Development of a Real-time PCR test for porcine group A rotavirus diagnosis / Desenvolvimento de um teste de PCR em Tempo Real para o diagnóstico de rotavírus suínos do Grupo A

Group A Rotavirus (RVA) is one of the most common causes of diarrhea in humans and several animal species. A SYBR-Green Real-Time polymerase chain reaction (PCR) was developed to diagnose RVA from porcine fecal samples, targeting amplification of a 137-bp fragment of nonstructural protein 5 (NSP5) gene using mRNA of bovine NADH-desidrogenase-5 as exogenous internal control. Sixty-five samples were tested (25 tested positive for conventional PCR and genetic sequencing). The overall agreement (kappa) was 0.843, indicating 'very good' concordance between tests, presenting 100% of relative sensitivity (25+ Real Time PCR/25+ Conventional PCR) and 87.5% of relative sensitivity (35- Real Time PCR/40- Conventional PCR). The results also demonstrated high intra- and inter-assay reproducibility (coefficient of variation ≤1.42%); thus, this method proved to be a fast and sensitive approach for the diagnosis of RVA in pigs...

Rotavírus do grupo A (RVA) é uma das causas mais frequentes de diarreias em humanos e várias espécies animais. Um teste de PCR em Tempo Real com SYBR-Green foi desenvolvido visando o diagnóstico de RVA a partir de fezes suínas, através da amplificação de um fragmento de 137 pares de bases do gene da proteína não estrutural 5 (NSP5) viral e de mRNA de NADH-desidrogenase-5 bovina como controle interno exógeno. Foram testadas 65 amostras (25 delas positivas por PCR convencional e sequenciamento nucleotídico). A concordância entre os testes foi de 0,843, considerada "muito boa", apresentando 100% de sensibilidade relativa (25+ PCR Tempo Real/25+ PCR convencional) e 87,5% de sensibilidade relativa (35- PCR Tempo Real/40- PCR convencional). Os resultados também demonstraram elevada reprodutibilidade inter e intra-ensaio (coeficiente de variação ≤ 1,42%); portanto, este método demonstrou ser uma rápida e sensível alternativa para o diagnóstico de RVA em suínos...

Detecção de rotavírus do grupo A em amostras fecais: adaptação do teste de aglutinação em látex para o emprego de anticorpos IgY anti-Rotavírus A / Detection of group a rotavirus in fecal samples: adaptation of latex agglutination test for anti-rotavirus a igy antibody employment

A infecção pelo rotavírus A (RVA) é responsável por cerca de 453.000 mortes anualmente e aproximadamente 40 por cento das hospitalizações por diarreia em crianças menores de cinco anos em todo o mundo, sendo o principal causador de gastroenterite aguda nesse grupo populacional. O desenvolvimento de um método de diagnóstico rápido, barato, sensível e específico para a detecção de RVA é importante do ponto de vista epidemiológico porque permite identificar surtos no local de ocorrência. O uso da imunoglobulina Y (IgY), anticorpo purificado a partir da gema de ovo, vem crescendo nos últimos anos, devido às características vantajosas quando comparada com a imunoglobulina G (IgG), como a obtenção de anticorpos de forma não invasiva e a produção de anticorpos em grandes concentrações. Este trabalho objetivou a adaptação de um teste de diagnóstico através da substituição do anticorpo de captura IgG pela IgY específica para o antígeno de RVA (LATEXY-ROTA). Para isto 09 frangas poedeiras foram imunizadas com o RVA, os ovos foram coletados e a IgY purificada a partir da gema do ovo por polietileno glicol 6.000, seguida da purificação adicional por cromatografia de troca iônicaA IgY anti-RVA purificada foi ligada covalentemente à partículas de poliestireno e usada como insumo na adaptação de um teste de aglutinação em látex, sendo testada em um painel de amostras fecais sabidamente positivas e negativas previamente selecionadas pelo Centro Regional de Referência para Rotaviroses do Laboratório de Virologia Comparada e Ambiental (LVCA/IOC-FIOCRUZ)...

