Sudha Chaturvedi

Cryptococcus gattii in AIDS patients, southern California.

A molecular analysis of pheromone genes showed a notable prevalence of Cryptococcus gattii isolates from AIDS patients in southern California.

Morphological and molecular characterizations of psychrophilic fungus Geomyces destructans from New York bats with white nose syndrome (WNS).

Background Massive die-offs of little brown bats (Myotis lucifugus) have been occurring since 2006 in hibernation sites around Albany, New York, and this problem has spread to other States in the Northeastern United States. White cottony fungal growth is seen on the snouts of affected animals, a prominent sign of White Nose Syndrome (WNS). A previous report described the involvement of the fungus Geomyces destructans in WNS, but an identical fungus was recently isolated in France from a bat that was evidently healthy. The fungus has been recovered sparsely despite plentiful availability of afflicted animals. Methodology/Principal Findings We have investigated 100 bat and environmental samples from eight affected sites in 2008. Our findings provide strong evidence for an etiologic role of G. destructans in bat WNS. (i) Direct smears from bat snouts, Periodic Acid Schiff-stained tissue sections from infected tissues, and scanning electron micrographs of bat tissues all showed fungal structures similar to those of G. destructans. (ii) G. destructans DNA was directly amplified from infected bat tissues, (iii) Isolations of G. destructans in cultures from infected bat tissues showed 100% DNA match with the fungus present in positive tissue samples. (iv) RAPD patterns for all G. destructans cultures isolated from two sites were indistinguishable. (v) The fungal isolates showed psychrophilic growth. (vi) We identified in vitro proteolytic activities suggestive of known fungal pathogenic traits in G. destructans. Conclusions/Significance Further studies are needed to understand whether G. destructans WNS is a symptom or a trigger for bat mass mortality. The availability of well-characterized G. destructans strains should promote an understanding of bat–fungus relationships, and should aid in the screening of biological and chemical control agents.

Cryptococcus gattii: a resurgent fungal pathogen.

Cryptococcus gattii and Cryptococcus neoformans are causal agents of cryptococcosis, which manifests as pneumonia and meningitis. C. gattii has recently received widespread attention owing to outbreaks in British Columbia, Canada and the US Pacific Northwest. The biology of this tree-dwelling yeast is relatively unexplored, and there are few clues about how it causes infections in humans and animals. In this review, we summarize recent discoveries about C. gattii genetics and its ecological niche and highlight areas ripe for future exploration. Increased focus on epidemiology, ecological modeling and host-pathogen interactions is expected to yield a better understanding of this enigmatic yeast, and ultimately lead to better measures for its control.

Extracellular Fibrils of Pathogenic Yeast Cryptococcus gattii Are Important for Ecological Niche, Murine Virulence and Human Neutrophil Interactions

Cryptococcus gattii, an emerging fungal pathogen of humans and animals, is found on a variety of trees in tropical and temperate regions. The ecological niche and virulence of this yeast remain poorly defined. We used Arabidopsis thaliana plants and plant-derived substrates to model C. gattii in its natural habitat. Yeast cells readily colonized scratch-wounded plant leaves and formed distinctive extracellular fibrils (40–100 nm diameter ×500–3000 nm length). Extracellular fibrils were observed on live plants and plant-derived substrates by scanning electron microscopy (SEM) and by high voltage- EM (HVEM). Only encapsulated yeast cells formed extracellular fibrils as a capsule-deficient C. gattii mutant completely lacked fibrils. Cells deficient in environmental sensing only formed disorganized extracellular fibrils as apparent from experiments with a C. gattii STE12α mutant. C. gattii cells with extracellular fibrils were more virulent in murine model of pulmonary and systemic cryptococcosis than cells lacking fibrils. C. gattii cells with extracellular fibrils were also significantly more resistant to killing by human polymorphonuclear neutrophils (PMN) in vitro even though these PMN produced elaborate neutrophil extracellular traps (NETs). These observations suggest that extracellular fibril formation could be a structural adaptation of C. gattii for cell-to-cell, cell-to-substrate and/or cell-to- phagocyte communications. Such ecological adaptation of C. gattii could play roles in enhanced virulence in mammalian hosts at least initially via inhibition of host PMN– mediated killing.

Molecular characterization, biofilm analysis and experimental biofouling study of Fusarium isolates from recent cases of fungal keratitis in New York State

BackgroundTo characterize Fusarium isolates from recent cases of fungal keratitis in contact lens wearers, and to investigate fungal association with MoistureLoc solution.MethodsWe studied six fungal isolates from recent cases of keratitis in New York State. The isolates were characterized by nucleotide sequencing and phylogenetic analyses of multiple genes, and then typed using minisatellite and microsatellite probes. Experimental fungal biofilm formation was tested by standard methods. MoistureLoc solutions were tested in biofouling studies for their efficacy in elimination of Fusarium contamination.ResultsFusarium solani – corneal ulcers (2 isolates), lens case (1 isolate), and F. oxysporum – corneal ulcer (1 isolate), eye (1 isolate), were recovered from five patients. An opened bottle of MoistureLoc solution provided by a patient also yielded F. solani. Two distinct genotypes of F. solani as well as of F. oxysporum were present in the isolated strains. Remarkably, F. solani strains from the lens case and lens solution in one instance were similar, based on phylogenetic analyses and molecular typing. The solution isolate of F. solani formed biofilm on contact lenses in control conditions, but not when co-incubated with MoistureLoc solution. Both freshly opened and 3-month old MoistureLoc solutions effectively killed F. solani and F. oxysporum, when fungal contamination was simulated under recommended lens treatment regimen (4-hr). However, simulation of inappropriate use (15 – 60 min) led to the recovery of less than 1% of original inoculum of F. solani or F. oxysporum.ConclusionTemporary survival of F. solani and F. oxysporum in MoistureLoc suggested that improper lens cleaning regimen could be a possible contributing factor in recent infections.

