Análise da ação de nanopartículas de prata sobre o biofilme de espécies de Candida
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Data
2012-10-24
Autores
Monteiro, Douglas Roberto [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
O objetivo deste estudo foi avaliar a atividade antifúngica de nanopartículas de prata (NP) contra os biofilmes de Candida albicans e Candida glabrata. NP foram sintetizadas por meio da redução do nitrato de prata com citrato de sódio e estabilizadas com amônia ou polivinilpirrolidona. Os testes de concentração inibitória mínima (CIM) das NP contra células de Candida foram baseados no método da microdiluição. NP foram aplicadas sobre os biofilmes de Candida (48 horas) e após 24 horas de contato sua atividade antifúngica foi determinada por meio da quantificação da biomassa total (coloração com violeta cristal (VC)) e por meio da enumeração das unidades formadoras de colônias (UFCs). Após o tratamento com NP, as matrizes dos biofilmes foram extraídas e analisadas em termos de proteínas, carboidratos e DNA, e a estrutura dos biofilmes foi analisada por meio da microscopia eletrônica de varredura e de epifluorescência. A atividade antibiofilme da combinação de NP com nistatina e clorexidina foi avaliada por meio dos ensaios de VC e UFCs. Leveduras viáveis foram recuperadas a partir dos biofilmes previamente tratados com NP e adicionadas às células epiteliais HeLa e aos poços de placas de poliestireno e, após 2 horas de contato, a adesão foi determinada usando VC. A eficácia de NP submetidas às variações de temperatura (50, 70 e 100ºC) e pH (5 e 9) também foi avaliada, assim como a susceptibilidade às NP dos biofilmes de Candida em diferentes fases de crescimento. Os resultados de CIM mostraram que NP foram fungicidas contra os isolados testados em concentrações baixas (0,4-3,3 μg/mL). NP foram mais efetivas na redução da biomassa para os biofilmes de C. glabrata (reduções de 90% na concentração de 108 μg/mL) do que para C. albicans, e promoveram reduções significativas no log10 do número de UFCs...
The aim of this study was to evaluate the antifungal activity of silver nanoparticles (SN) against Candida albicans and Candida glabrata biofilms. Colloidal suspensions of SN were synthesized by reducing silver nitrate with sodium citrate and stabilized with ammonia or polyvinylpyrrolidone. Minimal inhibitory concentrations (MIC) were performed for Candida cells grown in suspension following the microbroth dilution method. Candida biofilms (48 h) were treated with SN for 24 h and then the total biomass quantification (by crystal violet (CV) staining) and the colony forming units (CFUs) were determined. Also, after treating with SN, extracellular matrices were extracted from Candida biofilms and analyzed chemically in terms of proteins, carbohydrates and DNA. To investigate the biofilm structure, scanning electron microscopy and epifluorescence microscopy were carried out. The antibiofilm activity of SN in combination with nystatin and chlorhexidine was also assessed by CV and CFU. Moreover, viable yeasts were recovered from the biofilms pretreated with SN and added to HeLa epithelial cells or to empty wells of polystyrene plates and, after 2 h of contact, the adhesion capacity of the yeasts was determined by using CV staining. The antibiofilm efficacy of SN subjected to variations of temperature (50, 70 and 100ºC) and pH (5 and 9) was also evaluated. Finally, the susceptibility to SN of biofilms in different stages of growth was analyzed. MIC results showed that SN were fungicidal against the tested strains at very low concentrations (0.4- 3.3 μg/mL). SN were more effective in reducing the total biomass of C. glabrata (reductions around 90% at 108 μg/mL SN) than C. albicans biofilms, and provided significant log10 reduction of... (Complete abstract click electronic access below)
The aim of this study was to evaluate the antifungal activity of silver nanoparticles (SN) against Candida albicans and Candida glabrata biofilms. Colloidal suspensions of SN were synthesized by reducing silver nitrate with sodium citrate and stabilized with ammonia or polyvinylpyrrolidone. Minimal inhibitory concentrations (MIC) were performed for Candida cells grown in suspension following the microbroth dilution method. Candida biofilms (48 h) were treated with SN for 24 h and then the total biomass quantification (by crystal violet (CV) staining) and the colony forming units (CFUs) were determined. Also, after treating with SN, extracellular matrices were extracted from Candida biofilms and analyzed chemically in terms of proteins, carbohydrates and DNA. To investigate the biofilm structure, scanning electron microscopy and epifluorescence microscopy were carried out. The antibiofilm activity of SN in combination with nystatin and chlorhexidine was also assessed by CV and CFU. Moreover, viable yeasts were recovered from the biofilms pretreated with SN and added to HeLa epithelial cells or to empty wells of polystyrene plates and, after 2 h of contact, the adhesion capacity of the yeasts was determined by using CV staining. The antibiofilm efficacy of SN subjected to variations of temperature (50, 70 and 100ºC) and pH (5 and 9) was also evaluated. Finally, the susceptibility to SN of biofilms in different stages of growth was analyzed. MIC results showed that SN were fungicidal against the tested strains at very low concentrations (0.4- 3.3 μg/mL). SN were more effective in reducing the total biomass of C. glabrata (reductions around 90% at 108 μg/mL SN) than C. albicans biofilms, and provided significant log10 reduction of... (Complete abstract click electronic access below)
Descrição
Palavras-chave
Biofilme, Candida, Prata, Nanotecnologia, Biofilms
Como citar
MONTEIRO, Douglas Roberto. Análise da ação de nanopartículas de prata sobre o biofilme de espécies de Candida. 2012. 268 f. Tese (doutorado) - Universidade Estadual Paulista Júlio de Mesquita Filho, Faculdade de Odontologia de Araçatuba, 2012.