Foi obtida uma sensibilidade de 75 por cento e uma especificidade de 87,5 por cento quando o LATEXY-ROTA foi comparado com um teste imunoenzimático comercial disponível (padrão ouro). Quando comparado com dois testes comerciais de aglutinação em látex testados no painel de amostras utilizando a IgG, o LATEXY-ROTA obteve uma sensibilidade de 100 por cento e especificidade de 88,24 por cento. Baseado nos dados obtidos, sugerimos a viabilidade da substituição da IgG por IgY no ensaio de aglutinação pelo látex...

The infection by rotavirus (RV) is responsible for approximately 453,000 deaths annually and approximately 40 percent of hospitalizations by diarrhea in children under five years worldwide, being the major cause of acute gastroenteritis in this populationgroup. The development of a rapid method, inexpensive, sensitive and specific for rotavirus diagnosis is important from the disease because it allows the identification of outbreaks in the site of occurrence. The use of Immunoglobulin Y (IgY) antibodypurified from egg yolk, has been grown in recent years, due to the advantageous features compared to immunoglobulin G (IgG), as a noninvasive recovery of antibodies and production in high concentrations. The aim of this method was to adapt a diagnostic test by replacing the IgG capture antibody by specific IgY for RVAantigen (LATEXY-ROTA). For that, 09 laying hens were immunized with RVA, the eggs were collected and IgY purified from egg yolk by polyethylene glycol 6,000, followed by purification by ion exchange chromatography. The purified anti-IgY RVA was covalently bound to polystyrene particles, being tested in a panel of positive and negative fecal samples previously determined by the Rotavirus Regional ReferenceCenter of Comparative and Environmental Virology Laboratory (LVCA/IOCFIOCRUZ). A sensitivity of 75% and specificity of 85,7% was observed when the adapted test was compared to a commercial available enzyme immunoassay (goldenstandard). When compared to two commercial latex agglutination tests using the IgG tested on the panel of samples, the LATEXY-ROTA had a sensitivity of 100 percent and specificity of 88.2 percent. Based on the obtained data, we suggest the feasibility of replacing the IgG by IgY in the latex agglutination assay...

Rotavirus G2P[4] and G2P[4]+[6] infections during norovirus gastroenteritis outbreak: summer season 2010, Brazil

Introduction This study aimed to monitor the seasonality of rotavirus infection, and gain insight into the variability of Brazilian strains. Methods A total of 28 stool samples were analyzed from 698 revised cases of gastroenteritis during a norovirus outbreak in the summer of 2010 in Guarujá, Brazil. Diagnosis was performed using enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and sequencing. Results Rotavirus infection was detected in 17.9% (5/28) of samples; 4 samples were G2P[4] genotype, and one G2P[4]+P[6] genotype. G2 and P[4] sequences showed a genetic relationship to strains from India and Russia, respectively. Conclusions The seasonal pattern of rotavirus may be a consequence of human activity apart from climate factors. .

Síndrome diarreico por rotavirus en niños menores de 5 años inmunizados y no de la ciudad de Maracaibo, estado Zulia, Venezuela / Diarrheic syndrome due to rotavirus in immunized and non-immunized children under 5, in the city of Maracaibo, state of Zulia, Venezuela

La diarrea aguda es la causa más común de morbi-mortalidad infantil, donde el Rotavirus es uno de los principales agentes involucrados en las diarreas severas en niños menores de 5 años. El presente estudio tiene como objetivo determinar la presencia de rotavirus en una población infantil con cuadros diarreicos, vacunados y no vacunados que acuden a observación pediátrica del Hospital de Niños y Hospital Universitario de Maracaibo, estado Zulia en el período de Abril 2011 hasta Abril 2012, para ello se recolectaron 100 muestras de heces y para el diagnóstico se utilizó la técnica de Aglutinación Directa en partículas de Látex. Resultaron positivos un 43,6% (24/55) de niños no vacunados y 33,3% (10/30) de niños vacunados afectando principalmente a los menores de 1 año con predominio del sexo masculino con un 65,9% sobre el sexo femenino con 40%, la diarrea por el agente viral estudiado representó casi la mitad de las hospitalizaciones por gastroenteritis, encontrándose una diferencia estadística significativa de p= 0.027 en el numero de evacuaciones del cuarto día, entre niños no vacunados (5) y vacunados (3). Los resultados de esta investigación muestran la circulación de rotavirus en niños vacunados y no vacunados, es por ello importante considerar que el presente reporte tiene como fin último alertar a la comunidad médica a considerar en todo niño con gastroenteritis la posibilidad de que se trate de un cuadro por RV.

