Genetic and antigenic characterization of two diarrhoeicdominant rotavirus A genotypes G3P[12] and G14P[12] circulating in the global equine population.

Equine rotavirus species A (ERVA) G3P[12] and G14P[12] are two dominant genotypes that cause foal diarrhoea with a significant economic impact on the global equine industry. ERVA can also serve as a source of novel (equine-like) rotavirus species A (RVA) reassortants with zoonotic potential as those identified previously in 2013-2019 when equine G3-like RVA was responsible for worldwide outbreaks of severe gastroenteritis and hospitalizations in children. One hurdle to ERVA research is that the standard cell culture system optimized for human rotavirus replication is not efficient for isolating ERVA. Here, using an engineered cell line defective in antiviral innate immunity, we showed that both equine G3P[12] and G14P[12] strains can be rapidly isolated from diarrhoeic foals. The genome sequence analysis revealed that both G3P[12] and G14P[12] strains share the identical genotypic constellation except for VP7 and VP6 segments in which G3P[12] possessed VP7 of genotype G3 and VP6 of genotype I6 and G14P[12] had the combination of VP7 of genotype G14 and VP6 of genotype I2. Further characterization demonstrated that two ERVA genotypes have a limited cross-neutralization. The lack of an in vitro broad cross-protection between both genotypes supported the increased recent diarrhoea outbreaks due to equine G14P[12] in foals born to dams immunized with the inactivated monovalent equine G3P[12] vaccine. Finally, using the structural modelling approach, we provided the genetic basis of the antigenic divergence between ERVA G3P[12] and G14P[12] strains. The results of this study will provide a framework for further investigation of infection biology, pathogenesis and cross-protection of equine rotaviruses.

Escherichia coli LTB26 mutant enhances immune responses to rotavirus antigen VP8 in a mouse model.

Adjuvant is a major supplementary component of vaccines to boost adaptive immune responses. To select an efficient adjuvant from the heat-labile toxin B subunit (LTB) of E. coli, four LTB mutants (numbered LTB26, LTB34, LTB57, and LTB85) were generated by multi-amino acid random replacement. Mice have been intranasally vaccinated with human rotavirus VP8 admixed. Among the four mutants, enzyme-linked immunosorbent assay (ELISA) revealed that LTB26 had enhanced mucosal immune adjuvanticity compared to LTB, showing significantly enhanced immune responses in both serum IgG and mucosal sIgA levels. The 3D modeling analysis suggested that the enhanced immune adjuvanticity of LTB26 might be due to the change of the first LTB α-helix to a ß-sheet. The molecular mechanism was studied using transcriptomic and flow cytometric (FCM) analysis. The transcriptomic data demonstrated that LTB26 enhanced immune response by enhancing B cell receptor (BCR) and major histocompatibility complex (MHC) II+-related pathways. Furthermore, LTB26 promoted Th1 and Th2-type immune responses which were confirmed by detecting IFN-γ and IL-4 expression levels. Immunohistochemical analysis demonstrated that LTB26 enhanced both Th1 and Th2 type immunity. Therefore, LTB26 was a potent mucosal immune adjuvant meeting the requirement for use in human clinics in the future.

Immunogenicity of a Rotavirus VP8* Multivalent Subunit Vaccine in Mice.

Rotavirus remains a significant public health threat, especially in low-income countries, where it is the leading cause of severe acute childhood gastroenteritis, contributing to over 128,500 deaths annually. Although the introduction of the Rotarix and RotaTeq vaccines in 2006 marked a milestone in reducing mortality rates, approximately 83,158 preventable deaths persisted, showing ongoing challenges in vaccine accessibility and effectiveness. To address these issues, a novel subcutaneous vaccine formulation targeting multiple rotavirus genotypes has been developed. This vaccine consists of nine VP8* proteins from nine distinct rotavirus genotypes and sub-genotypes (P[4], P[6], P[8]LI, P[8]LIII, P[8]LIV, P[9], P[11], P[14], and P[25]) expressed in E. coli. Two groups of mice were immunized either with a single immunogen, the VP8* from the rotavirus Wa strain (P[8]LI), or with the nonavalent formulation. Preliminary results from mouse immunization studies showed promising outcomes, eliciting antibody responses against six of the nine immunogens. Notably, significantly higher antibody titers against VP8* P[8]LI were observed in the group immunized with the nonavalent vaccine compared to mice specifically immunized against this genotype alone. Overall, the development of parenteral vaccines targeting multiple rotavirus genotypes represents a promising strategy in mitigating the global burden of rotavirus-related morbidity and mortality, offering new avenues for disease prevention and control.

