Proyecto FONDECYT Adjudicado por el Dr. Ángel Oñate en el Concurso Regular FONDECYT 2013

Se transcribe el Resumen del Proyecto FONDECYT obtenido por el Dr. Ángel Oñate Contreras, luego que 9 grupos de investigación de la Facultad de Ciencias Biológicas se adjudicaron proyectos presentados al Concurso de Proyectos Regular Fondecyt  2013.
Título  del  Proyecto

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"Genomic Isle 3 (GI3) of Brucella abortus, a potential source of genes for development of vaccines"

Abstract

Bacterial genomes can change through a variety of processes, including, as one of its principa mechanisms, horizontal genetic transfer. In some cases, this involves direct insertion of genetic elements transferred from another microorganism or from the environment, as is the case of Genomic Isles (GIs). Acquisition of these genomic segments can transform non-pathogenic bacteria into pathogenic ones. These Isles can be used as indicators of certain types of bacteria, especially those pathogenic with clinical or interest, and also allow the development of molecular diagnostic tools or been the target for the development of vaccines. In Brucella, causative agent of brucellosis, a disease with a worldwide high incidence and prevalence, nine Genomic Isles (GIs) have been defined. Among these nine Brucella  GIs, GI3 is highly conserved in B. abortus  and it includes 29 open reading frames (ORFs), only five of them with a known function.

The development of genomics and the access to sequences of complete genomes, as well as their complementation with proteomics analysis, allows the discovery of new virulence factors. Comparative genomics has also become an important tool to identify virulence factors and the identification of genes involved in the persistence of pathogens in the environment. All these tools become instrumental when working with “hypothetic al proteins”, as it is the case of the majority of the ones codified by GI3. By means of mutation studies, we have been able to identify genes that might be involved in the virulence of B. abortus  (BAB1_0278, BAB1_0260, BAB1_0273). These discoveries a have potential use in the search of genetic or proteic vaccines. By means of comparative informatics analysis, it has also been possible to associate certain hypothetical functions to some of the identified ORFs. Thus, the general objective of our Project is to determine the importance in virulence of Genomic Isle 3 (GI3) ORFs of Brucella abortus, characterizing their role in pathogenicity and theto the development of genetic and protein-based vaccines.

This Project, based on an in silico  analysis, will determine the possible functions of some of the proteins codified by genes of the GI3 of B. abortus : BAB1_0278, BAB1_0260 and BAB1_273. Their function and possible participation in the intracellular traffic of Brucella  will be investigated. We will also construct mutants for two ORFS (BAB1_0267 and BAB1_0270), both of interest as possible virulence factors. Survival capacity, replication and protection of B. abortus  mutagenized for these reading frames will be evaluated. Also, we propose to study the immune response (inmunogenicity) generated by the immunization with DNA vaccines designed from the above mentioned and identified genic sequences contained by the GI3. The humoral response will be evaluated by measuring antibody production and the cellular immune responsethe latter will be evaluated by lymphocyte proliferation and cytotoxic activity by T lymphocytes. Measurement of several cytokinas in cultures of lymphocytes and the protection induced by DNA vaccines, against the challenge with the wild pathogenic strain Brucella abortus 2308 in the murine model, will also be carred out. We will also evaluate the immunogenicity of chimeric multi-epitope vaccines developed from immune dominant epitopes detected in silico  for proteins BAB1_0278, BAB1_0260 and BAB1_273, by means of a comparative analysis between a DNA vaccine and a vaccine with the synthetic peptide. Finally, the immunogenicity of vaccines with chimeric proteins, including Brucella  SOD protein in their structure, a protein with already demonstrated immune capacity when associated, separately, to BAB1_0278,BAB1_0260 or BAB1_273 proteins, will be evaluated by comparing a DNA vaccine and a vaccine with a synthetic peptide.