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Simple Summary Specimen collection and preservation can be challenging, especially when suboptimal conditions occur. A relevant amount of time could be required from sampling to the tissue analysis, and therefore a high-quality conservation technique is vitally important for diagnostic purposes. The aim of this study was the identification of a reliable and economic method for tissue preservation to be used in complex “in-field” situations, suitable for both histological and molecular analysis. Punch biopsies were collected from six cattle livers. Comparison among methods of preservation using RNAlater, silica beads, or under-vacuum was carried out using different times and temperatures. Three days were assumed as a maximum time interval from sampling to laboratory and 4 °C and 24 °C chosen as references for refrigeration temperature and room temperature, respectively. Histological and biomolecular analyses were performed. RNAlater and silica beads poorly preserved the histological parameters evaluated; conversely, vacuum-sealed samples showed a good grade of preservation for 48 h. DNA quality was acceptable for each sample. RNA integrity showed promising results for samples preserved with silica beads. Abstract A high quality of samples is crucial for the success of the analysis and diagnostic purposes, and therefore the right method of conservation is vitally important for an optimal preservation of tissues. Indeed, the time to deliver the sample to the laboratory could be remarkably long, especially under suboptimal conditions, and the use of specific fixatives or cold storage may not be possible. Moreover, the portability and cost of storage equipment, their toxicity, and their ease of use play a central role when choosing the correct preservation method. The aim of this study was the identification of a reliable and economic method for tissue preservation, to be used in “in-field” sampling, suitable for both histological and molecular analysis. Punch biopsies were collected from six cattle livers. Comparisons among methods of preservation using RNAlater, silica beads, and under-vacuum was carried out. These methods were tested through considering different times and temperatures, assuming three days as a maximum time interval from sampling to laboratory and choosing 4 °C and 24 °C as references for refrigeration temperature and room temperature, respectively. Histologically, the integrity of nucleus, cytoplasm, preservation of liver structure, and easiness of recognition of inflammatory infiltrate were evaluated. The integrity of the extracted DNA and RNA was evaluated through PCR and by means of an automated electrophoresis station, respectively. RNAlater and silica beads poorly preserved the histological parameters evaluated, independently from the temperature. Conversely, the vacuum-sealed samples showed a good grade of preservation until 48 h. DNA quality was acceptable for each sample. RNA integrity showed promising results only for samples preserved with silica beads.

Background Radiofrequency Ablation (RFA) is rarely performed in veterinary medicine. A rationale exists for its use in selected cases of canine liver tumours. RFA induces ablation zones of variable size and geometry depending on the technique used and on the impedance of the targeted organ. Objectives (a) to describe the geometry and reproducibility of the ablation zones produced by three commercially available systems from a single company, using isolated swine liver parenchyma as a model for future veterinary applications in vivo; (b) to study the effects of local saline perfusion into the ablated parenchyma through the electrode tip and of single versus double passage of the electrode on size, geometry and reproducibility of the ablation zones produced. Methods Size, and geometry of ablation zones reproduced in six livers with one cooled and perfused (saline) and two cooled and non‐perfused systems, after single or double passage (n = 6/condition), were assessed macroscopically on digitalized images by a blinded operator. Longitudinal and transverse diameters, equivalent diameter, estimated volume and roundness index were measured. Reproducibility was assessed as coefficient of variation. Results and Conclusions Ablation zone reproducibility was higher when expressed in terms of ablation zone diameters than estimated volume. Local saline perfusion of the parenchyma through the electrode tip during RFA increased the ablation zone longitudinal diameter. Ablation zone estimated volume increased with saline perfusion only when double passage was performed. These data may provide useful information for those clinicians who intend to include RFA as an additive tool in veterinary interventional radiology., Percutaneous Radiofrequency Ablation (RFA) is a widely used technique in human interventional radiology, while its use in veterinary medicine is restricted to isolated cases. With this ex vivo study, we attempted to assess the potential of RFA as an applicable technique in hepatic oncology in dogs with at‐risk tumour presentation.

