Genetic diversity and antifungal susceptibility testing of Trichosporon asahii isolated of Intensive Care Units patients

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Molecular typing and antifungal susceptibility of clinical sequential isolates of Cryptococcus neoformans from São Paulo State, Brazil

The antifungal susceptibility profiles and the genetic variability of 83 sequential clinical isolates of Cryptococcus neoformans, including four Cryptococcus gattii isolates, obtained from 38 São Paulo AIDS patients with cryptococcal meningitis were assessed by electrophoretic karyotyping and random amplified polymorphic DNA (RAPD) analysis. The majority of the Cryptococcus neoformans isolates were highly susceptible to amphotericin B and fluconazole. Twenty percent of the minimum inhibitory concentration values for amphotericin B varied from 0.5 to 1 mu g mL(-1). For fluconazole, 22% occurred in the range 8-16 mu g mL(-1). Sequential isolates from nine patients showed a trend towards lower susceptibility to fluconazole, flucytosine, itraconazole and amphotericin B. The results of molecular typing by electrophoretic karyotyping and RAPD analysis showed the presence of 22 electrophoretic karyotypes (EK) and 15 RAPD profiles that were highly correlated. Our results provided evidence for the occurrence of genetic changes in some strains associated with microevolution during the course of infection. We also observed both microevolution and simultaneous coinfection with two distinct Cryptococcus neoformans strains in one patient. In some patients, we found changed EK- and RAPD patterns in association with increased MIC values.

Prevalence and antifungal susceptibility of Candida parapsilosis complex isolates collected from oral cavities of HIV-infected individuals

At present, few data are available on the prevalence and antifungal susceptibility of Candida parapsilosis complex isolates from HIV-infected individuals. The C. parapsilosis complex comprises three species, C. parapsilosis sensu stricto, C. metapsilosis and C. orthopsilosis. Fifteen of 318 Candida isolates were identified as members of the C. parapsilosis complex by PCR and restriction fragment length polymorphism (RFLP). The prevalence of C. parapsilosis complex isolates was 4.7 %, 2.2 % being identified as C. parapsilosis sensu stricto and 2.5% as C. metapsilosis, while no C. orthopsilosis was isolated. This is believed to be the first study that has identified isolates of C. metapsilosis obtained from the oral cavity of HIV-infected individuals. Antifungal susceptibility tests indicated that all the isolates were susceptible to amphotericin B (AMB), fluconazole (FLC), ketoconazole (KTC), itraconazole (ITC), voriconazole (VRC) and caspofungin (CASPO). Although isolates of C. parapsilosis sensu stricto and C. metapsilosis were susceptible to FLC, isolates of C. metapsilosis showed a tendency for higher MICs (>= 1.0 mu g ml(-1)). Based upon the frequency of candidiasis and the fact that certain isolates of the C. parapsilosis complex respond differently to FLC therapy, our data may be of therapeutic relevance with respect to susceptibility and potential resistance to specific antifungal agents. Our data suggest that C. metapsilosis can be a human commensal; its importance as a pathogen has yet to be confirmed.

Microdilution procedure for antifungal susceptibility testing of Paracoccidioides brasiliensis to amphotericin B and itraconazole

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

In vitro antifungal susceptibility of Candida spp. oral isolates from HIV-positive patients and control individuals

Oropharyngeal candidiasis is the most common fungal infection among HIV-positive patients. This condition can be treated with either systemic or topical antifungal agents; treatments are usually indicated empirically on the basis of clinical data. The knowledge of in vitro antifungal susceptibility is important to determine correct therapeutic guides for the treatment of fungal infections. Therefore, the objective of this study was to determine the antifungal susceptibility profile of oral Candida isolates from HIV-positive patients and control individuals. Amphotericin B, fluconazole, flucytosine, nystatin and ketoconazole were tested according to the methodology of microdilution proposed by the Clinical and Laboratory Standards Institute (CLSI); results were recorded in values of minimal inhibitory concentration (MIC). A total of 71 Candida isolates from HIV-positive patients were examined with the following species represented: C. albicans (59), C. tropicalis (9), C. glabrata (1), C. guilliermondii (1) and C. krusei (1). A total of 15 Candida isolates were evaluated from control individuals comprised of 11 C. albicans and 4 C. tropicalis samples. Our results demonstrated that the tested antifungal agents showed good activity for most isolates from both groups; however, variability in MIC values among isolates was observed.

Comparison between E-test and CLSI broth microdilution method for antifungal susceptibility testing of Candida albicans oral isolates

Trinta Candida albicans isoladas de pacientes portadores de candidose oral e 30 Candida albicans isoladas de indivíduos controle foram estudadas. Testes de susceptibilidade in vitro foram realizados com anfotericina B, fluconazol, 5-flucitosina e itraconazol pelo método do Clinical and Laboratorial Standars Institute (CLSI) e por E-test. Os resultados obtidos foram analisados e comparados. Os valores de CIM foram semelhantes para amostras isoladas de pacientes portadores de candidose oral e indivíduos controle. A concordância entre os dois métodos foi de 66,7% para a anfotericina B, 53,33% para o fluconazol, 65% para a flucitosina e 45% para o itraconazol. de acordo com estes resultados, o método do E-test poderia ser uma alternativa para a triagem de casos de rotina pela sua simplicidade. Entretanto, este método não pode ser considerado como um substituto para o método de referência do CLSI.

