Granulomatous dermatitis caused by Exserohilum mcginnisii infection in a cow.

A 5-y-old, Piedmontese cow had a 4-mo history of ongoing development of skin masses. This was the only cow affected in a herd of 20 cows. Up to 12, hairless, red-to-black, raised nodules-to-plaques were distributed along the dorsum and tail head. Biopsies were taken for histopathology and ancillary testing. An ulcerated skin section contained dermal infiltrates of eosinophils, plasma cells, neutrophils, macrophages, lymphocytes, and multinucleate giant cells, and pyogranulomas. Fungal hyphae were seen within the dermis, multinucleate giant cells, and pyogranulomas. In pyogranulomas, fungi were surrounded by a Splendore-Hoeppli reaction. Dematiaceous (pigmented) hyphae were rarely observed with H&E-stained and unstained (cleared and mounted) sections, but stained well with a Fontana-Masson stain. Exserohilum mcginnisii was identified by fungal culture, followed by PCR assay and sequencing. Exserohilum is a dematiaceous fungus that causes disease in humans and rarely in animals. The use of unstained sections and Fontana-Masson stain are important to demonstrate pigment because dematiaceous fungi have little melanin and appear as hyaline hyphae histologically. PCR assay and sequencing aid in the differentiation and classification of fungal species. To our knowledge, E. mcginnisii dermal granulomas have not been reported previously in cattle.

Evaluation of targeted next-generation sequencing for detection of equine pathogens in clinical samples.

Equine infectious disease outbreaks may have profound economic impact, resulting in losses of millions of dollars of revenue as a result of horse loss, quarantine, and cancelled events. Early and accurate diagnosis is essential to limit the spread of infectious diseases. However, laboratory detection of infectious agents, especially the simultaneous detection of multiple agents, can be challenging to the clinician and diagnostic laboratory. Next-generation sequencing (NGS), which allows millions of DNA templates to be sequenced simultaneously in a single reaction, is an ideal technology for comprehensive testing. We conducted a proof-of-concept study of targeted NGS to detect 62 common equine bacterial, viral, and parasitic pathogens in clinical samples. We designed 264 primers and constructed a bioinformatics tool for the detection of targeted pathogens. The designed primers were able to specifically detect the intended pathogens. Results of testing 27 clinical samples with our targeted NGS assay compared with results of routine tests (assessed as a group) yielded positive percent agreement of 81% and negative percent agreement of 83%, overall agreement of 81%, and kappa of 0.56 (moderate agreement). This moderate agreement was likely the result of low sensitivity of some primers. However, our NGS assay successfully detected multiple pathogens in the clinical samples, including some pathogens missed by routine techniques.

Evaluation of Targeted Next-Generation Sequencing for Detection of Bovine Pathogens in Clinical Samples.

The laboratory diagnosis of infectious diseases, especially those caused by mixed infections, is challenging. Routinely, it requires submission of multiple samples to separate laboratories. Advances in next-generation sequencing (NGS) have provided the opportunity for development of a comprehensive method to identify infectious agents. This study describes the use of target-specific primers for PCR-mediated amplification with the NGS technology in which pathogen genomic regions of interest are enriched and selectively sequenced from clinical samples. In the study, 198 primers were designed to target 43 common bovine and small-ruminant bacterial, fungal, viral, and parasitic pathogens, and a bioinformatics tool was specifically constructed for the detection of targeted pathogens. The primers were confirmed to detect the intended pathogens by testing reference strains and isolates. The method was then validated using 60 clinical samples (including tissues, feces, and milk) that were also tested with other routine diagnostic techniques. The detection limits of the targeted NGS method were evaluated using 10 representative pathogens that were also tested by quantitative PCR (qPCR), and the NGS method was able to detect the organisms from samples with qPCR threshold cycle (CT ) values in the 30s. The method was successful for the detection of multiple pathogens in the clinical samples, including some additional pathogens missed by the routine techniques because the specific tests needed for the particular organisms were not performed. The results demonstrate the feasibility of the approach and indicate that it is possible to incorporate NGS as a diagnostic tool in a cost-effective manner into a veterinary diagnostic laboratory.

Evaluation of novel oral vaccine candidates and validation of a caprine model of Johne's disease.

