Biology subjects

Glazier, V. E.

Publications and source records attributed to Glazier, V. E..

3 recordsLinked to original sources

bioRxiv2026

A Novel Mycovirus Identified in Clinical Isolates of Cryptococcus neoformans

Mycoviruses are widespread throughout the fungal kingdom and are known to infect diverse fungal taxa including fungal species that are important plant and human pathogens. Although many mycoviruses have been found to have minimal effects on their host, several viruses have been found to modulate fungal physiology, and as a result impact fungal virulence. Screens for mycoviruses in clinically relevant fungi have identified numerous mycoviruses within several important human pathogens, however mycoviruses remain uncharacterized in the clinically relevant human pathogen Cryptococcus neoformans. C. neoformans is an opportunistic encapsulated yeast responsible for life-threatening cryptococcal meningitis, a leading cause of mortality among immunocompromised individuals, particularly those with HIV/AIDS. We performed a search for viral RNA-dependent RNA Polymerase (RdRP) signatures in publicly available C. neoformans transcriptomic data. This search identified Totiviridae viral genomes within six clinical isolates of C. neoformans from Botswana. All six isolates originated from the CSF of HIV positive individuals with cryptococcal meningitis. Reverse transcription PCR (RT-PCR) independently validated the continued presence of the virus in three of these clinical isolates. Subsequent analysis of the viral genome identified two genotypes of a single species of Totivirus. This new species possesses canonical features of the Totiviridae family, including a slippery heptamer and a predicted RNA pseudoknot structure involved in programmed -1 ribosomal frameshifting for RdRP expression. Taken together, these results provide evidence of a mycovirus capable of infecting C. neoformans.

microbiology↗

bioRxiv2023

The Candida albicans reference strain SC5314 contains a rare, dominant allele of the transcription factor Rob1 that modulates biofilm formation and oral commensalism

Candida albicans is a diploid human fungal pathogen that displays significant genomic and phenotypic heterogeneity over a range of virulence traits and in the context of a variety of environmental niches. Here, we show that the effects of Rob1 on biofilm and filamentation virulence traits is dependent on both the specific environmental condition and the clinical strain of C. albicans. The C. albicans reference strain SC5314 is a ROB1 heterozygote with two alleles that differ by a single nucleotide polymorphism at position 946 resulting in a serine or proline containing isoform. An analysis of 224 sequenced C. albicans genomes indicates that SC5314 is the only ROB1 heterozygote documented to date and that the dominant allele contains a proline at position 946. Remarkably, the ROB1 alleles are functionally distinct and the rare ROB1946Sallele supports increased filamentation in vitro and increased biofilm formation in vitro and in vivo, suggesting it is a phenotypic gain-of-function allele. SC5314 is amongst the most highly filamentous and invasive strains characterized to date. Introduction of the ROB1946Sallele into a poorly filamenting clinical isolate increases filamentation and conversion of an SC5314 laboratory strain to a ROB1946S homozygote increases in vitro filamentation and biofilm formation. In a mouse model of oropharyngeal infection, the predominant ROB1946P allele establishes a commensal state while the ROB1946S phenocopies the parent strain and invades into the mucosae. These observations provide an explanation for the distinct phenotypes of SC5314 and highlight the role of heterozygosity as a driver of C. albicans phenotypic heterogeneity. ImportanceCandida albicans is a commensal fungus that colonizes human oral cavity and gastrointestinal tracts but also causes mucosal as well as invasive disease. The expression of virulence traits in C. albicans clinical isolates is heterogenous and the genetic basis of this heterogeneity is of high interest. The C. albicans reference strain SC5314 is highly invasive and expresses robust filamentation and biofilm formation relative to many other clinical isolates. Here, we show that SC5314 derivatives are heterozygous for the transcription factor Rob1 and contain an allele with a rare gain-of-function SNP that drives filamentation, biofilm formation, and virulence in a model of oropharyngeal candidiasis. These finding explain, in part, the outlier phenotype of the reference strain and highlight the role of heterozygosity plays in the strain-to-strain variation of diploid fungal pathogens.

microbiology↗

bioRxiv2020

Puf4 Mediates Post-transcriptional Regulation of Caspofungin Resistance in Cryptococcus neoformans

Echinocandins have been on the market for 20 years, yet they are the newest class of antifungal drugs. The human fungal pathogen Cryptococcus neoformans is intrinsically resistant to the echinocandin antifungal drug caspofungin, which targets the {beta}-1,3-glucan synthase encoded by the FKS1. Analysis of a C. neoformans puf4{Delta} mutant, lacking the pumilio/FBF RNA binding protein family member Puf4, revealed exacerbated caspofungin resistance. In contrast, overexpression of PUF4 resulted in caspofungin sensitivity. The FKS1 mRNA contains three Puf4-binding elements (PBEs) in its 5 untranslated region. Puf4 binds with specificity to this region of the FKS1. The FKS1 mRNA was destabilized in the puf4{Delta} mutant, and the abundance of the FKS1 mRNA was reduced compared to wild type, suggesting that Puf4 is a positive regulator FKS1 mRNA stability. In addition to FKS1, the abundance of additional cell wall biosynthesis genes, including chitin synthases (CHS3, CHS4, CHS6) and deacetylases (CDA1, CDA2, CDA3) as well as a {beta}-1,6-glucan synthase gene (SKN1) was regulated by Puf4 during a caspofungin time course. The use of fluorescent dyes to quantify cell wall components revealed that the puf4{Delta} mutant had increased chitin content, suggesting a cell wall composition that is less reliant on {beta}-1,3-glucan. Overall, our findings suggest a mechanism by which caspofungin resistance, and more broadly, cell wall biogenesis, is regulated post-transcriptionally by Puf4. ImportanceCryptococcus neoformans is an environmental fungus that causes pulmonary and central nervous system infections. It is also responsible for 15% of AIDS-related deaths. A major contributor to the high morbidity and mortality statistics is the lack of safe and effective antifungal therapies, especially in resource-poor settings. Yet, antifungal drug development has stalled in the pharmaceutical industry. Therefore, it is of importance to understand the mechanism by which C. neoformans is resistant to caspofungin in order to design adjunctive therapies to potentiate its activity toward this important pathogen.

microbiology↗