Species distribution and antifungal susceptibility patterns of Candida spp. bloodstream isolates from a Brazilian tertiary care hospital

In this work, we collect data from surveys of bloodstream Candida isolates performed in Brazil from 1996 to 2004. Besides, we analyzed the species distribution of bloodstream Candida isolates together with potential risk factors for candidemia and the susceptibility profile of these isolates in patients from Hospital das Clinicas in Goiaonia city, Brazil. Blood samples were collected in the admission day and on every 7 days, in the intensive care unit (ICU) of a tertiary hospital. Candida isolates were identified by standard protocols that included germ tube formation, chlamydoconidia production on cornmeal agar and sugar fermentation and assimilation tests. Data of patients were recorded and analyzed according to age at the time of diagnosis, gender and presence of potential risk factors. Statistical analysis was used to determine if the time of hospital permanence increased Candida colonization in ICU patients' blood. The antifungal susceptibility testing was performed by broth microdilution method according to document NCCLS/CLSI M27-A2. Among the 345 blood samples cultured, candidemia was recovered in 33 patients, which were isolated 51.5% of Candida non-albicans. Fungemia was associated with long-term hospitalization. Fluconazole, itraconzole, voriconazole and amphotericin B exhibited a potent activity against all isolates of Candida. Voriconazole MICs were much low for all isolates tested. This work confirms data of increase of Candida non-albicans species in bloodstream in ICU and shows that voriconazole in vitro activity was higher than those of itraconazole, fluconazole and amphotericin B.

Prevalence and in vitro antifungal susceptibility of Candida spp isolated from clinical specimens in São Paulo, Brazil

We examined the prevalence and the in vitro susceptibility to antifungal drugs of Candida spp isolated from clinical specimens at our university hospital in São Paulo, Brazil. Among 6,417 samples studied, positive cultures, were obtained from 222 (3.5%) most of them (68%) from the pediatric unit and nursery. Candida albicans and Candida parapsilosis were the most frequent species and the susceptibility patterns of a panel of 130 isolates to amphotericin B, ketoconazole and fluconazole. showed that the order of antifungal efficacy was amphotericin B > ketoconazole > fluconazole.

In vitro antifungal susceptibility of Candida spp. isolates from patients with chronic periodontitis and from control patients

Superinfection by Candida can be refractory to conventional periodontal treatments in specific situations, such as in immunocompromised patients. In these cases, the systemic therapy with antifungal drugs could be indicated. The aim of this study was to analyse antifungal susceptibility of Candida spp. strains isolated from chronic periodontitis patients and from control individuals. A total of 39 C. albicans isolates, 9 C. tropicalis, 2 C. glabrata and 5 Candida spp. from control individuals and 30 C. albicans, 3 C. tropicalis and 2 C. glabrata from periodontitis patients were tested. In the control group, 1 isolate of C. glabrata was resistant to ketoconazole and 1 Candida spp. was resistant to amphotericin B, ketoconazole and miconazole. Among the isolates of periodontitis group, 1 (3.33%) C. albicans isolate was resistant to flucytosine and ketoconazole. According to the obtained results, it could be concluded that fluconazole was the most effective drug against the several Candida species studied. There were not expressive differences in the susceptibility of isolates from periodontitis patients or from control individuals.

Candiduria in adults and children: prevalence and antifungal susceptibility in outpatient of Jataí-GO

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

In vitro antifungal susceptibility of candida albicans isolates from patients with chronic periodontitis and diabetes

Alterations that lead to deficiency of the immune system, such as diabetes mellitus, may promote proliferation of Candida albicans and selection of strains which have greater ability to adhere and to penetrate the host tissues. Recent studies indicate an increase of the antifungal resistance of C. albicans isolates in periodontal pockets, suggesting that the oral cavity could be a reservoir of resistant yeast to antifungal agents. Moreover, oral cavity can act as a reservoir of certain pathogens that may cause systemic infections. The periodontal pocket is an ecological niche suitable to host microorganisms that could act as opportunistic pathogens. The aim of this study is to contribute to the understanding of resistance to conventional antifungal against C. albicans isolates from patients with periodontitis and diabetes. The determination of the minimal inhibitory concentrations (MIC) was evaluated according to M27S3 of the CLSI (2008), with modifications. The results showed that 48.8% of the studied strains were resistant to one or more antifungals and 6.6% were resistant to fluconazole and voriconazole. These results suggest an increasing resistance to conventional antifungal agents among Candida species, suggesting that the oral cavity could host pathogen fungi.

Antifungal Efficacy during Candida krusei Infection in Non-Conventional Models Correlates with the Yeast In Vitro Susceptibility Profile

The incidence of opportunistic fungal infections has increased in recent decades due to the growing proportion of immunocompromised patients in our society. Candida krusei has been described as a causative agent of disseminated fungal infections in susceptible patients. Although its prevalence remains low among yeast infections (2-5%), its intrinsic resistance to fluconazole makes this yeast important from epidemiologic aspects. Non mammalian organisms are feasible models to study fungal virulence and drug efficacy. In this work we have used the lepidopteran Galleria mellonella and the nematode Caenorhabditis elegans as models to assess antifungal efficacy during infection by C. krusei. This yeast killed G. mellonella at 25, 30 and 37°C and reduced haemocytic density. Infected larvae melanized in a dose-dependent manner. Fluconazole did not protect against C. krusei infection, in contrast to amphotericin B, voriconazole or caspofungin. However, the doses of these antifungals required to obtain larvae protection were always higher during C. krusei infection than during C. albicans infection. Similar results were found in the model host C. elegans. Our work demonstrates that non mammalian models are useful tools to investigate in vivo antifungal efficacy and virulence of C. krusei. © 2013 Scorzoni et al.