Evaluation of the immunogenicity of a recombinant HSV-1 vector expressing human group C rotavirus VP6 protein

Group C Rotavirus (RVC) has been associated globally with sporadic outbreaks of gastroenteritis in children and adults. RVC also infects animals, and interspecies transmission has been reported as well as its zoonotic potential. Considering its genetic diversity and the absence of effective vaccines, it is important and necessary to develop new generation vaccines against RVC for both humans and animals. The aim of the present study was to develop and characterize an HSV-1-based amplicon vector expressing a human RVC-VP6 protein and evaluate the humoral immune response induced after immunizing BALB/c mice. Local fecal samples positive for RVC were used for isolation and sequencing of the vp6 gene, which phylogenetically belongs to the I2 genotype. We show here that cells infected with the HSV[VP6C] amplicon vector efficiently express the VP6 protein, and induced specific anti-RVC antibodies in mice immunized with HSV[VP6C], in a prime-boost schedule. This work highlights that amplicon vectors are an attractive platform for the generation of safe genetic immunogens against RVC, without the addition of external adjuvants.Fil: Rota, Rosana Paola. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Palacios, Carlos Adolfo. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencia y Tecnología "Dr. César Milstein". Fundación Pablo Cassará. Instituto de Ciencia y Tecnología "Dr. César Milstein"; ArgentinaFil: Temprana, Carlos Facundo. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Argüelles, Marcelo Horacio. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Mandile, Marcelo Gastón. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Mattion, Nora Marta. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Ciencia y Tecnología "Dr. César Milstein". Fundación Pablo Cassará. Instituto de Ciencia y Tecnología "Dr. César Milstein"; ArgentinaFil: Laimbacher, Andrea S.. Universitat Zurich; SuizaFil: Fraefel, Cornell. Universitat Zurich; SuizaFil: Castello, Alejandro Andrés. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Almallo de Glikmann, Graciela. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentin

Pre-vaccine rotavirus surveillance in Buenos Aires, Argentina: Characterization of an emergent G1P[8] strain associated to fatal cases in 2014

Group A rotaviruses (RVA) are the most frequent etiological agents causing severe diarrhea in infants and surveillance of genotype, and genetic characteristics of circulating strains are necessary in order to evaluate vaccine programs. The objectives of this work were to describe G and P genotype from 2012 through 2014 in Buenos Aires, Argentina completing an overview of 19 years of genotype surveillance in our region and to characterize an emerging G1P[8] strain associated with severe cases and five fatalities in 2014. We performed genotyping by RT-PCR. The sequencing of several genes, phylogenetic analyses, and comparative epidemiological data were used to know the origin and phylogenetic relationships of the emerging G1P[8] strain. Along with this report, 19 years of continuous RVA genotype surveillance in Argentina in the pre-vaccine era was covered. During the last year of this surveillance, 2014, a significantly increased incidence of RVA associated gastroenteritis was related to the reemergence of G1P[8] strains, being these ones detected in low frequency in the last nine years. Interestingly, the patients affected were significantly older when compared with those from the last six seasons. Additionally, phylogenetic analysis of several genes infer that these G1P[8] strains were closely related to Asian strains circulating during 2012 and 2013. In addition to this, the suggested extra continental origin for the 2014 G1P[8] strains and the very low circulation of G1 type during nine years probably explain the increased incidence and severity in the gastroenteritis cases and the particular epidemiologic characteristics. In conclusion, this work gives us a whole panorama of the pre-vaccine era of the RVA molecular epidemiology in the most populated region of Argentina. In this way, this work inspires us to continue with this type of studies in the post-vaccination era.Fil: Mandile, Marcelo Gastón. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Argüelles, Marcelo Horacio. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Temprana, Carlos Facundo. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Peri Ibañez, Estefania Soledad. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Silvestre, Dalila. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Musto, Alejandra Beatriz. Gobierno de la Provincia de Buenos Aires. Hospital Provincial Evita Pueblo.; ArgentinaFil: Rodríguez Pérez, Alberto. Hospital General de Agudos Dr. Alberto A. Eurnekian; ArgentinaFil: Mistchenko, Alicia Susana. Gobierno de la Ciudad Autónoma de Buenos Aires. Hospital General de Niños "Ricardo Gutiérrez". Departamento de Medicina; Argentina. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas; ArgentinaFil: Almallo de Glikmann, Graciela. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; ArgentinaFil: Castello, Alejandro Andrés. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Universidad Nacional Arturo Jauretche; Argentin

Surveillance of group A Rotavirus in Buenos Aires 2008-2011, long lasting circulation of G2P[4] strains possibly linked to massive monovalent vaccination in the region.

