Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-04-25T22:57:38Z No. of bitstreams: 1 LuandaBarbaraFerreiraCanarioDeSouza_DISSERT.pdf: 2996890 bytes, checksum: 9a5f76941d2b2ee6ded886c004b1965d (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-04-27T23:22:44Z (GMT) No. of bitstreams: 1 LuandaBarbaraFerreiraCanarioDeSouza_DISSERT.pdf: 2996890 bytes, checksum: 9a5f76941d2b2ee6ded886c004b1965d (MD5) / Made available in DSpace on 2016-04-27T23:22:44Z (GMT). No. of bitstreams: 1 LuandaBarbaraFerreiraCanarioDeSouza_DISSERT.pdf: 2996890 bytes, checksum: 9a5f76941d2b2ee6ded886c004b1965d (MD5) Previous issue date: 2015-03-30 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico (CNPq) / A candid?ase ? uma importante manifesta??o oral em pacientes transplantados renais. Candida spp. possuem fatores de virul?ncia que contribuem para o processo infeccioso, incluindo a capacidade de aderir a c?lulas epiteliais e a forma??o de biofilme em superf?cies bi?ticas e abi?ticas. O extrato obtido a partir das folhas de Eugenia uniflora [acetona: ?gua (7:3, v/v)] tem demonstrado atividade antif?ngica contra Candida spp. Este estudo avaliou a influ?ncia do extrato de E. uniflora na ades?o ?s C?lulas Epiteliais Bucais Humanas (CEBH) e forma??o de biofilme de 42 cepas de Candida spp. isoladas da cavidade oral de pacientes transplantados renais. As cepas de Candida spp. pertencentes a um banco de microorganimos foram reativadas e reidentificadas fenotipicamente atrav?s da metodologia cl?ssica e por m?todos moleculares (genotipagem ABC e RAPD), quando necess?rio, para a conclus?o da identifica??o em n?vel de esp?cie. Para os ensaios de virul?ncia avaliados in vitro, as leveduras foram cultivadas na presen?a e aus?ncia de 1000 ?g/ mL de extrato. Uma propor??o de 10:1 c?lulas (c?lulas Candida spp. x CEBHs) foi incubada durante 1 hora a 37 ?C, 200 rpm, fixadas com formalina a 10% e o n?mero de c?lulas de Candida aderidas a 150 CEBH foram determinadas em microsc?pio ?ptico. Os biofilmes foram formados em microplacas de poliestireno na presen?a e aus?ncia do extrato. A quantifica??o foi realizada com colora??o por cristal violeta a 570 nm. Todos os isolados apresentaram-se vi?veis e na reidentifica??o fenot?pica apresentaram caracter?sticas sugestivas de cada esp?cie identificada. Duas cepas identificadas presuntivamente como Candida dubliniensis foram confirmadas como pertencentes a esta esp?cie atrav?s da genotipagem ABC, e as cepas identificadas como pertencentes ao complexo Candida parapsilosis foram diferenciadas atrav?s da genotipagem por RAPD. Candida albicans destacou-se como a esp?cie com maior potencial de ades?o ao epit?lio bucal, enquanto que C. tropicalis apresentou expressiva forma??o de biofilme. Detectamos que o extrato de E. uniflora foi capaz de reduzir a ades?o ao CEBH tanto para Candida albicans como para as esp?cies de Candida n?o-Candida albicans. Por outro lado, apenas 16 cepas de Candida spp. (36%) apresentaram a forma??o de biofilme reduzida na presen?a do extrato. No entanto, duas cepas de C. tropicalis altamente produtoras de biofilme tiveram uma redu??o expressiva na capacidade de forma??o de biofilme. Este estudo refor?a a ideia de que, al?m da inibi??o do crescimento, E. uniflora pode interferir com a express?o de alguns fatores de virul?ncia de Candida spp., e pode ser eventualmente aplicada no futuro como um poss?vel agente antif?ngico. / Candidiasis is a major oral manifestation in kidney transplant patients. Candida spp. possess essential virulence factors which contribute for the infectious process, including the ability to adhere to epithelial cells and biofilm formation. The extract obtained from the leaves of Eugenia uniflora [acetone: water (7:3, v/v)] has demonstrated antifungal activity against Candida spp. This study evaluated the influence of the extract of E. uniflora in adhesion to human buccal epithelial cells (HBEC) and biofilm formation of 42 strains of Candida spp. isolated from the oral cavity of kidney transplant patients. Candida spp. strains belonging to a culture collection were reactivated and phenotypically re-identified by classical and molecular methods (genotyping ABC and RAPD), when necessary, to complete the identification to the species level. For the virulence tests evaluated in vitro, yeasts were grown in the presence and absence of 1000 ?g/mL of the extract. A ratio of 10: 1 (Candida spp. cells x HBECs) was incubated for 1 hour at 37 ? C, 200 rpm, fixed with 10% formalin and the number of Candida cells adhered to 150 HBEC determined by optical microscope. Biofilms were formed on polystyrene microplates in the presence or absence of the extract. The quantification was performed with crystal violet staining at 570 nm. All isolates were viable and exhibited phenotypic characteristics suggestive of each species identified. Two strains presumptively identified as Candida dubliniensis belonged to this species as determined with genotyping ABC, while strains identified as belonging to the Candida parapsilosis species complex were differentiated by RAPD genotyping. Candida albicans was found to be the most adherent species to the buccal epithelia, while C. tropicalis showed remarkable biofilm formation.We could detect that the extract of E. uniflora was able to reduce adhesion to HBEC for both Candida albicans and non-Candida albicans Candida species. On the other hand, only 16 Candida spp. strains (36 %) showed reduced biofilm formation. However, two highly biofilm producer strains of C. tropicalis had an expressive reduction in biofilm formation. This study reinforces the idea that besides growth inhibition, E. uniflora may interfere with the expression of some virulence factors of Candida spp., and may be possibly applied in the future as a novel antifungal agent.