Acute diarrhea is the most common cause of morbidity and mortality in infants, where rotavirus is one of the main agents involved in severe diarrhea for children under five years. The present study aims to determine the presence of rotavirus in a vaccinated and unvaccinated pediatric population with acute diarrhea that came to the Children’s Hospital and the University Hospital of Maracaibo, State of Zulia, for observation from April 2011 to April 2012. One-hundred stool samples were collected and direct agglutination of latex particles was used for diagnosis. 43.6% of the unvaccinated children were positive (24/55) as were 33.3% (10/30) of the vaccinated children. Principally, children under 1 year were affected, predominantly males with 65.9% and females with 40%. Diarrhea due to the viral agent under study accounted for nearly half the hospitalizations for gastroenteritis. A statistically significant difference of p = 0.027 was found between the number of evacuations on the fourth day in unvaccinated (5) and vaccinated (3) children. Results of this research show rotavirus circulating in vaccinated and unvaccinated children. Therefore, the purpose of this report is to alert the medical community to consider that every child with gastroenteritis could possibly have RV.

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...

An unusual case of concomitant infection with chicken astrovirus and group A avian rotavirus in broilers with a history of severe clinical signs

A molecular study of intestinal samples from 21 broiler flocks with a history of enteritis revealed that 23.8% and 14.3% were positive for chicken astrovirus (CAstV) and avian rotavirus (ARV), respectively. CAstV and group A ARV were simultaneously detected in only one broiler flock. Birds in this group developed the significant intestinal lesions characterized by frothy contents, paleness, and thin intestinal walls. In this report we present an unusual case of runting stunting syndrome (RSS) with a history of high mortality and growth retardation in broiler chickens. We also make the first identification of CAstV and group A ARV in broiler chickens in Korea.

Uso de una prueba rápida de rotavirus en la prescripción de antibióticos en diarrea aguda pediátrica: un estudio observacional, aleatorizado y controlado / Use of rotavirus rapid test in the prescription of antibiotics in acute pediatric diarrhea: an observational study, randomized controlled

OBJETIVO: Evaluar el efecto de la disponibilidad de un método diagnóstico rápido y preciso de rotavirus sobre la frecuencia en el uso de antibióticos en niños menores de 5 años con diarrea aguda de menos de 5 días de duración del Hospital Nacional Arzobispo Loayza; Lima, Perú. MATERIALES Y MÉTODOS: se realizó un estudio observacional, prospectivo, aleatorizado, controlado, entre Julio 2008 y Enero 2009. Se asignó aleatoriamente 101 pacientes al Grupo A (con prueba de leucocitos fecales) y 100 al Grupo B (con prueba de leucocitos fecales y prueba rápida de rotavirus/adenovirus). Se evaluó en cada grupo la decisión clínica de administrar o no antibióticos. RESULTADOS: No existieron diferencias significativas en el tiempo de enfermedad, edad ni peso entre ambos grupos. Los casos de rotavirus tuvieron leucocitos fecales positivos en un 46.9%. El grado de positividad de los leucocitos fecales se asoció de manera directamente proporcional al uso de antibióticos (P<0.0001). El grupo positivo a rotavirus presentó un mayor riesgo de hospitalización que el grupo negativo a este agente (21.9% vs. 6.3%, P<0.0001). No se registraron muertes atribuibles a diarrea. CONCLUSIONES: La disponibilidad de un diagnóstico rápido y preciso de rotavirus en menores de 5 años con diarrea aguda, redujo significativamente el uso de antibióticos.

OBJECTIVE: To determine the impact of a rapid and accurate rotavirus test in the emergency ward on the reduction of antibiotic prescription in children under 5 years old with acute diarrhea at ôArzobispo Loayza National Hospitalõ, Lima, Peru. MATERIAL AND METHODS: We performed an observational prospective randomized controlled study, from July 2008 to January 2009. Stool samples from patients with diarrhea lasting less than 5 days were analyzed. Out of 201 cases, 101 were classified in Group A (with fecal leukocytes test performed) and 100 in Group B (with fecal leukocytes test and rotavirus/adenovirus test performed). We aimed to associate the signs and symptoms with the decision of prescribing antibiotics and with hospitalization risk. RESULTS: Both groups were comparable with regard to age, weight and illness duration. In patients with rotavirus infection, fecal leukocytes were positive in 46.9% of cases. Frequency of antibiotic use was directly associated with the number of fecal leukocytes (P<0.0001). There was a higher risk of admission in the group positive to rotavirus than in the group negative to this agent (21.9% vs. 6.3, P<0.0001). No diarrhea-attributable deaths were reported. CONCLUSION: The use of rotavirus test in the pediatric emergency room decreased antibiotic prescription in children with diarrhea.

Rotavirus infection in a tertiary hospital: laboratory diagnosis and impact of immunization on pediatric hospitalization

BACKGROUND AND OBJECTIVES: Rotavirus (RV) is the main etiological agent of diarrhea in childhood; its laboratory diagnosis is crucial to guide the clinical management and prevention of its spread. RV immunization was introduced in Brazilian 6-month-old children in 2006. The present study was aimed to evaluate three methodologies used for human RV detection in stool samples obtained from patients hospitalized due to gastroenteritis in a teaching hospital and report the impact of RV immunization in hospitalization by diarrhea. METHODS: 293 stool samples collected in the 2001-2008 period were analyzed by enzyme immunoassay (EIA), latex agglutination (LA) and polyacrylamide gel electrophoresis (PAGE). RESULTS: Rotavirus was detected in 34.8 percent of samples by LA assay, 28.3 percent of samples by EIA assay and in 25.6 percent of samples by PAGE assay. Considering the PAGE method as gold standard, the sensitivity, specificity and accuracy of EIA were 94.6 percent, 94.4 percent and 94.5 percent, and to LA were 82.6 percent, 81.6 percent and 81.9 percent, respectively. CONCLUSION: These results indicate that antigen detection by EIA is a rapid, sensitive and specific method, and could be used in large-scale applications for screening stool samples suspected of RV infection. This study showed decreased incidence of RV infection in hospitalized children prior to the implementation of the national immunization program against RV.

Detection and characterization of human rotavirus in hospitalized patients in the cities of Ponta Grossa, Londrina and Assai - PR, Brazil

Acute diarrheal disease is still one of the major public health problems worldwide. Rotaviruses (RV) are the most important viral etiologic agents and children under five years of age are the target population. OBJECTIVE: To investigate the rate of RV infection in hospitalized patients due to acute diarrhea in the cities of Ponta Grossa, Londrina and Assai - Paraná. METHODS: Latex agglutination (LA); immunochromatography (ICG); polyacrylamide gel electrophoresis (PAGE) and negative staining electron microscopy (ME) tests were used to detect the virus. For the genotyping, RT-PCR and RT-PCR-ELISA were used, respectively, for NSP4 and VP4/VP7. RESULT: Out of 124 samples there were 69 positive stool samples for RV, for at least one of the used tests, 67 of them being RV group A (RV-A). Overall, most of the RV positive stool samples came from children under thirteen years of age. However, 12 positive cases occurred in patients aged 13 years or above, including an 81-year old patient. CONCLUSION: The data showed similar electropherotypes and genotypes G, P and NSP4 of the inland wild circulating strains of RV.