Background: Group A rotaviruses (RVA) are the most frequent single etiological agents of severe diarrhea in infants. Since 2006 RVA vaccines have been introduced in national schedules of middle and high income countries with substantial declines in rotavirus associated disease burden. However, surveillance must be maintained to, eventually, detect emerging types or variants selected by the new pressure imposed by vaccination. Objectives: To analyze the molecular epidemiology of group A rotavirus after vaccine introduction in the region in the context of data from more than 15 years of continuous surveillance in Buenos Aires. Study design: RVA positive diarrhea samples collected in Buenos Aires from 2008 to 2011 were genotyped by RT-PCR. Selected samples were sequenced to gain insight on evolution of common and globally emerging human RVA strains. Results: Lineage III G12P[8] strain emerged in 2008 in Buenos Aires and shared co-dominancy with G3 strains during 2009. An atypical long lasting circulation of G2P[4] strains since 2004 reached rates around 80% in 2011 in Buenos Aires. Sequencing of the VP7 and VP4 genes of representative G2P[4] isolates suggests Brazil as the origin of the 2010?2011 strains. Conclusions: Globally emergent G12 lineage III strains could be established as dominant strains in a very populated area in two years since emergence. In this work it was also shown that the persistence of G2P[4] strains during 8 years could be related to massive immunization with the monovalent vaccine in the region.Fil: Mandile, Marcelo Gastón. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Esteban, Laura Emilia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; ArgentinaFil: Argüelles, Marcelo Horacio. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; ArgentinaFil: Mistchenko, Alicia Susana. Gobierno de la Ciudad de Buenos Aires. Hospital General de Niños ; ArgentinaFil: Almallo de Glikmann, Graciela. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; ArgentinaFil: Castello, Alejandro Andrés. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Virologia; Argentin

Production of recombinant NS1 protein and its possible use in encephalitic flavivirus differential diagnosis

Saint Louis encephalitis virus (SLEV) and West Nile virus (WNV) are two of the major causes of arboviral encephalitis in the Americas. The co-circulation of related flaviviruses in the Americas and prior vaccination against flaviviruses pose problems to the diagnostic specificity of serological assays due to the development of cross-reactive antibodies. An accurate diagnosis method capable of differentiating these related viruses is needed. NS1 is a glycosylated, nonstructural protein, of about 46 kDa which has a highly conserved structure. Anti-NS1 antibodies can be detected within 4?8 days after the initial exposure and NS1 is the least cross-reactive of the flaviviral antigens. This study was aimed to generate SLEV and WNV NS1 recombinants proteins for the development of a flavivirus diagnostic test. Local Argentinian isolates were used as the source of NS1 gene cloning, expression, and purification. The protein was expressed in Escherichia coli as inclusion bodies and further purified by metal-chelating affinity chromatography (IMAC) under denaturing conditions. Human sera from SLEV and WNV positive cases showed reactivity to the recombinant NS1 proteins by western blot. The unfolded NS1 proteins were also used as immunogens. The polyclonal antibodies elicited in immunized mice recognized the two recombinant proteins with differential reactivity.Fil: Lorch, Matías Sebastián. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; ArgentinaFil: Collado, Maria Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Inmunología, Genética y Metabolismo. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Inmunología, Genética y Metabolismo; Argentina. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; ArgentinaFil: Argüelles, Marcelo Horacio. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Rota, Rosana Paola. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Spinsanti, Lorena Ivana. Universidad Nacional de Córdoba. Facultad de Medicina. Instituto de Virología Dr. J. M. Vanella; ArgentinaFil: Lozano, Mario Enrique. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Goñi, Sandra Elizabeth. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Ingeniería Genética y Biología Molecular y Celular; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin