Feline Panleukopenia Virus in Dogs from Italy and Egypt.

Canine parvovirus and feline panleukopenia virus (FPV) are variants of Carnivore protoparvovirus 1. We identified and characterized FPV in dogs from Italy and Egypt using genomic sequencing and phylogenetic analyses. Cost-effective sequencing strategies should be used to monitor interspecies spread, evolution dynamics, and potential host jumping of FPV.

Identification of a Novel α-herpesvirus Associated with Ulcerative Stomatitis in Donkeys.

An outbreak of ulcerative stomatitis was observed in a donkey (Equus asinus) dairy herd. Similar lesions were also observed on the dams' udders and, sporadically, in genital areas. The lesions typically resolved in 1-3 weeks. An α-herpesvirus, Varicellovirus, genetically related to equid herpesvirus type 3, was identified.

Isolation and characterization of bovine alphaherpesvirus 2 strain from an outbreak of bovine herpetic mammillitis in a dairy farm.

BACKGROUND: Bovine alphaherpesvirus type 2 (BoHV-2) belongs to family Herpesviridae, subfamily Alphaherpesviridae and can cause two distinct, well-defined conditions: a generalized benign skin infection that somewhat mimics lumpy skin disease (LSD), referred to as Pseudo-Lumpy Skin Disease (PSLD) and a localized ulcerative mammillitis, referred to as Bovine Herpetic Mammillitis (BHM). BHM is a localized form of BoHV-2 infection that causes erosive-ulcerative self-limiting lesions on breast and nipples. BHM is chiefly a disease of lactating dairy cows and has been described sporadically in several countries. In this study we describe an outbreak of bovine herpetic mammillitis caused by BoHV-2 occurred in a dairy farm in Southern Italy. Clinical signs were observed in 26/59 lactating cows with the age ranging between 2 and 6 years. The affected animals were afebrile, showed lesions on the skin of nipples, breast and ventral surface of the abdomen, near the mammary veins and spontaneously recovered within 2 months. RESULTS: BoHV-2 DNA was detected in the crust samples by pan-herpes PCR and real-time quantitative PCR. The virus was isolated on bovine kidney cells and was characterised by deep sequencing technologies. The nucleotide identity to BoHV-2 of the strain ITA/2018/468 retrieved in this study ranged from 98.83 to 100%. Phylogenetic analyses based on three full-length gene (glycoprotein B, thymidine kinase and glycoprotein G) sequences confirmed the close relatedness of the strain ITA/2018/468 to BoHV-2 sequences. CONCLUSIONS: The report represents a significant outbreak of BHM in a dairy farm 50 years after the last description in Italy. However, outbreaks of PLSD have been described in Europe recently, indicating that the virus is present in European territories. Improving the diagnostic algorithms and enacting specific surveillance plans could be useful to understand better the epidemiological and pathogenetic patterns of BoHV-2 infection in livestock animals, and to develop, eventually, effective prophylaxis plans.

Validation of a new immunofluorescence antibody test for the detection of Leishmania infantum infection in cats.

The prevalence data of Leishmania infantum infection in cats are characterized by a large variability mainly attributed to the differences in diagnostic techniques. In the absence of consensus about the method of choice for diagnosing feline leishmaniosis, the performance of a new immunofluorescence antibody test (IFAT) was herein analytically described by the comparison with IFAT commonly used for the diagnosis of canine leishmaniosis (i.e., IFAT-OIE) and a laboratory enzyme-linked immunosorbent assay (ELISA). Sera of cats living in visceral leishmaniosis-endemic (n = 105) and visceral leishmaniosis-non-endemic (n = 50) areas were tested by the above methodologies and real-time PCR (qPCR). The most frequent result was represented by triple negativity to the three tests (IFAT-OIE, ELISA, and qPCR) in 42.9% and 80% cats from endemic and non-endemic areas, respectively. Bayes latent class analysis gave an output probability of 34.1% (posterior standard deviation, psd = 5.4%) of true L. infantum cases (TCL) which represent the true estimated prevalence of infection. The sensitivity of each variable contributing to define the TCL was 24% (psd = 6.3%) for qPCR, 78.8% (psd = 8.7%) for ELISA and 91.8% (psd = 5.2%) for IFAT-OIE. The probability to be a TCL was 94.5% for the sample from an endemic area. The cross-validation of the new IFAT by a logistic model correctly identified as positive 80.7% of subjects defined as TCL and negative 89.9% as not TCL, respectively, by the Bayesian model. The study results estimate a good accuracy of the IFAT in predicting cats exposed to L. infantum. Therefore, this procedure may be beneficial for screening cat populations for a better understanding of the epidemiology of feline leishmaniosis.

