Seroepidemiological survey to cell culture rabies vaccines (CCRV) in Brazil.

Rabies is a contagious viral disease that can be easily transmitted by the saliva and brain/nervous system tissues of the infected animals, causing severe and fatal encephalitis in both animals and humans. Vaccination campaigns are crucial to combat and prevent rabies's spread in dogs and humans. The Modified Fuenzalida & Palicios vaccines have been widely used since the 70s and have proven effective in producing a solid serological response. Since 2008, the Brazilian Ministry of Health has introduced a Cell Culture Rabies Vaccine (CCRV) for all dog mass vaccination campaigns in Brazil. However, to date, there is limited evidence on the immunologic response of dogs to this type of vaccine in field conditions. The present study evaluated the serological response in dogs vaccinated with CCRV from blood samples of 724 dogs using the Simplified Fluorescence Inhibition Microtest - SFIMT. Dogs with a titer equal to 0.5 IU/mL or above were considered seropositive. The results revealed that 59.12% (428/724) of all dogs tested and 48.49% (32/66) of primo-vaccinated animals were seropositive. The percentage of seronegative animals was higher than seropositive for animals that received a single dose during their life (p < 0.05). The opposite was observed in animals with five or more doses. The results of this study demonstrated that the CCRV vaccines elicit a satisfactory immunological response in field conditions and can constitute an essential population-level preventive strategy as part of annual canine rabies vaccination campaigns. Although its effectiveness has been studied, there is limited evidence of its immunological response in dogs under field conditions. This paper evaluates the serological response to CCRV in dogs vaccinated during mass vaccination campaigns from 2012 to 2017.

Rabies virus isolated from insectivorous bats induces different inflammatory responses in experimental model.

Rabies virus (RABV) is a neurotropic virus that causes fatal neuroinflammation in mammals. The insectivorous bat RABV strains are less pathogenic for mice than strains associated with other reservoirs. We characterized the tissue inflammatory response in the CNS of RABV isolated from insectivorous bats. Eptesicus furinalis (EPBRV)-infected mice had a robust inflammatory response and a greater amount of IL-1ß, IL-6 and TNF-α, while Myotis nigricans (MNBRV)-infected mice showed a higher expression of IL-17 and greater activation of IFN-ß. New approaches to understand the inflammatory response to different mechanisms of action may provide insights for the development of novel therapies for rabies.

Use of histamine-2 receptor antagonists and risk of inflammatory bowel diseases: A systematic review and meta-analysis of observational studies.

WHAT IS KNOWN AND OBJECTIVE: The effect of histamine-2 receptor antagonists (H2RAs) use causing inflammatory bowel diseases (IBD) has been reported in few isolated observational studies; however, pooled estimation of IBD risk has not been done. The present study was conducted to estimate the risk of IBD [Crohn's disease (CD), ulcerative colitis (UC) and microscopic colitis (MC)], among H2RAs users. METHODS: Databases such as MEDLINE/PubMed, Scopus and Cochrane Library were searched from inception to January 2021. A bibliographic search of selected articles, random search in Google Scholar and ResearchGate were also performed for any additional studies. The observational studies which assessed the incidence or risk of IBD in H2RA users published in the English language were considered. Modified Downs and Black Checklist was used for quality assessment. Two independent reviewers were involved in study selection, data extraction and quality assessment; any discrepancies were settled through consensus or by consulting a third reviewer. RESULTS: Four studies out of 2,658 articles were included for this meta-analysis. The meta-analysis of 4 studies with 8939 participants revealed a significantly higher risk of IBD (OR: 2.27; 95% CI: 1.70-3.02; p < 0.0001) in H2RA users compared to non-users. Similar significant relationships were observed in the subgroup analysis of adults (p < 0.0001) and paediatrics (p = 0.04). The quality of included studies was observed to be fair to good. WHAT IS NEW AND CONCLUSION: Our findings indicate a significantly higher IBD risk among those who used H2RA compared to non-users both in adults and in paediatrics. Further observational studies involving large populations are required to strengthen these results and to generalize these findings.

A comparative review of serological assays for the detection of rabies virus-specific antibodies.

