Orientadores: Pedro Luiz Rosalen, Marta Cristina Teixeira Duarte / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-16T16:26:49Z (GMT). No. of bitstreams: 1 Queiroz_VanessaCristinaPinPiazentin_M.pdf: 4355114 bytes, checksum: 2e7009da410cba032963dee4cd9aa7ac (MD5) Previous issue date: 2010 / Resumo: A candidíase oral é uma infecção fúngica que tem sido relatada como problema de saúde pública e em decorrência de um crescente índice de cepas resistentes de Candida spp., tem-se aumentado o interesse em produtos naturais como fonte de novos compostos bioativos. Neste sentido, a própolis brasileira apresenta importantes propriedades farmacológicas como antimicrobiana e poderia ser estudada quanto à sua atividade antifúngica. Assim, o objetivo deste trabalho foi avaliar a atividade anti-Candida de três tipos de própolis brasileiras contra formas planctônicas e em biofilme de Candida spp. de maiores incidências em patologias da cavidade oral. Foram utilizados três tipos de própolis selecionadas por suas propriedades antimicrobianas em estudos prévios, os quais estão classificadas na literatura como: tipo 3 (interior do estado do Paraná), tipo 6 (região de mata atlântica do estado da Bahia) e tipo 13 (região de mangue da cidade de Maceió, estado de Alagoas). Para cada tipo de própolis foram obtidos o extrato etanólico (EEP) e suas respectivas frações com diferentes gradientes de polaridade (hexano, diclorometano e acetato de etila). Aos EEP e respectivas frações foram aplicadas as seguintes metodologias para avaliação da atividade antifúngica contra Candida spp. de forma a conduzir um estudo bioguiado: 1- Determinação da Concentração Inibitória Mínima (CIM) e Concentração Fungicida Mínima (CFM) dos EEP dos tipos 3, 6 e 13 e frações hexânica (fr-hex), diclorometânica (fr-dicloro) e acetato de etila (fr-acet) contra as formas planctônicas das leveduras, obtidas de um banco internacional de cepas Centraalbureau voor Schimmelcultures (CBS): Candida albicans (CBS 562), Candida dubliniensis (CBS 7987), Candida glabrata (CBS 07), Candida krusei (CBS 573), Candida parapsilosis (CBS 604), Candida tropicalis (CBS 94), pelo método da microdiluição. 2- Determinação da CIM das própolis 3 e 13 (EEP e respectivas frações diclorometânicas) sobre biofilmes em pré-adesão. 3- Verificação de possíveis alterações ou danos na estrutura celular das leveduras quando submetidas às própolis 3 e 13 (EEP e respectivas frações diclorometânicas) pela Microscopia Eletrônica de Varredura (MEV) em biofilmes em pré-adesão e formados. De maneira geral, as própolis dos tipos 3 e 13 demonstraram forte atividade anti-Candida contra as leveduras analisadas (valores mínimos e máximos de CIM 4,0 e 31,3 µg/mL para EEP da própolis 3 e 1,0 e 7,8 µg/mL para EEP da própolis 13) em relação a do tipo 6 que demonstrou baixa atividade. O EEP tipo 13 e respectiva fr-dicloro se destacaram, por apresentarem efeito fungicida contra todas as cepas analisadas com valores mínimos e máximos de CFM 125 e 500 µg/mL para a fr-dicloro tipo 13. As própolis 3 e 13 (EEP e respectivas fr-dicloro) apresentaram atividade anti-biofilme em pré-adesão com valores máximos de 250 µg/mL para EEP tipo 3 (C. parapsilosis) e 250 µg/mL para fr-dicloro tipo 3 (C. parapsilosis) e CIM 125 µg/mL para EEP tipo 13 (C. tropicalis) e 31,3 µg/mL para fr-dicloro tipo 13 (C. parapsilosis). As imagens de MEV mostram a interferência dos EEP tipos 3 e 13 e respectivas fr-dicloro sobre biofilmes em pré-adesão, entretanto as própolis não foram capazes de desagregar completamente os biofilmes formados de Candida spp se comparadas ao grupo controle (Nistatina). Conclui-se que os três tipos de própolis estudadas apresentaram atividade anti-Candida tanto em organização planctônica quanto em biofilme, entretanto, a fração diclorometânica da própolis do tipo 13 é a fração ativa, que contém importantes compostos efetivos contra Candida spp. e pode ser fonte de novos princípios bioativos para prevenção ou tratamento da candidíase oral, a qual deve ser objeto de estudos futuros / Abstract: Oral candidiasis is a fungal infection that has been reported as a public health problem and due to increased rate of resistant strains of Candida spp. has increased the interest in natural products as sources for new bioactive compounds. Brazilian propolis have important biological properties such as antimicrobial and anticariogenic activity, thus antifungal activity could also be evaluated. The aim of this study was to evaluate the anti-Candida activity of three Brazilian propolis against planktonic and biofilm forms of Candida spp. of higher incidences in oral cavity diseases.Three types of propolis were selected for antimicrobial properties in previous studies, which are classified in the literature as type 3 (within the state of Parana state), type 6 (Atlantic