Chimeric Viruses Enable Study of Antibody Responses to Human Rotaviruses in Mice.

The leading cause of gastroenteritis in children under the age of five is rotavirus infection, accounting for 37% of diarrhoeal deaths in infants and young children globally. Oral rotavirus vaccines have been widely incorporated into national immunisation programs, but whilst these vaccines have excellent efficacy in high-income countries, they protect less than 50% of vaccinated individuals in low- and middle-income countries. In order to facilitate the development of improved vaccine strategies, a greater understanding of the immune response to existing vaccines is urgently needed. However, the use of mouse models to study immune responses to human rotavirus strains is currently limited as rotaviruses are highly species-specific and replication of human rotaviruses is minimal in mice. To enable characterisation of immune responses to human rotavirus in mice, we have generated chimeric viruses that combat the issue of rotavirus host range restriction. Using reverse genetics, the rotavirus outer capsid proteins (VP4 and VP7) from either human or murine rotavirus strains were encoded in a murine rotavirus backbone. Neonatal mice were infected with chimeric viruses and monitored daily for development of diarrhoea. Stool samples were collected to quantify viral shedding, and antibody responses were comprehensively evaluated. We demonstrated that chimeric rotaviruses were able to efficiently replicate in mice. Moreover, the chimeric rotavirus containing human rotavirus outer capsid proteins elicited a robust antibody response to human rotavirus antigens, whilst the control chimeric murine rotavirus did not. This chimeric human rotavirus therefore provides a new strategy for studying human-rotavirus-specific immunity to the outer capsid, and could be used to investigate factors causing variability in rotavirus vaccine efficacy. This small animal platform therefore has the potential to test the efficacy of new vaccines and antibody-based therapeutics.

Reverse vaccinology-based multi-epitope vaccine design against Indian group A rotavirus targeting VP7, VP4, and VP6 proteins.

Rotavirus, a primary contributor to severe cases of infantile gastroenteritis on a global scale, results in significant morbidity and mortality in the under-five population, particularly in middle to low-income countries, including India. WHO-approved live-attenuated vaccines are linked to a heightened susceptibility to intussusception and exhibit low efficacy, primarily attributed to the high genetic diversity of rotavirus, varying over time and across different geographic regions. Herein, molecular data on Indian rotavirus A (RVA) has been reviewed through phylogenetic analysis, revealing G1P[8] to be the prevalent strain of RVA in India. The conserved capsid protein sequences of VP7, VP4 and VP6 were used to examine helper T lymphocyte, cytotoxic T lymphocyte and linear B-cell epitopes. Twenty epitopes were identified after evaluation of factors such as antigenicity, non-allergenicity, non-toxicity, and stability. These epitopes were then interconnected using suitable linkers and an N-terminal beta defensin adjuvant. The in silico designed vaccine exhibited structural stability and interactions with integrins (αvß3 and αIIbß3) and toll-like receptors (TLR2 and TLR4) indicated by docking and normal mode analyses. The immune simulation profile of the designed RVA multiepitope vaccine exhibited its potential to trigger humoral as well as cell-mediated immunity, indicating that it is a promising immunogen. These computational findings indicate potential efficacy of the designed vaccine against rotavirus infection.

Design of multi-epitope vaccine against porcine rotavirus using computational biology and molecular dynamics simulation approaches.