Rabies is a major public health problem with a fatality rate close to 100%, caused by a virus of the Lyssavirus genus, of which rabies virus (RABV) is the prototype. Nonetheless, the complete prevention can be achieved by the induction of neutralizing antibodies by pre- or post-exposure prophylaxis. According to the world health organization (WHO) and World Organization for animal health (OIE), serum titers of rabies virus neutralizing antibodies (RVNA) that are higher or equal to 0.5 international units (IU)/ml indicate adequate immune response after vaccination against rabies. Currently, RFFIT and FAVN are the gold standard tests recommended by both WHO and OIE for detecting and quantitating RVNA in biological samples from individuals or animals previously vaccinated and/or subjects suspected of having been infected by RABV. Although the tests RFFIT and FAVN are efficient, they are time-consuming, labor-intensive manual tests and not cost-effective for routine use. Following the previously mentioned, approaches with alternative methods have been developed to detect RVNA or rabies-specific antibodies in human or animal serum, but with variable success. This work summarizes the advances in the serological assays for the detection of neutralizing antibodies or rabies antibodies and assesses the individual immune status after vaccination against rabies, as well as the mechanisms of RABV neutralization mediated by antibodies. Therefore, the main alternative methods for the determination of RABV or rabies-specific antibodies are exposed, with promising results, besides being easy to execute, of low cost, and representing a possibility of being applied, according to the proposal of each test to the network of Rabies Surveillance Laboratories.

Rabies virus isolated from insectivorous bats induces different inflammatory responses in experimental model

Rabies virus (RABV) is a neurotropic virus that causes fatal neuroinflammation in mammals. The insectivorous bat RABV strains are less pathogenic for mice than strains associated with other reservoirs. We characterized the tissue inflammatory response in the CNS of RABV isolated from insectivorous bats. Eptesicus furinalis (EPBRV)-infected mice had a robust inflammatory response and a greater amount of IL-1β, IL-6 and TNF-α, while Myotis nigricans (MNBRV)-infected mice showed a higher expression of IL-17 and greater activation of IFN-β. New approaches to understand the inflammatory response to different mechanisms of action may provide insights for the development of novel therapies for rabies.

Rabies Virus Exposure in Wild Lowland Tapirs (Tapirus terrestris) from Three Brazilian Biomes.

We evaluated the presence of antibodies for rabies virus in 177 serum samples from 125 wild lowland tapirs (Tapirus terrestris) from three different Brazilian biomes. The rapid fluorescent focus inhibition test was performed. No antibody titers suggesting the circulation of the rabies virus in tapir habitat were detected.

Glycosylation is required for the neutralizing activity of human IgG1 antibodies against human rabies induced by pre-exposure prophylaxis.

Rabies lyssavirus (RABV) neutralizing IgG antibodies confer protection after rabies vaccination, although how the RABV-specific antibodies neutralize the virus is still unknown. As changes in the antibody's carbohydrate chain can interfere with its effector functions, we compared the glycosylation patterns of both neutralizing and non-neutralizing IgG1 induced by pre-exposure prophylaxis to human rabies and analyzed their influence on in vitro antibody neutralizing activities. Specific IgG1 were purified from human serum using affinity chromatography. Purity and avidity were analyzed by SDS-PAGE and indirect ELISA using NH4SCN respectively. The N-linked oligosaccharide chain of the purified IgG antibody was evaluated using a lectin-based ELISA assay with a panel of seven lectins. The activity of purified IgG1 and neutralizing IgG1 deglycosylated by PNGase F enzyme were analyzed using the rapid fluorescent focus inhibition test. The purified IgG1 showed an electrophoretic pattern compatible with human IgG. All of the antibodies recognized RABV, although neutralizing IgG1 had a higher avidity (RAI = 80%) than non-neutralizing IgG1 (RAI = 30%). The neutralizing IgG1 also showed higher binding to WFA, ECA, WGA, and ConA lectins, indicating possible different N-acetylgalactosamine, galactose, N-acetylglucosamine, and mannose contents. Non-neutralizing IgG1, on the other hand, showed strong binding at UEA-1 and SNA, which bind to fucose and sialic acid residues respectively. Different glycosylation profiles were also observed in Fab and Fc fragments from neutralizing and non-neutralizing IgG1, although the deglycosylated IgG1 lost its neutralizing activity. Our results suggest that antibody glycosylation is important for neutralizing RABV in vitro, since neutralizing IgG1 has a different glycosylation profile than non-neutralizing IgG1. Further research will be needed to better evaluate the differential glycosylation patterns between IgG1 antibodies following vaccination.