Rainforest of Bahia state) and type 13 (mangrove region, Maceió city, Alagoas state). For each type of propolis were obtained the ethanolic extract (EEP) and respective fractions with different gradient polarity (hexane, dichloromethane and ethyl acetate). The following methodologies were applied on EEP and correspondent fractions to evaluate antifungal activity against Candida spp. by a bioguided study: 1 - Determination of Minimum Inhibitory Concentration (MIC) and Minimum Fungicidal Concentrations (MFC) of EEP types 3, 6 and 13 and hexane (hex-fr), dichloromethane (dichlo-fr) and ethyl acetate (ethyl-fr) against planktonic forms of yeasts from an international bank Centraalbureau voor Schimmelcultures (CBS): Candida albicans (CBS 562), Candida dubliniensis (CBS 7987), Candida glabrata (CBS 07), Candida krusei (CBS 573), Candida parapsilosis (CBS 604), Candida tropicalis (CBS 94) using the microdilution method. 2 - Determination of MIC of propolis 3 and 13 (EEP and dichloromethane fractions) on preformed biofilm. 3 - Verification of possible changes or damages of the yeast cell structure when subjected to 3 and 13 propolis (EEP and dichloromethane fractions) by Scanning Electron Microscopy (SEM) in preformed and mature biofilms. In most cases, propolis types 3 and 13 demonstrated strong anti-Candida activity against the yeasts tested (minimum and maximum MIC values 4.0 and 31.3 µg/mL for EEP type 3 and 1.0 and 7.8 µg/mL for EEP type 13 compared to the low effect of propolis type 6. EEP type 13 and respective dichlo-fr had the best fungicidal effect against all strains (minimum and maximum MFC values 125 and 500 µg/mL). EEP type 3 and 13 and their dichlo-fr presented anti-biofilm activity 250 µg/mL for EEP type 3 (C. parapsilosis), 250 µg/mL for dichlo-fr type 3 (C. parapsilosis) and (maximum MIC 125 µg/mL for EEP type 13 (C. tropicalis), 31.3 µg/mL for dichlo-fr type 13 (C. parapsilosis). SEM images show the influence of propolis samples on preformed biofilms, however these samples were not capable of completely disrupting the mature biofilms, if compared to control (Nistatin). Thus, we concluded that the three types of propolis studied presented anti-Candida activity both in organization and in planktonic biofilm, however, the dichloromethane fraction of propolis type 13 is the active fraction, which contains important compounds effective against Candida spp. and could be a source of new bioactive principles for prevention or alternative treatment of oral candidiasis, which should be the object of future studies / Mestrado / Farmacologia, Anestesiologia e Terapeutica / Mestre em Odontologia

Made available in DSpace on 2014-06-11T19:28:38Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-03-28Bitstream added on 2014-06-13T20:58:44Z : No. of bitstreams: 1 carmello_jc_me_arafo.pdf: 701741 bytes, checksum: 55eb9d8f5746ef34ede24573f0c2455a (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Este trabalho teve por objetivo avaliar a efetividade da terapia fotodinâmica (PDT) mediada pelo fotossensibilizador (FS) Photodithazine® (PDZ), associado à luz do tipo LED (660nm). Para tanto, foram utilizados 55 camundongos com aproximadamente 6 semanas de vida, os quais foram submetidos a indução de candidose no dorso da língua. Inicialmente os animais foram imunossuprimidos e no dia seguinte se realizou a inoculação de C. albicans na língua dos mesmos por meio de swabs embebidos na suspensão (107 Ufc/mL). Para a realização da terapia fotodinâmica o FS foi avaliado nas concentrações de 75, 100, 125 e 150mg/L. Tais grupos experimentais foram denominados de (P+L+ 75mg/L, P+L+ 100mg/L, P+L+ 125mg/L, P+L+ 150mg/L) associados a uma dose de luz de 37,5 J/cm2. Para a verificação apenas do efeito da PDZ, a mesma foi aplicada na língua dos animais, sem iluminação (grupos denominados de P+L- 75mg/L, P+L- 100mg/L, P+L- 125mg/L, P+L- 150mg/L). O efeito da luz foi avaliado por meio da iluminação das línguas com dose de luz de 37,5J/cm2, (grupo denominado de P-L+ 37,5J/cm2). Um grupo recebeu apenas inoculação por Candida (grupo P-L-, controle positivo), outro grupo não recebeu nenhum tratamento e nem inoculação fúngica (grupo CN – controle negativo). Após os experimentos realizou-se a recuperação de C. albicans das línguas dos animais por meio de swabs que foram esfregados sobre as mesmas durante 1 minuto. Esses swabs foram embebidos em tubos de ensaio com 1mL de solução salina, e diluições seriadas foram realizadas e colocadas em placas de petri com SDA. Após 48 horas de incubação a 37º C as células fúngicas foram quantificadas e o número de Ufc/mL foi determinado e analisado pelo teste ANOVA (P<.05). Os camundongos foram sacrificados e tiveram as línguas removidas cirurgicamente para realização da análise histológica. Os resultados demonstraram... / The