Project description:Vancouver Island, Canada, reports the world's highest incidence of Cryptococcus gattii infection among humans and animals. To identify key biophysical factors modulating environmental concentrations, we evaluated monthly concentrations of C. gatti in air, soil, and trees over a 3-year period. The 2 study datasets were repeatedly measured plots and newly sampled plots. We used hierarchical generalized linear and mixed effect models to determine associations. Climate systematically influenced C. gattii concentrations in all environmental media tested; in soil and on trees, concentrations decreased when temperatures were warmer. Wind may be a key process that transferred C. gattii from soil into air and onto trees. C. gattii results for tree and air samples were more likely to be positive during periods of higher solar radiation. These results improve the understanding of the places and periods with the greatest C. gattii colonization. Refined risk projections may help susceptible persons avoid activities that disturb the topsoil during relatively cool summer days.
Project description:Cryptococcus gattii causes life-threatening infection of the pulmonary and central nervous systems in hosts with normal immunity and traditionally has been considered to be restricted geographically to tropical and subtropical climates. The recent outbreak of C. gattii in the temperate climate of Vancouver Island, BC, Canada, led to a collaborative investigation. The objectives of the current study were to ascertain the environmental source of the outbreak infections, survey the molecular types of the outbreak and environmental cryptococcal isolates, and determine the extent of genetic diversity among the isolates. PCR-fingerprinting and amplified fragment length polymorphism (AFLP) were used to examine the genotypes, and mating assays were performed to determine the mating type of the isolates. All outbreak and environmental isolates belonged to C. gattii. Concordant results were obtained by using PCR-fingerprinting and AFLP analysis. The vast majority of clinical and veterinary infections were caused by isolates of the molecular type VGII/AFLP6, but two were caused by molecular type VGI/AFLP4. All environmental isolates belonged to molecular type VGII/AFLP6. Two or three subtypes were observed within VGII/AFLP6 among outbreak and environmental isolates. All mating-competent isolates were of the alpha-mating type. The emergence of this usually tropical pathogen on Vancouver Island highlights the changing distribution of this genotype and emphasizes the importance of an ongoing collaborative effort to monitor the global epidemiology of this yeast.
Project description:BackgroundThe disturbance of colonized trees and soil, such as through forestry activities, has been proposed to disperse soil- and tree-inhabiting fungal pathogens. Cryptococcus gattii sensu lato is one such pathogen that was detected on Vancouver Island, British Columbia, Canada, beginning in 1999 and caused human and animal illness.ObjectivesOur aim was to determine if C. gattii s.l. human case incidence on Vancouver Island was correlated with the intensity of landscape-level tree harvesting occurring near human settlement areas.MethodsWe created buffers around human settlement areas with radii increments of 2.5km, from 2.5 to 20km, and summed the area of annual tree harvests occurring within each buffer zone. We then performed Spearman rank-order correlation to measure the association between case incidence and annual tree harvest intensity at each radius from 1998 through 2014.ResultsThe incidence of C. gattii was positively correlated with tree harvesting intensity only at distances of 7.5km (r=0.66, p=0.004) and 10km (r=0.64, p=0.005) from human settlement areas. As annual tree harvesting area increased between 1999 and 2003, so did annual C. gattii incidence in humans, before both plateaued around 2002 and decreased after 2007.DiscussionOur findings suggest that tree harvesting plays a role in the spread of C. gattii on Vancouver Island. This may be due to tree cutting or soil disturbance facilitating the aerosolization of spores to increase infection risk. This research also illustrates the contribution that geographic information systems can make to public health research on environmental disturbance and disease outbreaks. https://doi.org/10.1289/EHP12396.
Project description:Cryptococcus neoformans is a human fungal pathogen that exists as three distinct varieties or sibling species: the predominantly opportunistic pathogens C. neoformans var. neoformans (serotype D) and C. neoformans var. grubii (serotype A) and the primary pathogen C. neoformans var. gattii (serotypes B and C). While serotypes A and D are cosmopolitan, serotypes B and C are typically restricted to tropical regions. However, serotype B isolates of C. neoformans var. gattii have recently caused an outbreak on Vancouver Island in Canada, highlighting the threat of this fungus and its capacity to infect immunocompetent individuals. Here we report a large-scale analysis of the mating abilities of serotype B and C isolates from diverse sources and identify unusual strains that mate robustly and are suitable for further genetic analysis. Unlike most isolates, which are of both the a and alpha mating types but are predominantly sterile, the majority of the Vancouver outbreak strains are exclusively of the alpha mating type and the majority are fertile. In an effort to enhance mating of these isolates, we identified and disrupted the CRG1 gene encoding the GTPase-activating protein involved in attenuating pheromone response. crg1 mutations dramatically increased mating efficiency and enabled mating with otherwise sterile isolates. Our studies provide a genetic and molecular foundation for further studies of this primary pathogen and reveal that the Vancouver Island outbreak may be attributable to a recent recombination event.
