Verification of the rabies virus glycoprotein lower limit of immunogenicity by serological assay.

Rabies lethality is close to 100% and annually 15 million people receive post-exposure prophylaxis. Testing for vaccines against this zoonosis should ensure its quality. A standardized test by the National Institutes of Health (NIH) test, based on mice immunization and challenge, has been used to determine the potency of vaccine lots. It has several disadvantages, the main one being its significant variability. Several in vitro methods like an Enzyme-linked immunosorbent assay (ELISA) have been proposed based on the quality and quantity of glycoprotein (Glptn) of rabies virus, but may also present limitations such as low sensitivity, instability and imprecision. The estimate of immunogenicity based on neutralizing antibody titer (Nab) evaluated by a serological test (ST) such as the Modified Rapid Fluorescent Focus Inhibition Test (mRFFIT), is not yet efectively applied for human vaccine. Nevertheless, a Nab concentration can be used as a predictor of clinical efficacy of this product in vaccinated humans, so, that can be applied in estimating the vaccine potency. The aim of this study was to verify the lower limit of immunogenicity of the viral Glptn content in mice using mRFFIT. The lower Glptn content by ELISA able to induce Nab response was determined. The results were correlated and demonstrated that ST was able to determine the Glptn immunogenicity lower limit. Our findings suggest that a test based on rabies Nabs may represent an additional alternative for the evaluation of rabies vaccines.

Development and pre-validation of a quantitative multi-dose serological assay for potency testing of inactivated rabies vaccines for human use.

It is mandatory to ensure the quality of biological products used in the prevention of rabies, a zoonosis with nearly 100% lethality. Fifteen million people receive post-exposure prophylaxis yearly. The vaccine batches are assessed by the National Institutes of Health (NIH) test which has several disadvantages such as significant variability and animal welfare issues. The estimation of immunogenicity based on titration of neutralizing antibodies (NA) is not applied to the human vaccine yet. Despite this, a satisfactory concentration of NA (0.5 IU/ml) can be used as a predictor of the clinical efficacy and for estimating rabies vaccine potency. The objective of this study was to develop and pre-validate a Serological Potency Test (SPT) using the modified Rapid Fluorescent Focus Inhibition Test (mRFFIT) to determine the potency of rabies vaccines for human use, demonstrating its relevance and reliability. The results show good agreement between the potencies determined by the SPT and the NIH test. The assay was able to distinguish between potent and sub-potent lots of vaccines. The results demonstrated that SPT is a viable candidate for validation and inclusion in pharmacopeias as a reduction and refinement for the NIH test.

Circulation of the rabies virus in non-hematophagous bats in the city of Rio de Janeiro, Brazil, during 2001-2010.

INTRODUCTION: Rabies is one of the most known lethal zoonosis, responsible for 55,000 human deaths per year. It is transmitted to humans mainly by the bite of domestic or wild animals infected with the virus. This paper shows the circulation of this virus in non-hematophagous bats in the City of Rio de Janeiro, Brazil. METHODS: A survey was performed on the number of bats that had been sent for diagnosis by the Seção de Virologia of the Instituto Municipal de Medicina Veterinária Jorge Vaitsman and were positive for rabies. The positive animals were identified, and the isolated viruses were sent for antigenic typification with indirect immunofluorescence. The results were compared with the antigenic panel of the Centers for Disease Control and Prevention. RESULTS: During 2001-2010, the laboratory received 555 non-hematophagous bats for rabies diagnosis, with 198 (35.7%) from Rio de Janeiro City. A total of 11 (5.5%) animals were positive for this disease. Antigenic typification revealed the predominance of variant 3 in 9 (81.8%) of the isolated viruses; 1 virus was classified as variant 4 and 1 variant was identified that segregated with the viruses in insectivorous bats. CONCLUSIONS: The data obtained in this study showed the presence of the rabies virus in synanthropic populations of non-hematophagous bats in the City of Rio de Janeiro. The circulation of this agent in these animals represents a serious risk to human and animal health and requires attention and control measures by the authorities.

Circulation of the rabies virus in non-hematophagous bats in the city of Rio de Janeiro, Brazil, during 2001-2010 / Circulação do vírus da raiva em morcegos não-hematófagos na cidade do Rio de Janeiro no período entre 2001-2010

INTRODUCTION: Rabies is one of the most known lethal zoonosis, responsible for 55,000 human deaths per year. It is transmitted to humans mainly by the bite of domestic or wild animals infected with the virus. This paper shows the circulation of this virus in non-hematophagous bats in the City of Rio de Janeiro, Brazil. METHODS: A survey was performed on the number of bats that had been sent for diagnosis by the Seção de Virologia of the Instituto Municipal de Medicina Veterinária Jorge Vaitsman and were positive for rabies. The positive animals were identified, and the isolated viruses were sent for antigenic typification with indirect immunofluorescence. The results were compared with the antigenic panel of the Centers for Disease Control and Prevention. RESULTS: During 2001-2010, the laboratory received 555 non-hematophagous bats for rabies diagnosis, with 198 (35.7%) from Rio de Janeiro City. A total of 11 (5.5%) animals were positive for this disease. Antigenic typification revealed the predominance of variant 3 in 9 (81.8%) of the isolated viruses; 1 virus was classified as variant 4 and 1 variant was identified that segregated with the viruses in insectivorous bats. CONCLUSIONS: The data obtained in this study showed the presence of the rabies virus in synanthropic populations of non-hematophagous bats in the City of Rio de Janeiro. The circulation of this agent in these animals represents a serious risk to human and animal health and requires attention and control measures by the authorities.