Mentoria de acolhimento para alunos ingressantes no curso de Medicina / Welcome mentoring for new medicine students

Resumo: Introdução: As mudanças de vida do estudante quando ingressa na Faculdade de Medicina tornam os primeiros anos os mais vulneráveis para desenvolver estresse psicológico e suas consequências. Em 2020, essa condição se agravou devido à pandemia de Sars-CoV-2. Para dar suporte aos alunos, várias escolas médicas desenvolvem programas de mentoria, entretanto a adesão deles costuma ser baixa. Objetivo: Este estudo teve como objetivos criar um modelo de mentoria - denominada "Mentoria de Acolhimento" - como suporte psicossocial e pedagógico aos alunos ingressantes, e estudar seu processo de implementação e os resultados para maior adesão e efetividade. Método: Com um desenho metodológico do campo da ciência da implementação, utilizaram-se técnicas e instrumentos de pesquisas qualitativas e quantitativas para compor um estudo de métodos mistos. Obtiveram-se os dados qualitativos em reuniões com os mentores e os dados quantitativos em questionários eletrônicos anônimos para os alunos. Os encontros dos grupos de mentoria foram previstos na grade curricular obrigatória, em horários predeterminados nos quatro meses do primeiro semestre da graduação. Analisaram-se os dados qualitativos por meio da técnica de análise temática, e os dados quantitativos foram examinados de modo descritivo. Resultado: Participaram da Mentoria de Acolhimento 147 alunos (77% dos ingressantes). Identificaram-se os seguintes facilitadores de implementação: inclusão automática de todos os alunos nos grupos, inserção na grade, qualidade do mentor e disposição dos discentes para a mentoria. Observaram-se as seguintes barreiras: falhas na comunicação com os alunos, não realização de encontros por mentores, atividades extracurriculares no horário previsto para a mentoria e o modo presencial remoto. A experiência foi avaliada como positiva por alunos e mentores, verificando-se ganhos de desempenho acadêmico, em conhecer o cotidiano escolar e lidar com aspectos emocionais e relacionais da vida de estudante. Conclusão: A Mentoria de Acolhimento ajudou a integração do aluno ingressante na faculdade. Mostrou-se adequada em termos de modelo e resultados, mas requer aprofundamento dos estudos de impacto na formação médica.

Abstract: Introduction: Upon entering medical school the student undergoes life changes that make the first years the most crucial in terms of vulnerability to developing psychological stress and the associated consequences. In 2020, this condition was aggravated due to the SARS-COV-2 pandemic. To offer students more support, many medical schools have encouraged them to participate in mentoring programs, however student adherence is typically low. Objective: To create a mentoring model - called "Welcome Mentoring" - as a psychosocial and pedagogical support for incoming students, and study its implementation process and results, with the aim of achieving greater adherence and effectiveness. Methodology: Using a methodological design from the field of implementation science, qualitative and quantitative research techniques and instruments were combined to form a mixed-methods study. Qualitative data were obtained in meetings with mentors, and quantitative data through anonymous electronic questionnaires for students. The meetings of the mentoring groups were foreseen in the mandatory curricular schedule, at predetermined times during the four months of the first semester of the undergraduate course. Qualitative data were analyzed with thematic analysis technique, and quantitative data were analyzed descriptively. Results: 147 students participated in the Welcome Mentoring (77% participation). The factors that facilitated the implementation were: the automatic inclusion of all students in the groups, insertion in the curriculum, quality of the mentor, and students' availability for mentoring. The following barriers were observed: communication failures with students, no meetings with mentors, extracurricular activities during mentoring hours, and the remote mode for meetings. The experience was evaluated as positive by students and mentors, with gains in academic performance, in learning about daily school life, and in dealing with the emotional and relational aspects of student life. Conclusion: Welcome Mentoring helped integrate incoming student into university life. It proved adequate in terms of the model and results, but requires further studies on its impact on medical education.

