Prevalence of anisakid parasites in fish collected from Apulia region (Italy) and quantification of nematode larvae in flesh.

Anisakis spp. and Hysterothylacium spp. are nematodes that commonly parasitize several fish species. Nematode larvae can be recovered in coelomic cavity and viscera, but also in flesh and have an important economic and public health impact. A total of 1144 subjects of wild teleosts, 340 samples of cephalopods and 128 specimens of farmed fish collected from Apulia region were analysed for anisakid larvae detection by visual inspection of coelomic cavity and viscera and by digestion of the flesh. No nematode larvae were found in farmed fish and cephalopod molluscs. All examined wild-caught fish species were parasitized, except for 5 species for each of which only a few subjects belonging to the same batch were sampled, therefore the results are just indicative. A total of 6153 larvae were isolated; among these, 271 larvae were found in the muscular portion. Larvae were identified by morphological method as belonging to the genera Anisakis (97.2%) (type I and type II) and Hysterothylacium (2.8%). Both nematodes could be found in all fish species, except for round sardinella (Sardinella aurita), infected only by Hysterothylacium spp. and for Mediterranean scaldfish (Arnoglossus laterna), little tunny (Euthynnus alleteratus) and chub mackerel (Scomber japonicus) infected only with Anisakis spp.. A sample of 185 larvae was sent to the National Reference Centre for Anisakiasis (C.Re.N.A.) of Sicily for identification at the species level: 180 larvae belonged to the species A. pegreffii and 2 larvae to A. physeteris. The remaining 3 larvae were identified at genus level as Hysterothylacium. Statistical indices such as prevalence, mean intensity and mean abundance were calculated. Chub mackerel (S. japonicus) was the species with the highest prevalence and mean intensity. Moreover, the average and the median values of larvae per 100 g of edible part for each fish species were determined to estimate the consumer exposure to Anisakis spp.. The obtained values were then recalculated by referring to the edible part of all specimens (infected and non-infected) forming a single parasitized batch, getting more realistic and objective data useful for risk assessment. Our results indicate that the consumption of raw or undercooked wild fish caught off Apulian coasts could result in the acquisition of anisakiasis; on the contrary, farmed fish and cephalopods appear to be safer for the consumer.

Detection and Full-Length Genome Characterization of Novel Canine Vesiviruses.

Vesiviruses have been detected in several animal species and as accidental contaminants of cells. We detected vesiviruses in asymptomatic kennel dogs (64.8%) and symptomatic (1.1%) and asymptomatic (3.5%) household dogs in Italy. The full-length genome of 1 strain, Bari/212/07/ITA, shared 89%-90% nt identity with vesiviruses previously detected in contaminated cells.

Analysis of the ORF2 of human astroviruses reveals lineage diversification, recombination and rearrangement and provides the basis for a novel sub-classification system.

Canonical human astroviruses (HAstVs) are important enteric pathogens that can be classified genetically and antigenically into eight types. Sequence analysis of small diagnostic regions at either the 5' or 3' end of ORF2 (capsid precursor) is a good proxy for prediction of HAstV types and for distinction of intratypic genetic lineages (subtypes), although lineage diversification/classification has not been investigated systematically. Upon sequence and phylogenetic analysis of the full-length ORF2 of 86 HAstV strains selected from the databases, a detailed classification of HAstVs into lineages was established. Three main lineages could be defined in HAstV-1, four in HAstV-2, two in HAstV-3, three in HAstV-4, three in HAstV-5 and two in HAstV-6. Intratypic (inter-lineages) ORF2 recombinant strains were identified in type 1 (1b/1d) and type 2 (2c/2b) with distinct crossover points. Other potential intratypic recombinant strains were identified in type 3, type 5 and type 6. In addition, a type-1b strain with a large insertion (~600 bp) of heterologous RNA in the N-terminal region and a type-6 strain with a large RNA rearrangement in the hypervariable region were identified. The classification scheme was integrated in a novel nomenclature system suitable for designation of HAstV strains.

Full-genome analysis of a canine pneumovirus causing acute respiratory disease in dogs, Italy.

An outbreak of canine infectious respiratory disease (CIRD) associated to canine pneumovirus (CnPnV) infection is reported. The outbreak occurred in a shelter of the Apulia region and involved 37 out of 350 dogs that displayed cough and/or nasal discharge with no evidence of fever. The full-genomic characterisation showed that the causative agent (strain Bari/100-12) was closely related to CnPnVs that have been recently isolated in the USA, as well as to murine pneumovirus, which is responsible for respiratory disease in mice. The present study represents a useful contribution to the knowledge of the pathogenic potential of CnPnV and its association with CIRD in dogs. Further studies will elucidate the pathogenicity and epidemiology of this novel pneumovirus, thus addressing the eventual need for specific vaccines.