Transcription Factor STE12α Has Distinct Roles in Morphogenesis, Virulence, and Ecological Fitness of the Primary Pathogenic Yeast Cryptococcus gattii

ABSTRACT Cryptococcus gattii is a primary pathogenic yeast, increasingly important in public health, but factors responsible for its host predilection and geographical distribution remain largely unknown. We have characterized C. gattii STE12α to probe its role in biology and pathogenesis because this transcription factor has been linked to virulence in many human and plant pathogenic fungi. A full-length STE12α gene was cloned by colony hybridization and sequenced using primer walk and 3′ rapid amplification of cDNA ends strategies, and a ste12αΔ gene knockout mutant was created by URA5 insertion at the homologous site. A semiquantitative analysis revealed delayed and poor mating in ste12αΔ mutant; this defect was not reversed by exogenous cyclic AMP. C. gattii parent and mutant strains showed robust haploid fruiting. Among putative virulence factors tested, the laccase transcript and enzymatic activity were down regulated in the ste12αΔ mutant, with diminished production of melanin. However, capsule, superoxide dismutase, phospholipase, and urease were unaffected. Similarly, Ste12 deficiency did not cause any auxotrophy, assimilation defects, or sensitivity to a large panel of chemicals and antifungals. The ste12αΔ mutant was markedly attenuated in virulence in both BALB/c and A/Jcr mice models of meningoencephalitis, and it also exhibited significant in vivo growth reduction and was highly susceptible to in vitro killing by human neutrophils (polymorphonuclear leukocytes). In tests designed to simulate the C. gattii natural habitat, the ste12αΔ mutant was poorly pigmented on wood agar prepared from two tree species and showed poor survival and multiplication in wood blocks. Thus, STE12α plays distinct roles in C. gattii morphogenesis, virulence, and ecological fitness.

Rhizomucor variabilis var. regularior and Hormographiella aspergillata Infections in a Leukemic Bone Marrow Transplant Recipient with Refractory Neutropenia

ABSTRACT Rhizomucor variabilis and Hormographiella aspergillata rarely cause human infections. This report details a fatal case of a 14-year-old female with leukemia posthematopoietic cell transplant and relapse with refractory pancytopenia. The patient first developed anR. variabilis var. regularior palate infection and later developed a cutaneous H. aspergillata infection while on posaconazole and caspofungin therapy.

Clonal genotype of Geomyces destructans among bats with White Nose Syndrome, New York, USA.

The dispersal mechanism of Geomyces destructans, which causes geomycosis (white nose syndrome) in hibernating bats, remains unknown. Multiple gene genealogic analyses were conducted on 16 fungal isolates from diverse sites in New York State during 2008–2010. The results are consistent with the clonal dispersal of a single G. destructans genotype.

Real-Time PCR Assay for Identification of Blastomyces dermatitidis in Culture and in Tissue

ABSTRACT The TaqMan real-time PCR assay was developed from the Blastomyces dermatitidis BAD1 gene promoter. The assay identified all haplotypes of B. dermatitidis and five of six positive paraffin-embedded tissues. The assay sensitivity threshold was 1 pg genomic DNA of the mold form and 2 CFU of the yeast form of B. dermatitidis. No cross-reactivity was observed against other fungal DNA. The assay allowed rapid (5-h) identification of B. dermatitidis from culture and from clinical specimens.

Importance of Resolving Fungal Nomenclature: the Case of Multiple Pathogenic Species in the Cryptococcus Genus

Cryptococcosis is a major fungal disease caused by members of the Cryptococcus gattii and Cryptococcus neoformans species complexes. After more than 15 years of molecular genetic and phenotypic studies and much debate, a proposal for a taxonomic revision was made. ABSTRACT Cryptococcosis is a major fungal disease caused by members of the Cryptococcus gattii and Cryptococcus neoformans species complexes. After more than 15 years of molecular genetic and phenotypic studies and much debate, a proposal for a taxonomic revision was made. The two varieties within C. neoformans were raised to species level, and the same was done for five genotypes within C. gattii. In a recent perspective (K. J. Kwon-Chung et al., mSphere 2:e00357-16, 2017, https://doi.org/10.1128/mSphere.00357-16 ), it was argued that this taxonomic proposal was premature and without consensus in the community. Although the authors of the perspective recognized the existence of genetic diversity, they preferred the use of the informal nomenclature “C. neoformans species complex” and “C. gattii species complex.” Here we highlight the advantage of recognizing these seven species, as ignoring these species will impede deciphering further biologically and clinically relevant differences between them, which may in turn delay future clinical advances.