Experiences of Caregivers in a Home-Based Palliative Care Model - A Qualitative Study.

BACKGROUND: Family caregivers are of vital support to patients receiving home-based palliative care. AIMS AND OBJECTIVES: This study sought to identify and comprehend the challenges that caregivers face while taking care of a terminally ill patient in a home-based palliative care setting and the mechanisms that facilitated their coping. MATERIALS AND METHODS: A qualitative approach was employed to understand the perceptions of primary caregivers through 3 focus group discussions and 4 in-depth interviews, across 3 socioeconomic categories and 3 geographic zones of Mumbai. RESULTS: Caregivers expressed that they wished they had been introduced to palliative care earlier. Being trained on minor clinical procedures and managing symptoms, and receiving emotional support through counselling were found beneficial. Caregivers did not perceive the need for self-care as the period of active caregiving was often short. Bereavement counselling was felt to be of much help. CONCLUSION: The study helped understand the caregivers' perceptions about the factors that would help them in patient as well as self-care. Recommendations for designing interventions for future caregivers and recipients were also made.

Development of biotinylated polyclonal anti-ribonucleoprotein IgG for detection of rabies virus antigen by direct rapid immunohistochemical test.

The direct rapid immunohistochemical test (dRIT) has been recommended for laboratorial diagnosis of rabies, especially in developing countries. The absence of commercial primary antibodies, however, still represents a major limitation to its wider use in testing. We describe here the development of a biotinylated polyclonal antibody against Rabies lyssavirus (RABV) ribonucleoprotein (RNP) and its use as a primary reagent in dRIT. Anti-RNP polyclonal horse IgG was purified by ionic exchange chromatography followed by immunoaffinity column chromatography, and its affinity, diagnostic sensitivity, and specificity were evaluated. CNS samples (120) of suspected rabies cases in different animal species were tested by dRIT, with the positive (n = 14) and negative (n = 106) results confirmed by direct fluorescence antibody testing (dFAT). Comparing the results of dRIT and dFAT, we found that the biotinylated anti-RNP IgG delivered 100% diagnostic specificity and sensibility for rabies diagnosis. Our findings show that the biotinylated anti-RNP polyclonal IgG can be produced with the quality required for application in dRIT. This work represents an important step in efforts to diagnose rabies in developing countries.

Representações Sociais do Processo de Diagnóstico e Cura da Hanseníase / Social Representations of the Process of Diagnosis and Cure of Leprosy / Representaciones Sociales del Proceso de Diagnóstico y Curación de la Enfermedad de Hansen

Este estudo descritivo do tipo transversal objetivou identificar as representações sociais do processo de diagnóstico e cura da hanseníase em dois territórios com diferentes índices de detecção da doença. O software R possibilitou a identificação do núcleo da representação, sendo os elementos mais citados pelos participantes: Contagiosa, Cura, Doença de Pele, Lepra, Mancha, Tratamento. Conclui-se que as representações sociais que os indivíduos têm em relação à hanseníase, quando envoltas em estigmas e preconceitos sobre a doença, retardam a busca por ajuda médica, ocasionando atraso do correto diagnóstico e início do tratamento. Além disso, as incapacidades físicas decorrentes do diagnóstico tardio podem gerar comportamentos preconceituosos, que refletem o conhecimento popular arcaico e crendices arraigadas no que concerne ao contágio da doença. Tais comportamentos preconceituosos com relação à hanseníase estão na base de suas representações, levantando o isolamento dos doentes e a dificuldade relacionada ao diagnóstico, tratamento e cura.

