Comparison of PCR with stained slides of bone marrow and lymph nodes aspirates with suspect diagnosis for leishmaniasis.

Visceral leishmaniasis (VL), also known as kala-azar, is a disseminated protozoan infection caused by Leishmania donovani complex. Traditionally the definite diagnosis is made by amastigote detection in the tissue. The aim this study was to evaluate the PCR technique in stained slides of bone marrow and lymph nodes aspirates with suspect diagnosis for leishmaniasis. Slides were selected totaling 62 suspect cases (33 bone marrow samples and 29 lymph node samples) and 17 positive cases (8 bone marrow and 9 lymph node). From 62 suspect cases, 39 (62.90%) were confirmed to be positive being 17 (n=29) lymph node aspirates and 22 (n=33) bone marrow. This finding is in agreement with the higher sensitivity of the PCR assay compared to direct microscopic observation. In conclusion, the findings of this study supports the use of PCR on archive cytological preparation stained slides for the diagnosis of canine visceral leishmaniasis, emphasizing the higher sensitivity of this technique when compared to direct microscopic examination and mostly the use of the suspect status for the cytology samples that presents the previously mentioned particularities with focus on detecting the oligosymptomatic or assymptomatic dogs in endemic areas functioning as potential reservoirs for this disease.

Detection of the tumour suppressor gene TP53 and expression of p53, Bcl-2 and p63 proteins in canine transmissible venereal tumour.

Canine transmissible venereal tumour (CTVT) is a neoplasm transmitted among healthy dogs by direct contact with injured skin and/or mucous tissue. This study aimed to identify the TP53 gene, messenger RNA (mRNA) as well as the expression of p53, Bcl-2 and p63 proteins in histological sections of 13 CTVT samples at different stages of evolution. The in situ hybridization (ISH) and in situ reverse transcriptase polymerase chain reaction (RT-PCR) assays were used, which showed the DNA homologous to TP53 and its respective mRNA in 92.3% of the samples. We detected p53, p63 and Bcl-2 proteins in most of the cell samples in different grades of intensity. In addition, 46% of the samples were in the progressive and 54% in the regression phase. This is the first description of these proteins and a detailed study of their role in CTVT cells needs to be addressed in or to verify how these cells undergo apoptosis.

Malignant catarrhal fever-like lesions associated with ovine herpesvirus-2 infection in young calves (Bos indicus): a case report

Infection of susceptible ruminants, including domestic cattle (Bos taurus) and American bison (Bison bison), with ovine herpesvirus-2 (OvHV-2) may provoke the fatal vasculitis and lymphoproliferative syndrome, known as malignant catarrhal fever (MCF), reported worldwide. To the best of our knowledge, this is the first report of a clinical case of MCF-like lesions associated with ovine herpesvirus-2 (OvHV-2) infection in young calves (Bos indicus) including central nervous symptoms that occurred in Três Lagoas city, Mato Grosso do Sul state, a border town near São Paulo state, Brazil. The diagnosis was based on typical histological lesions characterized by systemic lymphohistiocytic and fibrinoid vasculitis, confirmed by polymerase chain reaction and subsequent phylogenetic analysis of detected OvHV-2 sequences. This finding indicates that MCF disease is spread among herds concentrated in border areas between Mato Grosso do Sul and São Paulo states.

Epidemiological aspects of astrovirus and coronavirus in poults in the South Eastern Region of Brazil / Aspectos epidemiológicos associados à prevalência de Astrovirus e Coronavirus em lotes de perus da Região Sudeste do Brasil

A survey of Turkey Coronavirus (TCoV) and Astrovirus (TAstV-2) prevalence was carried out from February to December during 2006 year in semiarid region of Brazil, from a turkey producer area, localized in South Eastern of Brazil. To asses the risk factor related to clinical material, climatic condition and type of RT-PCR applied, cloacal swabs (CS), faeces, sera, bursa of Fabricius (BF), thymus (TH) and spleen (SP) and ileum-caeca region were collected from 30-day-old poults suffering of enteritis episode characterized as poult enteritis mortality syndrome (PEMS). The PEMS clinical features were characterized by watery to foamy faeces, light brown-yellow in colour and low mortality rate. Meteorological data (rainfall and relative humidity) observed during along the study presented monthly average temperature ranging from 39.3 and 31.2ºC, precipitation in rainy season from 40 to 270.3 mm/month, and no rain during dry season. Simplex RT-PCR gave odds ratio (OR) values suggesting that ileum-caeca region is at higher chance (OR=1.9; p=0.9741) to have both viral RNA than faeces (OR=1.5; p=0.7319). However, multiplex RT-PCR showed 3.98 (p=0.89982) more chance to give positive results in faeces than CS at dry season. The major risk factors seem to be low rate of humidity and high temperatures at winter, probably responsible for spread, easily, the TCoV and TAstv-2 among the flocks. The positive results of both virus suggested that they can play an important role in enteric disorders, associated to low humidity and high temperatures frequently found in tropical countries.

