Publications by authors named "Christina A Cuomo"

135 Publications

Phenotypic Characterization and Comparative Genomics of the Melanin-Producing Yeast Reveals a Distinct Stress Tolerance Profile and Reduced Ribosomal Genetic Content.

J Fungi (Basel) 2021 Dec 15;7(12). Epub 2021 Dec 15.

Center for Biomolecular Sciences and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.

The black yeast of the order Chaetothyriales is notable for its ability to produce abundant quantities of DHN-melanin. While many other species are frequent causal agents of human infection, CBS 102400 lacks the thermotolerance requirements that enable pathogenicity, making it appealing for use in targeted functional studies and biotechnological applications. Here, we report the stress tolerance characteristics of , with an emphasis on the influence of melanin on its resistance to various forms of stress. We find that has a distinct stress tolerance profile that includes variation in resistance to temperature, osmotic, and oxidative stress relative to the extremophilic and pathogenic black yeast . Notably, the presence of melanin substantially impacts stress resistance in , while this was not found to be the case in . The cellular context, therefore, influences the role of melanin in stress protection. In addition, we present a detailed analysis of the genome, revealing key differences in functional genetic content relative to other ascomycetous species, including a significant decrease in abundance of genes encoding ribosomal proteins. In all, this study provides insight into how genetics and physiology may underlie stress tolerance and enhances understanding of the genetic diversity of black yeasts.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/jof7121078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8709033PMC
December 2021

In vivo emergence of high-level resistance during treatment reveals the first identified mechanism of amphotericin B resistance in Candida auris.

Clin Microbiol Infect 2021 Dec 13. Epub 2021 Dec 13.

Department of Pharmacy and Pharmaceutical Sciences, St. Jude Children's Research Hospital, Memphis, Tennessee, USA. Electronic address:

Objective: Candida auris has emerged as a healthcare-associated and multidrug-resistant fungal pathogen of great clinical concern. While as many as 50% of C. auris clinical isolates are reported to be resistant to amphotericin B, to date, no mechanisms contributing to this resistance have been identified. Here we describe the clinical case in which high-level amphotericin B resistance was acquired in vivo during therapy and undertake molecular and genetic studies to identify and characterize the genetic determinant of resistance.

Methods: Whole genome sequencing was performed on four C. auris isolates obtained from a single patient case. Cas9-mediated genetic manipulations were then used to generate mutant strains harboring mutations of interest, and these strains were subsequently subjected to amphotericin B susceptibility testing, and comprehensive sterol profiling.

Results: A novel mutation in C. auris sterol-methyltransferase gene, ERG6, was found to be associated with amphotericin B resistance, and this mutation alone conferred a >32-fold increase in amphotericin B resistance. Comprehensive sterol profiling revealed an abrogation of ergosterol biosynthesis and a corresponding accumulation of cholesta-type sterols in isolates and strains harboring the clinically-derived ERG6 mutation.

Conclusions: Together these findings definitively demonstrate mutations in C. auris ERG6 as the first identified mechanism of clinical amphotericin B resistance in C. auris and represent a significant step forward in the understanding of antifungal resistance in this emerging public health threat.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cmi.2021.11.024DOI Listing
December 2021

Diversity, multifaceted evolution, and facultative saprotrophism in the European Batrachochytrium salamandrivorans epidemic.

Nat Commun 2021 11 18;12(1):6688. Epub 2021 Nov 18.

Wildlife Health Ghent, Department of Pathology, Bacteriology and Avian Diseases, Faculty of Veterinary Medicine, Ghent University, 9820, Merelbeke, Belgium.

While emerging fungi threaten global biodiversity, the paucity of fungal genome assemblies impedes thoroughly characterizing epidemics and developing effective mitigation strategies. Here, we generate de novo genomic assemblies for six outbreaks of the emerging pathogen Batrachochytrium salamandrivorans (Bsal). We reveal the European epidemic currently damaging amphibian populations to comprise multiple, highly divergent lineages demonstrating isolate-specific adaptations and metabolic capacities. In particular, we show extensive gene family expansions and acquisitions, through a variety of evolutionary mechanisms, and an isolate-specific saprotrophic lifecycle. This finding both explains the chytrid's ability to divorce transmission from host density, producing Bsal's enigmatic host population declines, and is a key consideration in developing successful mitigation measures.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-021-27005-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8602665PMC
November 2021

Gene Expression of Diverse Cryptococcus Isolates during Infection of the Human Central Nervous System.

mBio 2021 12 2;12(6):e0231321. Epub 2021 Nov 2.