Johne's disease (JD) caused by Mycobacterium avium subspecies paratuberculosis (MAP) is a major threat to the dairy industry and possibly some cases of Crohn's disease in humans. A MAP vaccine that reduced of clinical disease and/or reduced fecal shedding would aid in the control of JD. The objectives of this study were (1) to evaluate the efficacy of 5 attenuated strains of MAP as vaccine candidates compared to a commercial control vaccine using the protocol proposed by the Johne's Disease Integrated Program (JDIP) Animal Model Standardization Committee (AMSC), and (2) to validate the AMSC Johne's disease goat challenge model. Eighty goat kids were vaccinated orally twice at 8 and 10 weeks of age with an experimental vaccine or once subcutaneously at 8 weeks with Silirum® (Zoetis), or a sham control oral vaccine at 8 and 10 weeks. Kids were challenged orally with a total of approximately 1.44 × 10(9) CFU divided in two consecutive daily doses using MAP ATCC-700535 (K10-like bovine isolate). All kids were necropsied at 13 months post-challenge. Results indicated that the AMSC goat challenge model is a highly efficient and valid model for JD challenge studies. None of the experimental or control vaccines evaluated prevented MAP infection or eliminated fecal shedding, although the 329 vaccine lowered the incidence of infection, fecal shedding, tissue colonization and reduced lesion scores, but less than the control vaccine. Based on our results the relative performance ranking of the experimental live-attenuated vaccines evaluated, the 329 vaccine was the best performer, followed by the 318 vaccine, then 316 vaccine, 315 vaccine and finally the 319 vaccine was the worst performer. The subcutaneously injected control vaccine outperformed the orally-delivered mutant vaccine candidates. Two vaccines (329 and 318) do reduce presence of JD gross and microscopic lesions, slow progression of disease, and one vaccine (329) reduced fecal shedding and tissue colonization.

Extra-adrenal retroperitoneal paraganglioma in a dog.

An extra-adrenal retroperitoneal paraganglioma was observed in a 10.5-year-old male Boxer dog. Additionally, the dog had an aortic base tumor, multiple thyroid adenomas, multiple testicular interstitial cell tumors, bilateral nodular adrenal cortical hyperplasia, and parathyroid gland hyperplasia. The hypothesis that the retroperitoneal mass represents a primary extra-adrenal paraganglioma rather than metastatic mass from the aortic body tumor is considered. Either primary or metastatic extra-adrenal retroperitoneal paragangliomas are rarely reported in dogs.

Cholesterol granuloma associated with otitis media in a cat.

An 8-year-old, male neutered Siamese cat was presented with Horner syndrome and right head tilt. A soft tissue mass was observed in the right tympanic cavity, and bulla osteotomy was performed. Tissue samples retrieved from the tympanic cavity were sent for histology, and a middle ear fluid swab was sent for bacterial culture and sensitivity. Histologic diagnosis was of otitis media associated with cholesterol granuloma (CG). Bacterial culture yielded Pasteurella multocida and Leifsonia (Corynebacterium) aquaticum. Middle ear CG is frequently seen in human beings and is associated with a variety of middle ear diseases including otitis media. Cholesterol granuloma of the middle ear has been experimentally induced in cats. The clinical and pathological findings of a spontaneous case of CG in the tympanic cavity of a cat with otitis media are described herein.

The occurrence of Bovine viral diarrhea virus in hunter-harvested white-tailed deer (Odocoileus virginianus) in the state of Georgia, southeastern United States.

The prevalence of Bovine viral diarrhea virus (BVDV) in free-ranging white-tailed deer (WTD, Odocoileus virginianus) in the state of Georgia was evaluated using ear notches collected from hunter-harvested deer during the hunting season of 2010-2011. From September to December 2010, 367 ear samples from WTD were collected from 37 counties in Georgia. The samples were from 178 (48.5%) female deer, 187 (51%) male deer, and 2 (0.5%) of unknown sex. The age of the animals varied from 6 months to 6.5 years. The age was not recorded in 34 animals (9.3%). Of the animals with known ages, 42% were under 2 years. Screening of 367 samples for BVDV using an antigen-capture enzyme-linked immunosorbent assay (AgELISA) resulted in 364 negative samples and 3 suspect samples. The 3 suspect samples were negative for BVDV reverse transcription polymerase chain reaction (RT-PCR), virus isolation, and immunohistochemistry. A subpopulation of samples (n = 89) selected from various geographical regions also tested negative for BVDV RT-PCR. In conclusion, although a few of the samples were suspect for the presence of BVDV by AgELISA, the presence of the virus within the deer population studied could not be confirmed further.

Meningoencephalitis caused by Mycoplasma edwardii in a dog.

A 6-week-old, female, mixed-breed dog with a clinical history of sudden onset of neurologic signs was presented for necropsy. The dog was diagnosed with suppurative and histiocytic meningoencephalitis based on necropsy findings and histopathology. Mycoplasma sp. was isolated in pure culture from the brain and meninges and was identified as Mycoplasma edwardii using DNA sequencing.

