Mutation analysis of the spike protein in Italian feline infectious peritonitis virus and feline enteric coronavirus sequences.

Feline coronavirus (FCoV) exists as two different genotypes, FCoV type I and II, each including two biotypes, feline enteric coronavirus (FECV) and feline infectious peritonitis virus (FIPV), the latter being a virulent variant originating from the former virus. Recently, two amino acid substitutions, M1058L and S1060A, within the spike protein have been associated to the FECV/FIPV virulence change. In this study, we have analysed the frequency of detection of such mutations in FIPV and FECV strains circulating in Italian cats and obtained information about their evolutionary relationships with reference isolates. A total of 40 FCoV strains, including 19 strains from effusions or tissue samples of FIP cats and 21 strains from faecal samples of non-FIP cats, were analysed. Mutation M1058L was detected in 16/18 FCoV-I and 1/1 FCoV-II strains associated with FIP, while change S1060A was presented by two FIPV strains. By phylogenetic analysis, FCoV sequences clustered according to the genotype but not according to the biotype, with FECV/FIPV strains recovered from the same animal being closely related. Further studies are needed to better define the genetic signatures associated with the FECV/FIPV virulence shift.

Evidence of exposure to SARS-CoV-2 in cats and dogs from households in Italy.

SARS-CoV-2 originated in animals and is now easily transmitted between people. Sporadic detection of natural cases in animals alongside successful experimental infections of pets, such as cats, ferrets and dogs, raises questions about the susceptibility of animals under natural conditions of pet ownership. Here we report a large-scale study to assess SARS-CoV-2 infection in 817 companion animals living in northern Italy, sampled at a time of frequent human infection. No animals tested PCR positive. However, 3.4% of dogs and 3.9% of cats had measurable SARS-CoV-2 neutralizing antibody titers, with dogs from COVID-19 positive households being significantly more likely to test positive than those from COVID-19 negative households. Understanding risk factors associated with this and their potential to infect other species requires urgent investigation. ONE SENTENCE SUMMARY: SARS-CoV-2 antibodies in pets from Italy.

Evidence of exposure to SARS-CoV-2 in cats and dogs from households in Italy.

SARS-CoV-2 emerged from animals and is now easily transmitted between people. Sporadic detection of natural cases in animals alongside successful experimental infections of pets, such as cats, ferrets and dogs, raises questions about the susceptibility of animals under natural conditions of pet ownership. Here, we report a large-scale study to assess SARS-CoV-2 infection in 919 companion animals living in northern Italy, sampled at a time of frequent human infection. No animals tested PCR positive. However, 3.3% of dogs and 5.8% of cats had measurable SARS-CoV-2 neutralizing antibody titers, with dogs from COVID-19 positive households being significantly more likely to test positive than those from COVID-19 negative households. Understanding risk factors associated with this and their potential to infect other species requires urgent investigation.

Bubaline alphaherpesvirus 1 induces a latent/reactivable infection in goats.

Latent infection is a common mechanism used by several alphaherpesviruses to persist in their host but it is not clear whether this mechanism is also triggered in heterologous infections. Cross-species infections have been documented repeatedly for alphaherpesviruses of ruminants, a group of closely related viruses. Herewith we report latent infection with bubaline alphaherpesvirus 1 (BuHV-1) in experimentally infected goats and subsequent virus reactivation after treatment with dexamethasone (DMS) at 10 months after infection. After DMS treatment, the virus was isolated in one such animal in the nasal swabs from day 3 to 9 post treatment and in the ocular swabs at day 6. The goat was euthanized 48 days after DMS treatment and viral DNA was detected by PCR in the trigeminal ganglia and in two cervical ganglia. Additionally, BuHV-1 DNA was detected by PCR in the trigeminal ganglia of the other 3 goats.

Evidence for Circulation of Bovine Viral Diarrhoea Virus Type 2c in Ruminants in Southern Italy.

