Tipagem molecular de Candida albicans isoladas de pacientes hospitalizados

SUMMARY Introduction: The majority of nosocomial fungal infections are caused by Candida spp. where C. albicans is the species most commonly identified. Molecular methods are important tools for assessing the origin of the yeasts isolated in hospitals. Methods: This is a study on the genetic profifiles of 39 nosocomial clinical isolates of C. albicans using two typing methods: random amplifified polymorphic DNA (RAPD) and microsatellite, two different primers for each technique were used. Results: RAPD provided 10 and 11 different profiles with values for SAB of 0.84 ± 0.126 and 0.88 ± 0.08 for primers M2 and P4, respectively. Microsatellite using two markers, CDC3 and HIS3, allowed the observation of six and seven different alleles, respectively, with combined discriminatory power of 0.91. Conclusions: Although genetic variability is clear, it was possible to identify high similarity, suggesting a common origin for at least a part of isolates. It is important to emphasize that common origin was proven from yeasts isolated from colonization (urine, catheter or endotracheal secretions) and blood culture from the same patient, indicating that the candidemia must have started from a site of colonization. The combination of RAPD and microsatellite provides a quick and efficient analysis for investigation of similarity among nosocomial isolates of C. albicans.RESUMO Introdução: A maioria das infecções fúngicas hospitalares são causadas por Candida spp. e C. albicans é a espécie mais comumente identificada. Métodos moleculares são ferramentas importantes para a avaliação da origem das leveduras isoladas em hospitais. Métodos: Este é um estudo sobre o perfil genético de 39 isolados clínicos nosocomiais de C. albicans através das técnicas de RAPD e microssatélite, foram usados dois diferentes iniciadores para cada técnica. Resultados: RAPD forneceu 10 e 11 diferentes perfis com valores de SAB 0,84 ± 0,126 e 0,88 ± 0,08 para os primers M2 e P4, respectivamente. A análise de microssatélites, usando os marcadores CDC3 e HIS3 permitiu a observação de seis e sete diferentes alelos respectivamente, com poder discriminatório combinado de 0,91. Conclusões: Embora seja clara a variabilidade genética, foi possível identificar alta similaridade, sugerindo origem comum para pelo menos parte deles. É importante enfatizar que foi comprovada origem comum de leveduras isoladas de colonização (urina, cateter ou secreção orotraqueal) e hemocultura do mesmo paciente, indicando que a candidemia deve ter iniciado a partir de um sítio de colonização. A combinação das técnicas RAPD e microssatélites fornece uma análise rápida e eficiente para investigação de similaridade entre isolados nosocomiais de C. albicans

EFFECT OF HIGHLY ACTIVE ANTIRETROVIRAL THERAPY ON VAGINAL Candida spp. ISOLATION IN HIV-INFECTED COMPARED TO HIV-UNINFECTED WOMEN