Haplosporidium pinnae associated with mass mortality in endangered Pinna nobilis (Linnaeus 1758) fan mussels.

The fan mussel, Pinna nobilis (Linnaeus 1758), is an endemic bivalve of the Mediterranean basin, protected by international legislation as an endangered species. In the early summer of 2018, a mass mortality event (MME) of P. nobilis was recorded in the Gulf of Taranto (Southern Italy, Ionian Sea). Moribund specimens of P. nobilis were collected by scuba divers and processed by bacteriological, parasitological, histopathological and molecular analyses to investigate the causes of this MME. Different developmental stages (i.e., plasmodia, spores and sporocysts) of a presumptive haplosporidian parasite were observed during the histological analysis in the epithelium and in the lumen of the digestive tubules, where mature spores occurred either free or in sporocysts. The spores presented an operculum and an ovoid shape measuring 4.4 µm (±0.232) in length and 3.6 µm (±0.233) in width. BLAST analysis of an 18SrRNA sequence revealed a high nucleotide similarity (99%) with the reference sequence of Haplosporidium pinnae available in GenBank database. Phylogenetic analysis clustered the sequence of the pathogen in a paraphyletic clade with the reference sequence of H. pinnae, excluding other haplosporidians (i.e., Bonamia and Minchinia genera). Based on data reported, H. pinnae was the causative agent of MME in the populations of P. nobilis sampled in the Ionian Sea, where the conservation of this endangered species is heavily threatened by such a protozoan infection. Further investigations should contribute to knowledge about the life cycle of H. pinnae in order to reduce spread of the pathogen and to mitigate the burden of the disease where P. nobilis is facing the risk of extinction.

Novel Orthopoxvirus and Lethal Disease in Cat, Italy.

We report detection and full-genome characterization of a novel orthopoxvirus (OPXV) responsible for a fatal infection in a cat. The virus induced skin lesions histologically characterized by leukocyte infiltration and eosinophilic cytoplasmic inclusions. Different PCR approaches were unable to assign the virus to a defined OPXV species. Large amounts of typical brick-shaped virions, morphologically related to OPXV, were observed by electron microscopy. This OPXV strain (Italy_09/17) was isolated on cell cultures and embryonated eggs. Phylogenetic analysis of 9 concatenated genes showed that this virus was distantly related to cowpox virus, more closely related to to ectromelia virus, and belonged to the same cluster of an OPXV recently isolated from captive macaques in Italy. Extensive epidemiologic surveillance in cats and rodents will assess whether cats are incidental hosts and rodents are the main reservoir of the virus. The zoonotic potential of this novel virus also deserves further investigation.

Novel Parvovirus Related to Primate Bufaviruses in Dogs.

A novel protoparvovirus species, related genetically to human bufaviruses, was identified in dogs with respiratory signs. The canine bufavirus was distantly related to the well-known canine protoparvovirus, canine parvovirus type 2, sharing low amino acid identities in the nonstructural protein 1 (40.6%) and in the capsid protein 1 (33.4%). By screening collections of fecal, nasal, and oropharyngeal samples obtained from juvenile dogs (<1 year of age), canine bufavirus DNA appeared as a common component of canine virome. The virus was common in the stool samples of dogs with or without enteric disease and in the nasal and oropharyngeal swab samples of dogs with respiratory signs. However, the virus was not detected in nasal and oropharyngeal swab samples from animals without clinical signs.