Recently, bovine viral diarrhoea virus type 2c (BVDV-2c) was responsible for a severe outbreak in cattle in northern Europe. Here, we present the results of an epidemiological survey for pestiviruses in ruminants in southern Italy. Pooled serum samples were obtained from 997 bovine, 800 ovine, 431 caprine and eight bubaline farms, and pestiviral RNA was detected by molecular methods in 44 farms consisting of 16 cattle and one buffalo herds and of 21 sheep and six goat flocks. Twenty-nine and 15 farms were infected by BVDV-1 and BVDV-2 strains, respectively. BVDV-1 strains were recovered mainly from cattle and were heterogeneous, belonging to the subtypes 1b, 1u, 1e, 1g and 1h. In contrast, all BVDV-2 viruses but two were detected in sheep or goats and were characterized as BVDV-2c by sequence analysis of 5'UTR. These strains displayed high genetic identity to BVDV-2c circulating in cattle in northern Europe and were more distantly related to a BVDV-2c isolate recovered from a cattle herd in southern Italy more than 10 years before. The circulation of a BVDV-2c in small ruminants suggests the need for a continuous surveillance for the emergence of pestivirus-induced clinical signs in southern Italian farms.

Circulation of multiple subtypes of bovine viral diarrhoea virus type 1 with no evidence for HoBi-like pestivirus in cattle herds of southern Italy.

Pestiviruses of cattle include bovine viral diarrhoea 1 (BVDV-1) and 2 (BVDV-2) plus an emerging group, named HoBi-like pestivirus. In the present paper, the results of an epidemiological survey for pestiviruses circulating in cattle in southern Italy are presented. Molecular assays carried out on a total of 924 bovine samples detected 74 BVDV strains, including 73 BVDV-1 and 1 BVDV-2 viruses. Phylogenetic analysis carried out on partial 5'UTR and Npro sequences revealed the presence of 6 different subtypes of BVDV-1 and a single BVDV-2c strain. BVDV-1 displayed a high level of genetic heterogeneity, which can have both prophylactic and diagnostic implications. In addition, the detection of BVDV-2c highlights the need for a continuous surveillance for the emergence of new pestivirus strains in cattle farms in southern Italy.

Heterogeneity within the hemagglutinin genes of canine distemper virus (CDV) strains detected in Italy.

Canine distemper virus (CDV) is a highly contagious viral pathogen causing lethal disease in dogs and other mammalians. A high degree of genetic variation is found between recent CDV strains and the old CDV isolates used in the vaccines and such genetic variation is regarded as a possible cause of the increasing number of CDV-related diseases in dogs. The H gene shows the greatest extent of genetic variation that allows for distinction of various lineages, according to a geographical pattern of distribution and irrespective of the species of identification. In the present study, hemagglutinin (H) genes obtained from field strains detected from clinical specimens of Italian dogs were analyzed genetically. Phylogenetic analysis revealed that a homogeneous group of CDV strains is widespread in Italian dogs, all which are included into the European lineage. Unexpectedly, strains 179/04 and 48/05 clustered along with CDVs of the Arctic lineage, the highest identity being to strain GR88 (98.0 and 98.4%aa, respectively). The full-length sequence of a red fox CDV strain, 207/00 was also determined and analyzed. The H protein of the fox CDV strain was unrelated to strains within the major European lineage. These results suggest that at least three different CDV lineages are present in Italy.

HoBi-Like Pestivirus and Its Impact on Cattle Productivity.

The clinical features and economic impact of the infection caused by an emerging group of pestiviruses, namely HoBi-like pestivirus, in a cattle herd of southern Italy are reported. In 2011, the virus was first associated with respiratory disease, causing an abortion storm after 1 year and apparently disappearing for the following 3 years after persistently infected calves were slaughtered. However, in 2014, reproductive failures and acute gastroenteritis were observed in the same herd, leading to a marked decrease of productivity. A HoBi-like strain closely related to that responsible for previous outbreaks was detected in several animals. Application of an intensive eradication programme, based on the detection and slaughtering of HoBi-like pestivirus persistently infected animals, resulted in a marked improvement of the productive performances.

Lights and shades on an historical vaccine canine distemper virus, the Rockborn strain.