Prevalência de rotavírus em crianças atendidas na rede pública de saúde do estado de Pernambuco / Rotavirus prevalence in infants and children in the public healthcare system of the state of Pernambuco

INTRODUÇÃO: Os rotavírus são considerados, importantes agentes etiológicos de gastroenterite aguda e ainda causa comum de hospitalização de crianças na faixa etária de zero a quatro anos de idade. No Brasil, a incidência de rotavírus nas crianças com gastrenterite é de 12 a 42 por cento, e a distribuição da infecção esta relacionada à sazonalidade que aparentemente ocorre em diferentes períodos e intensidade de acordo com a região de ocorrência. O estudo pesquisou rotavírus do grupo A em amostras fecais de crianças com caso suspeito atendidas na rede pública de saúde do Estado de Pernambuco. MÉTODOS: O diagnóstico foi realizado através de ensaios imunoenzimático ELISA e teste imunoquímico de aglutinação em látex. RESULTADOS: Foram estudadas 171 amostras. Destas, 33 (19,3 por cento) apresentaram positividade. Das amostras positivas 24 (72,7 por cento) eram do sexo masculino e 09 (27,3 por cento) do sexo feminino. Dentro da amostragem positiva 15,2 por cento eram vacinadas. Quando comparamos os resultados obtidos entre o teste Elisa e aglutinação pelo Látex, houve 100 por cento de concordância entre a positividade pelo Látex e o ELISA. CONCLUSÕES: A alta incidência desta infecção reforça necessidade de monitoramento desse vírus, definindo políticas de saúde relacionadas ao diagnóstico, profilaxia, melhores condições sócio-econômicas e aprimoramento da vacina.

INTRODUCTION: Rotaviruses are considered important etiological agents of acute gastroenteritis and a common cause of the hospitalization of children aged zero to four years-old. In Brazil, the incidence of rotavirus gastroenteritis in children is 12 to 42 percent and the distribution of infection is related to seasonality, which apparently occurs in different periods and intensity according to each region. The study investigated group A rotavirus in fecal samples of suspected children attended by the public health system of the State of Pernambuco. METHODS: Diagnosis was achieved by ELISA and the Latex agglutination test. RESULTS: Of the 171 samples studied, 33 (19.3 percent) presented positivity for Rotavirus A. Among positive samples, 72.7 percent belonged to male patients and 27.3 percent to female. Among the positive casuistic, 15.2 percent were vaccinated. When comparing the results obtained for the ELISA and Latex agglutination tests, 100 percent agreement between positivity by Latex agglutination and ELISA was verified. CONCLUSIONS: The high incidence of this infection reinforces the need to monitor this virus and define health policies concerning its diagnosis, prophylaxis, improvement in socioeconomic conditions and the refinement of the vaccine.

Prevalence of rotavirus in feces of children with acute diarrhea: clinical signs and associated symptoms / Prevalência de rotavírus em fezes de crianças com diarreia aguda: sinais e sintomas clínicos associados

Introduction: Rotavirus is a very common etiologic agent in acute diarrheas in children worldwide, affecting mainly children under five years of age. The overall annual mortality associated with rotavirus infection was estimated at 454000-705000 deaths annually, predominantly in the developing countries. The aim of this study was to describe the prevalence and serotypes of rotavirus in a region of Brazil, as well as to present a clinical evaluation of this diarrheal disorder in children. Methods: Cross-sectional study which selected 124 samples from the same number of children under five years of age with acute gastroenteritis, attended to in a Basic Health Unit. Results: Thirty-one samples were positive for rotavirus A, with higher occurrence from September to March (p <0.05). With respect to genotype G, 16 (80%) samples were rated as G1. P genotype showed that all strains were P[8]. Dehydration was found 2.45 times more often among children infected with rotavirus. Conclusion: It is hoped that determining the prevalence of infection and disease caused by rotavirus and the characterization of the profile of circulating viral strains may contribute to information on the molecular biology and epidemiology of infection by that agent

