RESUMO
Introducción. Los datos de frecuencia de los adenovirus entéricos, sapovirus y astrovirus en casos de gastroenteritis aguda esporádica en Argentina son escasos. Métodos. Diseño descriptivo sobre una selección de muestras de heces de menores de 5 años con diarrea remitidas durante el período 2010-2021, con resultado previo negativo para rotavirus y norovirus. Se estudió la presencia de adenovirus entéricos, sapovirus y astrovirus por métodos moleculares, con posterior genotipificación de las muestras positivas. Resultados. De 574 muestras seleccionadas, en 226 (39,4 %) se identificó al menos uno de los virus estudiados. En particular, se detectaron adenovirus, sapovirus y astrovirus en el 30,7 %, el 5,6 % y el 3,1 %, respectivamente. El adenovirus 41, los sapovirus GI.1 y GI.2, y el astrovirus 1 fueron los más frecuentemente detectados. Se identificaron dos muestras con astrovirus no clásicos. Conclusiones. A pesar de ser menos frecuentes, estos enteropatógenos son responsables de un número considerable de episodios de diarrea esporádica. Por lo tanto, su estudio y vigilancia contribuye significativamente a reducir la brecha de casos no diagnosticados.
Introduction. Data on the frequency of enteric adenoviruses, sapoviruses, and astroviruses in cases of sporadic acute gastroenteritis in Argentina are scarce. Methods. Descriptive design of a selection of fecal samples of children with diarrhea younger than 5 years referred between 2010 and 2021, with a previous negative result for rotavirus and norovirus. The presence of enteric adenovirus, sapovirus, and astrovirus was tested by molecular methods, with subsequent genotyping of positive samples. Results. At least 1 of the tested viruses was detected in 226 (39.4%) of the 574 selected samples. Specifically, adenovirus, sapovirus, and astrovirus were detected in 30.7%, 5.6%, and 3.1% of the samples, respectively. The most frequent viruses detected were adenovirus 41, sapoviruses GI.1 and GI.2, and astrovirus 1. Non-classic astroviruses were detected in 2 samples. Conclusions. Despite being less frequent, these enteropathogens are responsible for a large number of sporadic diarrhea events. Therefore, their study and surveillance contribute significantly to reduce the gap of undiagnosed cases.
Assuntos
Humanos , Lactente , Pré-Escolar , Diarreia/epidemiologia , Diarreia/virologia , Gastroenterite/epidemiologia , Gastroenterite/virologia , Argentina/epidemiologia , Astroviridae/isolamento & purificação , Astroviridae/genética , Sapovirus/isolamento & purificação , Sapovirus/genética , Fezes/virologia , GenótipoRESUMO
Introduction. Data on the frequency of enteric adenoviruses, sapoviruses, and astroviruses in cases of sporadic acute gastroenteritis in Argentina are scarce. Methods. Descriptive design of a selection of fecal samples of children with diarrhea younger than 5 years referred between 2010 and 2021, with a previous negative result for rotavirus and norovirus. The presence of enteric adenovirus, sapovirus, and astrovirus was tested by molecular methods, with subsequent genotyping of positive samples. Results. At least 1 of the tested viruses was detected in 226 (39.4%) of the 574 selected samples. Specifically, adenovirus, sapovirus, and astrovirus were detected in 30.7%, 5.6%, and 3.1% of the samples, respectively. The most frequent viruses detected were adenovirus 41, sapoviruses GI.1 and GI.2, and astrovirus 1. Non-classic astroviruses were detected in 2 samples. Conclusions. Despite being less frequent, these enteropathogens are responsible for a large number of sporadic diarrhea events. Therefore, their study and surveillance contribute significantly to reduce the gap of undiagnosed cases.
Introducción. Los datos de frecuencia de los adenovirus entéricos, sapovirus y astrovirus en casos de gastroenteritis aguda esporádica en Argentina son escasos. Métodos. Diseño descriptivo sobre una selección de muestras de heces de menores de 5 años con diarrea remitidas durante el período 2010-2021, con resultado previo negativo para rotavirus y norovirus. Se estudió la presencia de adenovirus entéricos, sapovirus y astrovirus por métodos moleculares, con posterior genotipificación de las muestras positivas. Resultados. De 574 muestras seleccionadas, en 226 (39,4 %) se identificó al menos uno de los virus estudiados. En particular, se detectaron adenovirus, sapovirus y astrovirus en el 30,7 %, el 5,6 % y el 3,1 %, respectivamente. El adenovirus 41, los sapovirus GI.1 y GI.2, y el astrovirus 1 fueron los más frecuentemente detectados. Se identificaron dos muestras con astrovirus no clásicos. Conclusiones. A pesar de ser menos frecuentes, estos enteropatógenos son responsables de un número considerable de episodios de diarrea esporádica. Por lo tanto, su estudio y vigilancia contribuye significativamente a reducir la brecha de casos no diagnosticados.
