Um total de 144 amostras fecais colhidas de leitões com diarréia provenientes de diversas granjas distribuídas por 10 municípios do Estado de São Paulo, Brasil, foram examinadas para a presença de rotavírus através de eletroforese em gel de poliacrilamida e ELISA, num esquema de triagem em paralelo. Destas, 24 e 39 amostras foram, respectivamente, positivas por estes testes e a caracterização dos genotipos P e G foi realizada em 43 amostras por nested RT-PCR, usando diferentes conjuntos de primers. Utilizando-se primers animais, o genotipo P[6] foi o mais freqüente, detectado em 25,58% das amostras, seguido pelo P[1] (11,62%) e P[7] (9,3%). Infecção concomitante de genotipos P[6]+P[7] (9,3%), P[1]+P[6] (4,65%), P[1]+P[6]+P[7] (2,32%) foi também observada. O genotipo G[5] foi o mais freqüente, detectado em 30,23% das amostras, seguido pelo G[10] (20,93%) e G[6] (4,65%). Infecção concomitante de genotipos G[5]+G[10] (18,6%) foi também observada. Utilizando-se primers humanos, somente o genotipo P[6] (65,11%) foi encontrado. Foram detectados os genotipos G[4] (27,9%), G[3] (13,95%) nas amostras, e as infecções concomitantes G[3]+G[4] (9,3%), G[2]+G[3]+G[4] (4,65%) e G[2]+G[3] (2,32%). A combinação mais freqüente foi G[5]P[6] e G[4]P[6], porém foram encontradas também infecções mistas, genotipos atípicos e reassortants. O seqüenciamento do gene a partir dos produtos de nested PCR dos genotipos animais P e G mostrou diversidade de nucleotídeos e aminoácidos dentro de um mesmo genotipo. As árvores filogenéticas utilizando o critério de maxima parcimônia e o algoritmo branch-and-bound, tiveram topologias nas quais as seqüências de nucleotídeos geradas neste trabalho concordam com as outras descritas anteriormente e pertencentes a um mesmo genotipo, suportadas por índices aceitáveis de \"bootstrap\". Os resultados deste estudo, além de contribuir para um melhor entendimento da epidemiologia dos rotavírus suínos, é relevante ao mostrar que além de haver uma diversidade de genotipos circulantes no campo, há também dentro dos mesmos genotipos, e isto deve ser considerado ao se utilizar e desenvolver métodos de diagnóstico, bem como ao se adotarem medidas preventivas específicas contra esta doença. / A total of 144 fecal specimens collected from piglets with diarrhea from many swine-producing units distributed among 10 municipalities of São Paulo State, Brazil, were examined for rotavirus by polyacrylamide gel electrophoresis and ELISA, in a parallel screening scheme. Twenty-four and thirty-nine samples, respectively, were positive by these tests and the characterization of the P and G genotypes was performed on 43 samples by a nested reverse transcription-PCR typing assay, using different sets of primers. Using the animal set of primers, the P[6] genotype was the most frequent, accounting for viruses in 25.58% of the samples, followed by P[1] (11.62%) and P[7] (9.3%). Concomitant infection of P[6]+P[7] (9.3%), P[1]+P[6] (4.65%), P[1]+P[6]+P[7] (2.32%) genotypes was also observed. The G[5] genotype was the most frequent, accounting for viruses in 30.23% of the samples, followed by G[10] (20.93%) and G[6] (4.65%). Concomitant infection of G[5]+G[10] (18.6%) genotypes was observed. Using human set of primers, only the P[6] (65.11%) genotype was found. It was detected the genotypes G[4] (27.9%), G[3] (13.95%) on the samples. Concomitant infection of the genotypes G[3]+G[4] (9.3%), G[2]+G[3]+G[4] (4.65%) and G[2]+G[3] (2.32%) was also observed. The more frequent combination were G[5]P[6] and G[4]P[6], but it was found mixed infections, atypical genotypes and reassortants. Gene sequencing of nested products of G and P animal genotypes showed a diversity of nucleotides and aminoacids under the same genotype. The phylogenetic trees for P and G genotypes under the maximum parsimony criterion, using the branch-and-bound algorithm, had topologies in which the nucleotide sequences generated by this work agree to others described elsewhere that have the same genotypes, supported by acceptable scores of bootstrapping. The results of the present study, while contributing to a better understanding of epidemiology of porcine rotaviruses, address the relevance that there is not only a diversity of genotypes circulating on the field, but inside the same genotypes, and this must be considered when using and developing diagnostic tests, as well when carrying out specific preventive measures against this disease.