Candidíase vulvovaginal (CVV) em mulheres infectadas pelo HIV contribuiu substancialmente para a diminuição da sua qualidade de vida. O objetivo deste estudo foi avaliar o efeito do uso de terapia anti-retroviral altamente ativa (HAART) no isolamento de Candida spp. vaginais em mulheres HIV positivas comparado às não infectadas por HIV. Este estudo transversal incluiu 178 mulheres infectadas pelo HIV (grupo HIV) e 200 mulheres não infectadas (grupo controle) acompanhadas no Serviço de Assistência Especializada (SAE) para as doenças sexualmente transmissíveis (DST)/AIDS da cidade de Maringá/Brasil, de 1 abril a 30 de outubro de 2011. As leveduras foram isoladas e identificadas por métodos fenotípicos e moleculares. A susceptibilidade in vitro aos antifúngicos fluconazol, itraconazol, nistatina e anfotericina B foi avaliada pelo método de referência de microdiluição. Nós encontramos maior frequência de isolamento vaginal total de Candida spp. no grupo HIV do que no grupo controle. Entretanto, foi observada frequência similar de colonização e CVV entre os dois grupos. Apesar de C. albicans ser a mais frequente e sensível a azólicos e polienos em mulheres infectadas pelo HIV e não infectadas, foi detectada emergente resistência de C. glabrata a AMB nas mulheres infectadas pelo HIV. Embora tenha sido observada maior frequência de isolamento vaginal de Candida spp. nas mulheres infectadas pelo HIV do que nas não infectadas, colonização e CVV apresentaram frequência similar em ambos os grupos, o que indica que HAART parece proteger contra colonização vaginal e CVV. Vulvovaginal candidiasis (VVC) in HIV-infected women contributed to the impairment of their quality of life. The aim of this study was to evaluate the effect of highly active antiretroviral therapy (HAART) use on the vaginal Candida spp. isolation in HIV-infected compared to HIV-uninfected women. This cross-sectional study included 178 HIV-infected (HIV group) and 200 HIV-uninfected women (control) that were studied at the Specialized Assistance Service (SAE) for sexually transmitted diseases (STD)/AIDS of the city of Maringá, Brazil, from April 1 to October 30, 2011. The yeasts were isolated and identified by phenotypic and molecular methods. The in vitro antifungal susceptibility to fluconazole, itraconazole, nystatin and amphotericin B was tested by the reference microdilution method. Higher frequencies of total vaginal Candida spp. isolation were found in the HIV-infected group than in the control group. However, both groups showed a similar frequency of colonization and VVC. Although C. albicans was the most frequent and sensitive to azolics and polyenes in both HIV-infected and uninfected women, the emerging resistance of C. glabrata to amphotericin B in the HIV-infected women was observed. Although higher frequency of vaginal Candida spp. isolation had been observed in the HIV-infected than in HIV-uninfected women, colonization and VVC showed similar frequency in both groups, indicating that HAART appears to protect against vaginal colonization and VVC

Candida albicans PROTEIN PROFILE CHANGES IN RESPONSE TO THE BUTANOLIC EXTRACT OF Sapindus saponariaL.

Candida albicans is an opportunistic human pathogen that is capable of causing superficial and systemic infections in immunocompromised patients. Extracts of Sapindus saponaria have been used as antimicrobial agents against various organisms. In the present study, we used a combination of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify the changes in protein abundance of C. albicans after exposure to the minimal inhibitory concentration (MIC) and sub-minimal inhibitory concentration (sub-MIC) of the butanolic extract (BUTE) of S. saponaria and also to fluconazole. A total of six different proteins with greater than 1.5 fold induction or repression relative to the untreated control cells were identified among the three treatments. In general, proteins/enzymes involved with the glycolysis (GPM1, ENO1, FBA1), amino acid metabolism (ILV5, PDC11) and protein synthesis (ASC1) pathways were detected. In conclusion, our findings reveal antifungal-induced changes in protein abundance of C. albicans. By using the previously identified components of the BUTE of S. saponaria(e.g., saponins and sesquiterpene oligoglycosides), it will be possible to compare the behavior of compounds with unknown mechanisms of action, and this knowledge will help to focus the subsequent biochemical work aimed at defining the effects of these compounds

Candida albicans PROTEIN PROFILE CHANGES IN RESPONSE TO THE BUTANOLIC EXTRACT OF Sapindus saponaria L.

Candida albicans is an opportunistic human pathogen that is capable of causing superficial and systemic infections in immunocompromised patients. Extracts of Sapindus saponaria have been used as antimicrobial agents against various organisms. In the present study, we used a combination of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) to identify the changes in protein abundance of C. albicans after exposure to the minimal inhibitory concentration (MIC) and sub-minimal inhibitory concentration (sub-MIC) of the butanolic extract (BUTE) of S. saponaria and also to fluconazole. A total of six different proteins with greater than 1.5 fold induction or repression relative to the untreated control cells were identified among the three treatments. In general, proteins/enzymes involved with the glycolysis (GPM1, ENO1, FBA1), amino acid metabolism (ILV5, PDC11) and protein synthesis (ASC1) pathways were detected. In conclusion, our findings reveal antifungal-induced changes in protein abundance of C. albicans. By using the previously identified components of the BUTE of S. saponaria(e.g., saponins and sesquiterpene oligoglycosides), it will be possible to compare the behavior of compounds with unknown mechanisms of action, and this knowledge will help to focus the subsequent biochemical work aimed at defining the effects of these compounds