Porcine Rotavirus (PoRV) is a significant pathogen affecting swine-rearing regions globally, presenting a substantial threat to the economic development of the livestock sector. At present, no specific pharmaceuticals are available for this disease, and treatment options remain exceedingly limited. This study seeks to design a multi-epitope peptide vaccine for PoRV employing bioinformatics approaches to robustly activate T-cell and B-cell immune responses. Two antigenic proteins, VP7 and VP8*, were selected from PoRV, and potential immunogenic T-cell and B-cell epitopes were predicted using immunoinformatic tools. These epitopes were further screened according to non-toxicity, antigenicity, non-allergenicity, and immunogenicity criteria. The selected epitopes were linked with linkers to form a novel multi-epitope vaccine construct, with the PADRE sequence (AKFVAAWTLKAAA) and RS09 peptide attached at the N-terminus of the designed peptide chain to enhance the vaccine's antigenicity. Protein-protein docking of the vaccine constructs with toll-like receptors (TLR3 and TLR4) was conducted using computational methods, with the lowest energy docking results selected as the optimal predictive model. Subsequently, molecular dynamics (MD) simulation methods were employed to assess the stability of the protein vaccine constructs and TLR3 and TLR4 receptors. The results indicated that the vaccine-TLR3 and vaccine-TLR4 docking models remained stable throughout the simulation period. Additionally, the C-IMMSIM tool was utilized to determine the immunogenic triggering capability of the vaccine protein, demonstrating that the constructed vaccine protein could induce both cell-mediated and humoral immune responses, thereby playing a role in eliciting host immune responses. In conclusion, this study successfully constructed a multi-epitope vaccine against PoRV and validated the stability and efficacy of the vaccine through computational analysis. However, as the study is purely computational, experimental evaluation is required to validate the safety and immunogenicity of the newly constructed vaccine protein.

Update on Early-Life T Cells: Impact on Oral Rotavirus Vaccines.

Rotavirus infection continues to be a significant public health problem in developing countries, despite the availability of several vaccines. The efficacy of oral rotavirus vaccines in young children may be affected by significant immunological differences between individuals in early life and adults. Therefore, understanding the dynamics of early-life systemic and mucosal immune responses and the factors that affect them is essential to improve the current rotavirus vaccines and develop the next generation of mucosal vaccines. This review focuses on the advances in T-cell development during early life in mice and humans, discussing how immune homeostasis and response to pathogens is established in this period compared to adults. Finally, the review explores how this knowledge of early-life T-cell immunity could be utilized to enhance current and novel rotavirus vaccines.

The synergy of recombinant NSP4 and VP4 from porcine rotavirus elicited a strong mucosal response.

Porcine rotavirus (PoRV) is one of the main pathogens causing diarrhea in piglets, and multiple genotypes coexist. However, an effective vaccine is currently lacking. Here, the potential adjuvant of nonstructural protein 4 (NSP4) and highly immunogenic structural protein VP4 prompted us to construct recombinant NSP486-175aa (NSP4*) and VP426-476aa (VP4*) proteins, combine them as immunogens to evaluate their efficacy. Results indicated that NSP4* enhanced systemic and local mucosal responses induced by VP4*. The VP4*-IgG, VP4*-IgA in feces and IgA-secreting cells in intestines induced by the co-immunization were significantly higher than those induced by VP4* alone. Co-immunization of NSP4* and VP4* also induced strong cellular immunity with significantly increased IFN-λ than the single VP4*. Summarily, the NSP4* as a synergistical antigen exerted limited effects on the PoRV NAbs elevation, but conferred strong VP4*-specific mucosal and cellular efficacy, which lays the foundation for the development of a more effective porcine rotavirus subunit vaccine.

Bifidobacterium longum and Chlorella sorokiniana Combination Modulates IFN-γ, IL-10, and SOCS3 in Rotavirus-Infected Cells.

Rotavirus is the main cause of acute diarrhea in children up to five years of age. In this regard, probiotics are commonly used to treat or prevent gastroenteritis including viral infections. The anti-rotavirus effect of Bifidobacterium longum and Chlorella sorokiniana, by reducing viral infectivity and improving IFN-type I response, has been previously reported. The present study aimed to study the effect of B. longum and/or C. sorokiniana on modulating the antiviral cellular immune response mediated by IFN-γ, IL-10, SOCS3, STAT1, and STAT2 genes in rotavirus-infected cells. To determine the mRNA relative expression of these genes, HT-29 cells were treated with B. longum and C. sorokiniana alone or in combination, followed by rotavirus infection. In addition, infected cells were treated with B. longum and/or C. sorokiniana. Cellular RNA was purified, used for cDNA synthesis, and amplified by qPCR. Our results demonstrated that the combination of B. longum and C. sorokiniana stimulates the antiviral cellular immune response by upregulating IFN-γ and may block pro-inflammatory cytokines by upregulating IL-10 and SOCS3. The results of our study indicated that B. longum, C. sorokiniana, or their combination improve antiviral cellular immune response and might modulate pro-inflammatory responses.

Histo-Blood Group Antigen-Producing Bacterial Cocktail Reduces Rotavirus A, B, and C Infection and Disease in Gnotobiotic Piglets.

The suboptimal performance of rotavirus (RV) vaccines in developing countries and in animals necessitates further research on the development of novel therapeutics and control strategies. To initiate infection, RV interacts with cell-surface O-glycans, including histo-blood group antigens (HBGAs). We have previously demonstrated that certain non-pathogenic bacteria express HBGA- like substances (HBGA+) capable of binding RV particles in vitro. We hypothesized that HBGA+ bacteria can bind RV particles in the gut lumen protecting against RV species A (RVA), B (RVB), and C (RVC) infection in vivo. In this study, germ-free piglets were colonized with HBGA+ or HBGA- bacterial cocktail and infected with RVA/RVB/RVC of different genotypes. Diarrhea severity, virus shedding, immunoglobulin A (IgA) Ab titers, and cytokine levels were evaluated. Overall, colonization with HBGA+ bacteria resulted in reduced diarrhea severity and virus shedding compared to the HBGA- bacteria. Consistent with our hypothesis, the reduced severity of RV disease and infection was not associated with significant alterations in immune responses. Additionally, colonization with HBGA+ bacteria conferred beneficial effects irrespective of the piglet HBGA phenotype. These findings are the first experimental evidence that probiotic performance in vivo can be improved by including HBGA+ bacteria, providing decoy epitopes for broader/more consistent protection against diverse RVs.

A randomized, double-blind, placebo-controlled phase I clinical trial of rotavirus inactivated vaccine (Vero cell) in a healthy adult population aged 18-49 years to assess safety and preliminary observation of immunogenicity.

We conducted a phase I, randomized, double-blind, placebo-controlled trial including healthy adults in Sui County, Henan Province, China. Ninety-six adults were randomly assigned to one of three groups (high-dose, medium-dose, and low-dose) at a 3:1 ratio to receive one vaccine dose or placebo. Adverse events up to 28 days after each dose and serious adverse events up to 6 months after all doses were reported. Geometric mean titers and seroconversion rates were measured for anti-rotavirus neutralizing antibodies using microneutralization tests. The rates of total adverse events in the placebo group, low-dose group, medium-dose group, and high-dose group were 29.17 % (12.62 %-51.09 %), 12.50 % (2.66 %-32.36 %), 50.00 % (29.12 %-70.88 %), and 41.67 % (22.11 %-63.36 %), respectively, with no significant difference in the experimental groups compared with the placebo group. The results of the neutralizing antibody assay showed that in the adult group, the neutralizing antibody geometric mean titer at 28 days after full immunization in the low-dose group was 583.01 (95 % confidence interval [CI]: 447.12-760.20), that in the medium-dose group was 899.34 (95 % CI: 601.73-1344.14), and that in the high-dose group was 1055.24 (95 % CI: 876.28-1270.75). The GMT of serum-specific IgG at 28 days after full immunization in the low-dose group was 3444.26 (95 % CI: 2292.35-5175.02), that in the medium-dose group was 6888.55 (95 % CI: 4426.67-10719.6), and that in the high-dose group was 7511.99 (95 % CI: 3988.27-14149.0). The GMT of serum-specific IgA at 28 days after full immunization in the low-dose group was 2332.14 (95 % CI: 1538.82-3534.45), that in the medium-dose group was 4800.98 (95 % CI: 2986.64-7717.50), and that in the high-dose group was 3204.30 (95 % CI: 2175.66-4719.27). In terms of safety, adverse events were mainly Grades 1 and 2, indicating that the safety of the vaccine is within the acceptable range in the healthy adult population. Considering the GMT and positive transfer rate of neutralizing antibodies for the main immunogenicity endpoints in the experimental groups, it was initially observed that the high-dose group had higher levels of neutralizing antibodies than the medium- and low-dose groups in adults aged 18-49 years. This novel inactivated rotavirus vaccine was generally well-tolerated in adults, and the vaccine was immunogenic in adults (ClinicalTrials.gov number, NCT04626856).

Wa-VP4* as a candidate rotavirus vaccine induced homologous and heterologous virus neutralizing antibody responses in mice, pigs, and cynomolgus monkeys.

Group A rotavirus (RVA) is the primary etiological agent of acute gastroenteritis (AGE) in children under 5 years of age. Despite the global implementation of vaccines, rotavirus infections continue to cause over 120,000 deaths annually, with a majority occurring in developing nations. Among infants, the P[8] rotavirus strain is the most prevalent and can be categorized into four distinct lineages. In this investigation, we expressed five VP4(aa26-476) proteins from different P[8] lineages of human rotavirus in E. coli and assessed their immunogenicity in rabbits. Among the different P[8] strains, the Wa-VP4 protein, derived from the MT025868.1 strain of the P[8]-1 lineage, exhibited successful purification in a highly homogeneous form and significantly elicited higher levels of neutralizing antibodies (nAbs) against both homologous and heterologous rotaviruses compared to other VP4 proteins derived from different P[8] lineages in rabbits. Furthermore, we assessed the immunogenicity of the Wa-VP4 protein in mice, pigs, and cynomolgus monkeys, observing that it induced robust production of nAbs in all animals. Interestingly, there was no significant difference between in nAb titers against homologous and heterologous rotaviruses in pigs and mankeys. Collectively, these findings suggest that the Wa-VP4* protein may serve as a potential candidate for a rotavirus vaccine.

Lactiplantibacillus plantarum surface-displayed VP6 (PoRV) protein can prevent PoRV infection in piglets.

Porcine rotavirus (PoRV) poses a threat to the development of animal husbandry and human health, leading to substantial economic losses. VP6 protein is the most abundant component in virus particles and also the core structural protein of the virus. Firstly, this study developed an antibiotic-resistance-free, environmentally friendly expression vector, named asd-araC-PBAD-alr (AAPA). Then Recombinant Lactiplantibacillus plantarum (L. plantarum) strains induced by arabinose to express VP6 and VP6-pFc fusion proteins was constructed. Subsequently, This paper discovered that NC8/Δalr-pCXa-VP6-S and NC8/Δalr-pCXa-VP6-pFc-S could enhance host immunity and prevent rotavirus infection in neonatal mice and piglets. The novel recombinant L. plantarum strains constructed in this study can serve as oral vaccines to boost host immunity, offering a new strategy to prevent PoRV infection.

Recombinant bivalent subunit vaccine combining truncated VP4 from P[7] and P[23] induces protective immunity against prevalent porcine rotaviruses.

Porcine rotaviruses (PoRVs) cause severe economic losses in the swine industry. P[7] and P[23] are the predominant genotypes circulating on farms, but no vaccine is yet available. Here, we developed a bivalent subunit PoRV vaccine using truncated versions (VP4*) of the VP4 proteins from P[7] and P[23]. The vaccination of mice with the bivalent subunit vaccine elicited more robust neutralizing antibodies (NAbs) and cellular immune responses than its components, even at high doses. The bivalent subunit vaccine and inactivated bivalent vaccine prepared from strains PoRVs G9P[7] and G9P[23] were used to examine their protective efficacy in sows and suckling piglets after passive immunization. The immunized sows showed significantly elevated NAbs in the serum and colostrum, and the suckling piglets acquired high levels of sIgA antibodies from the colostrum. Challenging subunit-vaccinated or inactivated-vaccinated piglets with homologous virulent strains did not induce diarrhea, except in one or two piglets, which had mild diarrhea. Immunization with the bivalent subunit vaccine and inactivated vaccine also alleviated the microscopic lesions in the intestinal tissues caused by the challenge with the corresponding homologous virulent strain. However, all the piglets in the challenged group displayed mild to watery diarrhea and high levels of viral shedding, whereas the feces and intestines of the piglets in the bivalent subunit vaccine and inactivated vaccine groups had lower viral loads. In summary, our data show for the first time that a bivalent subunit vaccine combining VP4*P[7] and VP4*P[23] effectively protects piglets against the diarrhea caused by homologous virulent strains.IMPORTANCEPoRVs are the main causes of diarrhea in piglets worldwide. The multisegmented genome of PoRVs allows the reassortment of VP4 and VP7 genes from different RV species and strains. The P[7] and P[23] are the predominant genotypes circulating in pig farms, but no vaccine is available at present in China. Subunit vaccines, as nonreplicating vaccines, are an option to cope with variable genotypes. Here, we have developed a bivalent subunit candidate vaccine based on a truncated VP4 protein, which induced robust humoral and cellular immune responses and protected piglets against challenge with homologous PoRV. It also appears to be safe. These data show that the truncated VP4-protein-based subunit vaccine is a promising candidate for the prevention of PoRV diarrhea.