Project description:Human and animal cryptococcosis due to an unusual molecular type of Cryptococcus gattii (VGII) emerged recently on Vancouver Island, Canada. Unlike C. neoformans, C. gattii causes disease mainly in immunocompetent hosts, despite producing a similar suite of virulence determinants. To investigate a potential relationship between the regulation of expression of a virulence gene composite and virulence, we took advantage of two subtypes of VGII (a and b), one highly virulent (R265) and one less virulent (R272), that were identified from the Vancouver outbreak. By expression microarray analysis, 202 genes showed at least a 2-fold difference in expression with 108 being up- and 94 being down-regulated in strain R265 compared with strain R272. Specifically, expression levels of genes encoding putative virulence factors (e.g. LAC1, LAC2, CAS3 and MPK1) and genes encoding proteins involved in cell wall assembly, carbohydrate and lipid metabolism were increased in strain R265, whereas genes involved in the regulation of mitosis and ergosterol biosynthesis were suppressed. In vitro phenotypic studies and transcription analysis confirmed the microarray results. Gene disruption of LAC1 and MPK1 revealed defects in melanin synthesis and cell wall integrity, respectively, where CAS3 was not essential for capsule production. Moreover, MPK1 also controls melanin and capsule production and causes a severe attenuation of the virulence in a murine inhalational model. Overall, this study provides the basis for further genetic studies to characterize the differences in the virulence composite of strains with minor evolutionary divergences in gene expression in the primary pathogen C. gattii, that have led to a major invasive fungal infection outbreak.
Project description:Cryptococcus gattii was isolated from a 1.5-year-old dog with systemic cryptococcosis in Oregon. The dog had no link to Vancouver Island or British Columbia, Canada. Samples from a nasal swab and from a granulomatous mass within the cranial cavity were pooled for culture. Colonies on Sabouraud dextrose agar were mucoid and exhibited bimorphic morphology, melanin-pigmented and unpigmented. Pigmented colonies were encapsulated budding spherical yeast, whereas unpigmented colonies were of unencapsulated ovoid budding yeast. In addition to defective melanin production, the unpigmented colony type exhibited defective mating. Genetic analysis by high-resolution multilocus sequence typing revealed that the 2 isolates are genetically identical at 8 unlinked loci tested and that the 2 isolates are both the VGIIa Vancouver Island major genotype. Findings are consistent with expansion of the Vancouver Island outbreak onto the mainland Pacific Northwest region of the United States.
Project description:Comparison of transcriptional profiles of a high (a VGIIa clinical strain, R265) and a low (a VGIIb clinical strain, R272) virulence strain from the Vancouver Island outbreak.
Project description:Cryptococcus neoformans frequently causes fungal meningitis in immunocompromised patients, whereas the related species C. gattii is restricted to tropical and subtropical regions,where it usually infects immunocompetent individuals.An outbreak of C. gattii infection that began in 1999 on Vancouver Island has resulted in endemic C. gattii infection and caused numerous human and veterinary infections; the outbreak's range has spread to mainland British Columbia. The outbreak-related isolates have been molecular type VGIIa, the major genotype, or VGIIb, the minor genotype. Since 2006, human and veterinary cases of C. gattii infection have emerged in Washington and Oregon. Multilocus sequence typing demonstrates the spread of C. gattii VGIIa and VGIIb from Vancouver Island to the Pacific Northwest. Clinical strains recovered in Oregon represent a unique VGIIc genotype.
Project description:Cryptococcus gattii, emergent on Vancouver Island, British Columbia (BC), Canada, in 1999, was detected during 2003-2005 in 3 persons and 8 animals that did not travel to Vancouver Island during the incubation period; positive environmental samples were detected in areas outside Vancouver Island. All clinical and environmental isolates found in BC were genotypically consistent with Vancouver Island strains. In addition, local acquisition was detected in 3 cats in Washington and 2 persons in Oregon. The molecular profiles of Oregon isolates differed from those found in BC and Washington. Although some microclimates of the Pacific Northwest are similar to those on Vancouver Island, C. gattii concentrations in off-island environments were typically lower, and human cases without Vancouver Island contact have not continued to occur. This suggests that C. gattii may not be permanently colonized in off-island locations.