Schmallenberg virus pathogenesis, tropism and interaction with the innate immune system of the host.

Schmallenberg virus (SBV) is an emerging orthobunyavirus of ruminants associated with outbreaks of congenital malformations in aborted and stillborn animals. Since its discovery in November 2011, SBV has spread very rapidly to many European countries. Here, we developed molecular and serological tools, and an experimental in vivo model as a platform to study SBV pathogenesis, tropism and virus-host cell interactions. Using a synthetic biology approach, we developed a reverse genetics system for the rapid rescue and genetic manipulation of SBV. We showed that SBV has a wide tropism in cell culture and "synthetic" SBV replicates in vitro as efficiently as wild type virus. We developed an experimental mouse model to study SBV infection and showed that this virus replicates abundantly in neurons where it causes cerebral malacia and vacuolation of the cerebral cortex. These virus-induced acute lesions are useful in understanding the progression from vacuolation to porencephaly and extensive tissue destruction, often observed in aborted lambs and calves in naturally occurring Schmallenberg cases. Indeed, we detected high levels of SBV antigens in the neurons of the gray matter of brain and spinal cord of naturally affected lambs and calves, suggesting that muscular hypoplasia observed in SBV-infected lambs is mostly secondary to central nervous system damage. Finally, we investigated the molecular determinants of SBV virulence. Interestingly, we found a biological SBV clone that after passage in cell culture displays increased virulence in mice. We also found that a SBV deletion mutant of the non-structural NSs protein (SBVΔNSs) is less virulent in mice than wild type SBV. Attenuation of SBV virulence depends on the inability of SBVΔNSs to block IFN synthesis in virus infected cells. In conclusion, this work provides a useful experimental framework to study the biology and pathogenesis of SBV.

Analysis of suspected wildlife crimes submitted for forensic examinations in Scotland.

This study describes the occurrence of suspected wildlife crimes submitted for forensic examination in Scotland in 2010. The study aims were to determine which types of crimes were committed, which species were targeted, and the outcome of investigations, in order to assess the contribution of forensic examinations in the prosecution of wildlife crimes. Information on suspected wildlife crimes submitted between January 1 and December 31, 2010 to the SAC Consulting: Veterinary Services Disease Surveillance Centers, Science and Advice for Scottish Agriculture, and to the University of Glasgow, was used. The location of suspected crimes, the species targeted, cause of death, and types of the 188 submitted wildlife crimes were summarized. More information regarding cases involving birds were submitted than cases involving mammals, and included 114 raptors, 14 waterfowl, and 22 "other bird species." Mammal cases (n = 38) included 12 badgers, 8 foxes, 7 deer, 4 hares, and 7 "other mammals." The cause of death was determined in 124 suspected crimes; malicious or accidental trauma was the most likely cause of death in 72, and 33 were poisoned. Forensic evidence supporting criminal activity was found in 53 cases, and poisoning was the most frequent crime recorded. At least five individuals were successfully prosecuted, representing 2.7 % of submissions. It was challenging to track cases from submission through to prosecution and laboratories conducting forensic investigations were often not informed of the outcome of prosecutions or court decisions.

Activation of Wnt signalling promotes development of dysplasia in Barrett's oesophagus.

Barrett's oesophagus is a precursor of oesophageal adenocarcinoma, via intestinal metaplasia and dysplasia. Risk of cancer increases substantially with dysplasia, particularly high-grade dysplasia. Thus, there is a clinical need to identify and treat patients with early-stage disease (metaplasia and low-grade dysplasia) that are at high risk of cancer. Activated Wnt signalling is critical for normal intestinal development and homeostasis, but less so for oesophageal development. Therefore, we asked whether abnormally increased Wnt signalling contributes to the development of Barrett's oesophagus (intestinal metaplasia) and/or dysplasia. Forty patients with Barrett's metaplasia, dysplasia or adenocarcinoma underwent endoscopy and biopsy. Mice with tamoxifen- and ß-naphthoflavone-induced expression of activated ß-catenin were used to up-regulate Wnt signalling in mouse oesophagus. Immunohistochemistry of ß-catenin, Ki67, a panel of Wnt target genes, and markers of intestinal metaplasia was performed on human and mouse tissues. In human tissues, expression of nuclear activated ß-catenin was found in dysplasia, particularly high grade. Barrett's metaplasia did not show high levels of activated ß-catenin. Up-regulation of Ki67 and Wnt target genes was also mostly associated with high-grade dysplasia. Aberrant activation of Wnt signalling in mouse oesophagus caused marked tissue disorganization with features of dysplasia, but only selected molecular indicators of metaplasia. Based on these results in human tissues and a mouse model, we conclude that abnormal activation of Wnt signalling likely plays only a minor role in initiation of Barrett's metaplasia but a more critical role in progression to dysplasia.

Caudal vena caval thrombosis following treatment of deep digital sepsis.

A diagnosis of caudal vena caval thrombosis was made by ultrasonography of a Holstein cow presented for lethargy and poor milk production. Medical treatment was unsuccessful and the cow was euthanized. The diagnosis was confirmed at necropsy and Fusobacterium necrophorum was isolated from the thrombus. This paper discusses potential novel sources of caval thrombosis in this case.

Classical swine fever virus can remain virulent after specific elimination of the interferon regulatory factor 3-degrading function of Npro.

Pestiviruses prevent alpha/beta interferon (IFN-alpha/beta) production by promoting proteasomal degradation of interferon regulatory factor 3 (IRF3) by means of the viral N(pro) nonstructural protein. N(pro) is also an autoprotease, and its amino-terminal coding sequence is involved in translation initiation. We previously showed with classical swine fever virus (CSFV) that deletion of the entire N(pro) gene resulted in attenuation in pigs. In order to elaborate on the role of the N(pro)-mediated IRF3 degradation in classical swine fever pathogenesis, we searched for minimal amino acid substitutions in N(pro) that would specifically abrogate this function. Our mutational analyses showed that degradation of IRF3 and autoprotease activity are two independent but structurally overlapping functions of N(pro). We describe two mutations in N(pro) that eliminate N(pro)-mediated IRF3 degradation without affecting the autoprotease activity. We also show that the conserved standard sequence at these particular positions is essential for N(pro) to interact with IRF3. Surprisingly, when these two mutations are introduced independently in the backbones of highly and moderately virulent CSFV, the resulting viruses are not attenuated, or are only partially attenuated, in 8- to 10-week-old pigs. This contrasts with the fact that these mutant viruses have lost the capacity to degrade IRF3 and to prevent IFN-alpha/beta induction in porcine cell lines and monocyte-derived dendritic cells. Taken together, these results demonstrate that contrary to previous assumptions and to the case for other viral systems, impairment of IRF3-dependent IFN-alpha/beta induction is not a prerequisite for CSFV virulence.

Disseminated cutaneous mast cell tumors with epitheliotropism and systemic mastocytosis in a domestic cat.

A 15-year-old female domestic, medium-haired cat presented to the referring veterinarian with a 2-month history of multiple, raised, disseminated, nodular skin lesions. A biopsy of 1 of the lesions was submitted to the Oklahoma Animal Disease Diagnostic Laboratory for evaluation. Histologically, there were multiple dermal nodules composed of sheets of neoplastic round cells. Multifocally, the neoplastic cells formed multiple small clusters of 3 to 5 cells within the epidermis. Distinct cytoplasmic granules were evident within the neoplastic cells with toluidine blue and Giemsa stains. The neoplastic cells were immunoreactive for c-KIT and lacked immunoreactivity for cluster of differentiation 3 with immunohistochemistry. Based on these findings, multiple epitheliotropic cutaneous mast cell tumors were diagnosed. The cat's health declined rapidly despite aggressive treatment, and the animal was humanely euthanatized. A complete necropsy revealed sheets of similar neoplastic mast cells within the spleen, liver, and individual cells scattered within the bone marrow. Exon 11 of the c-KIT messenger RNA from 1 of the cutaneous masses and the spleen was amplified with reverse transcription polymerase chain reaction, sequenced, and compared with the published c-KIT messenger RNA sequence from fetal cat tissues. The maximum identity was 100% for both tissue samples. To the authors' knowledge, the present report is the first to describe disseminated cutaneous mast cell tumors with epitheliotropism and systemic mastocytosis in a domestic cat.

Malignant melanoma of the subcutaneous tissues in an umbrella cockatoo (Cacatua alba).

A 24-year-old female umbrella cockatoo (Cacatua alba) was examined because of a subcutaneous swelling of the left shoulder. Radiographs of the left wing revealed boney proliferation of the left proximal humerus. At surgery, an unencapsulated infiltrative nonpigmented mass within the subcutaneous tissue and skeletal muscle was identified overlying the left humerus. The mass was surgically excised and submitted for histologic examination; however, the cockatoo died after excision. Histologically, the mass was composed of nests and sheets of pleomorphic neoplastic cells with a high mitotic index and extensive necrosis. Rare neoplastic cells contained fine brown cytoplasmic granules that stained black with Fontana-Masson, confirming the presence of melanin pigment. The histomorphologic diagnosis was consistent with an anaplastic malignant melanoma of subcutaneous tissues.

The effects of formalin fixation and tissue embedding of bovine liver on copper, iron, and zinc analysis.

