2019-02-222018-09-21LOPES, Paloma da Silva Gomes. Identificação de candidatos vacinais em potencial para Trichosporon asahii por vacinologia reversa. 2018. 126 f. Dissertação (Mestrado em Ciências Biológicas) - Universidade Federal de Alfenas, Alfenas, MG, 2018.https://repositorio.unifal-mg.edu.br/handle/123456789/1316Members of the Trichosporon spp. genus are Basidioumycetous yeasts that have emerged as opportunistic pathogens in immunocompromised patients. Trichosporon asahii already displays evidence of resistance against various antifungal agents and is one of the main cause of invasive infections. Despite its clinical importance, there are still no vaccines and immunotherapies to prevent the disease in the main risk groups. Thus, the present study aimed to identify potential vaccine candidates to direct immunobiologicals production that may confer protection in these patients, by using reverse vaccination technology, in which there is no need to cultivate the microorganism in the laboratory, favoring the creation of a vaccine faster. We performed in silico analyzes using the predicted proteome of T. asahii deposited in NCBI, containing 8.300 putative proteins. The results indicate that 268 proteins were predicted as extracellular by the CELLO v.2.5 and PSORT II programs. By proteome analyzes using the CELLO v.2.5 and PSORT II programs, 268 common proteins were predicted as extracellular. Immunogenicity prediction was performed in the Vaxijen v2.0 program, in which 205 proteins with immunogenic potential (score ≥ 0.5) were obtained. The selected proteins were compared to the human proteome, through BLASTp-NCBI. Of the 205 proteins analyzed, 137 were selected by the criterion of no similarity with human proteins. To identify candidates that could induce cross-protection, these selected proteins were compared with proteins of the Cryptococcus spp. and Candida spp. genera, by using BLASTp-NCBI. The results indicated the presence of 82 proteins with similarity to the proteins of Cryptococcus spp. and 11 with Candida spp. Then, we selected proteins that presented functional characterization and similarity and coverage between sequences > 50%. It was evidenced that only 5 Cryptococcus spp. proteins met the selection criteria. The remaining 4proteins that did not have any similarity to Trichosporon spp. related genera were analyzed to identify vaccine candidates that could induce protection only against T. asahii. The selection criterion in this analysis was to select only the proteins that had functional characterization by gene ontology with fungal genera that already had their proteomes annotated, with 48 proteins analyzed, but only 3 with functional characterization. Thus, the 8 selected proteins had their epitopes mapped through the Tepitool software to assess the ability of MHC II binding in T cells, and we detected that all proteins had epitopes that bound to the HLA alleles of MHC II. Next, all MHC II-binding epitopes were analyzed in the VaxiJen 2.0 program to evaluate their immunostimulatory capacity, with cutoff point > 0.5. We found a minimum of 20 immunostimulatory epitopes for protection against T. asahii in cutinase protein, and a maximum of 43 in the Curculin domain (mannose-binding) lectin protein, and for cross-protection, a minimum of 18 epitopes and a maximum of 54, in the Major allergen Asp F2 and Glioxal oxidase precursor proteins, respectively. Then, molecular modeling of the 8 proteins through the I-TASSER software was performed to facilitate the localization of the immunogenic epitopes, visualized in 3D through the VMD software. Finally, the world population coverage of the epitopes was performed to evaluate the different HLA binding specificities and to select those epitopes with greater coverage in the human population. A total of 71 immunogenic epitopes were selected for exclusive protection for T. asahii and 132 for cross protection of T. asahii and Cryptococcus spp., with a global coverage rate of 99.98%, indicating that some epitopes are excellent vaccine candidates for the creation of a vaccine with protection against T. asahii and cross-protection between the genus Cryptococcus spp. and T. asahii, respectively.application/pdfAcesso Abertohttp://creativecommons.org/licenses/by-nc-nd/4.0/EpitoposTricosporonoseTricosporonose -- Prevenção e controleBIOLOGIA E FISIOLOGIA DOS MICROORGANISMOS::MICOLOGIADissertaçãoPadovan, Ana Carolina Barbosa