BackgroundThis report describes a case of primary subcutaneous aspergillosis in a 7-year-old neutered male dromedary camel (Camelus dromedarius).Case presentationThe animal developed a large nodular lesion in the right scrotum two years after surgical intervention for neutering. The mass had a firm consistency and was painful at palpation. Histopathology revealed dermal granulomatous inflammation with a necrotic centre, surrounded by plasma cells, macrophages, neutrophils, and sparse fungal hyphae characterised by parallel cell walls, distinct septa, and dichotomous branching. Fungal culture was not performed, but a panel of mono- and polyclonal antibodies specific for different fungal genera identified the hyphae as Aspergillus sp.ConclusionsThe occurrence of subcutaneous lesions is a rare manifestation of aspergillosis in animals, and this appears to be the first case reported in the dromedary camel.

A necropsy was conducted on a female grey-headed parrot (Poicephalus robustus suahelicus) that died following signs of depression, ruffled feathers, and inappetence. Microscopic examination revealed the presence of hemoprotozoa in the liver. A nested polymerase chain reaction (PCR), targeting the mitochondrial cytochrome b gene of Haemoproteus species, Plasmodium species, and Leucocytozoon species, was performed on frozen tissue samples collected at necropsy. The hemoprotozoa were identified by PCR analysis as Leucocytozoon species. Hemoprotozoa are rarely reported in African parrots, and this is the first report of a Leucocytozooon species infection in a Poicephalus robustus suahelicus.

The goals of this study were to estimate the prevalence of feather picking (FP) in Italian pet parrots and evaluate the risk factors and possible ethological correlation. A web survey was created and addressed to owners of all species of companion parrots, distributed through on line parrots association websites, social networks and by mail. The survey was available for compilation from June to October 2014. In 292 compiled surveys, 20 different parrots genera were indicated; Psittacus spp. (24.3%), Agapornis spp. (19.5%), Nymphicus hollandicus (18.0%) and Amazona spp. (9.0%) were the most represented species. Our study showed a FP prevalence of 17.5%, with the highest prevalence in Agapornis spp. (26.3%) and Psittacus spp. (22.5%). Living with other parrots was positively associated to FP (p

Glucocorticoids (GCs) are illegally used as growth promoters in cattle, and the analytical methods officially applied most likely underestimate the precise frequency of the abuse. As a side effect, the administration of GCs causes fat infiltration, apoptosis, and atrophy of the thymus. However, gross and histological observations carried out previously showed that the thymus preserves an intrinsic ability to regenerate. The aim of this work was to study the transcriptional effects of GCs on genes likely involved in regeneration of the epithelial cell network in the cervical and thoracic thymus of beef cattle treated with dexamethasone (DEX) or prednisolone (PRD) in comparison with a control group. Moreover, the ratio of bax/bcl2 genes was examined to verify a possible antiapoptotic activity occurring at the same time. In the cervical thymus, DEX administration increased the gene expression of c-myc (P0.01), tcf3 (P0.05), tp63 (P0.01), and keratin 5 (krt5; P0.01). In the thoracic thymus of DEX-treated cattle, the gene expression of tcf3 (P0.01), tp63 (P0.01), and krt5 (P0.05) was increased. These results suggested that thymic regeneration is underway in the DEX-treated animals. However, the bax/bcl2 ratio was decreased in both cervical and thoracic thymus of DEX-treated cattle (P0.01 and P0.05, respectively), showing an antiapoptotic effect through the mitochondrial pathway. Conversely, PRD administration caused no change in the expression of all considered genes. These results sustain the hypothesis that regeneration occurs in the thymus parenchyma 6 d after the DEX treatment was discontinued. This hypothesis is also supported by the absence of alterations in the thymus of PRD-treated beef cattle. Indeed, previous studies showed the inability of PRD to induce macroscopic and microscopic lesions in the thymus. Therefore, in this context, it is not surprising that PRD induced no alteration of genes involved in the regeneration pathway.

