RESULTS:

Group A rotaviruses (RVAs) are a major cause of severe enteritis in humans and animals. RVAs have been identified in several animal species and their genetic diversity the segmented nature of their RNA genome and the ability to spill over from one species to another can generate new RVA strains. In this study we investigated the genome constellations of an unusual rare bovine RVA strain G15P[21] identified from a farm with neonatal diarrhoea of calves in 2006. In parallel the genome constellations of other RVA strains with different G/P types identified from the same farm in the same time span (2006–2008) were analysed. The genome constellation of strain K53 was G15-P[21]-I2-R2-C2-M2-A13-N2-T9-E2-H3 and was similar overall to that of the other bovine RVA strains (G6/10-P[11]-I2-R2-C2-M2-A13-N2-T6-E2-H3) with the exception of the NSP3 segment (T9 vs T6). This study describes RVA genomes with different genotype combinations isolated at a farm and also contributes to the understanding of the diversity and evaluation of rotavirus in a global context.

The family Caliciviridae includes viruses with single-stranded positive-sense RNA genomes of 7.4–8.3 kb. The most clinically important representatives are human noroviruses which are a leading cause of acute gastroenteritis in humans. Virions are non-enveloped with icosahedral symmetry. Members of seven genera infect mammals (Lagovirus Norovirus Nebovirus Recovirus Sapovirus Valovirus and Vesivirus) members of two genera infect birds (Bavovirus and Nacovirus) and members of two genera infect fish (Minovirus and Salovirus). This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the family Caliciviridae which is available at ictv.global/report/caliciviridae.

Group C rotaviruses (RVC) are enteric pathogens of humans and animals. Whole-genome sequences are available only for few RVCs leaving gaps in our knowledge about their genetic diversity. We determined the full-length genome sequence of two human RVCs (PR2593/2004 and PR713/2012) detected in Italy from hospital-based surveillance for rotavirus infection in 2004 and 2012. In the 11 RNA genomic segments the two Italian RVCs segregated within separate intra-genotypic lineages showed variation ranging from 1.9 % (VP6) to 15.9 % (VP3) at the nucleotide level. Comprehensive analysis of human RVC sequences available in the databases allowed us to reveal the existence of at least two major genome configurations defined as type I and type II. Human RVCs of type I were all associated with the M3 VP3 genotype including the Italian strain PR2593/2004. Conversely human RVCs of type II were all associated with the M2 VP3 genotype including the Italian strain PR713/2012. Reassortant RVC strains between these major genome configurations were identified. Although only a few full-genome sequences of human RVCs mostly of Asian origin are available the analysis of human RVC sequences retrieved from the databases indicates that at least two intra-genotypic RVC lineages circulate in European countries. Gathering more sequence data is necessary to develop a standardized genotype and intra-genotypic lineage classification system useful for epidemiological investigations and avoiding confusion in the literature.

Norovirus (NoV) is one of the major causes of diarrhoeal disease with epidemic outbreak and sporadic patterns in humans of all ages worldwide. NoVs of genotype GII.4 cause nearly 80–90 % of all NoV infections in humans. Periodically some GII.4 strains become predominant generating major pandemic variants. Retrospective analysis of the GII.4 NoV strains detected in Italy between 2007 and 2013 indicated that the pandemic variant New Orleans 2009 emerged in Italy in the late 2009 became predominant in 2010–2011 and continued to circulate in a sporadic fashion until April 2013. Upon phylogenetic analysis based on the small diagnostic regions A and C the late New Orleans 2009 NoVs circulating during 2011–2013 appeared to be genetically different from the early New Orleans 2009 strains that circulated in 2010. For a selection of strains a 3.2 kb genome portion at the 3′ end was sequenced. In the partial ORF1 and in the full-length ORF2 and ORF3 the 2011–2013 New Orleans NoVs comprised at least three distinct genetic subclusters. By comparison with sequences retrieved from the databases these subclusters were also found to circulate globally suggesting that the local circulation reflected repeated introductions of different strains rather than local selection of novel viruses. Phylogenetic subclustering did not correlate with changes in residues located in predicted putative capsid epitopes although several changes affected the P2 domain in epitopes A C D and E.

Group A rotaviruses (RVAs) are a leading cause of viral gastroenteritis in children with G2P[4] RVA being one of the most common human strains worldwide. The complete genome sequences of nine G2P[4] RVA strains selected from a 26-year archival collection (1985–2011) established in Palermo Italy were determined. A strain associated with a peak of G2P[4] RVA activity in 1996 resembled a reassortant strain identified in Kenya in 1982 and differed completely in genomic make up from more recent strains that circulated during 2004–2011. Conversely the 2004–2011 G2P[4] RVAs were genetically more similar to contemporary RVA strains circulating globally. Recent G2P[4] strains possessed either single or multiple genome segments (VP1 VP3 and/or NSP4) likely derived from ruminant viruses through intra-genotype reassortment. Amino acid substitutions were selected and maintained over time in the VP7 and VP8* antigenic proteins allowing the circulation of two contemporary G2P[4] variants to be distinguished. Altogether these findings suggest that major changes in the genomic composition of recent G2P[4] RVAs occurred in the early 2000s leading to the appearance of a novel variant of the DS-1-like genotype constellation. Whether the modifications observed in the neutralizing antigens and in the genome composition of modern G2P[4] RVAs may affect the long-term effectiveness of the vaccination programmes remains to be explored.

Astroviruses (AstVs) have been identified only occasionally in dogs. A canine AstV strain Bari/08/ITA was detected from a pup with gastroenteric signs and the virus was isolated in cell culture and characterized molecularly. In the full-length capsid protein the virus displayed genetic similarities (83.5 % aa identity) to another canine AstV strain although a high rate of variation occurred in the hypervariable domain which is related to AstV antigenic specificity. Specific antibodies were detected in the convalescent dog indicating seroconversion and in 59 % of a collection of dog serum samples. Using primers specific for canine AstV designed to detect a conserved region of ORF1b canine AstVs were detected in 24.5 % of young pups with gastroenteritis either alone or in mixed infections with other canine pathogens. In contrast AstVs were detected in only 9.3 % of asymptomatic pups. These findings indicate that canine AstVs are common in dogs and may suggest a possible role as canine enteric pathogens.

Two isolates of canine parvovirus (CPV) were obtained from dogs affected with severe haemorrhagic diarrhoea. Type 2b antigenic specificity was predicted by both antigenic analysis with monoclonal antibodies and PCR characterization with type-specific primers. Nevertheless sequence analysis of the capsid protein-encoding gene revealed two amino acid changes. One of the changes affected position 426 (Asp to Glu) in a major antigenic site of the viral capsid determining the replacement of a residue unique to CPV type 2b. The failure of established typing methods to distinguish this antigenic variant was overcome by the development of an RFLP assay.