Ligneous conjunctivitis secondary to a congenital plasminogen deficiency in a dog.

CASE DESCRIPTION: A 7-month-old 16.6-kg (36.5-lb) sexually intact female Golden Retriever was evaluated because of progressive severe bilateral membranous conjunctivitis, oral lesions, nasal discharge, and cough. CLINICAL FINDINGS: Histologic examination of conjunctival biopsy specimens revealed findings consistent with ligneous conjunctivitis. Circulating plasminogen activity was repeatedly low, and congenital plasminogen deficiency was identified as the underlying cause of the ocular, oral, and respiratory lesions. TREATMENT AND OUTCOME: Topical and subconjunctival administrations of fresh frozen plasma (FFP), topical administration of cyclosporine, and oral administration of azathioprine had no effect on the conjunctival membranes. Excision of the membranes followed by intensive treatment with topical applications of heparin, tissue plasminogen activator, corticosteroid, and FFP and IV administration of FFP prevented membrane regrowth. Intravenous administration of FFP increased plasma plasminogen activity to within reference limits, improved respiratory and oral lesions, and resulted in weight gain; discontinuation of this treatment resulted in weight loss, signs of depression, and worsening of lesions. After euthanasia because of disease progression, necropsy findings included mild hydrocephalus; multifocal intestinal hemorrhages; and fibrinous plaques in the oral cavity, nasopharynx, trachea, esophagus, and pericardium. Microscopically, the plaques were composed of fibrin and poorly organized granulation tissue. Fibrin thrombi were present within vessels in the lungs, oral cavity, and trachea. CLINICAL RELEVANCE: In dogs, congenital plasminogen deficiency can occur and may be the underlying cause of ligneous conjunctivitis. A combination of surgical and medical treatments may improve conjunctival membranes, and administration of FFP IV appears to be effective in treating nonocular signs of plasminogen deficiency.

Pathogenicity of Rhodococcus equi in mice, isolated from environment, human and horse clinical samples

Rhodococcus equi is a facultative intracellular pathogen associated with bronchopneumonia, mesenteric lymphadenitis and enterocolitis in foals. Although R. equi is likely to be found in every horse-breeding farm, the clinical disease is unrecognized in most of them. Capsule components, equi factor, micolic acid and some products encoded by the large 85-90Kb plasmid were described as virulence factors. However, the pathogenesis of R. equi infections and the sensibility of foals are not completely understood. The aim of this study was evaluate the virulence of R. equi isolated from human, horses and environment for mices. Nine strains carrying the 85-90Kb plasmid isolated from foal clinical specimens, one from immunodeficient human patient and six plasmidless strains (four isolated from feces, one from pasture and one from immunodeficient human patient) were inoculated in cyclophosphamide immunossuppressed mice. The pathological changes and viability of R. equi cells in the liver of mice was verified after the 3rd, 6th an 10th day after inoculation for horse and environmental isolates and for R. equi isolates from human patients on the 1st, 3rd and 6th day. During the necropsy procedures, infiltrate of macrophages and pyogranulomatous lesions were detected after the sixth pos-inoculation day in the liver and spleen. In horse isolates, only plasmid positive strains were virulent, but in human isolates both strains (plasmid positive e plasmid negative) were virulent. Both groups of the immunossupressed mice inoculated with R. equi isolated from environment showed pathological changes. All R. equi strains were unable to kill non imunossuppressed mice.

Rhodococ-cus equi é um patógeno intracelular facultativo associado com broncopneumonia, linfadenite mesentérica e enterocolite em potros. Apesar do patógeno ser amplamente distribuído no ambiente equino, a doença não é encontrada em todos os criatórios. Componentes capsulares, "fator equi", ácido micólico e alguns produtos codificados por um grande plasmídeo de 85-90Kb foram descritos como fatores de virulência. Entretanto, a patogênese da infecção e a susceptibilidade dos potros não são completamente entendidas. Nove cepas carreando o plasmídeo e isoladas de potros doentes, um isolado de paciente humano imunossuprimido, e seis cepas sem plasmídeo (4 de fezes, 1 de pastagem e 1 de paciente humano imunossuprimido) foram inoculadas em camundongos imunossuprimidos com ciclofosfamida. As alterações patológicas e a viabilidade das células de R. equi no fígado foram verificadas depois do terceiro, sexto e décimo dia após a inoculação para isolados clínicos e depois do primeiro, terceiro e sexto dia para isolados de pacientes humanos. Na necropsia foram detectados infiltrado de macrófagos e lesões piogranulomatosas no fígado e baço dos camundongos após o sexto dia da inoculação. Para isolados clínicos, somente os plasmídeo positivos foram virulentos, mas para os humanos, ambos os isolados (plasmídeo positivos e plasmídeo negativos) foram virulentos. Ambos os grupos de camundongos imunossuprimidos inoculados com isolados ambientais mostraram alterações patológicas. Todos os isolados foram incapazes de matar camundongos imunocompe-tentes.

