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Preclinical testing of novel therapeutics for chronic hepatitis B (CHB) requires suitable animal models. Equids host homologs of hepatitis C virus (HCV). Because coinfections of hepatitis B virus (HBV) and HCV occur in humans, we screened 2,917 specimens from equids from five continents for HBV. We discovered a distinct HBV species (Equid HBV, EqHBV) in 3.2% of donkeys and zebras by PCR and antibodies against EqHBV in 5.4% of donkeys and zebras. Molecular, histopathological, and biochemical analyses revealed that infection patterns of EqHBV resembled those of HBV in humans, including hepatotropism, moderate liver damage, evolutionary stasis, and potential horizontal virus transmission. Naturally infected donkeys showed chronic infections resembling CHB with high viral loads of up to 2.6 × 10(9) mean copies per milliliter serum for >6 mo and weak antibody responses. Antibodies against Equid HCV were codetected in 26.5% of donkeys seropositive for EqHBV, corroborating susceptibility to both hepatitis viruses. Deltavirus pseudotypes carrying EqHBV surface proteins were unable to infect human cells via the HBV receptor NTCP (Na(+)/taurocholate cotransporting polypeptide), suggesting alternative viral entry mechanisms. Both HBV and EqHBV deltavirus pseudotypes infected primary horse hepatocytes in vitro, supporting a broad host range for EqHBV among equids and suggesting that horses might be suitable for EqHBV and HBV infections in vivo. Evolutionary analyses suggested that EqHBV originated in Africa several thousand years ago, commensurate with the domestication of donkeys. In sum, EqHBV naturally infects diverse equids and mimics HBV infection patterns. Equids provide a unique opportunity for preclinical testing of novel therapeutics for CHB and to investigate HBV/HCV interplay upon coinfection.

Pseudorabies virus (PRV) or suid herpesvirus 1 (SHV-1) is the causative agent of Aujeszky's disease, a highly contagious viral infection which causes neurological fatal illness in mammals other than suids. Here we report a case of a young wolf (Canis lupus) of around 2 years found dead by a hunter in the province of Avellino, Campania Region. Necropsy showed pathological findings consistent with encephalitis and gastroenteritis. Organs were analysed by microbiological and molecular investigations following standard procedures to ascertain the possible cause of death. Real-time PCR revealed the presence of PRV in the brain and of canine parvovirus 2b in organs like intestine, liver, brain, kidney and pancreas. Death probably occurred very shortly after SHV-1 infection in an animal already weakened by parvovirosis.

Toxoplasma gondii is a widespread protozoan parasite (phylum Apicomplexa), which causes a zoonotic parasitic disease, known as toxoplasmosis. The aim of this study was to evaluate the occurrence and genotypes of T. gondii in wild boars of southern Italy and thus to assess the risk of infection for consumers. The boars were inspected during the hunting season within the regional project ‘Wild Boar Emergency Plan in Campania’, and molecular analyses were performed on 338 boars analysing a total number of 884 matrices (263 brains, 310 hearts and 311 masseter muscles). Toxoplasma gondii was detected in 134 out of 338 boars (39.6%). No significant statistical difference between genders was found (χ2 = 0.15 p = .70). The prevalence was 47.1%, 39.3% and 39.2% in piglets, yearlings and adults, respectively (χ2 = 0.41; p = .81). The highest prevalence of T. gondii was found in masseter muscles (74/311, 23.8%), followed by the heart (70/310, 22.6%) and brain (58/263, 22.0%), respectively. Microsatellite (MS) analysis of 11 samples revealed eleven T. gondii genotypes (nine atypical, one belonging to type II one to type III). Most of the genotypes found were thus atypical and may be virulent in humans. Hierarchical clustering analysis showed the presence of three distinct clusters, with the majority of atypical genotypes in the GII-GIII cluster. The high prevalence of infection in masseters highlights the potential risk for public health, considering that this muscle is commonly used to prepare raw meat products (‘guanciale’ and sausages), which may be a source of T. gondii infection in humans. Wild boars may act as an interface role between wildlife, livestock and humans. Our data highlight the urgent need to minimize the risk of infection for animals and humans by setting up a surveillance programme and preventive strategies in a One Health approach to wildlife species.

