Inadequacies of musculoskeletal medicine curriculum for undergraduate medical students: a cross-sectional study

ABSTRACT BACKGROUND: Musculoskeletal disorders account for up to one in four of general-practice consultations and almost one third of complaints in primary-care clinical practice. However, an insufficient amount of time and importance is given to their teaching in most medical schools. OBJECTIVE: To evaluate the acquisition of musculoskeletal competences in our institution, in order to identify flaws and propose changes to correct and improve the musculoskeletal curriculum. DESIGN AND SETTING: Cross-sectional study conducted in São Paulo, Brazil. METHODS: First to fifth-year medical students were enrolled in a survey using the Freedman and Bernstein musculoskeletal examination, in order to evaluate the acquisition of musculoskeletal competencies. Categorical data were analyzed using the chi-square test. Continuous data were analyzed using one-way analysis of variance (ANOVA). The level of significance was set as P < 0.05. RESULTS: A total of 545 students completed the questionnaire: from year 2, 115/167 (29.6%); from year 3, 118/138 (30.4%); from year 4, 98/130 (25.3%); and from year 5, 57/110 (14.7%). None of the students achieved the pass mark (established as 70%). The level of confidence in performing musculoskeletal examination was very low (3.7 ± 2.2; n = 386) and bore no relationship to the percentage of correct answers in the questionnaire (r = 0.331; 95% confidence interval, CI: 0.239-0.417; P < 0.001). CONCLUSION: Undergraduate teaching is the only exposure most general practitioners have to orthopedic problems. Universities are concerned about the adequacy of the musculoskeletal programs taught in their institutions. Student scores were found to be unsatisfactory in all the topics evaluated.

A evolução regulatória e os desafios na perspectiva dos laboratórios públicos produtores de vacinas no Brasil / La evolución regulatoria y los desafíos desde la perspectiva de los laboratorios públicos productores de vacunas en Brasil / Regulatory evolution and challenges from the perspective of public laboratiories vaccine producers in Brazil

A regulamentação para produtos biológicos vem evoluindo rapidamente ao longo dos últimos anos, seja motivada por questões de qualidade com impacto na vida das pessoas, seja pelo advento de novas tecnologias. As mudanças nas regulamentações que ditam como um produto deve ser registrado, produzido e monitorado são constantes. A responsabilidade de reguladores e fabricantes na garantia da qualidade, segurança e eficácia das vacinas torna-se ainda mais crítica, uma vez que essas substâncias são utilizadas, em sua maioria, em crianças e em pacientes saudáveis. Diante desse cenário, fabricantes precisam criar estratégias para manter seus produtos e instalações adequadas e um sistema da qualidade atualizado e operante. Por outro lado, as agências reguladoras têm o papel de garantir que os produtos que estão em uso atendam aos critérios estabelecidos, sem comprometer o fornecimento de medicamentos para a população.

La regulación para productos biológicos ha evolucionado rápidamente a lo largo de los últimos años, sea motivada por cuestiones de calidad con impacto en la vida de las personas, o por el advenimiento de nuevas tecnologías. Los cambios en las regulaciones que dictan como un producto debe ser registrado, producido y monitoreado son constantes. La responsabilidad de reguladores y fabricantes en la garantía de la calidad, seguridad y eficacia de las vacunas se convierte en algo todavía más crítico, ya que estas sustancias se utilizan, en su mayoría, en niños y pacientes saludables. Ante este escenario, los fabricantes necesitan crear estrategias para mantener sus productos e instalaciones de forma adecuada, además de un sistema de calidad actualizado y operativo. Por otro lado, las agencias reguladoras tienen el papel de garantizar que los productos que están en uso atiendan a los criterios establecidos, sin comprometer el suministro de medicamentos para la población.

The regulation of biological products has evolved rapidly in recent years due to quality issues impacting people's lives and the advent of new technologies, with constant changes in regulations that dictate how a product is registered, produced, and monitored. In the case of vaccines, the responsibility of regulators and manufacturers in guaranteeing quality, safety, and efficacy is even more critical, since vaccines are mostly used in children and healthy patients. In this scenario, manufacturers need to create strategies to keep their products and installations adequate and up-to-date with a fully operational quality system. Meanwhile, regulatory agencies have the role of guaranteeing that products meet the established criteria without compromising the supply of medicines to the population.