This descriptive study of the transversal type aimed to identify the social representations of the process of diagnosis and cure of leprosy in two territories with different rates of detection of the disease. The software R allowed the identification of the nucleus of the representation, being the elements most cited by the participants: Contagious, Cure, Skin Disease, Leprosy, Spot, Treatment. It is concluded that the social representations that individuals have in relation to leprosy, when surrounded by stigmas and prejudices about the disease, delay the search for medical help, causing a delay of correct diagnosis and initiation of treatment. In addition, the physical disabilities resulting from late diagnosis can generate prejudiced behaviors, which reflect archaic popular knowledge and deep-rooted beliefs regarding the disease's contagion. Such prejudiced behaviors with respect to leprosy are at the basis of their representations that raise the isolation of patients and the difficulty related to diagnosis, treatment, and cure.

Este estudio descriptivo del tipo transversal objetivó identificar las representaciones sociales del proceso de diagnóstico y cura de la enfermedad de Hansen en dos territorios con diferentes índices de detección de la dolencia. El software R posibilitó la identificación del núcleo de la representación, siendo los elementos más citados por los participantes: Contagiosa, Cura, Enfermedad de Piel, Lepra, Mancha, Tratamiento. Se concluye que las representaciones sociales que los individuos poseen con relación a la enfermedad de Hansen, cuando envueltas en estigmas y prejuicios sobre la enfermedad, retardan la búsqueda por ayuda médica ocasionando retraso del correcto diagnóstico e inicio del tratamiento. Además, las incapacidades físicas derivadas del diagnóstico tardío pueden generar comportamientos preconcebidos, que reflejan el conocimiento popular arcaico y creencias arraigadas en lo que concierne al contagio de la enfermedad. Tales comportamientos predispuestos con relación a la lepra están en la base de sus representaciones, levantando el aislamiento de los pacientes y la dificultad relacionada al diagnóstico, tratamiento y curación.

Detection of rabies virus antigen by the indirect rapid immunohistochemistry test in equines and comparisons with other diagnostic techniques.

Laboratory diagnosis of rabies in equines is essential for distinguishing the disease from other sources of encephalitis. Diagnosis by conventional techniques such as a direct fluorescent antibody test (dFAT) or viral isolation in mice or cell culture can be difficult, and the application of molecular biological methods may be necessary. We performed an indirect rapid immunohistochemistry test (iRIT) for the detection of the rabies virus (RABV) antigen in the central nervous system (CNS) of equines and compared the results with those of other diagnostic techniques. We reviewed result records from the Rabies Diagnosis Laboratory at Instituto Pasteur, São Paulo, Brazil, of 174 samples of equine CNS from July 2014 to June 2016, which were investigated by dFAT, rabies tissue culture infection test (RTCIT), mouse inoculation test (MIT) and reverse transcription-polymerase chain reaction (RT-PCR) followed by genetic sequencing. These samples, 29 presented divergent results among techniques and were selected for the performed in the iRIT. The detected positivity rate was 4/29 (14%) by dFAT, 5/28 (18%) by RTCIT, 10/29 (35%) by MIT and 26/27 (96%) by RT-PCR. We analysed 29 samples through imprints of the cortex, hippocampus, cerebellum and brainstem in slides fixed in 10% buffered formaldehyde. Eighteen samples were identified as positive (62%) by iRIT assay, representing a greater number of positive cases than that detected by dFAT, MIT and RTCIT but not by RT-PCR. Among the brain regions, the brainstem presented the highest positivity (78%), followed by the hippocampus (69%), cerebellum (67%) and cortex (67%). Our results provide evidence that iRIT can contribute to a rapid diagnosis of rabies in equines and that complementary tests should be used to improve diagnostic accuracy in this species.

Performance evaluation of the polyclonal anti-rabies virus ribonucleoprotein IgG antibodies produced in-house for use in direct fluorescent antibody test.

