Molecular characterization of Brazilian equid herpesvirus type 1 strains based on neuropathogenicity markers.

Partial nucleotide sequences of ORF72 (glycoprotein D, gD), ORF64 (infected cell protein 4, ICP4) and ORF30 (DNA polymerase) genes were compared with corresponding sequences of EHV-1 reference strains to characterize the molecular variability of Brazilian strains. Virus isolation assays were applied to 74 samples including visceral tissue, total blood, cerebrospinal fluid (CSF) and nasal swabs of specimens from a total of 64 animals. Only one CSF sample (Iso07/05 strain) was positive by virus isolation in cell culture. EHV-1 Iso07/05 neurologic strain and two abortion visceral tissues samples (Iso11/06 and Iso33/06) were PCR-positive for ORF33 (glycoprotein B, gB) gene of EHV-1. A sequence analysis of the ORF72, ORF64 and ORF30 genes from three EHV-1 archival strains (A3/97, A4/72, A9/92) and three clinical samples (Iso07/05, Iso11/06 and Iso33/06) suggested that among Brazilian EHV-1 strains, the amplified region of the gD gene sequence is highly conserved. Additionally, the analysis of ICP4 gene showed high nucleotide and amino acid identities when compared with genotype P strains, suggesting that the EHV-1 Brazilian strains belonged to the same group. All the EHV-1 Brazilian strains were classified as non-neuropathogenic variants (N752) based on the ORF30 analysis. These findings indicate a high conservation of the gD-, ICP4- and ORF30-encoding sequences. Different pathotypes of the EHV-1 strain might share identical genes with no specific markers, and tissue tropism is not completely dependent on the gD envelope, immediate-early ICP4 and DNA polymerase proteins.

Molecular characterization of Brazilian equid herpesvirus type 1 strains based on neuropathogenicity markers

Partial nucleotide sequences of ORF72 (glycoprotein D, gD), ORF64 (infected cell protein 4, ICP4) and ORF30 (DNA polymerase) genes were compared with corresponding sequences of EHV-1 reference strains to characterize the molecular variability of Brazilian strains. Virus isolation assays were applied to 74 samples including visceral tissue, total blood, cerebrospinal fluid (CSF) and nasal swabs of specimens from a total of 64 animals. Only one CSF sample (Iso07/05 strain) was positive by virus isolation in cell culture. EHV-1 Iso07/05 neurologic strain and two abortion visceral tissues samples (Iso11/06 and Iso33/06) were PCR-positive for ORF33 (glycoprotein B, gB) gene of EHV-1. A sequence analysis of the ORF72, ORF64 and ORF30 genes from three EHV-1 archival strains (A3/97, A4/72, A9/92) and three clinical samples (Iso07/05, Iso11/06 and Iso33/06) suggested that among Brazilian EHV-1 strains, the amplified region of the gD gene sequence is highly conserved. Additionally, the analysis of ICP4 gene showed high nucleotide and amino acid identities when compared with genotype P strains, suggesting that the EHV-1 Brazilian strains belonged to the same group. All the EHV-1 Brazilian strains were classified as non-neuropathogenic variants (N752) based on the ORF30 analysis. These findings indicate a high conservation of the gD-, ICP4- and ORF30-encoding sequences. Different pathotypes of the EHV-1 strain might share identical genes with no specific markers, and tissue tropism is not completely dependent on the gD envelope, immediate-early ICP4 and DNA polymerase proteins.

Phylogenetic analysis of rabies virus isolated from herbivores in Minas Gerais and São Paulo border (2000-2009), Brazil / Análise filogenética de isolados do vírus da raiva de herbívoros na fronteira de Minas Gerais e São Paulo (2000-2009), Brasil

Rabies transmitted by the hematophagous bat Desmodus rotundus represents a public health concern and a burden for the Brazilian livestock industry. Current evidence suggests that rabies occurrence is related to landscape characteristics, topography, hydrography, animal production systems and land use...

A raiva transmitida por morcegos hematófagos da espécie Desmodus rotundus representa uma preocupação de saúde pública e causa de importantes prejuízos para a pecuária brasileira. A evidência atual sugere que a ocorrência de raiva está relacionada às características da paisagem, topografia, hidrografia, sistemas de produção animal e usos da terra...

Pathogenicity of different rabies virus isolates and protection test in vaccinated mice.

This study was aimed to evaluate and compare the pathogenicity of rabies virus isolated from bats and dogs, and to verify the efficacy of a commercial rabies vaccine against these isolates. For evaluation of pathogenicity, mice were inoculated by the intramuscular route (IM) with 500MICLD50/0.03 mL of the viruses. The cross-protection test was performed by vaccinating groups of mice by the subcutaneous route and challenged through the intracerebral (IC) route. Isolates were fully pathogenic when inoculated by the IC route. When inoculated intramuscularly, the pathogenicity observed showed different death rates: 60.0% for the Desmodus rotundus isolate; 50.0% for dog and Nyctinomops laticaudatus isolates; 40.0% for Artibeus lituratus isolate; 9.5% Molossus molossus isolate; and 5.2% for the Eptesicus furinalis isolate. Mice receiving two doses of the vaccine and challenged by the IC route with the isolates were fully protected. Mice receiving only one dose of vaccine were partially protected against the dog isolate. The isolates from bats were pathogenic by the IC route in mice. However, when inoculated through the intramuscular route, the same isolates were found with different degrees of pathogenicity. The results of this work suggest that a commercial vaccine protects mice from infection with bat rabies virus isolates, in addition to a canine rabies virus isolate.