General and type 2 parvovirus-specific passive immune transfer in puppies - Evaluation by early growth.

Failure of passive immune transfer put puppies at a higher risk of neonatal and weaning mortality due to low immune protection against infectious agents. The aim of this study was to investigate the evolution of the general via serum IgG concentration (IgG) and the specific via serum maternally derived canine parvovirus type 2-specific antibody titer (CPV2 MDA) passive immune transfer within the first 4 weeks of age. Furthermore, the relationship between general and specific immune transfer and the possibility of non-invasive evaluation was assessed. Puppies (169) were weighed systematically between birth and Day 28. IgG and CPV2 MDA were assayed in serum samples at 2 and at 28 days of age. At Day 2, there was a positive correlation between IgG and CPV2 MDA (ρ = 0.71; p < 0.001). At Day 2, 17.9% (27/151) of puppies presented a deficit of passive immune transfer according to IgG result (defined as IgG < 2.3 g/L) and 25.8% (39/151) of puppies were under the minimal protective CPV2 MDA titer (defined as <1:160). No correlation between IgG and CPV2 MDA was observed at Day 28 (ρ = 0.14; p = 0.11). Growth rate within the first 48 hours <-2.7% allowed to distinguish puppies at high risk of the general and specific passive immune failure (Youden's index = 0.79 and 0.75, respectively). The threshold value of early growth rate, although applicable only in puppies non-supplemented with milk replacer, allows identifying via non-invasive way individuals requiring a special care. Further investigation of the mechanism of passive immune transfer in dogs is necessary to understand the relationship between the general and specific immunoglobulin levels.

Rhipicephalus turanicus, a new vector of Hepatozoon canis.

The distribution of Hepatozoon canis mainly encompasses areas where its main tick vector, Rhipicephalus sanguineus sensu lato, is present. However, the detection of this pathogen in dogs, foxes and golden jackals well outside the areas inhabited by this tick species reinforced the hypothesis that additional ixodids are involved in the life cycle and transmission of this protozoon. The present study provides, for the first time, data supporting the sporogonic development of H. canis in specimens of Rhipicephalus turanicus collected from a naturally infected fox from southern Italy. The epidemiological role of R. turanicus as a vector of H. canis is discussed, along with information on the potential use of cell cultures for the experimental infection with H. canis sporozoites. The in vitro infection of canine leucocytes by sporozoites from ticks is proposed as a potential tool for future in-depth studies on the biology of H. canis.

Pestivirus infection in cattle dairy farms: E2 glycoprotein ELISA reveals the presence of bovine viral diarrhea virus type 2 in northwestern Italy.

BACKGROUND: Bovine viral diarrhea virus (BVDV) types 1 and 2 are members of the Pestivirus genus of the Flaviviridae family. This genus also includes the HoBi-like virus, tentatively classified as BVDV type 3. BVDV-1 is widely distributed in Italy despite the extensive use of BVDV-1-based vaccines, while BVDV-2 and HoBi-like Pestivirus have been detected occasionally. Monitoring the occurrence of sporadic or atypical pestiviruses is a useful approach to evaluate the need for additional vaccine strains that can be used in BVDV control programs. RESULTS: In this study we developed a multiwell antibody ELISA based on the recombinant E2 protein of the three bovine pestiviruses. We evaluated the assay's applicability for surveillance purposes using pooled milk samples, each prepared from a maximum of 35 lactating cows and collected from 176 dairy herds. As expected, the majority of the pooled samples reacted to a greater extent against the BVDV-1 E2 antigen. All three milk pools from a single farm reacted to the BVDV-2 antigen, however. Further analysis using spot tests, antigen detection, and sequence analysis of the 5'-UTR region confirmed the presence of five persistently infected calves carrying a BVDV-2a strain. CONCLUSIONS: This study highlights for the first time that sporadic circulation of BVDV-2 can be predicted by immunoenzymatic methods in the absence of specific vaccination.

