Population Genetic Analysis Reveals a High Genetic Diversity in the Brazilian Cryptococcus gattii VGII Population and Shifts the Global Origin from the Amazon Rainforest to the Semi-arid Desert in the Northeast of Brazil.

Cryptococcus neoformans and Cryptococcus gattii are responsible globally for almost one million cryptococcosis cases yearly, mostly in immunocompromised patients, such as those living with HIV. Infections due to C. gattii have mainly been described in tropical and subtropical regions, but its adaptation to temperate regions was crucial in the species evolution and highlighted the importance of this pathogenic yeast in the context of disease. Cryptococcus gattii molecular type VGII has come to the forefront in connection with an on-going emergence in the Pacific North West of North America. Taking into account that previous work pointed towards South America as an origin of this species, the present work aimed to assess the genetic diversity within the Brazilian C. gattii VGII population in order to gain new insights into its origin and global dispersal from the South American continent using the ISHAM consensus MLST typing scheme. Our results corroborate the finding that the Brazilian C. gattii VGII population is highly diverse. The diversity is likely due to recombination generated from sexual reproduction, as evidenced by the presence of both mating types in clinical and environmental samples. The data presented herein strongly supports the emergence of highly virulent strains from ancestors in the Northern regions of Brazil, Amazonia and the Northeast. Numerous genotypes represent a link between Brazil and other parts of the world reinforcing South America as the most likely origin of the C. gattii VGII subtypes and their subsequent global spread, including their dispersal into North America, where they caused a major emergence.

Molecular identification of Coccidioides spp. in soil samples from Brazil.

BACKGROUND: Since 1991 several outbreaks of acute coccidioidomycosis (CM) were diagnosed in the semi-arid Northeast of Brazil, mainly related to disturbance of armadillo burrows caused by hunters while digging them for the capture of these animals. This activity causes dust contaminated with arthroconidia of Coccidioides posadasii, which, once inhaled, cause the mycosis. We report on the identification of C. posadasii in soil samples related to outbreaks of CM. RESULTS: Twenty four soil samples had their DNA extracted and subsequently submitted to a semi-nested PCR technique using specific primers. While only 6 (25%) soil samples were positive for C. posadasii by mice inoculation, all (100%) were positive by the molecular tool. CONCLUSION: This methodology represents a simple, sensitive and specific molecular technique to determine the environmental distribution of Coccidioides spp. in endemic areas, but cannot distinguish the species. Moreover, it may be useful to identify culture isolates. Key-words: 1. Coccidioidomycosis. 2. Coccidioides spp. 3. C. posadasii. 4. Semi-arid. 5. Semi-nested PCR.

Viability and molecular authentication of Coccidioides immitis strains from culture collection of the Instituto Oswaldo Cruz, Rio de Janeiro, Brazil.

Twenty Coccidioides immitis strains were evaluated. Only 5 of the 20 strains kept under mineral oil maintained their viability while all 5 subcultures preserved in water remained viable and none of the 13 subcultures kept in soil were viable. A 519 bp PCR product from the csa gene confirmed the identity of the strains.

Viability and molecular authentication of Coccidioides immitis strains from culture collection of the Instituto Oswaldo Cruz, Rio de Janeiro, Brazil

Vinte cepas de Coccidioides immitis foram avaliadas.Cinco das 20 cepas preservadas sob óleo mineral mantiveram-se viáveis, todas as 5 subculturas preservadas em água permaneceram viáveis e nenhuma das 13 subculturas mantidas em solo foi viável. Um produto de PCR de 519 pb do gene csa confirmou a identidade das cepas.

DNA fingerprinting pattern and susceptibility to antifungal drugs in Cryptococcus neoformans variety grubii isolates from Barcelona city and rural environmental samples.

Cryptococcus neoformans var. grubii (serotype A) was isolated from 12 soil samples mixed with pigeon droppings (16.9%) from 71 soil samples in Barcelona and rural areas of Catalonia. C. neoformans was not isolated from indoor dust and Eucalyptus debris. PCR fingerprinting was performed in 22 representative isolates and all of them corresponded to the VNI pattern. Susceptibility testing for the 22 isolates of C. neoformans var. grubii showed that all of them were susceptible to amphotericin B. Three isolates presented MICs (Minimal Inhibitory Concentrations) > or = 1 microg/ml to Itraconazole, five MICs > or = 1 microg/ml to ketoconazole and four were fluconazole resistant, (MICs > or = 64 microg/ml), while three of them were shown to have MICs > or = 1 microg/ml to voriconazole. In spite that all isolates presented the same DNA fingerprinting pattern, the susceptibility to antifungals is very variable. The possibility of acquiring cryptococcosis infection with primarily resistant environment strains is feasible.

Serotyping of 467 Cryptococcus neoformans isolates from clinical and environmental sources in Brazil: analysis of host and regional patterns.

Cryptococcus neoformans is an important zoopathogen, and it is one of the most prevalent lethal mycotic agents. Its polysaccharide capsule, synthesized in vivo and in vitro, is a virulence factor, contains predominantly glucuronoxylomannan, and is responsible for the antigenic differentiation of serotypes A, B, C, D, and AD. A total of 467 isolates of C. neoformans obtained from clinical and environmental sources from Brazilian regions were studied serologically by using the Crypto Check Iatron RM 304-K kit. Serotyping of the clinical isolates showed the following prevalences of the serotypes: A (77.95%), followed by B (18.2%), AD (1.3%), D (0.4%), C (0.2%), and untypeable (1.93%). The epidemiology of serotype A in the Brazilian southern and southeastern regions reproduces the picture observed worldwide. In contrast, serotype B was the most frequent agent of cryptococcosis in the northeastern region, occurring nearly equally in male and female healthy hosts. Among the isolates from environmental sources, serotypes A and B were found to occur in the hollows of tropical trees of the genera Cassia, Ficus, and MOQUILLEA: The few isolates from Eucalyptus camaldulensis debris were serotypes A and B and untypeable. Overall, no association with a specific host tree was identified for these serotypes, denoting a distinct ecoepidemiological regional pattern. The one serotype C isolate was recovered from a human immunodeficiency virus-negative host. Serotype AD predominated over serotype D among both clinical and environmental isolates.