Introdução: O rotavírus é agente etiológico muito comum em diarreias agudas em crianças, em todo o mundo, afetando principalmente crianças menores de cinco anos de idade. A mortalidade anual global associada a infecções por rotavírus foi estimada em 454.000 a 705.000 mortes por ano, predominantemente nos países em desenvolvimento. Os objetivos deste estudo foi descrever os sorotipos circulantes e a prevalência de rotavírus em uma região do Brasil, bem como, apresentar a caracterização clínica da doença diarréica em crianças. Métodos: Estudo transversal que selecionou 124 amostras de um mesmo número de crianças de até cinco anos de idade, com gastrenterite aguda, atendidas em uma Unidade Básica de Saúde. Resultados: Trinta e uma amostras foram positivas para rotavírus A com maior ocorrência de setembro a março (p < 0,05). Com relação ao genótipo G, 16 (80%) amostras foram classificadas como G1. O genótipo P mostrou que todas as cepas eram P[8]. A desidratação foi encontrada 2,45 vezes mais entre crianças infectadas com rotavírus. Conclusão: Espera-se que a determinação da prevalência da infecção e doença provocada pelo rotavírus e a caracterização do perfil de cepas virais circulantes possa contribuir para informação sobre a biologia molecular e a epidemiologia da infecção pelo referido agente

Application of a Diagnostic Method Using Reverse Transcription-PCR ELISA for the Diagnosis of Enteroviral Infections / 대한진단검사의학회지

BACKGROUND: Enteroviruses are known as major pathogen for aseptic meningitis. Although rapid diagnosis for enteroviruses is very essential to exclude bacterial infections in patients with meningitis, classical diagnostic method based on virus isolation is not practicable for timely treatment of patients due to its laborious and time-consuming procedure. Recently molecular methodologies as alternatives are routinely used for rapid and sensitive diagnosis for enteroviruses infections. METHODS: Reverse transcription (RT)-PCR ELISA kit for targeting 5'non-coding region (NCR) with highly conserved genetic identity among all genotypes of enteroviruses was introduced in this investigation. RT-PCR ELISA was evaluated about sensitivity and specificity through virus isolation using clinical specimens from patients suspected of enteroviral infections and enteroviral isolates comparing with conventional RT-PCR identifying them. RESULTS: The detection limit of the RT-PCR ELISA was up to 10-100 folds higher than virus isolation using cell culture and conventional RT-PCR. On comparison between above two methods, the detection rate of RT-PCR ELISA for clinical specimens from patients with aseptic meningitis was 7% higher than that of conventional RT-PCR targeting 5'NCR (P=0.016). CONCLUSIONS: Our results suggest that RT-PCR ELISA developed in this study could be an alternative diagnostic method for the detection of enteroviral genome with high sensitivity and specificity.

Evaluación de dos equipos inmunocromatográficos comerciales para el diagnóstico rápido de la infección por rotavirus / Evaluation of two immunochromatography kits for rapid diagnosis of rotavirus infections

Se realizó un estudio prospectivo para evaluar dos equipos comerciales inmunocromatográficos para el diagnóstico rápido de infección por rotavirus a partir de muestras fecales: VIKIA® Rota-Adeno, de bioMérieux, y Simple Rota- Adeno, de Operon. Como método de referencia se utilizó la transcripción reversa y reacción en cadena de la polimerasa (RT-PCR) con cebadores específicos del gen de la proteína VP7 de rotavirus del grupo A. La sensibilidad y la especificidad respecto de la RT-PCR fueron del 98,4% y 84,8% para el Simple Rota-Adeno, y del 100% y 24,2% para el VIKIA® Rota-Adeno. Es de destacar la baja especificidad de este último equipo diagnóstico, que presentó un elevado número de falsos positivos, por lo que el valor predictivo de un resultado positivo es sólo del 71,6%. Asimismo, se identificaron los genotipos de las cepas de rotavirus detectadas; la mayoría de ellas correspondieron al genotipo G9P(8) (65%), seguido de los genotipos G1P(8) (25,4%) y G2P(8) (3,2%).