Vaccines for neglected and emerging diseases in Brazil by 2030: the valley of death and opportunities for RD& I in Vaccinology 4. 0 / Vacunas en Brasil para enfermedades emergentes y olvidadas en 2030: el valle de la muerte y oportunidades para RD& I en Vacunología 4. 0 / Vacinas para doenças negligenciadas e emergentes no Brasil até 2030: o vale da morte e oportunidades para PD& I na Vacinologia 4. 0

Abstract: We examine the implications of the very low competitiveness of the Brazilian vaccine RD&I system, which precludes the development of all the important vaccines required by the National Immunization Program (NIP), severely impacting the healthcare of the population. In a country dramatically affected by COVID-19 pandemic and by an exponential increase in emerging and neglected diseases, particularly the poor, these RD&I constraints for vaccines become crucial governance issues. Such constraints are aggravated by a global scenario of limited commercial interest from multinational companies in vaccines for neglected and emerging diseases, which are falling into a "valley of death," with only two vaccines produced in a pipeline of 240 vaccines. We stress that these constraints in the global pipeline are a window of opportunity for vaccine manufacturers in Brazil and other developing countries in the current paradigm transition towards Vaccinology 4.0. We conclude with recommendations for a new governance strategy supporting Brazilian public vaccine manufacturers in international collaborations for a sustainable national vaccine development and production plan by 2030.

Resumen: Examinamos las implicaciones de la muy baja competitividad del sistema brasileño de ID&I de vacunas, que imposibilita el desarrollo de todas las vacunas importantes, requeridas por el Progrma Nacional de Inmunización (PNI), con impactos muy graves en la salud de la población de un país con 200 millones de habitantes. En un país gravemente afectado por la pandemia de COVID-19 y por enfermedades emergentes y olvidadas que afectan particularmente a los pobres, estas restricciones del ID&I para vacunas es, de hecho, un asunto crucial de gobierno. Estas limitaciones locales se han visto agravadas por un escenario global de interés comercial limitado, por parte de las compañías multinacionales, en vacunas para enfermedades emergentes y olvidadas, que están cayendo en un "valle de la muerte", con solamente dos vacunas producidas a nivel global frente a 240 vacunas. Identificamos en estas limitaciones globales una ventana de oportunidad para los fabricantes de vacunas en Brasil y otros países en desarrollo dentro del paradigma actual de transición hacia la Vacunología 4.0. Concluimos con recomendaciones de una nueva estrategia de gobierno que apoye a los fabricantes brasileños de vacunas públicas en colaboraciones internacionales para el plan nacional de desarrollo y producción sostenible de vacunas en 2030.

Resumo: Examinamos as implicações da competitividade tão baixa do sistema brasileiro de pesquisa, desenvolvimento e inovação (PD&I) de vacinas, que impede o desenvolvimento de todas as vacinas importantes requeridas pelo Programa Nacional de Imunizações (PNI), prejudicando gravemente a saúde da população. Em um país seriamente afetado pela pandemia de COVID-19 e por um aumento exponencial de doenças emergentes e negligenciadas, principalmente entre os brasileiros pobres, essas restrições de PD&I quanto às vacinas tornam-se questões cruciais de governança. Essas restrições são agravadas por um cenário global de interesse comercial limitado por parte das empresas multinacionais de vacinas para doenças negligenciadas e emergentes, que estão caindo em um "vale da morte", com apenas duas vacinas produzidas em um pipeline de 240 vacinas. Ressaltamos que essas restrições na produção global constituem uma janela de oportunidade para os fabricantes de vacinas no Brasil e em outros países em desenvolvimento na atual transição de paradigma para a Vacinologia 4.0. Concluímos com recomendações para uma nova estratégia de governança em suporte aos fabricantes públicos de vacinas no Brasil em colaborações internacionais para um plano nacional de desenvolvimento e produção de vacinas que seja sustentável até 2030.

A evolução regulatória e os desafios na perspectiva dos laboratórios públicos produtores de vacinas no Brasil / La evolución regulatoria y los desafíos desde la perspectiva de los laboratorios públicos productores de vacunas en Brasil / Regulatory evolution and challenges from the perspective of public laboratiories vaccine producers in Brazil

A regulamentação para produtos biológicos vem evoluindo rapidamente ao longo dos últimos anos, seja motivada por questões de qualidade com impacto na vida das pessoas, seja pelo advento de novas tecnologias. As mudanças nas regulamentações que ditam como um produto deve ser registrado, produzido e monitorado são constantes. A responsabilidade de reguladores e fabricantes na garantia da qualidade, segurança e eficácia das vacinas torna-se ainda mais crítica, uma vez que essas substâncias são utilizadas, em sua maioria, em crianças e em pacientes saudáveis. Diante desse cenário, fabricantes precisam criar estratégias para manter seus produtos e instalações adequadas e um sistema da qualidade atualizado e operante. Por outro lado, as agências reguladoras têm o papel de garantir que os produtos que estão em uso atendam aos critérios estabelecidos, sem comprometer o fornecimento de medicamentos para a população.