Assuntos
Diarreia , Gastroenterite , Humanos , Argentina/epidemiologia , Gastroenterite/virologia , Gastroenterite/epidemiologia , Lactente , Pré-Escolar , Diarreia/virologia , Diarreia/epidemiologia , Masculino , Feminino , Sapovirus/genética , Sapovirus/isolamento & purificação , Fezes/virologia , Genótipo , Recém-Nascido , Astroviridae/genética , Astroviridae/isolamento & purificaçãoRESUMO
ABSTRACT Human astrovirus (HAstV) 1-8 and highly divergent HAstVMLB1−3 genotypes have been detected in children both with and without acute gastroenteritis (AGE). One hundred and seventy fecal samples from children (≤5 years old) living in the Amazon region were evaluated for the presence of HAstV1-8, HAstV MLB1−3 and HAstVVA1−3, using an usual RT-PCR protocol and a new protocol with specific primers designed to detect HAstVMLB1−3. HAstVMLB1 and HAstV MLB2, as well as the HAstV3 and 5 genotypes were detected. HAstVMLB1−2 genotype was detected for the first time in Brazil at a frequency of 3.5% (6/170).
Assuntos
Criança , Humanos , Lactente , Mamastrovirus , Infecções por Astroviridae , Gastroenterite , Filogenia , Mamastrovirus/genética , Brasil , Infecções por Astroviridae/diagnóstico , Infecções por Astroviridae/epidemiologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fezes , Gastroenterite/diagnóstico , GenótipoRESUMO
Human astrovirus (HAstV) 1-8 and highly divergent HAstVMLB1-3 genotypes have been detected in children both with and without acute gastroenteritis (AGE). One hundred and seventy fecal samples from children (≤5 years old) living in the Amazon region were evaluated for the presence of HAstV1-8, HAstV MLB1-3 and HAstVVA1-3, using an usual RT-PCR protocol and a new protocol with specific primers designed to detect HAstVMLB1-3. HAstVMLB1 and HAstV MLB2, as well as the HAstV3 and 5 genotypes were detected. HAstVMLB1-2 genotype was detected for the first time in Brazil at a frequency of 3.5% (6/170).
Assuntos
Infecções por Astroviridae , Gastroenterite , Mamastrovirus , Infecções por Astroviridae/diagnóstico , Infecções por Astroviridae/epidemiologia , Brasil , Criança , Fezes , Gastroenterite/diagnóstico , Genótipo , Humanos , Lactente , Mamastrovirus/genética , Filogenia , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Human astroviruses (HAstVs) cause severe diarrhea and represent an important health problem in children under two years of age. Despite their medical importance, the study of these pathogens has been neglected. To better understand the astrovirus antigenic structure and the basis of protective immunity, in this work we produced a panel of neutralizing monoclonal antibodies (Nt-MAbs) to HAstV serotypes 1, 2, and 8 and identified the mutations that allow the viruses to escape neutralization. We first tested the capacity of the recombinant HAstV capsid core and spike domains to elicit Nt-Abs. Hyperimmunization of animals with the two domains showed that although both induced a potent immune response, only the spike was able to elicit antibodies with neutralizing activity. Based on this finding, we used a mixture of the recombinant spike domains belonging to the three HAstV serotypes to immunize mice. Five Nt-MAbs were isolated and characterized; all of them were serotype specific, two were directed to HAstV-1, one was directed to HAstV-2, and two were directed to HAstV-8. These antibodies were used to select single and double neutralization escape variant viruses, and determination of the amino acid changes that allow the viruses to escape neutralization permitted us to define the existence of four potentially independent neutralization epitopes on the HAstV capsid. These studies provide the basis for development of subunit vaccines that induce neutralizing antibodies and tools to explore the possibility of developing a specific antibody therapy for astrovirus disease. Our results also establish a platform to advance our knowledge on HAstV cell binding and entry.IMPORTANCE Human astroviruses (HAstVs) are common etiological agents of acute gastroenteritis in children, the elderly, and immunocompromised patients; some virus strains have also been associated with neurological disease. Despite their medical importance, the study of these pathogens has advanced at a slow pace. In this work, we produced neutralizing antibodies to the virus and mapped the epitopes they recognize on the virus capsid. These studies provide the basis for development of subunit vaccines that induce neutralizing antibodies, as well as tools to explore the development of a specific antibody therapy for astrovirus disease. Our results also establish a platform to advance our knowledge on HAstV cell binding and entry.