In this study we examined macroscopic and microscopic lesions in the mitral valves of 50 regularly slaughtered horses in Italy. Macroscopically the results were minor. Microscopic lesions to the valve were found in both young and elderly subjects, confirming that there is no correlation between age and onset of lesions. We set out to evaluate whether the lesions correspond to valve disorder during the life of the animal.

A juvenile female striped dolphin Stenella coeruleoalba live stranded on 4 March 2016 at Alassio, western Ligurian Sea coast, Italy. The dolphin died shortly after stranding, and a complete postmortem examination was performed. Necropsy revealed severe tracheal occlusion and unilateral bronchial stenosis with luminal accumulation of abundant green-yellow mucous-gelatinous material. Histological features suggestive of tracheobronchial aspergillosis were observed. Cultures of lung tissue and tracheo-bronchial exudate isolated Aspergillus fumigatus, identified by a Microseq D2 LSUrDNA fungal sequencing kit. A pan-Herpesvirus nested-PCR assay on frozen samples obtained from multiple organs was positive. Phylogenetic analysis on the partial DNA polymerase gene revealed that the striped dolphin isolate was closely related to known cetacean Alphaherpesvirus sequences from the same host species. Attempted virus isolation was unsuccessful. The tissue levels of different persistent organic pollutants and the toxicological stress, evaluated using a theoretical model, showed a severely impaired immune response. This study reports the first case of occlusive mycotic tracheobronchitis in a free-living cetacean and the first molecular identification of an Alphaherpesvirus in a free-ranging striped dolphin stranded on the coast of Italy.

A methodology for the absolute quantification of regucalcin gene through quantitative PCR was set up to confirm that the decrease of regucalcin gene expression in the testis is an effective biomarker for tracing sex steroid hormone treatment in bovine husbandry. On the basis of TaqMan technology, an external standard curve was generated. Using in vivo experiments, a ROC curve was developed to calculate the criterion value, specificity, and sensitivity for this potential biomarker. Then, regucalcin gene expression was assessed in veal calves and beef intended for human consumption. In 11 of 54 calves and in 5 of 70 beef cattle the regucalcin gene was expressed under their respective cutoff. Additionally, a mild decrease of regucalcin protein expression was revealed by immunohistochemistry in subjects tested positive via qPCR. These preliminary results suggest that this transcriptomics test may be employed as a novel diagnostic screening tool, improving significantly the overall efficacy of food control.

Blood parasites infect all vertebrates (Clayton and Moore 1997). Avian malaria parasites (Plasmodium spp., Plasmodiidae) are cosmopolitan in their distribution and are responsible for severe diseases in domestic and wild birds.In September 2009, nine raptorial birds that either arrived recently or were maintained as permanent residents at the Safaripark Pombia (northwest Italy) showed loss of stamina, developing listlessness, anorexia and regurgitation. Within one month three animals died and were necropsied.Following the diagnosis of Plasmodium infection all other raptorial birds were treated: clinical improvement was observed in all birds, and blood smears made after one month resulted negative for parasites.

A renal neoplasia was observed in a slaughtered 33 month old neutered Piedmontese male cattle. The neoplastic tissue was characterized by lardaceous features, and fibrotic tissue was evident with boundless interlacing, which separated the lesion in distinct diffusely irregular lobules. The lungs showed several macroscopical subpleuric nodules. Histological investigations showed multiform and distinct features of renal tumour. Anaplastic and embryonal aspects were represented by poorly differentiated or undifferentiated cells. Rarely, neoplastic epithelial elements were organized in pseudo-tubular formations. Immunohisto- chemical staining for detection of cytokeratins, vimentin, actin, S100, desmin and factor VIII was performed. Ultrastructural findings revealed two different populations of epithelial cells. Histological, immunohistochemical and ultrastructural findings in the lungs confirmed the diagnostic hypothesis of a metastatic tumour from the kidney.