Identificação microbiológica e sensibilidade in vitro de Candida isoladas da cavidade oral de indivíduos HIV positivos

A candidíase orofaríngea é a infecção fúngica mais comum entre os pacientes infectados pelo vírus da imunodeficiência humana e seu tratamento é realizado com antifúngicos tópicos ou sistêmicos, que são indicados empiricamente com base em dados clínicos. O objetivo deste estudo foi determinar a freqüência de leveduras em lavados bucais de indivíduos HIV positivos, comparando os resultados entre pacientes com diferentes condições de imunodeficiência e verificar o perfil de susceptibilidade das espécies isoladas frente aos antifúngicos visando avaliar se as opções de tratamento utilizadas na prática clínica atingem a maioria das espécies identificadas. Leveduras foram isoladas em 58% das amostras de lavado bucal coletadas e Candida albicans foi a espécie mais (93%) freqüente. Resistência ou susceptibilidade dose dependente, frente aos antifúngicos testados foi registrada em aproximadamente 17% das amostras. A importante variabilidade de resposta sugere limitações quanto à eficácia das terapias instituídas empiricamente

MOLECULAR TYPING OF Candida albicans ISOLATES FROM HOSPITALIZED PATIENTS

SUMMARY Introduction: The majority of nosocomial fungal infections are caused by Candida spp. where C. albicans is the species most commonly identified. Molecular methods are important tools for assessing the origin of the yeasts isolated in hospitals. Methods: This is a study on the genetic profifiles of 39 nosocomial clinical isolates of C. albicans using two typing methods: random amplifified polymorphic DNA (RAPD) and microsatellite, two different primers for each technique were used. Results: RAPD provided 10 and 11 different profiles with values for SAB of 0.84 ± 0.126 and 0.88 ± 0.08 for primers M2 and P4, respectively. Microsatellite using two markers, CDC3 and HIS3, allowed the observation of six and seven different alleles, respectively, with combined discriminatory power of 0.91. Conclusions: Although genetic variability is clear, it was possible to identify high similarity, suggesting a common origin for at least a part of isolates. It is important to emphasize that common origin was proven from yeasts isolated from colonization (urine, catheter or endotracheal secretions) and blood culture from the same patient, indicating that the candidemia must have started from a site of colonization. The combination of RAPD and microsatellite provides a quick and efficient analysis for investigation of similarity among nosocomial isolates of C. albicans

Antifungal susceptibility and distribution of Candida spp. isolates from the University Hospital in the municipality of Dourados, State of Mato Grosso do Sul, Brazil

Introduction Hospital infections caused by Candida spp. are a leading cause of morbidity and mortality in hospitalized patients, particularly those that are critically ill or immunocompromised. In this study, the distribution of Candida species in isolates from the University Hospital of the Federal University at Grande Dourados and their in vitro susceptibility to antifungal drugs were analyzed. Methods Yeasts were phenotypically identified using classical methodologies. Antifungal susceptibility tests to amphotericin B and fluconazole were performed using the broth microdilution technique. Results A total of 50 Candida isolates were obtained from hospitalized patients during the study period. We analyzed yeast isolates from urine (n=31; 62%), blood (n=12; 24%), and tracheal secretions (n=7; 14%). The following Candida species were identified: C. tropicalis (n=21; 42%), C. albicans (n=18; 36%), C. glabrata (n=10; 20%), and C. krusei (n=1; 2%). Antifungal susceptibility tests demonstrated that C. albicans was susceptible to both antifungal agents. However, 31.2% of the non-C. albicans Candida isolates displayed dose-dependent susceptibility to fluconazole, and 3.1% were resistant to amphotericin B. Conclusions In contrast to previous reports, our results indicated that C. tropicalis was the most commonly isolated yeast species among the hospital patients. The predominance of non-C. albicans Candida infections confirms the importance of species-level identification for implementing appropriate antifungal therapies