Fluorescein isothiocyanate (FITC) labelled anti-rabies virus ribonucleoprotein (RNP) antibodies can be used as immunoreagents in direct fluorescent antibody testing (dFAT) for rabies diagnoses. While in-house products are occasionally used by laboratories, most conjugates are commercial reagents. Commercial anti-RNP antibodies are only available for research purposes in Brazil, however, which contributes to the increasing use of in-house produced antibodies. Considering that conjugate quality may influence the results obtained during rabies diagnosis, we sought to analyze the performance requirements of in-house produced polyclonal anti-RNP IgG-FITC for application in dFAT. To that end, their reproducibility, diagnostic sensitivity, and specificity were evaluated. The titer of polyclonal anti-RNP IgG-FITC was initially determined and evaluated by dFAT, using central nervous system (CNS) samples of different animal species (dogs, cats, bovines, equines, bats, and non-human primates). As our main result, the polyclonal anti-RNP IgG-FITC reached a titer of 1:30/1:40 in dFAT, with 100% of diagnostic sensitivity and specificity. In terms of reproducibility, the antibodies, regardless the production batch, presented the same performances. In conclusion, the in-house produced polyclonal anti-RNP IgG-FITC proved suitable for rabies virus antigen detection by dFAT.

Evaluation of polyclonal anti-RNP IgG antibody for rabies diagnosis by indirect rapid immunohistochemistry test.

Rabies still represents a major public health threat and estimated to cause 60,000 human deaths annually, particularly in developing countries. Thus, adequate surveillance based on rapid and reliable rabies diagnosis for both humans and animals is essential. The WHO and OIE recommended gold standard diagnostic technique for rabies is the direct immunofluorescence assay (dFAT). However, dFAT is expensive and requires a high level of expertise. As an alternative, the rapid immunohistochemistry technique is a promise to be a simple and cost effective diagnostic tool for rabies, and can be performed on field conditions prevalent in developing countries. However, no validated commercial conjugate antibody for rabies is available to meet the laboratory demand. Here, we evaluated the polyclonal anti-rabies virus ribonucleoprotein (RNP) IgG antibody for Rabies lyssavirus (RABV) detection by indirect rapid immunohistochemistry test (iRIT). We tested polyclonal anti-RNP IgG antibody against a batch of 100 brain specimens representing a wide phylogenetic origin in the State of São Paulo, Brazil. The purified IgG obtained 100% of diagnostic specificity and sensibility for RABV antigen detection in iRIT compared with the gold standard dFAT. In conclusion, our results demonstrate that the polyclonal anti-RNP IgG antibody may be used as a diagnostic reagent for rabies using iRIT, with the expectation of increase in availability and cost reduction of the epidemiological surveillance for developing countries.

Evaluation of polyclonal anti-RNP IgG antibody for rabies diagnosis by indirect rapid immunohistochemistry test

Rabies still represents a major public health threat and estimated to cause 60,000 human deaths annually, particularly in developing countries. Thus, adequate surveillance based on rapid and reliable rabies diagnosis for both humans and animals is essential. The WHO and OIE recommended gold standard diagnostic technique for rabies is the direct immunofluorescence assay (dFAT). However, dFAT is expensive and requires a high level of expertise. As an alternative, the rapid immunohistochemistry technique is a promise to be a simple and cost effective diagnostic tool for rabies, and can be performed on field conditions prevalent in developing countries. However, no validated commercial conjugate antibody for rabies is available to meet the laboratory demand. Here, we evaluated the polyclonal anti-rabies virus ribonucleoprotein (RNP) IgG antibody for Rabies lyssavirus (RABV) detection by indirect rapid immunohistochemistry test (iRIT). We tested polyclonal anti-RNP IgG antibody against a batch of 100 brain specimens representing a wide phylogenetic origin in the State of São Paulo, Brazil. The purified IgG obtained 100% of diagnostic specificity and sensibility for RABV antigen detection in iRIT compared with the gold standard dFAT. In conclusion, our results demonstrate that the polyclonal anti-RNP IgG antibody may be used as a diagnostic reagent for rabies using iRIT, with the expectation of increase in availability and cost reduction of the epidemiological surveillance for developing countries.

