Genetic Diversity of the Cryptococcus gattii Species Complex in Mato Grosso State, Brazil.

Cryptococcosis is caused by fungi of the genus Cryptococcus. Owing to its importance, this study aimed to analyze the genetic diversity of C. gattii isolates from animals, humans, and the environment in Mato Grosso State (MT), Brazil, during November 2010-December 2017. All isolates of the C. gattii species complex were subjected to molecular genotyping via Restriction Fragment Length Polymorphism (PCR-RFLP) and Multi-locus Sequence Typing (MLST). PCR-RFLP analysis revealed that 21 isolates presented the genotype VGII, which is considered the most common and virulent genotype globally among. MLST analysis revealed the presence of 14 sequence types (STs), of which 5 are considered new genotypes. Clonal Complex (CC) CC182 (n = 5; 23,80%) and CC309 (n = 3; 14,28%) were the most frequent. CC distribution in relation to origin revealed that three CCs were found in animals with a predominance of CC182 (66,66%), while nine were found in humans, and two CCs were found in the environment. Extensive genetic variability was observed among the isolates in the State of Mato Grosso. STs belonging to the already described clonal complexes (CC) indicate the global expansion and adaptation of isolates in several other countries. Therefore, detection of clonal complexes and STs already described in other regions and the occurrence of new STs in the present study help further the current understanding of the geographic dispersion and genetic origin of the C. gattii species complex.

Molecular typing and in vitro antifungal susceptibility of Cryptococcus spp from patients in Midwest Brazil.

INTRODUCTION: Cryptococcosis is a systemic fungal infection that affects humans and animals, mainly due to Cryptococcus neoformans and Cryptococcus gattii. Following the epidemic of acquired immunodeficiency syndrome (AIDS), fungal infections by C. neoformans have become more common among immunocompromised patients. Cryptococcus gattii has primarily been isolated as a primary pathogen in healthy hosts and occurs endemically in northern and northeastern Brazil. We to perform genotypic characterization and determine the in vitro susceptibility profile to antifungal drugs of the Cryptococcus species complex isolated from HIV-positive and HIV-negative patients attended at university hospitals in Cuiabá, MT, in the Midwestern region of Brazil. METHODOLOGY: Micromorphological features, chemotyping with canavanine-glycine-bromothymol blue (CGB) agar and genotyping by URA5-RFLP were used to identify the species. The antifungal drugs tested were amphotericin B, fluconazole, flucytosine, itraconazole and voriconazole. Minimum inhibitory concentrations (MICs) were determined according to the CLSI methodology M27-A3. RESULTS: Analysis of samples yelded C. neoformans AFLP1/VNI (17/27, 63.0%) and C. gattii AFLP6/VGII (10/27, 37.0%). The MICs ranges for the antifungal drugs were: amphotericin B (0.5-1 mg/L), fluconazole (1-16 mg/L), flucytosine (1-16 mg/L), itraconazole (0.25-0.12 mg/L) and voriconazole (0.06-0.5 mg/L). Isolates of C. neoformans AFLP1/VNI were predominant in patients with HIV/AIDS, and C. gattii VGII in HIV-negative patients. The genotypes identified were susceptible to the antifungal drugs tested. CONCLUSION: It is worth emphasizing that AFLP6/VGII is a predominant genotype affecting HIV-negative individuals in Cuiabá. These findings serve as a guide concerning the molecular epidemiology of C. neoformans and C. gattii in the State of Mato Grosso.

Fatal Cryptococcus gattii genotype AFLP6/VGII infection in a HIV-negative patient: case report and a literature review.

Cryptococcus gattii, a species belonging to the Cryptococcus complex which occurs endemically in tropical and subtropical regions, has been reported as a causative agent of cryptococcosis in healthy individuals. We report a case of meningitis in HIV-negative patient from Cuiaba, MT, in the Midwestern region of Brazil. Cryptococcus gattii AFLP6/VGII was isolated from cerebrospinal fluid and molecular typing was performed by URA5-RFLP. The in vitro susceptibility profile was determined using the standard method according to the document M27A3, CLSI 2008. C. gattii AFLP6/VGII was shown to be susceptible to the antifungals tested. Treatment with 0.8 mg/kg of amphotericin B was initiated; however, the patient died 2 days after the onset of therapy.

[Mycological aspects and susceptibility in vitro the yeast of the genus Candida from HIV-positive patients in the State of Mato Grosso]. / Aspectos micológicos e suscetibilidade in vitro de leveduras do gênero Candida em pacientes HIV-positivos provenientes do Estado de Mato Grosso.