Taenia hydatigena cysticercosis is a widespread parasitic disease of wild and domestic animals. In Europe, the increase in wild boar population may potentially contribute to the spread of this parasitic infection. To determine the occurrence of cysticerci (metacestodes) in wild boar population from southern Italy, carcasses were inspected during three hunting seasons (2016–2018). Out of 3363 wild boar examined, 229 (6.8%) harboured cysticerci with 188 (82.1%) infected by a single cyst, vs 41 (17.9%) boars having more than one. Most of the positive animals (187; 81.7%) showed cysts on the liver, whereas a multiple localization of cysticerci was reported in 10 (4.4%) wild boar. The total number of cysts retrieved from positive animals was 301 (average 1.3). Molecular analysis revealed the occurrence of a common haplotype (Hap 8) shared between wild boar and domestic animals. Our findings suggest the presence of a T. hydatigena semi-domestic life cycle in which wild boar may play an important role, due to a large number of offal available to hunting dogs, wolves and foxes during hunting seasons. Hunters may be players in the management of wildlife species to control and prevent the circulation of parasitic diseases.

Acantocheilonema reconditum is a parasite transmitted by fleas, lice and ticks and is included among the filaroid species infecting dogs. Contrary to the more well-studied Dirofilaria immitis and Dirofilaria repens, A. reconditum is believed to be less pathogenic, as adult worms are localized in the subcutaneous tissues and in the perirenal fat. Although previous studies reported the absence of clinical symptoms in infected dogs, there are few data regarding the haematological and biochemical changes that could be potentially caused by this parasite. Because hunting dogs are frequently exposed to vector-borne pathogens, the aim of the present study was to assess the prevalence, risk factors and potential hematobiochemical abnormalities associated with A. reconditum infection in this specific canine population. Blood samples were collected from 3,020 hunting dogs living in Campania region, and were tested by a modified Knott technique to count and identify microfilariae. Out of 3,020 dogs tested, 84 were positive to A. reconditum, with an overall prevalence of 2.78% (95% CI: 2.19– 3.37%). The number of microfilariae/ml ranged from 1 to 442. After excluding dogs co-infected by different filarial worm species and/or other vector-borne pathogens common in Southern Italy, n. 74 dogs showed A. reconditum single infection. The main clinical features observed were dehydration (n. 1), fever (n. 1), congested mucous membranes (n. 2) and exercise intolerance (n. 2). Complete blood cell count results revealed leukocytosis (n. 16), anaemia (n. 2), thrombocytopenia (n. 8), eosinophilia (n. 1). Biochemical data showed increased serum values of total globulins (n. 14), albumins (n. 9), gamma glutamyl transferase (n. 2) and alkaline phosphate (n. 1), hypoalbuminemia was observed in one dog. Dogs co-infected with other vector-borne pathogens were not included in the statistical analysis. The living area (p

Problems with parasitic infections are common in zoological gardens and circuses. In some animals it can lead to several disorders such as systemic disease, reproductive disorders (abortions and neonatal mortality), and even to death if severe illness is untreated. Thus, the aim of this study was to evaluate the prevalence of three common parasites in 74 animals from three zoos, and four circuses in Southern Italy. Antibodies to Toxoplasma gondii, Neospora caninum, and Encephalitozoon cuniculi were detected in 51%, 12%, and 20% of animals, respectively. Co‐infections of T. gondii and N. caninum were reported in seven animals (9%) and co‐infection of T. gondii and E. cuniculi in one animal. T. gondii, N. caninum and E. cuniculi seroprevalence differed in type of diet (P ≤ 0.0001; P ≤ 0.037 and P ≤ 0.004, respectively). T. gondii and E. cuniculi seroprevalence also differed in animal families (P ≤ 0.0001) and according to type of housing (P ≤ 0.003), respectively. Statistical differences were not found in other characteristics (gender, age, country of birth, origin, and contact with cats or dogs). This is the first serological study focusing on protozoan and microsporidian parasites in zoo and circus animals from Southern Italy and the first detection of antibodies to E. cuniculi in camels in Europe.