Both egg- and cell-adapted canine distemper virus (CDV) vaccines are suspected to retain residual virulence, especially if administered to immuno-suppressed animals, very young pups or to highly susceptible animal species. In the early 1980s, post-vaccine encephalitis was reported in dogs from various parts of Britain after administration of a particular batch of combined CDV Rockborn strain/canine adenovirus type-1 vaccine, although incrimination of the Rockborn strain was subsequently retracted. Notwithstanding, this, and other reports, led to the view that the Rockborn strain is less attenuated and less safe than other CDV vaccines, and the Rockborn strain was officially withdrawn from the markets in the mid 1990s. By sequencing the H gene of the strain Rockborn from the 46th laboratory passage, and a commercial vaccine (Candur(®) SH+P, Hoechst Rousell Vet GmbH), the virus was found to differ from the commonly used vaccine strain, Onderstepoort (93.0% nt and 91.7% aa), and to resemble more closely (99.6% nt and 99.3% aa) a CDV strain detected in China from a Lesser Panda (Ailurus fulgens). An additional four CDV strains matching (>99% nt identity) the Rockborn virus were identified in the sequence databases. Also, Rockborn-like strains were identified in two vaccines currently in the market. These findings indicate that Rockborn-like viruses may be recovered from dogs or other carnivores with distemper, suggesting cases of residual virulence of vaccines, or circulation of vaccine-derived Rockborn-like viruses in the field.

Genetic analysis of feline panleukopenia viruses from cats with gastroenteritis.

Thirty-nine parvovirus strains contained in faecal samples collected in Italy (n=34) and UK (n=5) from cats with feline panleukopenia were characterized at the molecular level. All viruses were proven to be true feline panleukopenia virus (FPLV) strains by a minor groove binder probe assay, which is able to discriminate between FPLV and the closely related canine parvovirus type 2. By using sequence analysis of the VP2 gene, it was found that the FPLV strains detected in Italy and UK were highly related to each other, with a nucleotide identity of 99.1-100 and 99.4-99.8% among Italian and British strains, respectively, whereas the similarities between all the sequences analysed were 98.6-100%. Eighty-eight variable positions were detected in the VP2 gene of the field and reference FPLV strains, most of which were singletons. Synonymous substitutions (n=57) predominated over non-synonymous substitutions (n=31), and the ratio between synonymous and non-synonymous substitutions (dN/dS) was 0.10, thus confirming that evolution of FPLV is driven by random genetic drift rather than by positive selection pressure. Some amino acid mutations in the VP2 protein affected sites that are thought to be responsible for antigenic and biological properties of the virus, but no clear patterns of segregation and genetic markers, were identified, confirming that FPLV is in evolutionary stasis.

Identification of group A porcine rotavirus strains bearing a novel VP4 (P) Genotype in Italian swine herds.

The VP4 gene of a G5 Italian porcine rotavirus strain, 344/04-1, was nontypeable by PCR genotyping. The amino acid sequence of the full-length VP4 protein had low identity (

New approaches for the molecular characterization of canine parvovirus type 2 strains.

Characterization of the canine parvovirus type 2 (CPV-2) is sometimes ambiguous, frequently requiring more than one technique for definitive prediction of the viral type. Taking into account the single-nucleotide polymorphisms encountered in the VP2-protein gene between types 2a and 2b and between type 2b and Glu-426 mutant (type 2c), two different minor groove binder (MGB) probe assays were developed for rapid identification of the CPV-2 variants. A total of 315 samples collected from dogs with diarrhoea were screened for CPV-2 by a real-time polymerase chain reaction (PCR) assay capable of detecting all CPV-2 types. In order to compare the type-specific assays with the traditional techniques [haemagglutination inhibition with monoclonal antibodies, PCR-restriction fragment-length polymorphism (RFLP), sequence analysis] for prediction of CPV-2 antigen specificity, the 203 samples tested CPV-2 positive were analysed using the different methods. The results showed a 100% concordance between the MGB probe assays and the combined conventional methods, with 116 samples characterized as type 2a, 32 as type 2b and 55 as type 2c. Therefore, the MGB probe assays represent a quick, reliable tool for prediction of CPV-2 antigen specificity, with regard to the more time-consuming assays currently used.