Evaluation of polyclonal anti-RNP IgG antibody for rabies diagnosis by indirect rapid immunohistochemistry test

Rabies still represents a major public health threat and estimated to cause 60,000 human deaths annually, particularly in developing countries. Thus, adequate surveillance based on rapid and reliable rabies diagnosis for both humans and animals is essential. The WHO and OIE recommended gold standard diagnostic technique for rabies is the direct immunofluorescence assay (dFAT). However, dFAT is expensive and requires a high level of expertise. As an alternative, the rapid immunohistochemistry technique is a promise to be a simple and cost effective diagnostic tool for rabies, and can be performed on field conditions prevalent in developing countries. However, no validated commercial conjugate antibody for rabies is available to meet the laboratory demand. Here, we evaluated the polyclonal anti-rabies virus ribonucleoprotein (RNP) IgG antibody for Rabies lyssavirus (RABV) detection by indirect rapid immunohistochemistry test (iRIT). We tested polyclonal anti-RNP IgG antibody against a batch of 100 brain specimens representing a wide phylogenetic origin in the State of São Paulo, Brazil. The purified IgG obtained 100% of diagnostic specificity and sensibility for RABV antigen detection in iRIT compared with the gold standard dFAT. In conclusion, our results demonstrate that the polyclonal anti-RNP IgG antibody may be used as a diagnostic reagent for rabies using iRIT, with the expectation of increase in availability and cost reduction of the epidemiological surveillance for developing countries.

Nyctinomops laticaudatus bat-associated rabies virus causes disease with a shorter clinical period and has lower pathogenic potential than strains isolated from wild canids

Rabies is a lethal viral disease that can affect a wide range of mammals. Currently, Rabies virus (RABV) in some European and American countries is maintained primarily in wild species. The regulation of viral replication is one of the critical mechanisms involved in RABV pathogenesis. However, the relationship between replication and the pathogenesis of RABV isolated from wild animals remains poorly understood. In the present study, we evaluated the pathogenicity of the street viruses Nyctinomops laticaudatus bat-associated RABV (NYBRV) and Cerdocyon thous canid-associated RABV (CECRV). Infection of mice with NYBRV led to 33% mortality with rapid disease evolution and marked histopathological changes in the CNS. In contrast, infection with CECRV led to 67% mortality and caused mild neuropathological lesions. The proportion of RABV antigen was significantly higher in the cytoplasm of neuronal cells of the cerebral cortex and in the meninges of mice infected with CECRV and NYBRV, respectively. Moreover, the replication rate of NYBRV was significantly higher (p < 0.001) than that of CECRV in neuroblastoma cells. However, CECRV replicated to a significantly higher titer in epithelial cells. Our results indicate that NYBRV infection results in rapid disease progression accompanied by frequent and intense histopathological alterations in the CNS in mice, and in a high replication rate in neuroblastoma cells. Although, CECRV is more pathogenic in mice, it caused milder histopathological changes in the CNS and replicated more efficiently in epithelial cells. Our data point to a correlation between clinical aspects of disease and the replication of RABV in different cell lines. (AU)

In situ histopathological and immunohistochemical characterization of rabies in the brains of naturally infected equines and bovines / Caracterização histopatológica e imuno-histoquímica In situ do vírus da raiva em cérebros de equinos e bovinos naturalmente infectados

The present study sought to characterize the phenomena involved in the histopathology of rabies and to assess the presence and amount of viral antigen in situ in different brain regions of naturally infected equines and bovines. The histopathological examination showed several changes due to inflammation, being most often infected cells neurons. The neuronal degeneration involved 100% of cases, in addition to a diffuse lymphocytic Infiltration and gliosis, characterized by vasculitis and perivasculitis. The presence of Negri bodies was in most cases in discreet, and the fragments with higher concentrations of antigen by both techniques employed were the cerebellum and the brain stem. Immunohistochemistry test (IHC) demonstrated greater sensitivity when applied to samples of bovines. Our results showed that in 37.5% of the total number of fragments analyzed, viral inclusions were not observed, however, was the presence of inflammatory process. In relation to the species, the fragments from bovine's animals showed a slight increase when examined under this feature. These findings highlight the importance of submitting samples from suspected animals for laboratory diagnostic, even when there are no apparent abnormal histological findings. (AU)