Babesia caballi and Theileria equi are tick-borne pathogens causing equine piroplasmosis infecting the Equidae family in which they cause significant sanitary and economic losses. Furthermore, equine piroplasmosis is included in the World Animal Health Organization (OIE) notifiable diseases list with possible movement restrictions for positive horses. Thirty-nine EDTA and whole-blood samples collected during 2013 and 2014 from symptomatic and asymptomatic horses of Central-Southern Italy were included in the present study either because of their strongly positive results in Real Time (RT) PCRs targeting the 18S rRNA gene specific for each piroplasm and/or due to their serological ELISA/18S rRNA RT-PCR discordant results. A nested PCR amplifying the hypervariable V4 region of the 18S rRNA gene of both piroplasms was performed on all samples. T. equi positive samples were also analysed with a PCR targeting the equi merozoite antigen 1-gene (EMA-1). The sequences obtained were thirty for T. equi, 25 of which were for the hypervariable V4 region of the 18S rRNA gene and 13 for the EMA-1 gene, with eight samples positive for both targets, while only six 18S rRNA gene sequences were retrieved for B. caballi. The phylogenetic analysis results are as follows: T. equi sequences of the 18S rRNA gene clustered in three different phylogenetic groups, respectively in the A (15), B (9) and C (1) while those of B. caballi in the A (1), B1 (3) and B2 (2) groups. T. equi sequences for EMA-1 gene clustered in A (11) and in B (2). This analysis confirms that both T. equi and B. caballi sequences present a genetic heterogeneity independently of their geographical location, similar to that reported by other authors. Statistical associations were evaluated between phylogenetic groups of T. equi 18S rRNA gene and each of the following variables, using Fisher's exact test: clinical signs, serological ELISA/18S rRNA RT-PCR discordant results and T. equi EMA-1 negativity. The different groups were found to be statistically related to the presence of signs (less present in group B samples), to ELISA negativity/18S rRNA RT-PCR positivity (more seronegative samples in group B). No statistical analysis was performed for the B. caballi as the number of sequences available was insufficient and for the EMA −1 sequences which almost all grouped in the same cluster. The results here obtained provide additional information about T. equi and B. caballi sequences, which could also be used to verify the performance of serological and molecular diagnostic methods.

Canine vector-borne diseases (CVBDs) result from a range of pathogens transmitted to dogs by arthropods. The present study aimed to examine a population of hunting dogs from southern Italy using both serological and molecular diagnostic methods for evidence of infection by known pathogens of CVBDs. Blood samples were collected from hunting dogs (n. 1,191) in the Avellino and Salerno provinces of Campania region of southern Italy. Serological testing for Ehrlichia canis, Anaplasma spp., and Borrelia burgdorferi sensu lato-specific antibodies and Dirofilaria immitis-specific antigen was performed using a commercial in-clinic enzyme-linked immunosorbent assay kit, SNAP® 4Dx® Plus. Molecular diagnostic testing for the blood-borne presence of Ehrlichia canis, Ehrlichia chaffeensis, Anaplasma platys, Anaplasma phagocytophilum, Babesia canis, Babesia vogeli, Babesia gibsoni, Dirofilaria immitis, Dirofilaria repens, Hepatozoon canis, and Leishmania spp. was performed by real time polymerase chain reaction (RT-PCR) assays. Seropositive rates for the four individual pathogens were: E.canis 7.7%; Anaplasma spp. 4.2%; D.immitis,1.4%; and B.burgdorferi s.l. 0.3%. PCR positive rates for eight detected pathogens were: H.canis 16.8%; A.platys 2.4%; E.canis 1.9%; B.vogeli 1.6%; Leishmania spp. 0.4%; B.canis 0.2%; D.repens 0.2%; and D.immitis 0.1%. Co-infections were detected at a rate of 2.3% by serology and 2.7% by RT-PCR with Anaplasma/E.canis and E.canis/H.canis co-infections occurring most frequently by serology and PCR, respectively. The results of present survey indicate that hunting dog populations are at risk of CVBDs in southern Italy. Further studies are needed to evaluate the role of hunting dogs in the epidemiology of vector-borne agents and the relationship to sympatric populations of wild animals.

A national survey to obtain information of helminth control practices on horses and farm management was carried out in 2014 in Italy. Data were collected using 387 face-to-face questionnaire with questions regarding type and size of farms, use of horses, grazing management, parasite control protocol, interval worming, time of treatments, number of treatments per year, anthelmintic class used, specific treatments (tapeworms and encysted cyathostomins). In tolal, data of 7,616 horses were obtained In Italy, most control protocols were strategic. Sixty eight percent of respondents administered anthelmintics at particular times of the year. With respect to worming intervals, most of them (45%) treated animals at 6 months intervals and 4% wormed horses bi-monthly. Six percent of respondents declared that they left animals untreated. With regard to the time of treatment administration, most reported that dosing was carried out independently from the season, although someone emphasized a greater attention to dosing during autumn and spring months. The mean number of treatments per year was 2.6 (range 1-12). Over half (64%) of respondents indicated that ivermectin and moxidectin or their associations (e.g plus praziquantel) were the most commonly used anthelmintics in the previous 12 months. Treatments against cyathostomin encysted larvae and tapeworms were administered by 11% and 21% of respondents, respectively. Most owners obtained information on parasite control from veterinarians (93%), in addition to specialized journals and internet forums in decreasing order of importance. This is the first national survey investigating parasite control practices and anthelmintic use in horses