A prospective study was conducted to evaluate two immunochromatography (ICG) commercial kits for diagnosis of rotavirus infection, VIKIA® Rota-Adeno (bioMérieux) and Simple Rota-Adeno (Operon). Reverse transcriptase and polymerase chain reaction (RT-PCR) with specific primers for the VP7 gene of group A rotavirus was used as the reference method. The sensitivity and specificity of the ICG tests compared with those of the reference method were 98.4% and 84.8%, respectively, for Simple Rota-Adeno (Operon), and 100% and 24.2% for VIKIA® Rota-Adeno (bioMérieux). It is remarkable the low specificity of the latter method, which yields a high number of false positive results. The predictive value of a positive result by this method was only 71.6%. Most of the detected rotavirus strains corresponded to genotype G9P(8) (65%), followed by G1P(8) (25.4%) and G2P(8) (3.2%).

Hospitalization due to norovirus and genotypes of rotavirus in pediatric patients, state of Espírito Santo

Viruses are the leading cause for hospitalization due to gastroenteritis worldwide. Group A rotaviruses (RV) are the most prevalent and are assorted in glycoproteins (G) and protease sensitive (P) dual genotypes based on polymorphic genes that encode the external VP7 and VP4 capsid proteins, respectively. Noroviruses (NoV) have increasingly answered by sporadic gastroenteritis. This study aimed to determine the prevalence of NoV and RV in 68 hospitalized children, between July 2004 and November 2006, at a pediatric hospital in Vitória city, state of Espírito Santo, Southeastern Brazil. Nucleic acid was extracted from fecal suspension following the guanidine-silica procedure. Reverse transcriptase-polymerase chain reaction (RT-PCR) and polyacrylamide gel electrophoresis were employed for NoV and RV detection, respectively. RV genotyping was accomplished using RT-PCR followed by heminested multiplex PCR with specific primers for the most prevalent types of G and P. Fecal samples were positive for NoV and RV in 39.7 percent (27/68) and 20.5 percent (14/68), respectively and together were responsible for 60 percent (41/68) of the cases. RV genotypes were: 50 percent G9P[8], 28.7 percent G2P[4], 7.1 percent G1P[8], G2P[8] and G?P[8]. Vomit was a prominent manifestation observed in 92 percent and 85 percent of the NoV and RV cases, respectively. The median hospitalization was 5 and 5.5 days for the patients infected with NoV and RV, respectively. The data showed that NoV prevailed over RV and it also corroborated the emergence of RV G9 genotype followed by G2P[4], reinforcing the need for RV genotype surveillance.

Rotavirus infection in children and adult patients attending in a tertiary Hospital of São Paulo, Brazil

During the period of January 2003 to December 2005, 3,768 stool samples were received in the Microbiology Laboratory for rotavirus antigen detection from outpatients and inpatients of Albert Einstein Hospital, SP. Fresh stool samples from children and adults were analyzed by two methodologies: during 2003 and 2004 by latex agglutination (Slidex Rotavirus, Biomerieux) and 2005 by an immunochromatographic assay for the combined detection of rotavirus and adenovirus (Vikia Rota-Adeno, Biomerieux). Rotavirus group A was detected in 755 (20 percent) samples. The annual prevalence was 19.8 percent in 2003, 21.7 percent in 2004, and 18.7 percent in 2005. Rotavirus was detected every month during the period of the study, with peak of positivity between June and August (>35 percent). The prevalence in hospitalized patients was 26.1 percent (352/1,350) and in outpatients was 16.7 percent (403/2,418). For hospitalized patients most of the rotavirus infections were diagnosed in Pediatric setting, age range of 0 to 10 years (prevalence of 55.3 percent, 295/534). Overall positivity was up to 30 percent in patients between six months and five years of age (67 percent of all positive patients), all other age groups had at least 10 percent positive tests. Rotavirus infection is common in Sao Paulo, and besides the expected higher frequency in children it is also frequent in adults.