As diarréias neonatais constituem-se em um dos mais importantes fatores econômico e sanitário nas granjas suínas, quer pela mortalidade, quer pelas perdas agregadas ao atraso no desenvolvimento dos leitões, à profilaxia e ao manejo. Os rotavírus ocupam lugar de destaque pela rápida disseminação dentro do plantel, bem como pelo potencial zoonótico, dada a probabilidade de rearranjo ou recombinação genética entre amostras humanas e animais O objetivo deste trabalho foi detectar a presença de rotavírus a partir de 277 amostras fecais de leitões com quadro clínico de diarréia, provenientes dos Estados do Rio Grande do Sul (RS), Santa Catarina (SC), Paraná (PR), Mato Grosso do Sul (MS), Mato Grosso (MT), São Paulo (SP), Rio de Janeiro (RJ) e Minas Gerais (MG) entre os anos de 2009 e 2011 e analisar o perfil eletroforético de migração dos segmentos genômicos bem como diferenças de eletroferótipos nas amostras positivas, pela técnica de eletroforese em gel de poliacrilamida (PAGE). Das 277 amostras fecais diarréicas de leitões analisadas, 25 foram positivas (25/277= 9%). De conformidade com os Estados de origem, foram verificadas as frequências de 20% (1/5) no RS, 11,1% (1/9) em SC, 12,5% (1/8) no PR, 15,3% (6/39) em MS, 14,2% (3/21) em MT, 6,7% (5/74) em SP, 0% (0/7) no RJ e 7% (8/114) em MG. Pela análise da migração eletroforética dos segmentos genômicos, todas as 25 amostras positivas apresentaram perfil eletroforético compatível com o RV-A, tal como a amostra padrão NCDV com migração característica em quatro classes ou agregados [4-2-3- 2]. Foram observadas pequenas diferenças na velocidade de migração de um ou mais segmentos dentro da mesma classe. Estes resultados evidenciam a importância da PAGE como metodologia de diagnóstico e de investigações epidemiológicas nas rotaviroses suínas. / The neonatal diarrhea constitute one of the most important economic and health factors in pig farms either by death or by the aggregate losses to the delay in the development of piglets, prophylaxis and management. Rotaviruses have a prominent role by the rapid spread within the herd, as well as the zoonotic potential, given the likelihood of genetic reassortment or recombination between human and animal samples. The objective of this study was to detect the presence of rotavirus from fecal samples from 277 piglets with clinical cases of diarrhea, from the states of Rio Grande do Sul (RS), Santa Catarina (SC), Paraná (PR), Mato Grosso do Sul (MS), Mato Grosso (MT), São Paulo (SP), Rio de Janeiro (RJ) e Minas Gerais (MG) between the years 2009 and 2011 and to analyze the electrophoretic migration of genomic segments and electropherotype differences in positive samples, by polyacrylamide gel electrophoresis (PAGE) technique. From 277 piglets diarrheal stool samples analyzed, 25 were positive (25/277 = 9%). In accordance with the States of origin, were observed frequencies of 20% (1/5) in the RS, 11.1% (1/9) in SC, 12.5% (1/8) in PR, 15.3 % (6/39) in MS, 14.2% (3/21) in MT, 6.7% (5/74) in SP, 0% (0/7) in Rio de Janeiro and 7% (8/114) in MG. For the analysis of the electrophoretic migration of genome segments, all 25 positive samples showed electrophoretic profile compatible with the RV-A as a standard sample NCDV, showing characteristic fourth class or aggregates [4-2-3-2]. We observed small differences in the migration speed of one or more segments within the same class. These results highlight the importance of PAGE as a method of diagnosis and epidemiological investigations in the porcine rotavirus.