In Italy despite the increasing use of donkeys in leisure activities, onotherapy and milk production, there are no epidemiological data regarding the parasites affecting this species. A survey was conducted to determine the prevalence of the principal endoparasitic infections in 50 donkey farms. Between March 2012-2015, faecal samples were collected from 961 donkeys and examined individually using a modified McMaster technique. In addition a centrifugation/flotation technique (Proudman test) and a sedimentation technique were used for the diagnosis of Anoplocephalidae and Fasciola hepatica eggs, respectively. On each farm pooled coprocoltures were made. Strongyles were the most common parasitic species found, with a prevalence of 88.5%. 11.4% of donkeys were not infected and infection intensity was lower than the donkey cut-off selective therapy of 300 eggs per gram (epg) in 33.4% of animals. The other epg values were 300–500 in 14.3%, 501–1,000 in 26.0%, 1,001-2,000 in 12.8% and > 2,000 in 2.1% of donkeys. In all tested farms, coprocoltures revealed the presence of Cyathostomes (100%) followed by Strongylus vulgaris in 13 farms (26%), Poteriostomum spp. (24%), Triodontophorus spp. (6%) and S. edentatus (4%). Other parasites found in studied donkeys were: Oxyuris equi (7.8%), Dictyocaulus arnfieldi (4.4%), Parascaris equorum (2.8%), Anoplocephala perfoliata (1.1%) Eimeria leuckarti (0.6%) and Strongyloides westeri (0.1%). No positivity was detected for F. hepatica. This study represents the first large-scale survey in Italy and added data to the parasitological scenario of donkey population.

Moxidectin (MOX) has a wide range of ecto-endoparasitic activity in many species including horses. There is a paucity of data on the efficacy of anthelmintics used in donkeys. The aims of the present study were to evaluate the field efficacy and Egg Reappearance Period (ERP) of MOX up to 84 days at horse dose against natural infection of Cyathostominae in donkeys. The trial was conducted in northern Italy, twenty-nine donkeys, were selected on the basis of faecal egg counts (FEC) > 300 EPG. Animals were treated on Day 0 with an oral gel containing MOX, at the manufacturer’s recommended horse dose (0.4mg/kg BW). FECs were performed on each donkey at least every fortnight during 84 days after treatment using a modified McMaster technique with a sensitivity 10 EPG. The efficacy and ERP values were determined following the AAEP Parasite Control Guidelines. The percentage reductions in FEC were 99.7% at the day 14, 99.5% at the day 28, 97.7% at the day 56 and 73.3% on day 84 post treatment. This trial demonstrates that MOX at horse dose was effective and safe for the treatment of Cyathostominae in donkeys, however with a shortened ERP this could be the first indication of developing anthelmintic resistance. Considering that the donkeys have a greater capacity to metabolize certain drugs compared with horses, a higher dosage or shorter intervals could be required for maintaining effective drug concentrations. The shorter ERP observed in the studied donkeys could be associated with a sub-therapeutic level of MOX.

Therapeutics are administered to donkeys based on dosage and intervals recommended for horses because few drugs have donkey-specific indications. Although, mebendazole (MBZ) has been licensed for use in horses there are no data available regarding its pharmacokinetic disposition and effectiveness in donkeys. The plasma disposition, milk excretion and efficacy of a paste formulation of MBZ in donkeys were examined in a field trial. Two groups of six donkeys received MBZ paste at horse dosage 10 mg/kg body weight (MBZ 1) and at double horse dosage 20 mg/kg body weight (MBZ 2) in animals naturally infected by Cyathostominae. Heparinised blood and milk samples were collected at times between 1 and 120 h post treatment. The samples were analysed by high performance liquid chromatography. Individual FECs (Faecal Egg Count) were performed on each animal before the treatment (day -3) and weekly from day 7 until day 56 post treatment using a modified McMaster technique with a sensitivity of 10 EPG. The results of FECRT for both MBZ dosages were efficient (>95% efficacy) until day 28. The plasma concentrations and systemic exposure of MBZ in donkeys were relatively lower compared with the other benzimidazoles. Dose-dependent plasma dispositions were observed at the increased dose (10 vs 20 mg/kg). This study indicated that oral administration of MBZ has a minimal disposition rates into the milk (ratio of AUC milk/plasma was 0.18±0.02) and may be used safely in lactating donkeys with zero milk-withdrawal period. Therefore, horse dose of MBZ could be used for parasite control in donkeys.