INTRODUÇÃO: A raiva é uma das mais letais zoonoses conhecidas, responsável pelo óbito de 55 mil pessoas anualmente. É transmitida ao homem principalmente pela mordida de animais, domésticos ou silvestres, infectados pelo vírus. O presente estudo mostra a circulação deste agente em morcegos não-hematófagos no município do Rio de Janeiro, Brasil. MÉTODOS: Na Seção de Virologia do Instituto Municipal de Medicina Veterinária Jorge Vaitsman, foi realizado um levantamento do número de morcegos recebidos positivos para o diagnóstico. Os animais positivos foram identificados, e o vírus isolado foi submetido à tipificação de variantes antigênicas pela técnica de imunofluorescência indireta, com os resultados comparados com o painel antigênico utilizado pelo Centro de Controle de Doenças (Center of Disease Control -CDC). RESULTADOS: Entre 2001-2010, o laboratório recebeu 555 morcegos nãohematófagos para o diagnóstico da raiva, sendo 135 (35,5%) do município do Rio de Janeiro. Um total de 11 (5,5%) animais foram positivos para a doença. A tipificação antigênica revelou a predominância da variante 3, com nove (81,9%) vírus isolados, um pertencente a variante 4 e outro de uma variante que segrega com a presente em morcegos insetívoros. CONCLUSÕES: Os dados observados no presente estudo demonstraram a presença do vírus da raiva em populações sinantrópicas de morcegos não-hematófagos no município do Rio de Janeiro. A circulação deste agente nesses animais representa um grave risco a saúde humana e animal, e requer atenção e medidas de controle e prevenção por parte das autoridades competentes.

Comparação entre conjugados in house e comerciais pela técnica de rápida inibição de focos fluorescentes (RFFIT) na avaliação de anticorpos antirrábicos / Comparison of in house and commercial conjugates by rapid fluorescent focus inhibition test (RFFIT) in rabies antibodies evaluation

A raiva é uma zoonose letal transmitida ao homem pela inoculação do vírus rábico, principalmente pela mordedura de animais infectados. Em 2005, o Ministério da Saúde brasileiro gastou cerca de R$66 milhões com ações de vigilância epidemiológica, empregados em campanha de vacinação e na aquisição de imunobiológicos. O controle sorológico é exigência básica para a correta avaliação da pessoa vacinada. Neste trabalho, 91 soros de 34 indivíduos foram submetidos à titulação de anticorpos pela técnica de rápida inibição de focos fluorescentes, adaptada a microplacas de 96 poços, para comparar um conjugado produzido in house com outro comercial. Do total, 74 soros (82,2por cento) apresentaram título maior ou igual a 0,5UI/mL e 12 soros (13,33por cento), título menor que 0,5UI/mL. Estes resultados mostram que a rápida inibição de focos fluorescentes utilizando o conjugado produzido in house foi tão sensível quanto com o conjugado comercial. As diferenças entre os conjugados não foram significativas e os títulos de anticorpos apresentaram elevada correlação (r = 0,94).

[Rabies virus in Nyctinomops laticaudatus bats in the City of Rio de Janeiro: isolation, titration and epidemiology]. / Vírus rábico em morcego Nyctinomops laticaudatus na Cidade do Rio de Janeiro, RJ: isolamento, titulação e epidemiologia.

The first case report of rabies in bats of the species Nyctinomops laticaudatus, in the city of Rio de Janeiro City, is presented. Virus isolation and titration were performed in different tissues, and high titers were found in the brain and salivary glands. Rabies occurrence in such an infrequent species in this state suggests that the disease may be more prevalent than it appears to be.

Vírus rábico em morcego Nyctinomops laticaudatus na Cidade do Rio de Janeiro, RJ: isolamento, titulação e epidemiologia / Rabies virus in Nyctinomops laticaudatus bats in the City of Rio de Janeiro: isolation, titration and epidemiology

Apresenta-se o primeiro relato de raiva em morcego da espécie Nyctinomops laticaudatus, na Cidade do Rio de Janeiro, RJ. Foram realizados isolamento e titulação viral em diferentes tecidos, encontrando-se altos títulos no cérebro e glândulas salivares. A ocorrência de raiva em uma espécie pouco freqüente neste estado sugere que a doença pode ser mais prevalente do que aparenta.

The first case report of rabies in bats of the species Nyctinomops laticaudatus, in the city of Rio de Janeiro City, is presented. Virus isolation and titration were performed in different tissues, and high titers were found in the brain and salivary glands. Rabies occurrence in such an infrequent species in this state suggests that the disease may be more prevalent than it appears to be.