Human cytomegalovirus seropositivity and its influence on oral rotavirus vaccine immunogenicity: a specific concern for HIV-exposed-uninfected infants.

Oral rotavirus vaccines demonstrate diminished immunogenicity in low-income settings where human cytomegalovirus infection is acquired early in childhood and modulates immunity. We hypothesized that human cytomegalovirus infection around the time of vaccination may influence immunogenicity. We measured plasma human cytomegalovirus-specific immunoglobulin M antibodies in rotavirus vaccinated infants from 6 weeks to 12 months old and compared rotavirus immunoglobulin A antibody titers between human cytomegalovirus seropositive and seronegative infants. There was no evidence of an association between human cytomegalovirus serostatus at 9 months and rotavirus-specific antibody titers at 12 months (geometric mean ratio 1.01, 95% CI: 0.70, 1.45; P = 0.976) or fold-increase in RV-IgA titer between 9 and 12 months (risk ratio 0.999, 95%CI: 0.66, 1.52; P = 0.995) overall. However, HIV-exposed-uninfected infants who were seropositive for human cytomegalovirus at 9 months old had a 63% reduction in rotavirus antibody geometric mean titers at 12 months compared to HIV-exposed-uninfected infants who were seronegative for human cytomegalovirus (geometric mean ratio 0.37, 95% CI: 0.17, 0.77; P = 0.008). While the broader implications of human cytomegalovirus infections on oral rotavirus vaccine response might be limited in the general infant population, the potential impact in the HIV-exposed-uninfected infants cannot be overlooked. This study highlights the complexity of immunological responses and the need for targeted interventions to ensure oral rotavirus vaccine efficacy, especially in vulnerable subpopulations.

Caracterização molecular dos genótipos do rotavírus em pacientes pediátricos imunossuprimidos e não imunossuprimidos / Molecular characterization of rotavirus genotypes in immunosuppressed and non-immunosuppressed pediatric patients

OBJETIVO: Descrever a variabilidade genotípica do rotavírus grupo A (RVA) encontrado em pacientes pediátricos imunocompetentes e imunocomprometidos tratados no Hospital de Clínicas/Universidade Federal do Paraná (HC/UFPR), Curitiba, Paraná. MÉTODOS: Foi realizado um estudo transversal com 1.140 amostras de fezes coletadas, de abril de 2001 a dezembro de 2008, em pacientes ambulatoriais e pacientes hospitalizados com gastroenterite aguda encaminhados ao hospital. As técnicas usadas foram o método da aglutinação do látex e imunoensaio enzimático para diagnóstico de RVA. Foi realizada transcrição reversa, seguida por PCR multiplex semi-nested e sequência de nucleotídeos para caracterização do genótipo. Foram relatados dados de combinações de genótipos, clínicos, epidemiológicos, laboratoriais e sobre a presença de infecções hospitalares. RESULTADOS: Foi analisado um total de 80 amostras de fezes positivas para rotavírus. As associações mais frequentes entre os genótipos G e P foram: G4 P[8] (38,9%), G1 P[8] (30,5%), G9 P[8] (13,9%), G2 P[4] (6.9 %) e G3 P[8] 1,4%). O genótipo prevalente foi G2 P[4] depois da implementação da vacina nos anos de 2006 e 2008. Verificou-se que um total de 62,5% das crianças com idade abaixo de 12 meses estavam infectadas. Destas, 55,6% tinham grave desidratação, e 26,7% precisaram de cuidados intensivos. Encontrou-se uma frequência de 12,5% de infecções hospitalares. Não se observou correlação entre o genótipo e a gravidade da infecção nos pacientes estudados. CONCLUSÃO: As infecções por RVA podem associar-se a manifestações clínicas graves e é crucial a vigilância da variabilidade genotípica desse vírus para monitorizar a emergência de novas cepas e o impacto da imunização nesses pacientes.