La regulación para productos biológicos ha evolucionado rápidamente a lo largo de los últimos años, sea motivada por cuestiones de calidad con impacto en la vida de las personas, o por el advenimiento de nuevas tecnologías. Los cambios en las regulaciones que dictan como un producto debe ser registrado, producido y monitoreado son constantes. La responsabilidad de reguladores y fabricantes en la garantía de la calidad, seguridad y eficacia de las vacunas se convierte en algo todavía más crítico, ya que estas sustancias se utilizan, en su mayoría, en niños y pacientes saludables. Ante este escenario, los fabricantes necesitan crear estrategias para mantener sus productos e instalaciones de forma adecuada, además de un sistema de calidad actualizado y operativo. Por otro lado, las agencias reguladoras tienen el papel de garantizar que los productos que están en uso atiendan a los criterios establecidos, sin comprometer el suministro de medicamentos para la población.

The regulation of biological products has evolved rapidly in recent years due to quality issues impacting people's lives and the advent of new technologies, with constant changes in regulations that dictate how a product is registered, produced, and monitored. In the case of vaccines, the responsibility of regulators and manufacturers in guaranteeing quality, safety, and efficacy is even more critical, since vaccines are mostly used in children and healthy patients. In this scenario, manufacturers need to create strategies to keep their products and installations adequate and up-to-date with a fully operational quality system. Meanwhile, regulatory agencies have the role of guaranteeing that products meet the established criteria without compromising the supply of medicines to the population.

Yellow fever outbreak in Brazil: the puzzle of rapid viral spread and challenges for immunisation

We discuss the complex eco-social factors involved in the puzzle of the unexpected rapid viral spread in the ongoing Brazilian yellow fever (YF) outbreak, which has increased the reurbanisation risk of a disease without urban cases in Brazil since 1942. Indeed, this rapid spatial viral dissemination to the Southeast and South regions, now circulating in the Atlantic Forest fragments close to peri-urban areas of the main Brazilian megalopolises (São Paulo and Rio de Janeiro) has led to an exponential increase in the number of yellow fever cases. In less than 18 months, 1,833 confirmed cases and 578 deaths were recorded most of them reported in the Southeast region (99,9%). Large epizooties in monkeys and other non-human primates (NHPs) were communicated in the country with 732 YF virus (YFV) laboratory confirmed events only in the 2017/2018 monitoring period. We also discuss the peculiarities and similarities of the current outbreak when compared with previous great epidemics, examining several hypotheses to explain the recent unexpected acceleration of epizootic waves in the sylvatic cycle of the YFV together with the role of human, NHPs and mosquito mobility with respect to viral spread. We conclude that the most feasible hypothesis to explain this rapidity would be related to human behavior combined with ecological changes that promoted a significant increase in mosquito and NHP densities and their contacts with humans. We emphasize the urgent need for an adequate response to this outbreak such as extending immunisation coverage to the whole Brazilian population and developing novel strategies for immunisation of NHPs confined in selected reserve areas and zoos. Finally, we stress the urgent need to improve the quality of response in order to prevent future outbreaks and a catastrophic reurbanisation of the disease in Brazil and other South American countries. Continuous monitoring of YFV receptivity and vulnerability conditions with effective control of the urban vector Aedes aegypti and significant investments in YF vaccine production capacity and research and development for reduction of adverse effects are of the highest priority.

Prevalence and titers of yellow fever virus neutralizing antibodies in previously vaccinated adults

ABSTRACT Introduction: The World Health Organization (WHO) recommends one single dose of the Yellow Fever (YF) vaccine based on studies of antibody persistency in healthy adults. We assessed the prevalence and titers of YF virus neutralizing antibodies in previously vaccinated persons aged ≥ 60 years, in comparison to younger adults. We also evaluated the correlation between antibody titers and the time since vaccination among participants who received one vaccine dose, and the seropositivity among participants vaccinated prior to or within the past 10 years. Methods: previously vaccinated healthy persons aged ≥ 18 years were included. YF virus neutralizing antibody titers were determined by means of the 50% Plaque Reduction Neutralization Test. Results: 46 persons aged ≥ 60 years and 48 persons aged 18 to 59 years were enrolled. There was no significant difference in the prevalence of YF virus neutralizing antibodies between the two groups (p = 0.263). However, titers were significantly lower in the elderly (p = 0.022). There was no correlation between YF virus neutralizing antibody titers and the time since vaccination. There was no significant difference in seropositivity among participants vaccinated prior to or within the past 10 years. Conclusions: the clinical relevance of the observed difference in YF virus neutralizing antibody titers between the two groups is not clear.