Assuntos
Anticorpos Neutralizantes/isolamento & purificação , Antígenos Virais/imunologia , Infecções por Astroviridae/imunologia , Mamastrovirus/imunologia , Animais , Anticorpos Monoclonais/isolamento & purificação , Anticorpos Antivirais/isolamento & purificação , Antígenos Virais/genética , Infecções por Astroviridae/virologia , Células CACO-2 , Proteínas do Capsídeo/genética , Proteínas do Capsídeo/imunologia , Variação Genética , Humanos , Imunização , Mamastrovirus/genética , Camundongos , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/imunologiaRESUMO
Human astrovirus is an important etiological agent of acute gastroenteritis in young children. Despite advances in the characterization of the structure of the virion by cryo-electron microscopy and of capsid proteins by x-ray crystallography, the definition of the minimal polypeptide composition of infectious virus particles has been elusive. In this work we show that mature infectious particles are composed by only two proteins; VP34 that forms the core domain of the virus, and VP27 that constitutes the 30 dimeric spikes present on the virus surface. Our results also indicate that during the transition of immature (90 spikes) to mature (30 spikes) virus particles, that occur during trypsin activation, the viral protein VP25, that most likely forms the 60 spikes that are lost during maturation, detaches from the virus particle. This information is relevant to better understand the biology of virus entry and also for the efficient development of subunit vaccines.
Assuntos
Proteínas do Capsídeo/química , Capsídeo/química , Mamastrovirus/química , Peptídeos/química , Células CACO-2 , Microscopia Crioeletrônica , Cristalografia por Raios X , Humanos , Modelos Moleculares , Vírion , Internalização do VírusRESUMO
Classic Human Astrovirus (Classic HAstV) are one of the most important causes of pediatric acute gastroenteritis (AGE), after rotaviruses and arguably caliciviruses. The aim of this study was to determine the molecular epidemiology of Classic HAstV from 175 clinical samples, being 153 stools and 22 vomits, collected from pediatric hospitalized patients with AGE in Salto city, Uruguay, from January 2011 to December 2012. Classic HAstV were detected and genotyped by using a qualitative Retro Transcription-Polimerase Chain Reaction (RT-PCR) directed to the Open Reading Frame-2 (ORF2) region C. Amplicons were sequenced and phylogenetic analyses were carried out in order to determine genotypes and lineages. Classic HAstV were detected in 18 out of 175 analyzed samples (10.3%) and 14 of them (78.0%) were successfully sequenced being 6 (42.8%) classified as HAstV-1 (1a lineage), 4 (28.6%) as HAstV-2 (2c lineage), and 4 (28.6%) as HAstV-3 (3c lineage). A higher detection of Classic HAstV infections was observed in autumn for both years of surveillance, and the majority of the positive cases were observed in 2011. The group of children between 2 and 5 years old presented the higher percentage of infections. To our knowledge, the present study represents the first report of astrovirus from acute gastroenteritis cases in Uruguay, evidencing its role as a relevant etiologic agent in severe cases of this disease.
Assuntos
Infecções por Astroviridae/epidemiologia , Gastroenterite/epidemiologia , Variação Genética , Mamastrovirus/crescimento & desenvolvimento , Doença Aguda , Criança Hospitalizada , Pré-Escolar , Fezes/virologia , Feminino , Gastroenterite/virologia , Genótipo , Humanos , Masculino , Mamastrovirus/classificação , Mamastrovirus/genética , Mamastrovirus/isolamento & purificação , Fases de Leitura Aberta , Filogenia , RNA Viral/genética , Rotavirus/genética , Rotavirus/isolamento & purificação , Uruguai/epidemiologiaRESUMO
AIMS: The aim of this study was to investigate the presence of the recently identified human astrovirus (HAstV) and to increase the knowledge of the molecular epidemiology of classical HAstV detected in Uruguay. METHODS AND RESULTS: Recently identified and classical HAstV genotypes were investigated by RT-PCR targeting the ORF1b and ORF2 genome regions in 20 samples obtained between September 2011 and April 2013 in two cities of the eastern region of Uruguay. Four of 20 samples (20%) were identified as MLB-1 genotype and it was found a new MLB-1 classification through the segregation of the worldwide reported MLB-1 strains in two genetic lineages proposed and named: MLB-1a and MLB-1b. Fourteen (70%) samples were positive for classical HAstV and 12 of them were successfully sequenced and genotyped as: HAstV-1 (n = 10), HAstV-2 and HAstV-5 (one sample each). CONCLUSION: These results constitute the first report in the Latin American region concerning the molecular detection and characterization of MLB-1 HAstV strains in environmental samples. SIGNIFICANCE AND IMPACT OF THE STUDY: This study highlights the benefits of an environmental surveillance to study emerging enteric viruses circulating in human societies.