OBJECTIVE: To describe the genotypic variability of group A rotavirus (RVA) found in immunosuppressed and non-immunosuppressed pediatric patients treated at the Hospital de Clínicas da Universidade Federal do Paraná (HC-UFPR), Curitiba, Paraná. METHODS: A cross-sectional study was conducted with 1,140 stool samples collected from April, 2001 to December, 2008 in outpatients and hospitalized patients with acute gastroenteritis referred to the hospital. RVA diagnosis was performed through the latex agglutination method and enzyme immunoassay. Reverse transcription followed by multiplex hemi-nested polymerase chain reaction (PCR) and nucleotide sequencing were used for genotype characterization. Genotype combinations, clinical data, epidemiological data, laboratory data, and presence of hospital-acquired infections were reported. RESULTS: A total of 80 rotavirus-positive stool samples were analyzed. The most frequent associations between genotypes G and P were: G4 P[8] (38.9%), G1 P[8] (30.5%), G9 P[8] (13.9%), G2 P[4] (6.9%), and G3 P[8] (1.4%). G2 P[4] was the most prevalent genotype after the vaccine implementation in the years 2006 and 2008. A total of 62.5% of children aged less than 12 months were found to be infected. Of these, 55.6% had severe dehydration and 26.7% needed intensive care. A frequency of 12.5% of nosocomial infections was found. No correlation was observed between genotype and severity of infection in the study patients. CONCLUSION: RVA infections can be associated with severe clinical manifestations, and the surveillance of genotypic variability of this virus is crucial to monitor the emergence of new strains and the impact of the immunization in these patients.

Efetividade da vacina oral contra rotavírus humano espécie A e monitoramento das amostras v irais circulantes em Belém, Pará, Brasil / Effectiveness of oral vaccine against human rotavirus type A and monitoring of samples v irais circulating in Belém, Pará, Brazil

Globalmente, os rotavírus da espécie A (RVA) são a principal causa de doença diarreica aguda grave em crianças abaixo de cinco anos de idade, sendo responsáveis por mais de um terço de todas as hospitalizações por diarreia e 453.000 óbitos a cada ano, principalmente nos países em desenvolvimento. Ensaios clínicos multicêntricos envolvendo aproximadamente 100.000 crianças na América Latina, Europa, África e Ásia demonstraram a segurança e a eficácia da vacina monovalente contra RVA de origem humana (Rotarix ® , GlaxoSmithKline, Bélgica), em prevenir gastroenterite grave causada por esse agente viral em crianças. No Brasil essa vacina foi introduzida no Programa Naci onal de Imunizações em 6 de março de 2006, sob a denominação de Vacina Oral contra Rotavírus Humano (VORH). Realizou-se estudo caso-controle de base hospitalar que avaliou a efetividade da vacina através da vigilância diária das hospitalizações por gastroenterite ocorridas entre crianças nascidas após seis de março de 2006, em quatro clínicas selecionadas em Belém, Pará. Consentimento por escrito foi obtido dos pais/responsável legal pela criança antes de sua inclusão no estudo. Após a hospitalização, amostras de fezes dessas crianças foram coletadas e enviadas à Seção de Virologia do Instituto Evandro Chagas para detecção dos RVA por ensaio imunoenzimático (ELISA). As amostras positivas foram posteriormente genotipadas por reação em cadeia da polimerase precedida de transcrição reversa (RT-PCR). No primeiro ano (2008-2009), 538 crianças foram incluídas no estudo como casos (gastroenterite grave por RVA) e pareadas, de acordo com a idade, a 507 controles hospitalares e 346 domiciliares; estes, sem quaisquer sintomas de gastroenterite. Haviam recebido esquema vacinal completo quanto a VORH (duas doses) 54 por cento, 61 por cento e 74 por cento dos casos, controles hospitalares e domiciliares, respectivamente...