Hyperthermic syndrome in dairy cattle associated with consumption of ergots of Claviceps purpurea in southern Brazil.

A hyperthermic syndrome (HS) associated with consumption of ration contaminated with Claviceps purpurea is described in 4 dairy cattle herds from different counties in the state of Rio Grande do Sul, southern Brazil. Outbreaks of this mycotoxicosis occurred during the summer of 1999-2000 and autumn of 2002 when 25% to 70% of cattle from those herds were affected. The disease occurred after the introduction of a new batch of feed contaminated with sclerotia of C. purpurea. First clinical signs were observed up to 3 mo after the ration was introduced and consisted of pyrexia, intense salivation and drooling, open-mouth, difficult breathing, in some cases with the tongue protruded, decrease in milk yield, decrease in food intake and weight loss. Affected cattle compulsively sought shade or water ponds. Signs were more pronounced during daytime. The severity of the clinical picture varied according to changes in environmental temperature during the day. There was complete recovery approximately 60 d after feeding with the contaminated ration was discontinued. Abortions occurred during the outbreak and soon after the remission of the clinical signs associated with HS. In one of the herds, reproductive performance and fertility were seriously impaired even after clinical signs of HS completely disappeared. Diagnosis was based on epidemiology, clinical signs and pathology. The pathogenetic mechanisms involved in the occurrence of HS are briefly discussed.

Síndrome do abscesso pituitário em bezerros associada ao uso de tabuleta nasal para desmame interrompido / Pituitary abscess syndrome in calves following injury of the nasal septum by a plastic device used to prevent suckling

Descrevem-se surtos e casos esporádicos de síndrome do abscesso pituitário em bovinos no Estado do Rio Grande do Sul. A doença ocorreu em 8 propriedades de gado de corte, nos municípios de Cachoeira do Sul, Lavras do Sul, Bagé, Osório e Vila Nova do Sul, no período de 1998 a 2002. De um total de 2.438 bezerros submetidos ao desmame interrompido com o uso da tabuleta nasal, aproximadamente 35 (1,4 por cento) animais adoeceram e 24 (0,98 por cento) morreram. A idade dos bezerros afetados variava entre 3 e 12 meses. Os animais doentes apresentavam corrimento nasal, depressäo, febre, incoordenaçäo motora, andar em círculos, desvio lateral da cabeça, hipermetria, exoftalmia, disfagia, mandíbula caída, protusäo lingual, dificuldade de mastigaçäo e sialorréia. Em alguns casos, observou-se também cegueira, acompanhada ou näo de turvaçäo dos humores do globo ocular, exoftalmia e opacidade da córnea. Nas fases terminais, ocorriam decúbito lateral, convulsöes, nistagmo, opistótono, coma e morte. Os principais achados de necropsia consistiam em abscessos únicos pituitários ou parapituitários que comprimiam dorsalmente o tronco encefálico e nervos cranianos próximos à pituitária. Em alguns casos, havia osteomielite envolvendo o osso baso-esfenóide com a formaçäo de abscessos na substância encefálica, leptomeningite na superfície ventral do encéfalo e medula espinhal cervical e rinite necrosante ou abscedativa associada às lesöes traumáticas provocadas pela colocaçäo da tabuleta nasal. Histologicamente, os abscessos correspondiam a grandes agregados de neutrófilos e restos celulares circundados por células mononucleares e proliferaçäo de tecido conjuntivo. Meningite fibrinopurulenta nas leptomeninges do cerebelo, tronco encefálico e medula espinhal cervical também foi observada. Em alguns casos, a inflamaçäo purulenta se estendia para o parênquima da pituitária. Arcanobacterium (Actinomyces) pyogenes foi isolado dos abscessos. O diagnóstico de síndrome do abscesso pituitário foi baseado nos dados epidemiológicos, sinais clínicos, achados macroscópicos, histológicos e microbiológicos

Intoxicaçäo espontânea por Senecio brasiliensis (Asteraceae) em ovinos no Rio Grande do Sul / Spontaneous poisoning in sheep by Senecio brasiliensis (Asteraceae) in southern Brazil