INTRODUCTION: Candidiasis is one of the most common fungal infections among patients infected by human immunodeficiency virus. The present study aimed to characterize yeasts of the genus Candida from distinct clinical samples from HIV-positive patients and determine the in vitro susceptibility profile to five antifungal drugs. METHODS: Characterization of Candida sp was achieved using the classic methodology: biochemical (zymogram and auxanogram) and micromorphology (germinative tube growth test and slide microculture) tests. Genotypic technique (PCR) and identification by the commercial method API 20C AUX (Biomeriéux) were also performed. To determine the in vitro susceptibility profile, five antifungal drugs were used (ketoconazole, fluconazole, itraconazole, voriconazole and amphotericin-B) following a commercially available method, the Etest. RESULTS: The procedure isolated 105 yeasts of the genus Candida from 102 HIV-infected patients. Of these, 82 (78.1%) were characterized as Candida albicans, 8 (7.6%) as C. parapsilosi s, 8 (7.6%) C. tropicalis, 4 (3.8%) C. krusei, 2 (1.9%) C. glabrata, and 1 (1%) as C. guiilliermondii. CONCLUSIONS: Considering the general profile of sensitivity, 60% of isolates were susceptible to all the antifungal drugs tested; however, the species C. tropicalis and C. krusei showed a tendency toward higher MICs to azoles than those obtained for C. albicans, suggesting resistance.

Aspectos micológicos e suscetibilidade in vitro de leveduras do gênero Candida em pacientes HIV-positivos provenientes do Estado de Mato Grosso / Mycological aspects and susceptibility in vitro the yeast of the genus Candida from HIV-positive patients in the State of Mato Grosso

INTRODUÇÃO: A candidíase é uma das infecções fúngicas mais frequentes entre os pacientes infectados pelo vírus da imunodeficiência humana. O presente estudo objetivou a caracterização das leveduras do gênero Candida de distintas amostras clínicas, provenientes de pacientes HIV - positivos, assim como a determinação do perfil de suscetibilidade in vitro a cinco drogas antifúngicas. MÉTODOS: A caracterização dos isolados de Candida sp foi realizada através da metodologia clássica, testes bioquímicos (zimograma e auxanograma) e morfológicos (prova do tubo germinativo e microcultivo em lâmina). Também, foram realizadas a técnica genotípica (PCR) e identificação pelo método comercial API 20C AUX (BioMeriéux). Para a determinação do perfil de suscetibilidade in vitro, foram utilizadas cinco drogas antifúngicas (cetoconazol, fluconazol, itraconazol, voriconazol e anfotericina B), através do método comercialmente disponível - Etest. RESULTADOS: Foram identificados 105 isolados de leveduras do gênero Candida provenientes de 102 pacientes infectados pelo vírus HIV. Destes, foram caracterizadas 82 (78,1 por cento) Candida albicans, 8 (7,6 por cento) Candida parapsilosis, 8 (7,6 por cento) Candida tropicalis, 4 (3,8 por cento) Candida krusei, 2 (1,9 por cento) Candida glabrata e 1 (1 por cento) Candida guilliermondii. CONCLUSÕES: Considerando o perfil geral de sensibilidade, 60 por cento dos isolados foram suscetíveis a todos os antifúngicos testados, porém as espécies C. tropicalis e C. krusei demonstraram uma tendência a valores mais elevados de CIMs para os azóis do que os encontrados paraC. albicans, sugerindo resistência.

INTRODUCTION: Candidiasis is one of the most common fungal infections among patients infected by human immunodeficiency virus. The present study aimed to characterize yeasts of the genus Candida from distinct clinical samples from HIV-positive patients and determine the in vitro susceptibility profile to five antifungal drugs. METHODS: Characterization of Candida sp was achieved using the classic methodology: biochemical (zymogram and auxanogram) and micromorphology (germinative tube growth test and slide microculture) tests. Genotypic technique (PCR) and identification by the commercial method API 20C AUX (Biomeriéux) were also performed. To determine the in vitro susceptibility profile, five antifungal drugs were used (ketoconazole, fluconazole, itraconazole, voriconazole and amphotericin-B) following a commercially available method, the Etest. RESULTS: The procedure isolated 105 yeasts of the genus Candida from 102 HIV-infected patients. Of these, 82 (78.1 percent) were characterized as Candida albicans, 8 (7.6 percent) as C. parapsilosi s, 8 (7.6 percent) C. tropicalis, 4 (3.8 percent) C. krusei, 2 (1.9 percent) C. glabrata, and 1 (1 percent) as C. guiilliermondii. CONCLUSIONS: Considering the general profile of sensitivity, 60 percent of isolates were susceptible to all the antifungal drugs tested; however, the species C. tropicalis and C. krusei showed a tendency toward higher MICs to azoles than those obtained for C. albicans, suggesting resistance.