Pathogenicity of different rabies virus isolates and protection test in vaccinated mice / Patogenicidade de diferentes isolados do vírus da raiva e teste de proteção em camundongos vacinados

This study was aimed to evaluate and compare the pathogenicity of rabies virus isolated from bats and dogs, and to verify the efficacy of a commercial rabies vaccine against these isolates. For evaluation of pathogenicity, mice were inoculated by the intramuscular route (IM) with 500MICLD50/0.03mL of the viruses. The cross-protection test was performed by vaccinating groups of mice by the subcutaneous route and challenged through the intracerebral (IC) route. Isolates were fully pathogenic when inoculated by the IC route. When inoculated intramuscularly, the pathogenicity observed showed different death rates: 60.0 percent for the Desmodus rotundus isolate; 50.0 percent for dog and Nyctinomops laticaudatus isolates; 40.0 percent for Artibeus lituratus isolate; 9.5 percent Molossus molossus isolate; and 5.2 percent for the Eptesicus furinalis isolate. Mice receiving two doses of the vaccine and challenged by the IC route with the isolates were fully protected. Mice receiving only one dose of vaccine were partially protected against the dog isolate. The isolates from bats were pathogenic by the IC route in mice. However, when inoculated through the intramuscular route, the same isolates were found with different degrees of pathogenicity. The results of this work suggest that a commercial vaccine protects mice from infection with bat rabies virus isolates, in addition to a canine rabies virus isolate.

O estudo avaliou e comparou as propriedades patogênicas de cinco isolados do vírus da raiva de morcegos e um isolado do vírus da raiva de cão e analisou a eficácia de vacina comercial contra estes isolados, em camundongos. Para o estudo de patogenicidade camundongos foram inoculados pela via IM com 0,1 mL contendo 500MICLD50/0,03mL das amostras de vírus. Quando inoculados pela via IC, os isolados do vírus da raiva provocaram a morte de 100 por cento dos camundongos. No entanto, 500MICLD50/0,03mL das mesmas amostras, inoculadas pela via IM, ocasionaram mortalidade de: 60,0 por cento quando a amostra era de Desmodus rotundus; 50,0 por cento de cão e de Nyctinomops laticaudatus; 40,0 por cento de Artibeus lituratus; 9,5 por cento de Molossus molossus; e 5,2 por cento de Eptesicus furinalis. Camundongos que receberam duas doses de vacina foram protegidos quando desafiados pela via IC, com todas as amostras testadas. Quando os camundongos receberam uma dose da mesma vacina, houve proteção parcial daqueles desafiados com a amostra de cão. Todos os isolados do vírus da raiva testados foram patogênicos para camundongos, inoculados pela IC. No entanto, pela via IM, os mesmos isolados mostraram diferentes graus de patogenicidade. Concluiu-se também que a vacina comercial contra raiva protegeu os camundongos desafiados com amostras de vírus isolados de morcegos e de cão.

Low genetic diversities of rabies virus populations within different hosts in Brazil.

The low rates of nonsynonymous evolution observed in natural rabies virus (RABV) isolates are suggested to have arisen in association with the structural and functional constraints operating on the virus protein and the infection strategies employed by RABV within infected hosts to avoid strong selection by the immune response. In order to investigate the relationship between the genetic characteristics of RABV populations within hosts and the virus evolution, the present study examined the genetic heterogeneities of RABV populations within naturally infected dogs and foxes in Brazil, as well as those of bat RABV populations that were passaged once in suckling mice. Sequence analyses of complete RABV glycoprotein (G) genes showed that RABV populations within infected hosts were genetically highly homogeneous whether they were infected naturally or experimentally (nucleotide diversities of 0-0.95x10(-3)). In addition, amino acid mutations were randomly distributed over the entire region of the G protein, and the nonsynonymous/synonymous rate ratios (d(N)/d(S)) for the G protein gene were less than 1. These findings suggest that the low genetic diversities of RABV populations within hosts reflect the stabilizing selection operating on the virus, the infection strategies of the virus, and eventually, the evolutionary patterns of the virus.

[Isolation of rabies virus in Artibeus fimbriatus bat in the State of Sao Paulo, Brazil]. / Isolamento do vírus da raiva em Artibeus fimbriatus no Estado de São Paulo.

This is the first report of the isolation and identification of the rabies virus in the frugivorous bat Artibeus fimbriatus in the city of Sao José do Rio Preto, Sao Paulo State, Brazil. The virus was isolated from an animal found in an urban area. The animal was found on the ground under a tree, still alive. Diagnosis was made by direct immunofluorescence and intracerebral inoculation of mice.