Veterinary analytical chemistry laboratories might be called upon to analyze formalin-fixed or paraffin-embedded tissue samples for trace minerals. The purpose of this study was to determine whether concentrations of copper (Cu), iron (Fe), and zinc (Zn) are comparable among fresh or frozen, formalin-fixed, and paraffin-embedded bovine liver samples on an as-received basis. Three liver sample subtypes (fresh or frozen, formalin-fixed, and paraffin-embedded) from 12 cows were collected and analyzed for Cu, Fe, and Zn concentrations. Concentrations were measured by using inductively coupled argon plasma atomic-emission spectroscopy. There was no significant difference in mineral measurements between fresh or frozen and formalin-fixed samples for Cu and Zn (both P > or = 0.052). The median concentration of Fe was lower in the fresh or frozen samples than in the formalin-fixed samples. However, for every pair of fresh or frozen and paraffin-embedded samples for all 3 minerals, the fresh or frozen sample had a lower measurement than the paraffin-embedded sample (all P = 0.005). Differences in mineral measurements associated with tissue processing did not result in differences in classification (within or outside the reference range) for Fe. However, the classification of Cu and Zn was different up to 25% of the time with fresh or frozen versus formalin-fixed or embedded liver. Although Cu, Fe, and Zn concentrations attained from processed tissue may be useful, they must be evaluated with caution.

Parvovirus infection in domestic companion animals.

Parvovirus infects a wide variety of species. The rapid evolution, environmental resistance, high dose of viral shedding, and interspecies transmission have made some strains of parvovirus infection difficult to control within domestic animal populations. Some parvoviruses in companion animals, such as canine parvovirus (CPV) 1 and feline parvovirus, have demonstrated minimal evolution over time. In contrast, CPV 2 has shown wide adaptability with rapid evolution and frequent mutations. This article briefly discusses these three diseases, with emphasis on virus evolution and the challenges to protecting susceptible companion animal populations.

Characterization of rupture of abdominal artery aneurysm in dairy cattle.

In this retrospective study, the authors describe the gross and histologic changes associated with rupture of an abdominal artery aneurysm in 33 mature female Holstein cattle between January 1980 and June 2005 from 29 farms in upstate New York and northern Pennsylvania. Over this period, there was an increase in the number of cases submitted for necropsy per year, and a seasonal trend did not exist. Affected animals ranged from 2.5 to 5.5 years of age. Grossly affected cattle exhibited marked hemoabdomen. There was marked dilation and rupture of the abdominal aorta or one of its branches, including the mesenteric, left gastric, celiac artery, right ruminal artery, or left ruminal artery. Histologically, the tunica media of the affected arteries was often thin and irregular in width with disrupted, fragmented, and coiled elastin. Granulation tissue and hemorrhage was localized at the site of rupture. There was mild-to-marked hyperplasia of the tunica intima that was irregular and disorderly with adjacent smooth-muscle hyperplasia within the tunica media. In addition, in a fraction of cases, there was mild-to-moderate mucinous change (48%) and mineralization (30%) within the tunica media. To the authors' knowledge, this is the first description of the gross and histologic changes in Holstein cows with abdominal artery aneurysm and rupture.

Magnetic Resonance Imaging and Clinical Findings Associated with Choroid Plexus Spinal Cord "Drop" Metastases.

A 5 yr old castrated male whippet presented with a unique presentation of ambulatory paraparesis and subsequent diagnosis of primary intracranial choroid plexus carcinoma, with metastases to the cervical, thoracic, lumbar, and sacral spinal cord segments. Magnetic resonance imaging was performed initially of the thoracolumbar vertebral column and was followed by MRI sequences of the brain for confirmation of the presence of a primary intracranial tumor. The dog was euthanized immediately following diagnostic imaging due to the severity of clinical signs and poor prognosis. Postmortem examination confirmed the presence of a primary choroid plexus carcinoma, with diffuse metastatic lesions to the spinal cord. To the authors' knowledge, this is the first description of MRI findings of spinal cord metastasis of a choroid plexus carcinoma in a dog. Additionally, this case demonstrates primary clinical signs arising from metastatic lesions. This confirms the importance of extensive neuroimaging investigations when suspecting choroid plexus tumors and evaluating response to treatment regimens.

Acquired cervical scoliosis attributed to Parelaphostrongylus tenuis infection in an alpaca.