Burden of rotavirus gastroenteritis among under five years children at Al-Alwaiya paediatric hospital

Rotavirus is the main cause of diarrhea in <5 years children causing high rates of hospitalizations and high mortality. To estimate the burden of rotavirus gastroenteritis among less than five years children attending Al-Alwaiya Pediatric Hospital. To examin the validity of ELISA and Latex screening tests and to find out the concordane rates. Across sectional study involved under five years[1-59months] children with acute gastroenteritis attending Al-Alwaiya Pediatric Hospital for the period from 1[st] of January 2008 to 31[st] of May 2008. The questionnaire used depended on WHO protocol of the National Rotavirus Gastroenteritis Surveillance Program. LATEX and ELISA tests were used for diagnosis. The period of study included two seasons: Winter and Spring. Stool samples of 342 children with acute gastroenteritis were investigated by general stool examination, stool culture, LATEX and ELISA tests for the presence of any parasite, bacteria and/ or rotavirus antigen respectively. Rotavirus was detected in 162[47.4%] of children with acute gastroenteritis either by LATEX, ELISA or both. Total concordance rate between the two tests was as high as 85.4%. Females were 61[37.7%] and males were 101[62.3%]. Age group mostly affected was [>6-12] months which was [31.4%]. Rotavirus cause 47.4% of acute gastroenteritis cases of the studied sample with no significant statistical difference between rotavirus and other causes of gastroenteritis regarding demographic features [except for gender, more in males], clinical features, and type of treatment and course of disease [except time of onset].Concordance rate between ELISA and Latex was found to be high confirming their validity

[Diagnosis of rotavirus and adenovirus type 40/41 in children with acute diarrhea by immunochromatography in Mofid Children Hospital]

Acute diarrhea is one of the leading causes of worldwide mortality and morbidity; four millions annual deaths. The aim of this study is to investigate the prevalence of rotavirus and adenovirus among children with acute diarrhea and to evaluate their clinical and laboratory findings. 199 children aged 1 to 168 months with acute diarrhea were studied in "Mofid" Children Hospital for a one year period. Clinical and laboratory findings including stool culture and WBC and RBC counts in their stool samples were analyzed. From 199 children with diarrhea, 14 cases [7%] were positive for viruses; 11 [5.5%] rotaviruses and 3[1.5%] adenoviruses. In Rotavirus and adenovirus, male to female ratio were 1.2/1 and 2/1, and the mean age were 40.8 and 20.4 months, respectively. Clinical findings comprised frequency, dehydration, fever, vomiting and watery diarrhea. Stool culture was negative and no RBC and WBC were detected. High prevalence of rotavirus and adenovirus in former studies and low prevalence of rotavirus in this study seems to be caused by the diagnostic method we used. Since a fast laboratory process is necessary for immunochromatography, other methods as EIA and PCR are recommended

Rotavirus and adenovirus in Rondônia

Acute gastroenteritis is one of the most common diseases in humans worldwide. Viral gastroenteritis is a global problem in infants and young children. In this study the incidence of diarrhea was assessed in 877 hospitalized children under five years old, over a period of 24 months and distributed in 470 cases of diarrhea and 407 age-matched group with other pathologies, as control group. Two antigen detection techniques based on enzyme immunoassay (EIA) and latex particles were used for detection of rotavirus and adenovirus. Rotavirus A was a major cause of gastroenteritis with 23.6 percent of cases, being 90 percent of these cases in young children. Adenovirus infections was detected by EIA with frequency of 6.4 percent. Rotavirus and adenovirus were detected in 10.1 and 1.7 percent of stools from control group, respectively. Interestingly, the frequency of the youngest children in the control group excreting Rotavirus A was comparable to that detected in stools from diarrheic children. We cannot rule out the existence of other enteric viruses because the etiology of 171 cases of diarrhea was not determined and active search for astrovirus and calicivirus was not done. This is the first study that shows the presence of enteric viruses in the infantile population from Western Brazilian Amazonia and it was important to help physicians in the treatment of viral gastroenteritis.