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.

Identification of a Bovine Enteric Calicivirus, Kirklareli Virus, Distantly Related to Neboviruses, in Calves with Enteritis in Turkey.

A calicivirus was detected in neonatal calves with enteritis in Kirklareli, Thrace, Turkey. In the full-length genome, Kirklareli virus was related (48% nucleotide identity) to bovine enteric caliciviruses (Nebovirus genus). The virus was also detected in a herd in Ankara, Central Anatolia, but not in other Turkish prefectures.

Seroprevalence of norovirus genogroup IV antibodies among humans, Italy, 2010-2011.

Noroviruses (NoVs) of genogroup IV (GIV) (Alphatron-like) cause infections in humans and in carnivorous animals such as dogs and cats. We screened an age-stratified collection of serum samples from 535 humans in Italy, using virus-like particles of genotypes GIV.1, circulating in humans, and GIV.2, identified in animals, in ELISA, in order to investigate the prevalence of GIV NoV-specific IgG antibodies. Antibodies specific for both genotypes were detected, ranging from a prevalence of 6.6% to 44.8% for GIV.1 and from 6.8% to 15.1% for GIV.2 among different age groups. These data are consistent with a higher prevalence of GIV.1 strains in the human population. Analysis of antibodies against GIV.2 suggests zoonotic transmission of animal NoVs, likely attributable to interaction between humans and domestic pets. This finding, and recent documentation of human transmission of NoVs to dogs, indicate the possibility of an evolutionary relationship between human and animal NoVs.

Genome sequencing identifies genetic and antigenic divergence of porcine picobirnaviruses.

The full-length genome sequence of a porcine picobirnavirus (PBV) detected in Italy in 2004 was determined. The smaller (S) genome segment was 1730 nt, coding for a putative RNA-dependent RNA polymerase. Two distinct subpopulations of larger (L) genome segment (LA and LB) were identified in the sample, with the sizes ranging from 2351 to 2666 nt. The ORF1, coding for a protein of unknown function, contained a variable number of repetitions of the ExxRxNxxxE motif. The capsid protein-coding ORF2 spanned nt 810-2447 in the LB variants and started at nt 734 in the LA variants. However, a termination codon was present only in one of all the LA segment variants. Three-dimensional modelling of the porcine PBV capsids suggested structural differences in the protruding domain, tentatively involved as antigens in the humoral immune response. Altogether, these findings suggest the simultaneous presence of two different PBV strains sharing the same S segment but displaying genetically diverse L segments. In addition, the sample probably contained a mixture of PBVs with aberrant RNA replication products. Altered structure in the L segments could be tolerated and retained in the presence of functionally integer-cognate genes and represents a mechanism of virus diversification.

Mucosal disease-like syndrome in a calf persistently infected by Hobi-like pestivirus.

A calf persistently infected with Hobi-like pestivirus displayed severe clinical signs and subsequently died. Gross lesions and histopathological changes were suggestive of hemorrhagic and necrotic inflammation involving several tissues. A Hobi-like pestivirus pair was isolated from the dead calf, i.e., cytopathogenic (CP) and noncytopathogenic (NCP) strains strictly related to each other and to Italian prototype isolates at the genetic level. Two biotype-specific real-time reverse transcription-PCR assays determined the time of the emergence of the CP virus as 1 month before the calf's death. This highest RNA titers were reached in lymphoid and nervous system tissues, whereas only traces of CP viral RNA were found in blood. In contrast, great NCP virus loads were present in all tissues and biological fluids. The present report provides new insights into the pathogenesis and molecular mechanisms of this emerging group of pestiviruses.

Mechanism of surfactant interactions with feline coronavirus: A physical chemistry perspective.