CASE DESCRIPTION: A 2-year-old alpaca was evaluated because of acute onset of cervical scoliosis. CLINICAL FINDINGS: Physical examination revealed severe scoliosis of the caudal portion of the cervical vertebral column with a C-shaped curvature to the right side. No gait deficits were observed. Cervical radiography confirmed severe curvature of C4 to C6 but did not reveal any bony changes. Cerebrospinal fluid had high total protein concentration and extremely high nucleated cell count with a high proportion of eosinophils, suggesting parasitic infection. TREATMENT AND OUTCOME: The alpaca was treated for suspected parelaphostrongylosis with ivermectin, fenbendazole, flunixin, vitamin E, thiamine, physical therapy, and a custom-made neck brace. The alpaca's condition continued to deteriorate, and it developed tetraparesis and ataxia and was euthanized after approximately 1 month. Microscopic evaluation of the cervical spinal cord revealed marked vacuolar changes in the left medial portion of the ventral funiculus, mild lymphoplasmacytic infiltration, and multifocal granulomas. The lesions were continuous from C1 to C7 and were compatible with parasite migration. CLINICAL RELEVANCE: To the authors' knowledge, this is the first report of acquired scoliosis in an alpaca, which appears to represent an unusual manifestation of parelaphostrongylosis that was reported in horses.

Mycoplasma otitis in California calves.

A retrospective study of Mycoplasma otitis in California calves submitted for necropsy between 1993 and 2002 was conducted to characterize the demographic features of the disease and the pathologic findings associated with infection. Sixty-one confirmed cases of Mycoplasma otitis were identified among 20,525 necropsied cattle. All affected animals were calves, ranging in age from 2 weeks to 4 months and with a median age of 1.5 months. Ninety-two percent of the cases were dairy breeds. A higher percent of necropsied calves with Mycoplasma otitis were males (0.45%) than females (0.23%). The proportion of cases that had Mycoplasma otitis increased from 1993 to 2002, and there was a significant (P < 0.05) seasonal distribution, with the highest proportion in the spring and the lowest in the summer months. Infections involved both the middle and inner ear and were characterized by a suppurative inflammatory response with extensive bony involvement. Three species of Mycoplasma were isolated from the ears: M. bovis, M. bovirhinis, and M. alkalescens. Concurrent pneumonia occurred in 47 cases (77%), and Mycoplasma was isolated from the lungs of 30 of those cases. The increasing proportion of Mycoplasma otitis cases in the past 10 years emphasizes the importance of identifying risk factors that could be modified to lower the incidence of this disease in calves.

A rare case of hepatic T-cell rich B-cell lymphoma (TCRBCL) in a juvenile dog.

A 7-month-old castrated male French Bull dog was presented with vomiting, lethargy, anorexia and weight loss of 2 weeks duration. The patient's history and clinical manifestations of suspected hepatopathy were subjected to ultrasonography, radiography, biochemical investigations and cytology of hepatic lesion. The cytologic impression was hepatic lymphoma, which was later confirmed by histopathology. The neoplastic cells were strongly diffusely immunoreactive for PAX5, but not immunoreactive for CD3, and B lymphocyte specific clonal proliferation was detected using by assay of antigen receptor rearrangement. Large numbers of immunoreactive mature non-neoplastic lymphocytes were admixed with the neoplastic cell population. Therefore, the immunohistochemical results were definitively consistent with a T-cell rich B-cell lymphoma (TCRBCL). This is the first description of a hepatic TCRBCL in a juvenile dog showing a poor response to aggressive chemotherapy.

Outcome and prognostic indicators in 20 cats with surgically treated primary lung tumors.

The purpose of this retrospective study of 20 client-owned cats was to describe the clinical signs, surgical interventions, histological features, stage and treatments of primary lung tumors removed by surgical excision, and to determine which factors significantly influence survival. Any cat that underwent surgical resection of a primary lung tumor between 2000 and 2007 was included in the study. Patient records were reviewed and signalment, clinical signs, preoperative diagnostics, surgical findings and histopathological results recorded. Histological reports were reviewed and scored using World Health Organization criteria. The Kaplan-Meier test was used to evaluate each potential prognostic factor with survival. Twenty cats met the inclusion criteria. The presence of clinical signs (such as dyspnea) at the time of diagnosis (P = 0.032), pleural effusion (P = 0.046), stage M1 (P = 0.015), and moderately and poorly differentiated tumors on histopathology (P = 0.011) were factors that were significantly correlated with reduced survival times. The median survival time of the 20 cats was 11 days. Cats presenting with no clinical signs had a median survival time of 578 days post-surgery vs 4 days post-surgery when presented with clinical signs. Cats staged T1N0M0 lived longer than cats at other stages (P = 0.044). Of the cats that survived to the time of suture removal, median survival time was 64 days. The results indicate that the presence of clinical signs, pleural effusion, moderately and poorly differentiated tumors on histopathology, evidence of metastasis and any stage beyond T1N0M0 are negative prognostic indicators for cats with primary lung tumors. The findings demonstrate that cats that presented with clinical signs, pleural effusion, any stage other than T1N0M0, or moderately and poorly differentiated tumors on histopathology had a poor prognosis. Therefore, extensive preoperative diagnostics, including computed tomography scans, should be performed before considering surgical intervention in these cats. These findings may be used to guide therapeutic decision-making in cats diagnosed with primary lung tumors.