Os resultados obtidos nesse estudo demonstraram a boa efetividade da vacina VORH frent e aos casos graves de gastroenterite causada por RVA em condições reais na população est udada, inclusive contra genótipo distinto daquele contido na composição da vacina. A lém disso, permitiu o monitoramento da circulação de amostras virais no período de três anos consecutivos em Belém, Pará, demonstrando variação nos genótipos circulantes ao longo do estudo; o que corrobora a hipótese de flutuação natural das amostras circulantes ao longo do tempo...

Vacinação contra rotavírus e intussuscepção intestinal em crianças: artigo de revisão / Rotavirus vaccination and bowel intussusception in children: review article

A doença causada pelo rotavírus é responsável por elevada taxa de hospitalização e de mortes em crianças. O desenvolvimento de vacina eficaz contra o vírus assumiu caráter prioritário para organismos como a Organização Mundial de Saúde (OMS). A intussuscepção intestinal, entretanto, representa manifestações relacionadas ao uso da vacina que ainda constitui preocupação quanto ao seu uso disseminado. No Brasil, encontram-se disponíveis as vacinas do tipo RV1 e RV5, não sendo observado aumento de risco significativo de intussuscepção nas crianças com menos de sete dias de vida. Os benefícios que as vacinações trouxeram, pelas reduções de hospitalizações e mortalidade infantil no mundo, continuam a indicar a sua vacinação. (AU)

The overall impact of the disease caused by rotavirus is still a major cause of hospitalizations and deaths in children. The development of an effective vaccine against the virus became a priority for organizations like WHO (World Health Organization). Intestinal intussusception is one manifestation of which was related to vaccine use. In Brazil, are available vaccines RV1and RV5 type, and there was no significant increase in the risk of intussusception in children less than seven days. The benefits brought vaccinations, by reductions in hospitalizations and mortality in the world, continue to indicate the worldwide vaccination against rotavirus by WHO. (AU)

Avaliação da ação neutralizante e da reatividade de anticorpos IgA e IgG anti-rotavírus SA-11 em soro de adultos saudáveis / Evaluation of neutralizing ability and reactivity of anti-rotavirus SA-11 IgA and IgG antibodies in serum samples from healthy adults

O rotavírus é a principal causa de diarréia em crianças em todo o mundo. Infecta também adultos, mas não há dados completos sobre a sua incidência nesse grupo nem sobre o papel de anticorpos preexistentes na proteção contra o vírus. O objetivo do trabalho foi avaliar a presença de anticorpos IgA e IgG anti-rotavírus SA-11, por ELISA, em amostras de soro de adultos saudáveis e sua ação neutralizante frente ao vírus, em ensaios de neutralização. Por Immunoblotting foi avaliado o reconhecimento de proteínas virais pelos anticorpos séricos. Observou-se que os títulos das amostras foram muito variáveis, sendo os de IgG superiores aos de IgA. Todas as amostras mostraram-se capazes de neutralizar o vírus em diferentes níveis, porém não foi possível estabelecer uma correlação com os títulos de anticorpos. Foi observado que anticorpos da classe IgG reconhecem mais proteínas virais que os da classe IgA. Este trabalho pode ser considerado mais um passo na elucidação do papel dos anticorpos séricos IgA e IgG anti-rotavírus na infecção em adultos.

Rotavirus has been considered the leading cause of diarrhea in children worldwide. The virus also infects adults but there is no conclusive data neither on the incidence of infection on this group nor on the role of pre-existing antibodies. The aim of the work was to evaluate the presence of anti-rotavirus SA-11 IgA and IgG by ELISA in serum samples of healthy adults and the serum neutralizing ability against the virus by neutralization assays. Immunoblotting was used to evaluate viral proteins recognition by serum antibodies. The antibody titers were extremely variable where IgG titers are greater than IgA ones. All samples were able to neutralize the virus in different levels but it was not possible to establish a correlation between antibody titers and neutralization ones. Immunoblotting assays revealed that IgG antibodies recognize more viral proteins than IgA did. This work can be considered a valuable step for elucidating the role of serum anti-rotavirus IgG and IgA antibodies in adults infection.