Descreve-se a ocorrência de um surto de intoxicaçäo espontânea por Senecio brasiliensis em ovinos em um estabelecimento do município de Mata, Estado do Rio Grande do Sul, em meados de janeiro de 1997. De um total de 94 ovinos, 51 (54,25 por cento) animais adoeceram e 50 (53,2 por cento) morreram. Esse rebanho permaneceu durante aproximadamente 7 meses (de junho de 1996 a janeiro de 1997) em piquetes de pastagem nativa onde havia grande quantidade de S. brasiliensis. O quadro clínico manifestado pelos animais afetados consistia em fotossensibilizaçäo, emagrecimento progressivo, apatia, fraqueza, perturbaçöes neurológicas como depressäo, andar a esmo e desequilibrado, icterícia e hemoglobinúria. Houve melhora das lesöes de pele naqueles ovinos que desenvolveram fotossensibilizaçäo hepatógena depois que foram retirados do sol. As principais lesöes macroscópicas observadas em 9 dos 10 ovinos necropsiados incluíam fígado diminuído de tamanho, firme, difusamente marrom amarelado ou esverdeado, com quantidades variáveis de nódulos de 1-3 mm de diâmetro, bem circunscritos, salientes na cápsula, amarelados, distribuídos aleatoriamente por todo o parênquima. A vesícula biliar estava repleta e preenchida por bile verde escura e espessa. Havia também derrames cavitários (hidropericárdio e ascite). Crise hemolítica aguda fatal associada à intoxicaçäo crônica hepatógena por cobre foi observada em cinco ovinos. Além das lesoes hepáticas macroscópicas já mencionadas, foi observada icterícia generalizada da carcaça, rins tumefeitos, friáveis, difusamente escurecidos ou com fino pontilhado enegrecido; a urina era marrom escura (hemoglobinúria). As principais lesöes microscópicas foram observadas no fígado e consistiam em hepatomegalocitose, proliferaçäo de ductos biliares (hiperplasia ductal) e fibrose periportal moderada acompanhada de infiltrado inflamatório mononuclear. Macrófagos carregados de pigmento acastanhado formavam aglomerados nas tríades portais ou estavam dispersos entre os hepatócitos remanescentes. O material armazenado no citoplasma desses macrófagos correspondia a ceróide e cobre, positivo nas técnicas de PAS e rodanina, respectivamente. Nos rins de cinco animais, havia nefrose hemoglobinúrica caracterizada por degeneraçäo e necrose do epitélio tubular, presença de hemoglobina e hemossiderina no citoplasma das células epiteliais dos túbulos contorcidos e cilindros de hemoglobina na luz tubular. Evidência morfológica de encefalopatia hepática incluía...

Síndrome distérmica (hipertermia) em bovinos associada à intoxicaçäo por Claviceps purpurea / Dysthermic syndrome (hyperthermia) in cattle associated with poisoning by Claviceps purpurea

Descrevem-se três surtos de síndrome distérmica (hipertermia) associada à intoxicaçäo por Claviceps purpurea, em bovinos de leite durante o veräo de 1999-2000, em três estabelecimentos do Rio Grande do Sul. De um total de 66 bovinos que ingeriram a raçäo contaminada com o fungo, 37 (56 por cento) adoeceram até 3 meses após a introduçäo da raçäo contaminada. Os principais sinais clínicos foram temperatura retal elevada, pêlos compridos, longos e sem brilho, salivaçäo intensa, respiraçäo ofegante, com a boca aberta e, em alguns casos, com a língua para fora da cavidade oral. Os animais acometidos procuravam sombra ou permaneciam dentro d'água. Houve diminuiçäo de 10 a 30 por cento no consumo de alimentos e perda de peso. A reduçäo na produçäo de leite foi de 30 a 50 por cento. Os sinais clínicos se intensificavam durante o dia e eram diretamente proporcionais à elevaçäo da temperatura ambiental. Os achados de necropsia em um bovino que foi eutanasiado, incluíram leve enfisema pulmonar, principalmente na regiäo dorsal dos lobos pulmonares diafragmáticos. Histologicamente havia moderada hipertrofia da musculatura lisa dos bronquíolos e ruptura de septos alveolares formando cotos alveolares em clava. Nos três estabelecimentos onde ocorreram os surtos, escleródios de C. purpurea foram observados nas amostras de raçäo fornecida aos bovinos. Os animais afetados recuperaram-se após aproximadamente 60 dias da retirada da raçäo contaminada. O diagnóstico baseou-se em dados epidemiológicos, sinais clínicos, na presença de escleródios de C. purpurea na raçäo fornecida aos animais, nos achados de necropsia e na histopatologia. A patogenia e o quadro clínico-patológico observados sao discutidos e comparados com outros relatos dessa enfermidade