Genital prevalence of HPV types and co-infection in men

Introduction: HPV infection is a highly prevalent sexually transmitted disease and there is evidence of the relationship of HPV infection and the development of genital warts, penile intraepitelial neoplasia, invasive penile carcinoma and cervical cancer. However, there is sparse data regarding the prevalence of HPV types and co-infection of different HPV types among men. Objectives: To assess the prevalence of HPV subtypes infections and rates of co-infection among men. Materials and Methods: 366 men were evaluated from March to October 2010. Men were referred to our institution for HPV diagnostic evaluation based on the following criteria: 1. presence of a genital wart; 2. presence of an atypical genital lesion; 3. absence of symptoms and a partner with a HPV diagnosis; 4. absence of symptoms and a desire to undergo a full STD diagnostic evaluation. Genital samples were collected from the urethra, penile shaft, scrotum and anus with Digene® collection and preservation kit and submitted to HPV genotype microarray detection (Papillocheck®). All men were tested for the low-risk HPV types 6-11-40-42-43-44 and for the high-risk HPV types 16-18-31-33-35-39-45-51-52-53-56-58-59-66-68-70-73-82. Results: Of the 366 men, 11 were tested inconclusive and were excluded from the analysis. 256 men (72.1% of the men from the cohort referred to our institution) tested positive with genotype micro-array detection and 99 tested negative. The most prevalent HPV-subtypes in the studied population were 6, 42, 51 and 16. Co-infection was found in 153 men. Of those, 70 (19.7%) had a co-infection by 2 types, 37 (10.4%) by 3 types; 33 men (9.2%) by 4 types; 8 men (2.2%) by 5 types; 1 man (0.3%) by 6 types; 1 man (0.3%) by 7 types; 2 men (0.6%) by 8 types and 1 man (0.3%) by 9 types. Conclusion: The most frequent HPV types were 6, 16, 42 and 51. Co-infection was found in 59% of our patients. This information is vital to drive future public health ...

Declining prevalence of hepatitis A virus antibodies among children from low socioeconomic groups reinforces the need for the implementation of hepatitis A vaccination in Brazil

Age-related seroprevalence studies that have been conducted in Brazil have indicated a transition from a high to a medium endemicity of hepatitis A virus (HAV) infection in the population. However, most of these studies have focused on urban populations that experience lower incidence rates of HAV infection. In the current study, the prevalence of anti-HAV antibodies was investigated in children with a low socioeconomic status (SES) that live on the periphery of three capital cities in Brazil. A total of 1,162 dried blood spot samples were collected from individuals whose ages ranged from one-18 years and tested for anti-HAV antibodies. A large number of children under five years old (74.1-90%) were identified to be susceptible to HAV infection. The anti-HAV antibody prevalence reached ≥ 50% among those that were 10-14 years of age or older. The anti-HAV prevalence rates observed were characteristics of regions with intermediate level of hepatitis A endemicity. These data indicated that a large proportion of children with a low SES that live at the periphery of urban cities might be at risk of contracting an HAV infection. The hepatitis A vaccine that is currently offered in Brazil is only available for high-risk groups or at private clinics and is unaffordable for individuals with a lower SES. The results from this study suggest that the hepatitis A vaccine should be included in the Brazilian National Program for Immunisation.

Dengue-2 and yellow fever 17DD viruses infect human dendritic cells, resulting in an induction of activation markers, cytokines and chemokines and secretion of different TNF-α and IFN-α profiles

Flaviviruses cause severe acute febrile and haemorrhagic infections, including dengue and yellow fever and the pathogenesis of these infections is caused by an exacerbated immune response. Dendritic cells (DCs) are targets for dengue virus (DENV) and yellow fever virus (YF) replication and are the first cell population to interact with these viruses during a natural infection, which leads to an induction of protective immunity in humans. We studied the infectivity of DENV2 (strain 16681), a YF vaccine (YF17DD) and a chimeric YF17D/DENV2 vaccine in monocyte-derived DCs in vitro with regard to cell maturation, activation and cytokine production. Higher viral antigen positive cell frequencies were observed for DENV2 when compared with both vaccine viruses. Flavivirus-infected cultures exhibited dendritic cell activation and maturation molecules. CD38 expression on DCs was enhanced for both DENV2 and YF17DD, whereas OX40L expression was decreased as compared to mock-stimulated cells, suggesting that a T helper 1 profile is favoured. Tumor necrosis factor (TNF)-α production in cell cultures was significantly higher in DENV2-infected cultures than in cultures infected with YF17DD or YF17D/DENV. In contrast, the vaccines induced higher IFN-α levels than DENV2. The differential cytokine production indicates that DENV2 results in TNF induction, which discriminates it from vaccine viruses that preferentially stimulate interferon expression. These differential response profiles may influence the pathogenic infection outcome.

Atualização em vacinas, imunizações e inovação tecnológica / Vaccines, immunization and technological innovation: an update

A erradicação da varíola foi a maior conquista da saúde pública mundial. E o binômio vacinas e imunizações continua a demonstrar alto desempenho na prevenção e no controle de outras doenças imunopreveníveis. As novas iniciativas globais em vacinação, como a GAVI, vêm possibilitando a introdução de novas vacinas e salvando vidas de milhares de crianças nos países mais pobres do mundo. O Programa Nacional de Imunizações (PNI) do Brasil também vem sendo fortalecido com a incorporação de novas vacinas no seu calendário de imunizações, como a vacina contra rotavírus, pneumococos conjugada, meningite meningocócica do sorogrupo C conjugada, além do H1N1 para as populações de maior risco. Com o descobrimento de novas vacinas de alto valor agregado, os grandes laboratórios multinacionais despertaram para este segmento farmacêutico e buscam a liderança da área, investindo maciçamente em inovação tecnológica, além de realizar fusões, aquisições e parcerias tecnológicas. O Brasil também vem se fortalecendo, tendo criado marcos reguladores e financiando projetos de inovação tecnológica e modernização da infraestrutura de produção.

The smallpox worldwide eradication was the major world public health achievement. The binomial vaccines and immunization continues to demonstrate very high performance in the prevention and control of other diseases preventable by vaccination. The new global initiatives on vaccination, such as GAVI, have made possible the introduction of new and important vaccines preventing million of children deaths in the poorest countries in the world. The National Immunization Program of Brazil is also being strengthened, with the introduction of several new vaccines into the basic calendar as rotavirus, pneumococcal and meningococcal conjugated and H1N1 in national campaign, covering the population at risk. With the discovery of high valued vaccines, the big pharmaceutical companies became interested in this area, investing heavily in technological innovation, making fusions, acquisitions and technological partnerships. Brazil has also established a new innovation policy, creating new laws as well as subsidizing projects in technological innovation and modernization of production infra-structure.

Epidemiologic survey of traumatic dental injuries in children seen at the Federal University of Rio de Janeiro, Brazil

This epidemiologic survey aimed at assessing the prevalence of traumatic dental injuries in children seen at the Federal University of Rio de Janeiro, Brazil. The records of a total of 111 children (aged 0 to 6 years) seen from 2004 to 2006 in the dental trauma clinic were surveyed, comprising a total of 201 traumatized primary teeth. Data pertaining to the child and to the trauma such as age, gender, etiology, teeth involved, type of traumatic injury, time elapsed between the trauma and seeking care, and the presence and kind of clinical and radiographic sequelae in the first visit were collected from the dental records. All variables studied were assessed by means of frequency analysis and the Chi-square test (p < 0.05). A higher prevalence of trauma was observed in boys (56.7 percent) and in the age group from 0-3 years (73.8 percent). The most affected teeth were the central incisors (84.7 percent) and the most common trauma etiology was a fall from the child's own height (63.0 percent). The supporting tissues were the most affected. Lateral luxation was the most frequent alteration observed (33.4 percent), followed by concussion (21.0 percent). Coronal discoloration (17.7 percent) and external resorption (18.3 percent) were, respectively, the most prevalent clinical and radiographic sequelae. Gender had no influence on the clinical (p = 0.54) and radiographic (p = 0.55) sequelae. Even though age had no influence on radiographic sequelae (p = 0.41), clinical sequelae were more prevalent in children aged 0 to 3 years (p = 0.03). In conclusion, traumatisms in primary teeth were more prevalent in boys, and in 0-3-year-old children. Luxation was the most frequent traumatic lesion, and coronal discoloration and external resorption were the most prevalent sequelae.

Limited replication of yellow fever 17DD and 17D-Dengue recombinant viruses in rhesus monkeys

For the development of safe live attenuated flavivirus vaccines one of the main properties to be established is viral replication. We have used real-time reverse transcriptase-polymerase chain reaction and virus titration by plaque assay to determine the replication of yellow fever 17DD virus (YFV 17DD) and recombinant yellow fever 17D viruses expressing envelope proteins of dengue virus serotypes 2 and 4 (17D-DENV-2 and 17D-DENV-4). Serum samples from rhesus monkeys inoculated with YFV 17DD and 17D-DENV chimeras by intracerebral or subcutaneous route were used to determine and compare the viremia induced by these viruses. Viral load quantification in samples from monkeys inoculated by either route with YFV 17DD virus suggested a restricted capability of the virus to replicate reaching not more than 2.0 log10 PFU mL-1 or 3.29 log10 copies mL-1. Recombinant 17D-dengue viruses were shown by plaquing and real-time PCR to be as attenuated as YF 17DD virus with the highest mean peak titer of 1.97 log10 PFU mL-1 or 3.53 log10 copies mL-1. These data serve as a comparative basis for the characterization of other 17D-based live attenuated candidate vaccines against other diseases.

Uma das principais propriedades a serem estabelecidas para o desenvolvimento de vacinas seguras e atenuadas de flavivirus,é a taxa de replicação viral. Neste trabalho, aplicamos a metodologia de amplificação pela reação em cadeia da polimerase em tempo real e titulação viral por plaqueamento para determinação da replicação do vírus 17DD (FA 17DD) e recombinantes, expressando proteínas do envelope de dengue sorotipos 2 e 4 (17D-DENV-2 e 17D-DENV-4). As amostras de soros de macacos inoculados por via intracerebral ou subcutânea com FA 17DD ou 17D-DENV foram usadas para determinar e comparar a viremia induzida por estes vírus. A quantificação da carga viral em amostras de macacos inoculados por ambas as vias com FA 17DD sugere restrita capacidade de replicação com taxa não superior a 2,0 log10 PFU mL-1 ou 3,29 log10 cópias/mL-1. Os vírus recombinantes 17D-DENV mostraram-se tão atenuados quanto o vírus 17DD, tanto porRT-PCR em tempo real quanto por plaqueamento, com título médio máximo de 1,97 log10 PFU mL-1 ou 3,53 log10 cópias/mL-1. Estes dados servem como base comparativapara caracterização de outros vírus vivos atenuados, derivados do vírus 17D, candidatos a vacinas contra outras doenças.

Influence of culture conditions on Vero cell propagation on non-porous microcarriers

Animal cell cultures are widely employed for the production of viral vaccines and for recombinant protein expression. The cell line Vero is a continuous, adherent cell line, which has been recommended by the World Health Organization for the production of human vaccines. For the large-scale production of vaccines, microcarriers, which are microspheres that serve as support for the cells, are being increasingly used. The use of microcarriers in stirred bioreactors allows high cell densities and, consequently, high virus titres to be achieved. With the aim of selecting appropriate culture conditions for the cultivation of Vero cells at high cell densities, in this work the influence of several variables (agitation rate, ratio of inoculated cells to microcarrier mass and fetal bovine serum concentration) on cell growth on Cytodex 1 microcarriers was studied. Under the best conditions determined, a comparison with Vero cell cultivation on Cytodex 3 microcarriers was carried out.

Reactogenicity of yellow fever vaccines in a randomized, placebo-controlled trial

OBJETIVO: Comparar a reatogenicidade de três vacinas contra a febre amarela (FA) das sub-cepas WHO-17D e 17DD (diferentes lotes-semente), e placebo. MÉTODOS: Foram recrutados 1.087 adultos elegíveis para vacinação contra FA no Rio de Janeiro, RJ, Brasil. Vacinas produzidas por Bio-Manguinhos, Fiocruz (Rio de Janeiro, RJ) foram administradas ("dia zero") seguindo procedimentos adaptados para alocação randômica em blocos e "cega" para o tipo de vacina. Eventos adversos pós-vacinação foram registrados em questionários e diários preenchidos pelos participantes. Enzimas hepáticas foram medidas nos dias 0, 4-20 e 30 do estudo. A viremia foi medida nos dias 4-20. A resposta imune foi verificada em testes sorológicos nos dias 0 e 30. RESULTADOS: Os participantes eram predominantemente homens jovens. A taxa de soroconversão foi superior a 98% no grupo soronegativo antes da vacinação. Comparado ao placebo, a diferença de risco de eventos adversos locais variou de 0,9% a 2,5%, e de 3,5% a 7,4% para eventos adversos sistêmicos nos grupos vacinados. A diferença de risco desses eventos com assistência médica e/ou falta ao trabalho variou de 2,0% a 4,5%. Viremia foi detectada em 3% a 6% dos vacinados até 10 dias após a vacinação. As variações nos níveis de enzimas hepáticas pós-vacinação foram semelhantes nos grupos vacinados e placebo. CONCLUSÕES: Foi demonstrada pela primeira vez a semelhança do perfil de reatogenicidade das vacinas contra FA das cepas 17D e 17DD, comparados entre si e com placebo. As variações das enzimas hepáticas constituem evidência contra o potencial de viscerotropismo do vírus vacinal.

Immunogenicity of WHO-17D and Brazilian 17DD yellow fever vaccines: a randomized trial

OBJETIVO: Comparar a imunogenicidade de três vacinas contra febre amarela ) das subcepas WHO-17D e 17DD brasileira (diferentes lotes-semente). MÉTODOS: Trata-se de ensaio de equivalência envolvendo 1.087 adultos no Rio de Janeiro, RJ. As vacinas foram produzidas em Bio-Manguinhos, Fiocruz (Rio de Janeiro, Brasil) e foram administradas seguindo procedimentos adaptados para randomização em blocos, com tipos de vacinas codificados ("duplo-cego"). Anticorpos neutralizantes contra febre amarela foram dosados antes e depois da vacinação. Definiu-se equivalência como diferença nas taxas de soroconversão não superior a cinco pontos percentuais, e razão de títulos médios geométricos superior (TMG) a 0,67. RESULTADOS: As taxas de soroconversão foram iguais ou maiores do que 98 por cento nos participantes previamente soronegativos. Na coorte completa (incluindo os previamente soropositivos) a soroconversão foi igual ou superior a 90 por cento. As diferenças na soroconversão variaram de -0,05 por cento a -3,02 por cento entre os grupos de comparação. A intensidade da resposta imune também foi semelhante nos grupos: 14,5 UI/mL a 18,6 UI/mL. As razões de TMG variaram de 0,78 a 0,93. Considerando o grupo placebo, as vacinas explicaram 93 por cento da soroconversão. Viremia foi detectada entre os dias três e sete em 2,7 por cento dos participantes vacinados. CONCLUSÕES: A equivalência na imunogenicidade das vacinas contra a febre amarela das subcepas 17D e 17DD foi demonstrada pela primeira vez em ensaio clínico randomizado, duplo-cego, controlado com placebo. O estudo completou o processo de validação clínica do novo lote-semente de vacina, além de ampliar as bases para utilização da vacina brasileira em outros países e de trazer alternativas de subcepas para o produtor da vacina no Brasil.

Estudo sobre a propagação de vírus da febre amarela vacinal 17D e produção de vacina em culturas primárias de fibroblastos / Study on the propagation of virus of the vaccine yellow fever vaccine 17D and production in primary cultures of fibroblastos

A tecnologia básica para a produção de vacinas contra a febre amarela em embriões de galinha permanece inalterada,desde 1940.Descreve um método econômico e eficiente,de produção de vacina contra febre amarela da cepa 17DD,em culturas de fibroblasto de embrião de galinha(CEF),com títulos de 6,3 a 6,7 log10 PFU/mL.A metodologia foi utilizada com sucesso na produção de lotes sementes de vírus 17D/204/DD a partir de cDNA clonado e na produção de lotes vacinais com o vírus 17DD.A termoestabilidade de 2 diferentes formulações(5 e 50 doses)da vacina produzida em CEF com o vírus 17DD demonstraram ser tão alta quanto a das vacinas atualmente em uso.A produção em CEF não levou à seleção de variações genéticas.O vírus 17DD produzido em CEF também foi indistinguível do seu vírus semente parental em termos de tamanho de placa de lise e imunogenicidade em camundongos e macacos.A comparação do vírus produzido em CEF com o vírus do lote semente preparado em embrião,no teste em macacos revelou um escore clínico mais elevado para o vírus de CEF.As diferenças nos escores histológicos do sistema nervoso central para macacos inoculados com o vírus de embrião e com a vacina experimental de CEF ficaram no limite da significância estatística.O sistema do interferon foi identificado como sendo o fator de interferência levando à baixa produtividade viral em CEF.Esta identificação foi realizada através de diversas abordagens experimentais utilizando-se culturas de CEF em alta e baixa densidade e incluíram:a influência de sobrenadantes de culturas de CEF infectadas com vírus 17DD,tratados ou não por exposição ao ácido ou calor,na replicação de vírus Sindbis,a análise da atividade da enzima 2’-5’oligoadenilato sintetase,cuja expressão é controlada ao nível transcricional pelo sistema de interferon e a detecção por transcrição reversa seguida de amplificação por reação em cadeia da polimerase(RT-PCR)da expressão de genes de interferon e do gene da própria 2’-5’oligoadenilato sintetase.Embora experimentos com culturas de CEF em baixa densidade não tenham sido extensivamente realizados,os resultados sugerem que tanto em alta como em baixa densidade o IFN está sendo produzido.Devido à diluição do IFN no meio e ao espaçamento celular na cultura de baixa densidade,este não é capaz de estabelecer o estado anti-viral nas células ainda não infectadas a tempo de impedir a replicação do vírus.

Low frequency of side effects following an incidental 25 times concentrated dose of yellow fever vaccine

In August/1999, a group of 14 adults from the staff of a private hospital in Contagem -- Minas Gerais State, Brazil, received unintentionally a 25 times concentrated dose of the 17-DD yellow fever vaccine (Bio-Manguinhos), due to a mistake at the reconstitution step. All patients were clinically and laboratorially evaluated at days 5, 13 and 35 post vaccination. Frequency of side effects and clinical observations of this group of individuals were not different from the observed in recipients immunized with normal doses of the vaccine. At the second and third evaluation none of the subjects reported symptoms. None of the patients presented abnormalities at the physical examination at none of the time points and in all cases the blood examination was normal, except for a reduced number of platelets that was detected in one subject at the first and second evaluation and reverted to normal at third evaluation. At the first evaluation point, 8 subjects were serum negative and 6 serum positive for yellow fever at the plaque reduction neutralization test. In 5 subjects the observed titre was 10 times higher as the baseline of 2.36 Log10 mUI/ml. The samples collected at second and third evaluation (13th and 35th days) demonstrated that all subjects responded to the vaccination with the exception of one that did not present a positive result in any of the samples collected. This evaluation confirms the safety of the 17-DD yellow fever vaccine