O presente estudo buscou caracterizar os fenômenos envolvidos na histopatologia da raiva e avaliar a presença e quantidade de antígeno viral in situ nas diferentes regiões cerebrais de equinos e bovinos naturalmente infectados. O exame histopatológico demonstrou várias mudanças devido à inflamação, sendo mais frequentemente infectadas as células neuronais. A degeneração neuronal foi observada em 100% dos casos, além de uma infiltração linfocitária difusa e gliose, caracterizada por vasculite e perivasculite. A presença de corpúsculos de Negri foi observada na maioria dos casos de maneira discreta, e os fragmentos com maior concentração de antígeno, por ambos os testes empregadas foram o cerebelo e o tronco encefálico. O teste de Imuno-histoquímica (IHC) demonstrou maior sensibilidade quando aplicada em amostras de bovinos. Nossos resultados demostraram que em 37,5% do número total de fragmentos analisados, inclusões virais não foram observadas, no entanto, havia processo inflamatório. Em relação à espécie, os fragmentos de bovinos demonstraram um ligeiro aumento quando examinado sob este aspecto. Esses achados destacam a importância de submeter amostras de animais suspeitos para diagnóstico laboratorial, mesmo quando não houver nenhum achado histopatológico anormal.Palavras-chave: raiva, equinos, bovinos, imuno-histoquímica, IFD, alterações histopatológicas. (AU)

Purification of IgG against ribonucleoprotein by a homemade immunoaffinity chromatography column for rabies diagnosis

Polyclonal or monoclonal antibodies against rabies virus ribonucleoprotein (RNP) conjugated to fluorescein isothiocyanate (FITC) have been employed for Rabies virus (RABV) antigen detection by the direct fluorescent antibody test (DFA). To date, these biomolecules have been purified by traditional methods such as precipitation by ammonium sulfate or ion exchange chromatography followed by ammonium sulfate precipitation, which allows only for partial detection of the protein of interest. In this study, we aimed to purify anti-RNP polyclonal horse IgG antibodies by cation-exchange chromatography in combination with a homemade immunoaffinity chromatography on RNP immobilized (RNP-IAC). Furthermore, to evaluate the accuracy of the prepared anti-RNP IgG fluorescent antibody in diagnostic purposes, DFA was applied for RABV antigen detection in suspected brain samples of different animal species. The combination of these two techniques made it possible to obtain antibodies with high selectivity and purity. Compared with the performance of the traditional method, anti-RNP IgG antibodies purified by RNP-IAC can be obtained from a smaller volume of hyperimmune serum and with greater avidity. Furthermore, the results obtained by DFA analyses revealed that the prepared anti-RNP IgG fluorescent antibody achieved 100% diagnostic specificity and sensitivity for RABV antigen detection. Thus, two-technique chromatographic, including RNP-IAC technology could be appropriate methods for the purification of polyclonal anti-RNP IgG for the use as a diagnostic reagent for rabies.(AU)

Nyctinomops laticaudatus bat-associated Rabies virus causes disease with a shorter clinical period and has lower pathogenic potential than strains isolated from wild canids.

Rabies is a lethal viral disease that can affect a wide range of mammals. Currently, Rabies virus (RABV) in some European and American countries is maintained primarily in wild species. The regulation of viral replication is one of the critical mechanisms involved in RABV pathogenesis. However, the relationship between replication and the pathogenesis of RABV isolated from wild animals remains poorly understood. In the present study, we evaluated the pathogenicity of the street viruses Nyctinomops laticaudatus bat-associated RABV (NYBRV) and Cerdocyon thous canid-associated RABV (CECRV). Infection of mice with NYBRV led to 33% mortality with rapid disease evolution and marked histopathological changes in the CNS. In contrast, infection with CECRV led to 67% mortality and caused mild neuropathological lesions. The proportion of RABV antigen was significantly higher in the cytoplasm of neuronal cells of the cerebral cortex and in the meninges of mice infected with CECRV and NYBRV, respectively. Moreover, the replication rate of NYBRV was significantly higher (p < 0.001) than that of CECRV in neuroblastoma cells. However, CECRV replicated to a significantly higher titer in epithelial cells. Our results indicate that NYBRV infection results in rapid disease progression accompanied by frequent and intense histopathological alterations in the CNS in mice, and in a high replication rate in neuroblastoma cells. Although, CECRV is more pathogenic in mice, it caused milder histopathological changes in the CNS and replicated more efficiently in epithelial cells. Our data point to a correlation between clinical aspects of disease and the replication of RABV in different cell lines.