Primeira descrição da caracterização fenotípica e susceptibilidade in vitro a drogas de leveduras do gênero Cryptococcus spp isoladas de pacientes HIV positivos e negativos, Estado de Mato Grosso / First description of phenotypic profile and in vitro drug susceptibility of Cryptococcus spp yeast isolated from HIV-positive and HIV-negative patients in State of Mato Grosso

Foram avaliados 37 isolados de 10 pacientes HIV negativos e 26 positivos, em Mato Grosso. Exame direto, cultura e quimiotipagem de espécies foram realizados. Cetoconazol, itraconazol, voriconazol, fluconazol e anfotericina B foram avaliados. Foram identificadas 37 leveduras do gênero Cryptococcus spp sendo 26 de pacientes HIV- positivos (25 Cryptococcus neoformans e um Cryptococcus gattii) e 10 de HIV- negativos (cinco Cryptococcus neoformans e cinco Cryptococcus gattii). Considerando isolados clínicos (Cryptococcus neoformans) de HIV positivos observou-se resistência (8 por cento e 8,7 por cento) e susceptibilidade dose-dependência (20 por cento e 17,4 por cento) para fluconazol e itraconazol respectivamente. Para isolados de Cryptococcus neoformans oriundos de pacientes HIV negativos, observou-se susceptibilidade dose-dependência (40 por cento) ao fluconazol. Os isolados de Cryptococcus gattii provenientes de pacientes HIV- negativos mostraram-se susceptíveis a todos os antifúngicos, exceto um isolado de Cryptococcus gattii que foi susceptível dose-dependente ao fluconazol (20 por cento). O isolado proveniente do paciente HIV- positivo demonstrou resistência ao fluconazol (CIM > 256µg/mL) e itraconazol (CIM=3µg/mL).

Thirty-seven isolates from 10 HIV-negative and 26 HIV-positive patients in Mato Grosso were evaluated. Direct examination, culturing and chemotyping of species were performed. Ketoconazole, itraconazole, voriconazole, fluconazole and amphotericin B were evaluated. Thirty-seven yeasts of Cryptococcus spp were identified, of which 26 were from HIV-positive patients (25 Cryptococcus neoformans and one Cryptococcus gattii) and 10 from HIV-negative patients (five Cryptococcus neoformans and five Cryptococcus gattii). The Cryptococcus neoformans clinical isolates from HIV-positive patients showed resistance (8 percent and 8.7 percent) and dose-dependent susceptibility (20 percent and 17.4 percent) to fluconazole and itraconazole, respectively. Among the Cryptococcus neoformans isolates from HIV-negative patients, there was dose-dependent susceptibility (40 percent) to fluconazole. Cryptococcus gattii isolates from HIV-negative patients were shown to be susceptible to all antifungal agents, except for one isolate of Cryptococcus gattii that showed dose-dependent susceptibility to fluconazole (20 percent). The Cryptococcus gattii isolate from an HIV-positive patient showed resistance to fluconazole (MIC > 256 »g/ml) and itraconazole (MIC = 3 »g/ml).

[First description of phenotypic profile and in vitro drug susceptibility of Cryptococcus spp yeast isolated from HIV-positive and HIV-negative patients in State of Mato Grosso]. / Primeira descrição da caracterização fenotípica e susceptibilidade in vitro a drogas de leveduras do gênero Cryptococcus spp isoladas de pacientes HIV positivos e negativos, Estado de Mato Grosso.

Thirty-seven isolates from 10 HIV-negative and 26 HIV-positive patients in Mato Grosso were evaluated. Direct examination, culturing and chemotyping of species were performed. Ketoconazole, itraconazole, voriconazole, fluconazole and amphotericin B were evaluated. Thirty-seven yeasts of Cryptococcus spp were identified, of which 26 were from HIV-positive patients (25 Cryptococcus neoformans and one Cryptococcus gattii) and 10 from HIV-negative patients (five Cryptococcus neoformans and five Cryptococcus gattii). The Cryptococcus neoformans clinical isolates from HIV-positive patients showed resistance (8% and 8.7%) and dose-dependent susceptibility (20% and 17.4%) to fluconazole and itraconazole, respectively. Among the Cryptococcus neoformans isolates from HIV-negative patients, there was dose-dependent susceptibility (40%) to fluconazole. Cryptococcus gattii isolates from HIV-negative patients were shown to be susceptible to all antifungal agents, except for one isolate of Cryptococcus gattii that showed dose-dependent susceptibility to fluconazole (20%). The Cryptococcus gattii isolate from an HIV-positive patient showed resistance to fluconazole (MIC > or = 256 (1/4)g/ml) and itraconazole (MIC = 3 (1/4)microg/ml).