Characterization of liposomes coated with S-layer proteins from lactobacilli

The stability of liposomes coated with S-layer proteins from Lactobacillus brevis and Lactobacillus kefir was analyzed as a previous stage to the development of a vaccine vehicle for oral administration. The interactions of the different S-layer proteins with positively charged liposomes prepared with soybean lecithin or dipalmitoylphosphatidylcholine were studied by means of the variation of the Z potential at different protein-lipid ratios, showing that both proteins were able to attach in a greater extent to the surface of soybean lecithin liposomes. The capacity of these particles to retain carboxyfluorescein or calcein by exposure to bile salts, pancreatic extract, pH change and after a thermal shock showed that both S-layer proteins increased the stability of the liposomes in the same magnitude. The non-glycosylated protein from L. brevis protects more efficiently the liposomes at pH 7 than those from L. kefir even without treatment with glutaraldehyde.Centro de Investigación y Desarrollo en Criotecnología de Alimento

Characterization of liposomes coated with S-layer proteins from lactobacilli

The stability of liposomes coated with S-layer proteins from Lactobacillus brevis and Lactobacillus kefir was analyzed as a previous stage to the development of a vaccine vehicle for oral administration. The interactions of the different S-layer proteins with positively charged liposomes prepared with soybean lecithin or dipalmitoylphosphatidylcholine were studied by means of the variation of the Z potential at different protein-lipid ratios, showing that both proteins were able to attach in a greater extent to the surface of soybean lecithin liposomes. The capacity of these particles to retain carboxyfluorescein or calcein by exposure to bile salts, pancreatic extract, pH change and after a thermal shock showed that both S-layer proteins increased the stability of the liposomes in the same magnitude. The non-glycosylated protein from L. brevis protects more efficiently the liposomes at pH 7 than those from L. kefir even without treatment with glutaraldehyde.Centro de Investigación y Desarrollo en Criotecnología de Alimento

Characterization of liposomes coated with S-layer proteins from lactobacilli

The stability of liposomes coated with S-layer proteins from Lactobacillus brevis and Lactobacillus kefir was analyzed as a previous stage to the development of a vaccine vehicle for oral administration. The interactions of the different S-layer proteins with positively charged liposomes prepared with soybean lecithin or dipalmitoylphosphatidylcholine were studied by means of the variation of the Z potential at different protein-lipid ratios, showing that both proteins were able to attach in a greater extent to the surface of soybean lecithin liposomes. The capacity of these particles to retain carboxyfluorescein or calcein by exposure to bile salts, pancreatic extract, pH change and after a thermal shock showed that both S-layer proteins increased the stability of the liposomes in the same magnitude. The non-glycosylated protein from L. brevis protects more efficiently the liposomes at pH 7 than those from L. kefir even without treatment with glutaraldehyde.Centro de Investigación y Desarrollo en Criotecnología de Alimento

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

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

Antigen vehiculization particles based on the Z protein of Junin virus

Abstract Background Arenavirus matrix protein Z plays an important role in virus budding and is able to generate enveloped virus-like-particles (VLPs) in absence of any other viral proteins. In these VLPs, Z protein is associated to the plasma membrane inner surface by its myristoyl residue. Budding induction and vesicle formation properties can be exploited to generate enveloped VLPs platform. These structures can be designed to carry specific antigen in the inner side or on the surface of VLPs. Vaccines based on VLPs are a highly effective type of subunit vaccines that mimic the overall structure of virus particles in absence of viral nucleic acid, being noninfectious. In this work we assayed the capacity of Junin Z protein to produce VLPs carrying the green fluorescent protein (eGFP), as a model antigen. Results In this report the Junin Z protein ability to produce VLPs from 293T cells and its capacity to deliver a specific antigen (eGFP) fused to Z was evaluated. Confocal microscopy showed a particular membrane bending in cells expressing Z and a spot welded distribution in the cytoplasm. VLPs were detected by TEM (transmission electron microscopy) and were purified from cell supernatant. The proteinase protection assay demonstrated the VLPs integrity and the absence of degradation of the fused antigen, thus indicating its internal localization. Finally, immunization of mice with purified VLPs produced high titres of anti-eGFP antibodies compared to the controls. Conclusions It was proved that VLPs can be generated from cells transfected with a fusion Junin virus Z-eGFP protein in absence of any other viral protein, and the capacity of Z protein to support fusions at the C-terminal, without impairing its budding activity, allowing vehiculization of specific antigens into VLPs.</p

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