Orientador: Hyun Mo Yang / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Matematica, Estatistica e Computação Cientifica / Made available in DSpace on 2018-08-12T03:04:36Z (GMT). No. of bitstreams: 1 Pinheiro_Andressa_M.pdf: 1109369 bytes, checksum: 4cae928d860fd4d141699ffe961e8140 (MD5) Previous issue date: 2008 / Resumo: Os rotavírus são considerados, atualmente, um dos mais importantes agentes causadores de gastroenterites e óbitos em crianças com menos de 5 anos no mundo. Ocorrem globalmente cerca de 125 milhões de episódios diarréicos por rotavírus a cada ano, causando entre 600.000 e 870.000 óbitos. Esses números alarmantes estimularam a busca por um controle desse vírus, mas para combatê -lo é necessário estudar seu comportamento, como ele penetra no organismo humano, como age dentro dele e como se espalha. Nesse trabalho apresenta-se um breve estudo sobre a biologia do rotavírus e os mecanismos de defesa apresentados pelo sistema imunológico. O principal objetivo é, utilizando métodos quantitativos, estudar a interação entre o rotavírus e o sistema imunológico e avaliar, comparativamente, o desempenho das respostas imunológicas humoral e celular. Seguindo esse intuito apresenta-se um modelo matemático, composto de equações diferenciais ordinárias não lineares de primeira ordem, que descreve a ação do sistema imunológico a fim de eliminar o rotavírus. A partir deste modelo nós encontramos os pontos de equilíbrio trivial e não-trivial e analisamos sua estabilidade. Também discutimos sobre a ação das respostas imunológicas humoral e celular. / Abstract: In infants and young children, rotavirus is the major cause of severe inflammation of the intestine (gastroenteritis). Rotavirus infection frequently results in fever, vomiting and diarrhea, wich symptoms are so intense that they can lead to death. This virus causes nearly a million deaths each year worldwide, mostly in developing countries. Rotavirus attacks the epithelial cells of the thin intestine and replicates in the cytoplasm and do not fully uncoat during the process of replication. The reason for their failure to fully uncoat is that the coat is resistant to protease digestion, which prevents them from being completely destroyed by the infected cell and of readily being seen by the immune system. This complex biology of rotavirus and its interaction with the immune system are the motivation of this work, that presents a model for this interaction, structured by non-linear ordinary differential equations of first-order that describes the action of the innate immune system to eliminate rotavirus. From this model, we find the trivial and non-trivial equilibrium points and analyze their stabilities, as well we analyze, comparatively, the humoral and cellular responses. / Mestrado / Biomatematica / Mestre em Matemática Aplicada

No description available.

In view of the recent implementation of a publicly funded rotavirus vaccination program in Ontario, we undertook studies to help guide the design of a safety evaluation of the vaccine with respect to intussusception. We used administrative data to develop and validate an algorithm for intussusception, and quantified its incidence in Ontario. We also conducted a systematic review of study designs used to evaluate post-licensure vaccine safety, and discussed each design’s strengths and weaknesses. The validated algorithm for intussusception was sensitive (89.3%) and highly specific (>99.9%). We observed the highest mean incidence (34 / 100,000) in males <1 year of age. While other designs are more robust, the inability to ascertain individual vaccination status from Ontario’s administrative data dictated our selection of an ecological design for safety evaluation of rotavirus vaccine. Data assimilated from this thesis represent a critical step toward the timely evaluation of rotavirus vaccine safety in Ontario.

No description available.

Single particle electron microscopy (EM) allows us to examine the molecular world and gain insights into protein structures implicated in human disease. Visualizing the 3D architecture of the macromolecules can inform drug design and preventative care. While X-ray crystallography and NMR are able to resolve atomic structures, the methodology is better suited for smaller structures with limited flexibility. Single particle EM allows us analyze larger structures that have inherent flexibility. Protein structures can broadly be categorized as symmetry or asymmetric. There are computational advantages when analyzing symmetrical structures. Specifically, structural information can be extrapolated from fewer vantage points. Thus, symmetrical macromolecules are an advantageous for pioneering new methodologies in single particle EM. Rotavirus double layered particles (DLPs) are large macromolecular complexes that display icosahedral symmetry. Previous studies have led to a high resolution structure of transcriptionally inactive rotavirus frozen in time. However, to more fully understand rotavirus we need to examine the structure under transcriptionally active conditions. To expand our understanding, we first evaluated these viral assemblies using cryo-EM under active and inactive conditions. We found both internal and external structural differences. Based on these findings we sought to further our understanding of these nano-machines by developing a liquid cell environment to evaluate their dynamics over time. Our research not only developed a new methodology to evaluate active particles over time, we also found that the mobility of the DLPs were directly correlated to the level of transcriptional activity. When analyzing asymmetrical and flexible protein complexes previous studies have utilized methodologies to limit the proteins' conformational variability. While this does allow for a higher resolution structure, it limits our understanding to a specific orientation and compromises the biological insights. BRCA1 is an asymmetric protein containing a large flexible region and is important in the prevention of breast cancer. We utilize silicon nitride microchips with integrated wells and decorated with a lipid monolayer to capture and image BRCA1 complexes. This imaging platform minimizes heterogeneity and ensures the sample quality while not biasing confirmation. Thus, allowing for high resolution cryo-EM imaging of flexible native proteins. We were able to examine BRCA1 complexes from cells at both the primary and metastatic sites. Our ability to visualize these proteins in their native form provide insights into the variability of BRCA1 in disease progression. We found that BRCA1 complexes isolated from metastatic cells have additional density in the C-terminal domain. Our data suggests this density it due an interaction with p53. Overall, our methodologies highlight the power of single particle EM for studying protein complexes. Furthermore, our findings emphasize the importance of examining protein complexes in their native state. / PHD / Single particle electron microscopy (EM) allows us to examine the molecular world and gain insights into protein structures implicated in human disease. Visualizing the 3D architecture of macromolecules can inform drug design and preventative care. While X-ray crystallography and NMR are able to resolve atomic structures, the methodology is better suited for smaller structures with limited flexibility. Single particle EM allow us analyze larger structures that have inherent flexibility. Protein structures can broadly be categorized as symmetry or asymmetric. There are computational advantages when analyzing symmetrical structures. Specifically, structural information can be extrapolated from fewer vantage points. Thus, symmetrical macromolecules are an advantageous for pioneering new methodologies in single particle EM. Rotavirus double layered particles (DLPs) are large, highly symmetrical macromolecular complexes that represent an ideal model system for developing technology. Previous studies have led to a high resolution structure of inactive rotavirus DLP frozen in time. However, to more fully understand rotavirus we need to examine the structure under active conditions. To expand our understanding, we first evaluated these viral assemblies using cryo-EM under active and inactive conditions. We found structural differences. Based on these findings we sought to further our understanding of these nano-machines by developing a liquid cell environment to evaluate their dynamics over time. Our new methodology revealed new insights into the mobility of the DLPs. When analyzing asymmetrical and flexible protein complexes previous studies have utilized methodologies to limit the proteins’ movement. While this does allow for a higher resolution structure, it limits our understanding to a specific orientation and compromises the biological insights. BRCA1 is a highly flexible asymmetric protein implicated in the development of breast cancer. We utilize specialized microchips to capture and image BRCA1 complexes. This imaging platform ensures sample quality and allows for high resolution cryoEM imaging of flexible native proteins. We were able to examine BRCA1 complexes from cells at both the primary and metastatic sites. Our ability to visualize these proteins in their native form provide insights into the variability of BRCA1 in disease progression. Our data found that BRCA1 complexes isolated from metastatic cells are structurally different than those at the primary site. Overall, our methodologies highlight the power of single particle EM for studying protein complexes. Furthermore, our findings emphasize the importance of examining protein complexes in their native state.