O presente estudo foi conduzido para avaliar a prevalência do Coronavirus dos perus (TCoV) e Astrovirus tipo 2 (TAstV-2) entre os meses de Fevereiro a Dezembro de 2006, em uma região produtora localizada no semi-árido a Sudeste do Brasil. Os principais fatores de risco associado a prevalência foram material clínico analisado, condições climáticas e tipo de técnica molecular empregada. Os sinais clínicos foram caracterizados como intenso fluido intestinal e baixo crescimento em aves jovens, sendo o material coletado swabs cloacais, fezes, soros, bursa de Fabrícius, segmentos do intestino delgado, timo e baço. Os dados meteorológicos (índice pluviométrico e umidade relativa) desta região, durante o período de estudo, foram de temperatura média mensal variando de 39.3 a 31.2ºC, precipitação na época chuvosa variando de 40 a 270.3mm/mês e ausência de chuva na estação fria e seca. A técnica de simplex RT-PCR resultou em valores de odds ratio (OR) que sugerem que a região do intestino delgado (junção íleo-cecal) possui alta chance (1.9 vezes) de gerar resultados positivos na amplificação de RNA viral que as fezes (1.5 vezes) analisadas. A técnica de multiplex RT-PCR demonstrou ser 3.98 vezes mais eficiente em promover resultados positivos nas fezes que nos swabs cloacais, durante a época de inverno. Os maiores fatores de risco encontrados foram baixa umidade relativa associada a altas temperaturas, durante a estação seca, o que pode permitir uma maior disseminação aérea do ambos os vírus entre os lotes estudados. A alta prevalência detectada para dois vírus sugerem que, no Brasil, estes representam os maiores responsáveis pelos surtos de enterite viral nas regiões semi-áridas, associado a baixas umidades e altas temperaturas típicas de países tropicais.

Epidemiological aspects of astrovirus and coronavirus in poults in the South Eastern Region of Brazil.

A survey of Turkey Coronavirus (TCoV) and Astrovirus (TAstV-2) prevalence was carried out from February to December during 2006 year in semiarid region of Brazil, from a turkey producer area, localized in South Eastern of Brazil. To asses the risk factor related to clinical material, climatic condition and type of RT-PCR applied, cloacal swabs (CS), faeces, sera, bursa of Fabricius (BF), thymus (TH) and spleen (SP) and ileum-caeca region were collected from 30-day-old poults suffering of enteritis episode characterized as poult enteritis mortality syndrome (PEMS). The PEMS clinical features were characterized by watery to foamy faeces, light brown-yellow in colour and low mortality rate. Meteorological data (rainfall and relative humidity) observed during along the study presented monthly average temperature ranging from 39.3 and 31.2ºC, precipitation in rainy season from 40 to 270.3 mm/month, and no rain during dry season. Simplex RT-PCR gave odds ratio (OR) values suggesting that ileum-caeca region is at higher chance (OR=1.9; p=0.9741) to have both viral RNA than faeces (OR=1.5; p=0.7319). However, multiplex RT-PCR showed 3.98 (p=0.89982) more chance to give positive results in faeces than CS at dry season. The major risk factors seem to be low rate of humidity and high temperatures at winter, probably responsible for spread, easily, the TCoV and TAstv-2 among the flocks. The positive results of both virus suggested that they can play an important role in enteric disorders, associated to low humidity and high temperatures frequently found in tropical countries.

Brain and lung cryptococcoma and concurrent corynebacterium pseudotuberculosis infection in a goat: a case report

A four-year-old male goat with a history of neurological disorder was euthanized. It presented uncommon nodules in the brain and lungs associated with multiple abscesses, predominantly in the spleen and liver. Histological examination of brain and lung sections revealed yeast forms confirmed to be Cryptococcus gattii after a combination of isolation and polymerase chain reaction (PCR) procedures. Moreover, Corynebacterium pseudotuberculosis infection was diagnosed by PCR of samples from the lung, spleen and liver. The present report highlights the rare concurrent infection of C. gatti and C. pseudotuberculosis in an adult goat from São Paulo state, Brazil, and indicates the necessity of surveillance in the treatment of goats with atypical pulmonary infections associated with neurological disorders.(AU)

Experimental infectious bursal disease in the ostrich (Struthio camelus).

Clinically severe disease was produced in ostriches aged 4 weeks by oral infection with a virulent strain of infectious bursal disease virus (vIBDV), namely strain Faragher 52/70. Four days after infection the birds were humanely killed and tissue samples, including thymus, bursa of Fabricius (BF), brain and kidney were collected for examination. Histopathologically, the thymus and BF showed severe lymphoid depletion and necrosis, while immunolabelling with a polyclonal antibody demonstrated abundant viral antigen.

Detection of bovine Herpesvirus type 5 in formalin-fixed, paraffin-embedded bovine brain by PCR: a useful adjunct to conventional tissue-based diagnostic test of bovine encephalitis.

The aim of this study was to evaluate the application of PCR technique for the detection of BoHV-5 in routinely formalin-fixed, paraffin-embedded brain tissues in 20 naturally infected calves affected by fatal meningoencephalitis. Brains were divided into two halves, one kept fresh for virus isolation and PCR assay, targeting the glycoprotein C gene from BoHV-5 genome. The other half brain, corresponding to posterior cortex region, was submitted to formalin fixation and embedded into paraffin blocks for microscopic evaluation and total DNA isolation. Most of the slides showed severe multifocal non-supurative encephalitis with neuronal degeneration, neurophagia, and no acidophilic intranuclear inclusions could be found in neurons and glial. The 20 fresh samples were confirmed, by virus isolation and PCR assay, as having the BoHV-5 virus and, respective glicoprotein C sequence, while 15 of 20 formalin-fixed, paraffin-embedded samples were considered positive for the same analysis. The results revealed the first description of PCR efficiency, applied to formalin-fixed, paraffin-embedded brain collected from naturally infected calves, improving the detection of BoHV-5 from archival samples in South America.