The early use of yellow fever virus strain 17D for vaccine production in Brazil - a review

The use of yellow fever (YF) virus 17D strain for vaccine production adapted in Brazil since its introduction in 1937 was reviewed. This was possible due to the availability of official records of vaccine production. The retrieved data highlight the simultaneous use of several serially passaged 17D substrain viruses for both inocula and vaccine preparation that allowed uninterrupted production. Substitution of these substrain viruses became possible with the experience gained during quality control and human vaccination. Post-vaccinal complications in humans and the failure of some viruses in quality control tests (neurovirulence for monkeys) indicated that variables needed to be reduced during vaccine production, leading to the development of the seed lot system. The 17DD substrain, still used today, was the most frequently used substrain and the most reliable in terms of safety and efficacy. For this reason, it is possible to derive an infectious cDNA clone of this substrain combined with production in cell culture that could be used to direct the expression of heterologous antigens and lead to the development of new live vaccines

Estudo de soroconversäo com formulaçöes da vacina Biken CAM-70 contra sarampo / Comparison of seroconversion rates with CAM-70 measles vaccine at different dosages

Objetivo: Comparar a resposta sorológica induzida por formulaçöes com diferentes concentraçöes de vírus da vacina contra sarampo da cepa Biken CAM-70. Métodos: Crianças sadias de 9 a 18 meses de um centro de saúde do Rio de Janeiro, RJ, cujos responsáveis concordaram em participar, foram randomizadas em três grupos vacinados com concentraçöes de 5.000, 1.000 ou 200 CCID50 (50 por cento Tissue Culture Infective Dose). Os participantes e o pessoal da pesquisa ignoravam o tipo de vacina administrada. A avaliaçäo sorológica foi realizada pelo teste de reduçäo em plaque de lise. Duas análises intermediárias dos dados foram programadas. Resultados: Das 223 crianças recrutadas, 84 por cento completaram todos os procedimentos; 79 por cento tinham idade menor que 10 meses; 93 por cento näo tinham anticorpos contra sarampo no soro pré-vacinal. As proporçöes de soroconversäo (quadruplicaçäo das concentraçöes pré-vacinais) foram 82 por cento, 55 por cento e 37 por cento (p<0,0000), nos grupos vacinados com 5.000, 1.000 ou 200 CCID50, respectivamente. As diferenças nas concentraçöes médias de anticorpos pós-vacinais também foram substanciais e estatisticamente significativas (p<0,000). A soroconversäo (independente da formulaçäo da vacina) foi de 73 por cento nas crianças com 10 ou mais meses de idade e 53 por cento naquelas com menos de 10 meses. Conclusöes: Formulaçöes da vacina com concentraçöes inferiores a 5.000 CCID50 näo induziram soroconversäo satisfatória. O desempenho da vacina por faixa etária foi compatível com o observado em outros estudos com a vacina Biken CAM-70 e indica que uma proporçäo apreciável de crianças vacinadas aos 9 meses pode näo obter resposta imunológica plena

The yellow fever 17D vaccine virus as a vector for the expression of foreign proteins: development of new live flavivirus vaccines

The Flaviviridae is a family of about 70 mostly arthropod-borne viruses many of which are major public health problems with members being present in most continents. Among the most important are yellow fever (YF), dengue with its four serotypes and Japanese encephalitis virus. A live attenuated virus is used as a cost effective, safe and efficacious vaccine against YF but no other live flavivirus vaccines have been licensed. The rise of recombinant DNA technology and its application to study flavivirus genome structure and expression has opened new possibilities for flavivirus vaccine development. One new approach is the use of cDNAs encopassing the whole viral genome to generate infectious RNA after in vitro transcription. This methodology allows the genetic mapping of specific viral functions and the design of viral mutants with considerable potential as new live attenuated viruses. The use of infectious cDNA as a carrier for heterologous antigens is gaining importance as chimeric viruses are shown to be viable, immunogenic and less virulent as compared to the parental viruses. The use of DNA to overcome mutation rates intrinsic of RNA virus populations in conjunction with vaccine production in cell culture should improve the reliability and lower the cost for production of live attenuated vaccines. The YF virus despite a long period ignored by researchers probably due to the effectiveness of the vaccine has made a come back, both in nature as human populations grow and reach endemic areas as well as in the laboratory being a suitable model to understand the biology of flaviviruses in general and providing new alternatives for vaccine development through the use of the 17D vaccine strain