aim of this investigation was to evaluate the effectiveness of photodynamic therapy (PDT) mediated by photossensitizer Photodithazine® (PDZ) associated with LED light (660nm) for the photoinactivation of C. albicans in a murine model of oral candidosis. Fifty-five 6-week-old female Swiss mice were immunossuppressed and in the next day small cotton pads were soaked in a C. albicans cell suspension (107CFU/mL) and swabbed in the oral cavity of mice. PDT was performed and the PS was applicated topically on the dorsum of the tongue of mice at concentrations 75, 100, 125 and 150mg/L (P+L+75, P+L+100, P+L+125 and P+L+150mg/L) associated with LED at a fluence of 37,5J/cm2. The effect of PS only was tested by application of PDZ for the same period of pre-irradiation time and irradiation at the same concentration as that for the P+L+ group, without the LED illumination (P+L-75, P+L-100, P+L-125 and P+L-150mg/L). To verify the effect of the light only, the group was exposed to the same LED dose mentioned previously (P-L+ 37,5J/cm2), 1 group). The positive control did not receive any PS or light (P-L-). The negative control group (CN) of animals was evaluated and mice did not receive any treatment. After treatment the dorsum of the tongue was swabbed for 1 minute with a cotton pad to recover C. albicans cells and the microbiological evaluation was performed. The yeast colony counts were quantified and the number of CFU/mL was determined and analyzed by ANOVA test (P<.05). Animals were killed 24 hours after treatment and the tongue of all mice were surgically removed for histological analysis. The results of this investigation demonstrated that PDT was effective in reducing C. albicans recovered from the tongue of mice at concentrations 100, 125 and 150mg/L of PS when compared with the animals from the positive control group (P-L-) (P<0.05). There was no difference between these concentrations... (Complete abstract click electronic access below)

Magister Chirurgiae Dentium (MChD) / The rate at which the population is ageing is much faster than in the past. An increase in age results in an increase in oral diseases. One of the most common types of oral diseases in the elderly are fungal infections caused by Candida albicans. It has been noted that drug resistance to fungal pathogens is developing into a serious threat to public health and healthcare systems worldwide. This has consequently led to the need to develop effective and innocuous treatment modalities. The purpose of this study was to explore the antimicrobial and cytotoxic potential of silver nanoparticles (AgNPs) synthesised from Berzelia lanuginose, Helichrysum cymosum, and Searsia crenata.

Objectives: Candida glabrata is the second most common cause of oral candidiasis, second only to C. albicans. Incidence of antifungal resistance has shown a steady increase for C. glabrata. Iron has shown to modulate C. albicans pathogenesis and affect drug-susceptibility. Here, we assess the effect of iron on the growth, antifungal-susceptibility, biofilms, and cell wall of C. glabrata.Methods: Growth, minimal inhibitory concentration (MIC), and biofilm experiments were conducted using 96-well polystyrene plates. Yeast Nitrogen Base medium was used for growth experiments. Cultures of C. glabrata and C. albicans were grown over two nights in respective media containing varying iron concentrations. Rosewell Park Memorial Institute medium was used for MIC and biofilm experiments. Serial dilution was performed to obtain desired concentrations of antifungal drugs. For all experiments, growth was assessed at OD600nm over 24 hours using BioTek Synergy Multi Mode Reader. Paraformaldehyde treated cells and specific stains were used for cell wall studies. Results: Growth of C. glabrata declined significantly below 5μM iron, while C. albicans continued to grow at decreasing iron concentrations, up to 0.5μM. MIC experiments revealed 1.562μM, 1.562μM, and 4μM, as the MIC for Deferasirox, Nystatin, and Fluconazole, respectively. Drug synergy experiments revealed a 128-fold reduction in the amount of Nystatin and Fluconazole needed, with the addition of 1/8th of Deferasirox concentration. The biofilm experiments were inconclusive and the cell wall studies showed decreased levels of mannan, chitin, and an increased β-glucan exposure in high iron conditions. Conclusion: C. glabrata is more sensitive to alterations in environmental iron when compared to C. albicans. Drug synergy experiments underscore the importance of Deferasirox in lowering the MICs of Nystatin or Fluconazole. This can allow use of classical antifungals at lower doses, thereby limiting their side effects. Cell wall studies discuss the effect of iron on the virulence of the C. glabrata. / Oral Biology

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