Introduction: Despite advances in prevention and treatment of vaccine-preventable diseases, diarrhoeal and pneumococcal diseases remain a major source of morbidity and mortality among children worldwide. The introduction of vaccines has led to dramatic reductions in the burden of infectious diseases and mortality among children. South Africa was the first country in Africa to introduce rotavirus vaccine (RV) and pneumococcal conjugate vaccine (PCV) in 2008 as part of its national immunisation programme. Performance of immunization programmes is commonly measured by the coverage and uptake of vaccines, hence ensuring that every child is immunized at the earliest or appropriate age is an important public health goal. We therefore assessed proportions and factors associated with uptake of RV and PCV among infants who were followed during the routine demographic surveillance system of the Africa Centre Demographic Surveillance Area (DSA) in a rural South Africa setting. Methods: An open cohort of children resident in the DSA aged 12 months or below was prospectively followed between January 2008 and December 2011. Trained interviewers visited households and administered a standardised questionnaire. Mothers and caregivers were asked to show the interviewers the South African Road-To-Health (RTH) card for all children aged 12-23 months at the time of the visit or through maternal recall for children whose RTH card was not available. The RTH card includes dates of all routine vaccinations a child has received. Rotavirus vaccine doses are given at 6 and 14 weeks of age and PCV doses at 6 and 14 weeks and 9 months. Complete uptake was defined as “complete” if a child received all recommended doses of either RV or PCV and incomplete if a child did not receive any dose or received one dose of RV or PCV. Logistic regression models were used to assess factors associated with uptake of RV and PCV separately. Results: A total of 6,263 children were included in the analysis, of which 3,082 (49%) were females. At birth, 3,823 (61%) children were living in rural areas and about one-sixth of the children were living in households located far from a health facility (≥5km). The overall uptake of RV and PCV vaccines among children aged 12 months or below was 50% and 37% respectively. Infants who ever migrated outside the DSA had reduced odds of complete RV and PCV vaccination compared to infants who did not out migrate (adjusted OR=0.49, 95% CI 0.41-0.57) and (adjusted OR=0.52, 95% CI 0.43-0.63) respectively. Complete uptake of RV was associated with the increase in education levels of mothers compared secondary education (adjusted OR=1.70, 95 % CI 1.02-2.34) or tertiary education (adjusted OR=1.80, 95 % CI 0.97-2.44). Infants whose mothers were employed were less likely than infants whose mothers were not employed to have complete vaccination for RV or PCV (adjusted OR=0.71, 95 % CI 0.60-0.84) and (adjusted OR=0.81, 95% CI 0.68-0.96) respectively. Similarly, infants whose mothers were resident in the DSA were more likely than infants whose mothers were not resident to have complete vaccination for RV or PCV (adjusted OR=1.97, 95 % CI 1.49-2.60) and (adjusted OR=1.55, 95% CI 1.16-2.08) respectively. Conclusion and recommendation: The uptake of complete RV and PCV were generally low among children in rural South Africa within our study period. Child outmigration, maternal employment, maternal education and maternal residency in the DSA at child birth were associated with complete uptake of RV and PCV vaccines. Programmes targeting mothers of lower socio-economic status are required. Such programmes may include vaccine awareness and immunization campaigns at the community level to improve vaccine uptake and more targeted interventions in areas with low RV and PCV uptake.