HYPOTHESIS: Surfactants are inexpensive chemicals with promising applications in virus inactivation, particularly for enveloped viruses. Yet, the detailed mechanisms by which surfactants deactivate coronaviruses remain underexplored. This study delves into the virucidal mechanisms of various surfactants on Feline Coronavirus (FCoV) and their potential applications against more pathogenic coronaviruses. EXPERIMENTS: By integrating virucidal activity assays with fluorescence spectroscopy, dynamic light scattering and laser Doppler electrophoresis, alongside liposome permeability experiments, we have analyzed the effects of non-ionic and ionic surfactants on viral activity. FINDINGS: The non-ionic surfactant octaethylene glycol monodecyl ether (C10EO8) inactivates the virus by disrupting the lipid envelope, whereas ionic surfactants like Sodium Dodecyl Sulfate and Cetylpyridinium Chloride predominantly affect the spike proteins, with their impact on the viral membrane being hampered by kinetic and thermodynamic constraints. FCoV served as a safe model for studying virucidal activity, offering a faster alternative to traditional virucidal assays. The study demonstrates that physicochemical techniques can expedite the screening of virucidal compounds, contributing to the design of effective disinfectant formulations. Our results not only highlight the critical role of surfactant-virus interactions but also contribute to strategic advancements in public health measures for future pandemic containment and the ongoing challenge of antimicrobial resistance.

Genetic characterization of parvoviruses identified in stray cats in Nigeria.

Parvoviruses are a major cause of haemorrhagic gastroenteritis, leukopenia and high mortality in cats and dogs. In this study, the presence and genetic characteristics of parvoviruses circulating among cats in Nigeria are reported. Faecal samples of stray cats from live animal markets in southwestern (Oyo and Osun States) and north-central (Kwara State) Nigeria were screened for the presence of parvoviral DNA using a qPCR. Positive samples were further characterized using a qPCR based on minor groove binder probes. Overall, 85/102 (83.3 %) stray cats tested positive for feline panleukopenia virus (FPV) DNA and one cat was co-infected with canine parvovirus-2 type a. Sequence analysis of the complete capsid region of 15 Nigerian FPV strains revealed that they were up to 99.9 % similar to the American reference strain FPV-b at the nucleotide level, and three of them presented amino acid mutations in key capsid residues. This is the first report of identification and molecular characterization of FPV strains in cats in Nigeria. The high prevalence of the virus emphasizes the need for constant surveillance of the circulation of parvoviruses in Nigeria and underscores the need to deploy an effective vaccination strategy.

Persistent infection caused by Hobi-like pestivirus.

A calf persistently infected by Hobi-like pestivirus was monitored for about 6 months, displaying clinical signs typical of bovine viral diarrhea virus persistent infection and shedding the virus through all body secretions, with maximal titers detected in urine. This report provides new insights into the pathogenesis of the emerging pestivirus.

Clinical bovine piroplasmosis caused by Babesia occultans in Italy.

A clinical outbreak of bovine piroplasmosis was reported in Italy. The etiological agent was characterized as Babesia occultans, a parasite regarded as apathogenic and never detected before in continental Europe. This report paves the way for further studies to assess the occurrence of this tick-transmitted protozoan in other European regions.

European surveillance for pantropic canine coronavirus.

Highly virulent pantropic canine coronavirus (CCoV) strains belonging to subtype IIa were recently identified in dogs. To assess the distribution of such strains in Europe, tissue samples were collected from 354 dogs that had died after displaying systemic disease in France (n = 92), Hungary (n = 75), Italy (n = 69), Greece (n = 87), The Netherlands (n = 27), Belgium (n = 4), and Bulgaria (n = 1). A total of 124 animals tested positive for CCoV, with 33 of them displaying the virus in extraintestinal tissues. Twenty-four CCoV strains (19.35% of the CCoV-positive dogs) detected in internal organs were characterized as subtype IIa and consequently assumed to be pantropic CCoVs. Sequence and phylogenetic analyses of the 5' end of the spike protein gene showed that pantropic CCoV strains are closely related to each other, with the exception of two divergent French viruses that clustered with enteric strains.