Molecular epidemiological analysis of bat rabies viruses in Brazil.

A molecular epidemiological analysis was performed in 19 rabies viruses (RVs) isolated from haematophagous, frugivorous and insectivorous bats, in Sao Paulo, Brazil. The authors carried out RT-PCR for amplification of the RV nucleoprotein (N) gene, and determined 1,335 nucleotide sequences of N gene by direct sequencing method. Phylogenetic analysis, which was based on the N gene of Brazilian RV isolates identified presently and previously, revealed that RVs isolated from bats were genetically divided into four lineages had a tendency to depend on the host bat species. The first lineage consisted mainly of haematophagous bat (Desmodus rotundus) isolates, including frugivorous bat (Artibeus spp.) isolates. Other three lineages consisted of insectivorous bat isolates; mainly Eptesicus spp., Molossus spp. and Nyctinomops spp. isolates, respectively. These results indicate a possibility of that there are bat species-specific RV variants in Brazil.

Isolamento do vírus da raiva em Artibeus fimbriatus no Estado de São Paulo / Isolation of rabies virus in Artibeus fimbriatus bat in the State of Sao Paulo, Brazil

Descrevem-se, pela primeira vez, o isolamento e a identificação do vírus da raiva em morcego frugívoro Artibeus fimbriatus no município de São José do Rio Preto, Estado de São Paulo. O vírus foi isolado de exemplar encontrado em área urbana, caído sob uma árvore e ainda vivo. O diagnóstico foi realizado pelas técnicas de imunofluorescência direta e inoculação intracerebral em camundongos.

Genetic characterization of rabies viruses isolated from frugivorous bat (Artibeus spp.) in Brazil.

In Latin America, rabies cases related to frugivorous bats have been reported since 1930's. Recently, two viruses isolated from Artibeus lituratus were proved to be vampire bat variants by monoclonal antibodies panels [2], but their genetic information is not well known. In this report, four rabies viruses were isolated from frugivorous bats (Artibeus spp.) in Brazil and their nucleoprotein gene sequences were determined. These isolates were found to be genotype 1 of lyssavirus and showed the maximum nucleotide sequence homology of 97.6-99.4% with vampire bat-related viruses in Brazil [6]. These results indicate that the Brazilian frugivorous bat rabies viruses in this study are closely related to vampire bat-related viruses that play a main role in rabies virus transmission to livestock in Brazil.

Rabies laboratory diagnosis: peculiar features of samples from equine origin

Rabies laboratory diagnosis is performed by using microscopic examination for Negri bodies (MEN), fluorescent-antibody test (FAT) and mouse inoculation test (MIT). In the majority of cases, when specimens are properly collected and conserved and the laboratory worker has good experience, agreement among employed techniques is verified. Comparing the sensitivity of these three diagnosis techniques in 3,713 samples (hippocampus and brain stem) received during 1981-1994 period, being 3,010 from bovine (983 positives) and 703 from equine (111 positives) species, it was observed that in equine rabid samples, this agreement was not mantained. For the latter specie, only in few opportunities the Negri bodies could be observed. With respect to FAT, the test detected a lower porcentage of positive equine samples compared to bovie species. Statistical analysis demonstrated that the difference was significative. Mouse inoculation test proved to be more sensitive. However, a significant difference in mice incubation period was observed for samples from both species. The absence of inclusion bodies and the longer incubation period for equine samples suggest that rabies pathogenesis studies for equine species have to be intensified.

Isolamento do vírus rábico em Molossus ater (Chiroptera: Molossidae) no Estado de Säo Paulo / Rabies in insectivorous bat - Molossus ater in S. Paulo State, Brazil

Descreve-se o isolamento e a identificaçäo do vírus rábico em morcegos insetívoros Molossus ater, no Estado de Säo Paulo, nos municípios de Araçatuba, Penápolis e Säo José do Rio Preto. A maioria dos exemplares foi capturada ainda com vida, näo havendo, porém, contato com pessoas ou animais. O diagnóstico foi realizado pelas provas de imunofluorescência direta e inoculaçäo intracerebral em camundongos

A açäo programática na visäo dos profissionais de saúde em uma Unidade Básica no Município do Rio de Janeiro / Health profissionals' social representations of a Municipal Health Center in Rio de Janeiro city, Brazil

Tem por objetivo compreender as representações sociais de profissionais de saúde de um Centro Municipal de Saúde (CMS) do Rio de Janeiro em relação a educação em saúde. Foram realizadas doze entrevistas semi-estruturadas com profissionais sobre: trajetória profissional, visão do cargo ocupado, desempenho dos programas executados e prioridades, visão do modelo atual de saúde, metas e objetivos em relação as atividades desenvolvidas. Conclui que: as modificações legais que culminaram com a implantação do Sistema Único de Saúde, onde os CMS teriam sua relevância reconhecida, não alteraram o sentimento de desvalorização por parte dos entrevistados tanto em relação às atividades desenvolvidas quanto da saúde pública em geral. A falta de consenso em relação à percepção de saúde e de educação em saúde, bem como a precária integração entre as equipes do CMS, interferiram negativamente na atuação daquela unidade