Hobi-like pestivirus: both biotypes isolated from a diseased animal.

A Hobi-like pestivirus pair consisting of cytopathogenic (cp) and non-cytopathogenic (noncp) strains, Italy 83/10cp and Italy 83/10ncp, was isolated from the lung of a heifer that died of respiratory disease. The noncp and cp viruses were isolated on Madin-Darby bovine kidney cells and separated by plaque purification and end point dilution. Analysis of the nearly full-length genomes revealed that the two viruses were very closely related to each other and to the noncp Hobi-like strain Italy 1/10-1, which had been isolated a few weeks earlier from the same herd. One major difference between noncp and cp viruses concerned the presence of a cellular Jiv sequence in the 3' domain of the NS2-encoding region of the cp strain. This is the first study, to our knowledge, reporting the isolation and molecular characterization of a Hobi-like virus pair.

Discovery and genomic characterization of noroviruses from a gastroenteritis outbreak in domestic cats in the US.

Norovirus (NoV) RNA was detected in the stools of 6 out 14 (42.8%) 8-12-week-old cats with enteritis from a feline shelter, in New York State. Upon sequence analysis of the complete capsid, the six NoVs were found to be identical, suggesting the spread of a unique NoV strain in the shelter. The full-length genomic sequence (7839 nt) of one feline NoV, CU081210E/2010/US, was determined. In the capsid protein VP1 region, the virus displayed the highest amino acid identity to animal genogroup IV genotype 2 (GIV.2) NoVs: lion/Pistoia-387/06/IT (97.9%) and dog/Bari-170/07/IT (90.4%). These findings document the discovery of a novel feline calicivirus, different from vesiviruses, and extend the spectrum of NoV host range. Epidemiological studies using feline NoV-specific diagnostic tools and experimental infection of cats are required to understand whether NoVs have a pathogenic role in this species.

Hobi-like pestivirus in aborted bovine fetuses.

An outbreak of abortion affecting multiparous cows was associated with Hobi-like pestivirus infection. Viral RNA and antigens were detected in the tissues of two aborted fetuses. Molecular assays for other common abortogenic agents tested negative. At the genetic level, the Hobi-like pestivirus displayed the closest relatedness to Italian, Australian, and South American viruses, whereas it diverged from the prototype Thai isolate. These findings may have important implications for the pestivirus control/eradication programs in cattle herds.

Astroviruses in rabbits.

By screening rabbits with enterocolitis or enteritis complex and asymptomatic rabbits, we identified a novel astrovirus. The virus was distantly related (19.3%-23.7% aa identity) in the capsid precursor to other mammalian astroviruses within the Mamastrovirus genus. By using real-time reverse transcription PCR, with specific primers and probes and targeting a conserved stretch in open reading frame 1b, we found rabbit astrovirus in 10 (43%) of 23 samples from animals with enteric disease and in 25 (18%) of 139 samples from asymptomatic animals in Italy during 2005-2008. The mean and median titers in the positive animals were 10(2)× and 10(3)× greater, respectively, in the symptomatic animals than in the asymptomatic animals. These findings support the idea that rabbit astroviruses should be included in the diagnostic algorithm of rabbit enteric disease and animal experiments to increase information obtained about their epidemiology and potential pathogenic role.

Canine noroviruses.

Noroviruses are recognized as emerging enteric pathogens of humans and have been identified in recent years in a number of mammalian species. The role of noroviruses as pathogens in immune-competent animals and under natural conditions remains uncertain, although both homologous and heterologous animal models are now available to investigate the pathogenesis, the immune response, and the molecular mechanism regulating norovirus infection. Recently, evidence has been gathered that noroviruses may also circulate in domestic carnivores. The zoonotic implications of these novel viruses deserve more attention, due to the strict social interactions between humans and pets.

Mixed infection by Feline astrovirus and Feline panleukopenia virus in a domestic cat with gastroenteritis and panleukopenia.

Astroviruses are important pathogens of human beings and animals. Feline astroviruses have been identified by electron microscopy in the feces of either asymptomatic or symptomatic cats, and experimental infection with one isolate was shown to induce enteric clinical signs and virus excretion, thus suggesting a possible role as enteric pathogen. However, due to the lack of specific diagnostic assays, feline astroviruses are not included in the diagnostic algorithms of feline infectious diseases, and their role as feline pathogens remains unclear. The present report describes a dual infection by Feline astrovirus and Feline panleukopenia virus in a 4-month-old cat with severe gastroenteritis and panleukopenia. Upon sequence analysis, the Feline astrovirus strain was found to be genetically related to astroviruses identified in human beings and felids.

Detection and characterization of canine astroviruses.

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.