Les rotavirus et les norovirus sont les principaux agents étiologiques des gastro-entérites en Tunisie. Pendant l’hiver 2011-2012, nous avons collecté les selles et les salives de 114 enfants âgés de moins de 6 ans, souffrant de gastro-entérites et admis à l’Hôpital Fattouma Bourguiba de Monastir. L’analyse des salives a montré que la cohorte se répartissait entre 79% de sécréteur et 21% de non-sécréteur (absence d’antigène dans les salives). Parmi les sécréteurs, les individus du groupe O étaient les plus représentés (42%) suivis des groupes A (30%), B (21%) et AB (7%) alors que 96% des patients étaient positifs pour l’antigène Lewis. Pour 98 patients, l’analyse génétique du sang a montré que le gène FUT2 se répartissait entre 77.6% de sécréteur (Se+/Se+ et Se+/se-) et 22.4% de non-sécréteurs (se-/se-, N=22).L’analyse des fèces a montré que les rotavirus et les norovirus étaient responsables respectivement de 22% et 31% des cas, les infections mixtes représentant 6% des cas. Parmi les norovirus, le génotype GII.3 était prédominant (69% de tous les NoV) tandis que le génotype G9P[8] était le plus fréquemment détecté de tous les rotavirus (37,5%). Les rotavirus ont été détectés chez les individus sécréteurs (N=28) mais aussi chez 4 patients non-sécréteurs (3 souches G9P[8] et une souche G3P[8]). Nous n’avons pas observé de distribution particulière des rotavirus en fonction des antigènes A, B et H parmi les enfants sécréteurs. En revanche, nous avons constaté que l'infection à rotavirus ne s’était produite que chez les individus positifs pour l’antigène Lewis (P=0.017). La présence de génotype P[8] chez des non-sécréteurs est inédite, elle a été confirmée par le séquençage du segment correspondant à VP8* de ces rotavirus.La majorité des infections à norovirus a été détectée chez les patients sécréteurs et cela sans distribution particulière en fonction des antigènes A, B, H et Lewis. Cinq GII.3, un GII.1 et un norovirus de génotype GII.7 ont été détectés chez les non-sécréteurs, Lewis-positifs. La production de particules de synthèse (VLP) de norovirus GII.3 en baculovirus à partir des selles d’un des patients non-sécréteurs nous a permis de tester les échantillons salivaires de toute la cohorte. L’absence d’attachement de ces VLP sur les salives des non-sécréteurs montre que la présence ou l’absence des antigènes de groupe ne reflète pas nécessairement le pouvoir infectieux des norovirus chez les jeunes enfants. Les résultats obtenus sur les rotavirus et les norovirus suggèrent qu’ils existent des voies alternatives aux antigènes de groupe sanguin pour l’attachement des rotavirus et des norovirus dans l’intestin. / Rotavirus and norovirus are the main aetiological agents of gastroenteritis in Tunisia. Stool specimens and saliva were collected from children younger than 6 years of age, admitted to the Fattouma Bourguiba Hospital (Monastir, Tunisia) for gastroenteritis during the winter 2011-12. Saliva analysis showed that 79% and 21% patients had secretor and non-secretor phenotypes, respectively. Group O blood type was predominant (42%) followed by groups A (30%), B (21%) and AB (7%), whilst 96% of the patients were positive for Lewis antigen. For 98 patients, blood samples were available and were used for FUT2 genotyping. 77.6% of the cohort were secretor (Se+/Se+ and Se+/se-) and 22.4% were non-secretor (se-/se-).Rotavirus and norovirus were found alone in 22% and 31% of the stool specimens, respectively. Mixed rotavirus-norovirus infections accounted for 6% of the cases. GII.3 noroviruses were predominant among the noroviruses whilst the G9P[8] genotype was predominant for the rotaviruses.Rotaviruses were detected in secretor (N=28) as well as in non-secretor individuals (three G9P[8] strains and one G3P[8]). No significant association was found between ABO antigens or the secretor status and RV infection. Inversely, we observed that RV infection always occurred in Lewis-positive patients (P=0.017). The presence of the P[8] genotype was confirmed by sequencing part of the VP8* coding region.There was no significant association between norovirus infection and ABO antigens and the FUT2 genotype. Five GII.3, one GII.1 and one GII.7 noroviruses were found in Lewis-positive non-secretor patients. Virus-like particles from a GII.3 norovirus infecting a non-secretor patient from the cohort were expressed in baculovirus and used for binding assay with the 114 saliva samples of the study group. VLP binding with non-secretor saliva was negative and suggested that saliva binding assay might not reflect norovirus infectivity. Overall, our data suggested that rotavirus and norovirus infection might involve non-HBGA binding pathways as well as the canonical HBGA ligands.