Background: Haemoparasites in feral pigeons have been studied in several countries but no data are available from Italy. The aim of this work was to evaluate the prevalence and diversity of Haemoproteus spp./Plasmodium spp. and Leucocytozoon spp. in feral pigeons from northwest Italy, as well as the association between infection and host age or sex. Methods: Feral pigeons were collected during a regional culling programme from the Piedmont region (northwest Italy) and subjected to necropsy. Infections were detected from DNA extracted from the spleen following a nested PCR protocol. The association between sex or age and infection status was evaluated using the chi-squared test for independence or Fisher’s exact test. Results: Out of 51 animals, 15 were positive for Haemoproteus/Plasmodium spp. and eight for Leucocytozoon spp., with a significant difference between haemoparasites prevalence. There was no significant association between age or sex and infection status. The coinfection with different haemoparasites was very significant (p < 0.01), showing a greater relative risk to be infected by a second haemoparasite in birds already infected, in particular in male and in adult pigeons. DNA sequencing of Leucocytozoon spp. showed six different lineages in pigeons, and one of Haemoproteus and Plasmodium, respectively. Conclusions: Blood parasites are continuously circulating around the world, and the results presented in the paper suggest that cross infection of feral pigeons with haemoparasites typical of other migratory or nonmigratory bird species is possible. Moreover, the geographical location of Italy along the main migratory routes is a crucial factor to be considered for migratory birds, because they can be affected by blood parasites detected in feral pigeons, and vice versa.

Spontaneous odontogenic tumors are neoplasms characterized by a mixed odontogenic ectomesenchymal and odontogenic epithelial origin; they are rare in both humans and animals. A 3-year-old male Alpine Chamois (Rupicapra rupicapra) was found dead in north-west Italy, and was referred for the necropsy to the Department of Veterinary Sciences of the University of Turin (Italy). At the external examination a 10 × 8 cm, exophytic, red-pink, smooth, firm and ulcerated mass was observed on the inferior lip. Histologically the tumor was characterized by spindle shaped cells arranged in bundles in an abundant hyaline matrix. Multifocal and rare chords of odontogenic epithelium mixed with rare melanocytes that penetrate the neoplasia were visible. Immunohistochemistry showed a clear cytokeratin positivity of epithelial clusters. Macroscopical, histological and immunohistochemical findings were consistent with a diagnosis of locally infiltrative ameloblastic fibroma. To our best knowledge, this is the first report of this tumor in a wild ungulate and in Alpine Chamois.

In this work, we identified for the first time hepatitis E virus (HEV) in a pet house rabbit, an adult 7 years old female of domestic rabbit (Oryctolagus cuniculus). Importantly, the resulting phylogenetic tree showed that the HEV strain identified in the pet house rabbit was closely related to a human HEV sequence; this finding reawakens concerns regarding the zoonotic risk represented by HEV in animals and expands to house rabbit the spectrum of potential source of infection for humans. Potential for domestic transmission of HEV to humans should be taken into account.

We have recently demonstrated that an attenuated strain of Salmonella enterica serovar Typhimurium unable to synthesize the zinc transporter ZnuABC (S. Typhimurium ΔznuABC), is able to protect mice against systemic and enteric salmonellosis and is safe in pigs. Here, we have tested the protective effects of S. Typhimurium ΔznuABC in pigs. Resistance to challenge with the fully virulent strain S. Typhimurium ATCC 14028 was assessed in animals vaccinated with S. Typhimurium ΔznuABC (two dosages tested), in controls vaccinated with a formalin-inactivated virulent strain and in unvaccinated controls. Clinical signs of salmonellosis, faecal shedding and bacterial colonization of organs were used to assess vaccine-induced protection. After the challenge, pigs vaccinated with the attenuated S. Typhimurium ΔznuABC strain did not display clinical signs of salmonellosis (fever or diarrhoea). The vaccine also reduced intestinal tract colonization and faecal shedding of the fully virulent Salmonella strain, as compared to control groups. S. Typhimurium ΔznuABC represents a promising candidate vaccine against salmonellosis in pigs.

