspecies transmission of rotavirus strains and/or genes, and may indicate the occurrence of at least 3 separate rotavirus transmission events between pigs and humans, providing convincing evidence that evolution of human rotaviruses is tightly intermingled with the evolution of animal rotaviruses.

Genomic characterization of a novel group A lamb rotavirus isolated in Zaragoza, Spain

“…An isolate was propagated onto African green monkey kidney (MA-104) cells as described [21][22][23]. Evidence of viral replication was checked both by the appearance cytopathic effect and indirect immunofluorescence test, using a rabbit hyperimmune serum raised against a group A rotavirus (strain Bo/A-125/96 P7 [5],G6).…”

“…Cryolysates were extracted with Vertrel XF (Dupont, Wilmington, DE) as previously described [23] and clarified twice by centrifugation for 30 min at 4,800g. Subsequently, the supernatants were centrifuged for 8 h at 85,000g on a 30% sucrose layer in SW28 rotor (Beckman Coulter, Inc., Fullerton, CA).…”

“…Subsequently, the supernatants were centrifuged for 8 h at 85,000g on a 30% sucrose layer in SW28 rotor (Beckman Coulter, Inc., Fullerton, CA). After digestion of viral pellet with 1% sodium-dodecyl-sulfate (SDS) and proteinase K 1 lg/ml (Sigma-Aldrich, Milano, Italy), viral dsRNA was extracted and the e-type was visualized as described [21][22][23].…”

“…Virus Genes (2008) 37:250-265 251 RNA extraction and PCR amplification of the VP8*, VP7, VP6, NSP4, and NSP5/NSP6 genes Viral dsRNA from the fecal specimen was extracted by adsorption on cellulose CF11 as described previously [23]. Viral dsRNA was denatured in dimethyl-sulfoxide (SigmaAldrich, Milano, Italy) at 97°C for 5 min.…”

“…In recent years, epidemiological surveillance to monitor the appearance of novel rotavirus antigenic types has intensified throughout the world, yielding evidence for the increasing antigenic diversity of group A rotaviruses [21][22][23]. Although the number of characterized OVR strains is small, OVRs seem to represent a diverse group, suggesting that sheep may interchange rotaviruses readily.…”