Submitted by Anderson Silva (avargas@icict.fiocruz.br) on 2012-06-04T18:27:54Z No. of bitstreams: 1 ludmila_n_rocha_ioc_bp_0020_2010.pdf: 2174715 bytes, checksum: ebf8bfd1505512fa4a3904c3dc6b97f6 (MD5) / Made available in DSpace on 2012-06-04T18:27:54Z (GMT). No. of bitstreams: 1 ludmila_n_rocha_ioc_bp_0020_2010.pdf: 2174715 bytes, checksum: ebf8bfd1505512fa4a3904c3dc6b97f6 (MD5) Previous issue date: 2010 / Instituto Oswaldo Cruz, Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil. / Em todo o mundo as gastroenterites infantis agudas causadas por rotavírus do grupo A (RV-A) estão associadas a cerca de 611.000 mortes por ano. RV-A pertencem à família Reoviridae, gênero Rotavirus, e são constituídos de onze segmentos de RNA dupla-fita que codificam para seis proteínas estruturais (VPs) e seis proteínas não-estruturais (NSPs). Os genótipos de RV-A são definidos por dois genes que codificam para as duas proteínas do capsídeo externo, VP4 (P) e VP7 (G). Estudos epidemiológicos demonstraram que mundialmente cinco genótipos G são mais frequentes: G1-G4 e G9; em associação com os genótipos P [8], P [4] ou P [6]. Em 2005 houve a re-emergência do genótipo G3 em crianças internadas em um hospital público no Rio de Janeiro, em concordância com a emergência desse genótipo em diversas partes do mundo, principalmente nos países asiáticos. O objetivo do presente estudo é determinar a relação entre os genes VP4, VP7 e NSP4 de RV-A genótipo G3 isoladas no Rio de Janeiro entre 1996 e 2009. A análise filogenética realizada a partir das seqüências de nucleotídeos dos genes do VP7, VP4 e NSP4 demonstrou que as amostras que circularam em 1996 e 1997 no Rio de Janeiro são distintas daquelas que foram isoladas em 2005 e 2006, sugerindo uma possível introdução deste genótipo no Rio de Janeiro. Em 2009, mais amostras de RV-A G3 foram detectadas no Rio Grande do Sul. A análise dos três genes estudados demonstrou estreita relação genética com as amostras isoladas em 2005. Recentemente, um novo sistema de classificação de rotavírus foi proposto, em que todos os 11 segmentos do genoma de RNA são utilizados e valores de cut-off foram determinados para diferenciar os genótipos. Segundo essa nova classificação, todas as amostras desse estudo foram classificadas como pertencentes aos genótipos G3 – P[8] – E1 para VP7, VP4 e NSP4, respectivamente. Em todos os genes estudados foram observadas mutações pontuais em regiões antigênicas, porém são necessários mais estudos para avaliar a importância na estrutura e função das proteínas. Foram evidenciados eventos de reestruturação genômica entre amostras humanas para o gene que codifica para VP4 (VP8*), os quais podem estar relacionados a uma vantagem adaptativa para o vírus. Os resultados do presente estudo confirmam a importância do monitoramento contínuo e da caracterização molecular das amostras de RV-A a fim de obter melhor entendimento da epidemiologia e a evolução dos RV-A genótipo G3. / Acute gastroenteritis caused by group A rotaviruses (RV-A) are associated to nearly 611,000 deaths of children worldwide per year. RV-A belong to the family Reoviridae, genus Rotavirus, and are characterized by having a segmented genome, composed of 11 segments of double-stranded RNA that encodes six structural proteins (VPs) and six non-structural proteins (NSPs). The genotypes of RV-A are defined by two genes that codify the two outer proteins, VP4 (P) and VP7 (G). Epidemiological studies in several parts of the world have indicated that there are five most common G genotypes: G1-G4 and G9; in association with P [8], P [4] or P [6] genotypes. In 2005, genotype G3 has emerged in hospitalized children in a public hospital in Rio de Janeiro, corroborating the global emergence of this genotype, especially in Asian countries. The present study aims to determine the relationship between the genes VP4, VP7 and NSP4 of stool samples positive for RV-A genotype G3 isolated in Rio de Janeiro between 1996 and 2006. Phylogenetic analyses carried out on the VP7, VP4 and NSP4 genes demonstrated that the samples that circulated in 1996 and 1997 in Rio de Janeiro are distinct from those that were detected in 2005 and 2006, which suggests a possible entering of a new G3 variant in Rio de Janeiro. In 2009, new samples of G3 were detected in Rio Grande do Sul. The analysis of the three studied genes demonstrated a straight genetic relation with the samples identified in 2005. A new rotavirus classification system has been recently proposed, in which all the 11 RNA genome segments are utilized and cut-off values were determinated to differ the genotypes. According to this new classification, all the samples in this study were characterized as genotypes G3 – P[8] – E1 to VP7, VP4 and NSP4, respectively. In all studied genes there were amino acids substitutions in antigenic regions, although more studies are necessary to evaluate the importance in the structure and protein functions. Genomic restructuration events in the VP4 (VP8*) codifying gene between human samples have been been described, which might be related to an adaptive advantage for the virus. The results of the current study confirm the importance of continuous monitoring and molecular characterization of RV-A samples with the purpose of a better understanding of RV epidemiology and evolution.