A wild Beech Marten ( Martes foina ), was referred for necropsy to the Department of Animal Pathology of the University of Turin (Italy). At gross examination, whitish and firm masses, 10-mm in diameter, were found on the heart and in the kidney. Spleen showed lighter color and greater consistency, and the cut surface of the liver appeared scattered with whitish-yellow coalescing foci homogeneously distributed. Amyloid deposits were present in the perivascular and intercellular spaces of the visceral organs, such as the heart, liver, and kidneys. Amyloid stained positively with Congo red with and without 5% potassium permanganate pretreatment and showed green birefringence observable under polarized light. A diagnosis of systemic AL amyloidosis was made. This is the first description of systemic AL amyloidosis in a wild Stone Marten.

During the years 2009–2011, 7 Siberian tigers ( Panthera tigris altaica), aged between 2 and 14 years, from the Safaripark of Pombia were referred for necropsy to the Department of Animal Pathology of the University of Turin (Italy). Three tigers, aged 10 (2 animals) and 14 years, had multifocal, irregularly distributed, white, soft, subpleural, 3-mm nodules scattered throughout the lungs. Histologically, there was a marked infiltration of macrophages, with foamy cytoplasm, and multinucleate giant cells interspersed with numerous clusters of cholesterol clefts. A mild lymphocytic infiltration was localized around the lesion. The findings were consistent with endogenous lipid pneumonia, which was considered an incidental finding of no clinical significance.

An 18-year-old female lion (Panthera leo) was referred to the Department of Animal Pathology of the University of Turin (Italy). At necropsy, multiple nodular, 4-20-mm, confluent white firm nodules were scattered throughout the pleural surfaces of the thoracic wall and of the lungs. Histological lesions were represented by proliferations of papillary structures lined by cuboidal basophilic mesothelial cells with large, oval nuclei and abundant granular eosinophilic cytoplasm. Immunohistochemistry revealed immunoreactivity for pancytokeratin and vimentin. None of the cells expressed calretinin antigen. Asbestos fibers and asbestos bodies were not detected respectively by light microscopy and by Scanning Electron Microscope-Energy Dispersive Spectrometer investigations. On the contrary, chrysotile asbestos were identified in samples from shelter material. Histological and immunohistochemical findings were consistent with the diagnosis of an epithelial malignant mesothelioma. To our best knowledge, this is the first report of a pleural mesothelioma in a lion.

White lions ( Panthera leo krugeri) have never been common in the wild, and at present, the greatest population is kept in zoos where they are bred for biological and biodiversity conservation. During the years 2003 to 2008 in a zoological garden in northern Italy, 19 white lions were born to the same parents, who were in turn paternally consanguineous. Out of the 19 lions, 4 (21%) were stillborn, 13 (69%) died within 1 month, and 1 (5%) was euthanatized after 6 months because of difficulty with prehension of food. Six lions (32%) showed malformations involving the head (jaw, tongue, throat, teeth, and cranial bones). One lion (5%) still alive at 30 months revealed an Arnold–Chiari malformation upon submission for neurological evaluation of postural and gait abnormalities. Paternal consanguinity of the parents, along with inbreeding among white lions in general, could account for the high incidence of congenital malformations of the head in this pride of white lions.

This report describes an uncommon case of nonamyloidotic fibrillary glomerulonephritis. A 5-year-old female European cat was presented with nephrotic syndrome. Serum biochemistry and urinalysis revealed a mild increase in cholesterol, low total protein, severe hypoalbuminemia, and high proteinuria with a high protein-to-creatinine ratio. An histologic examination revealed an interstitial nephritis and a diffuse glomerulonephritis, with multifocal thickening of the Bowman's capsule. Transmission electron microscopy showed widespread fibrillary deposits in the glomerular basement membrane and in the mesangium. These fibrils ranged between 18 and 26 nm in diameter and were Congo red negative, which allowed their differentiation from amyloid. Immunohistochemistry demonstrated expression for immunoglobulin M (IgM) and immunoglobulin G (IgG) within the mesangium. Renal deposits of Congo red-negative amyloid-like fibrils have been described in humans, horses, monkeys, and dogs. This is the first report of noncongophilic fibrillary glomerulopathy in a cat.