Division of Infectious Diseases, Department of Medicine, Duke University School of Medicine, Durham, North Carolina, USA.

Cryptococcus neoformans is a major human central nervous system (CNS) fungal pathogen causing considerable morbidity and mortality. In this study, we provide the widest view to date of the yeast transcriptome directly from the human subarachnoid space and within cerebrospinal fluid (CSF). We captured yeast transcriptomes from C. neoformans of various genotypes in 31 patients with cryptococcal meningoencephalitis as well as several Cryptococcus gattii infections. Using transcriptome sequencing (RNA-seq) analyses, we compared the yeast transcriptomes to those from other environmental conditions, including growth on nutritious media or artificial CSF as well as samples collected from rabbit CSF at two time points. We ranked gene expressions and identified genetic patterns and networks across these diverse isolates that reveal an emphasis on carbon metabolism, fatty acid synthesis, transport, cell wall structure, and stress-related gene functions during growth in CSF. The most highly expressed yeast genes in human CSF included those known to be associated with survival or virulence and highlighted several genes encoding hypothetical proteins. From that group, a gene encoding the putative glycoprotein (CNAG_06000) was selected for functional studies. This gene was found to impact the virulence of Cryptococcus in both mice and the CNS rabbit model, in agreement with a recent study also showing a role in virulence. This transcriptional analysis strategy provides a view of regulated yeast genes across genetic backgrounds important for human CNS infection and a relevant resource for the study of cryptococcal genes, pathways, and networks linked to human disease. Cryptococcus is the most common fungus causing high-morbidity and -mortality human meningitis. This encapsulated yeast has a unique propensity to travel to the central nervous system to produce disease. In this study, we captured transcriptomes of yeasts directly out of the human cerebrospinal fluid, the most concerning site of infection. By comparing the RNA transcript levels with other conditions, we gained insights into how the basic machinery involved in metabolism and environmental responses enable this fungus to cause disease at this body site. This approach was applied to clinical isolates with diverse genotypes to begin to establish a genotype-agnostic understanding of how the yeast responds to stress. Based on these results, future studies can focus on how these genes and their pathways and networks can be targeted with new therapeutics and possibly classify yeasts with bad infection outcomes.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.02313-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8561399PMC
December 2021

Diagnostic Allele-Specific PCR for the Identification of Clades.

J Fungi (Basel) 2021 Sep 13;7(9). Epub 2021 Sep 13.

Laboratory of Molecular Cell Biology, Department of Biology, Institute of Botany and Microbiology, KU Leuven, 3001 Leuven, Belgium.

is an opportunistic pathogenic yeast that emerged worldwide during the past decade. This fungal pathogen poses a significant public health threat due to common multidrug resistance (MDR), alarming hospital outbreaks, and frequent misidentification. Genomic analyses have identified five distinct clades that are linked to five geographic areas of origin and characterized by differences in several phenotypic traits such as virulence and drug resistance. Typing of strains and the identification of clades can be a powerful tool in molecular epidemiology and might be of clinical importance by estimating outbreak and MDR potential. As has caused global outbreaks, including in low-income countries, typing strains quickly and inexpensively is highly valuable. We report five allele-specific polymerase chain reaction (AS-PCR) assays for the identification of and each of the five described clades of based on conserved mutations in the internal transcribed spacer (ITS) rDNA region and a clade-specific gene cluster. This PCR method provides a fast, cheap, sequencing-free diagnostic tool for the identification of , clades, and potentially, the discovery of new clades.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3390/jof7090754DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8471779PMC
September 2021

Whole-genome sequencing of multiple isolates of Puccinia triticina reveals asexual lineages evolving by recurrent mutations.

G3 (Bethesda) 2021 09;11(9)

USDA-ARS, Cereal Disease Laboratory, St. Paul, MN 55108, USA.

The wheat leaf rust fungus, Puccinia triticina Erikss., is a worldwide pathogen of tetraploid durum and hexaploid wheat. Many races of P. triticina differ for virulence to specific leaf rust resistance genes and are found in most wheat-growing regions of the world. Wheat cultivars with effective leaf rust resistance exert selection pressure on P. triticina populations for virulent race types. The objectives of this study were to examine whole-genome sequence data of 121 P. triticina isolates and to gain insight into race evolution. The collection included isolates comprising of many different race phenotypes collected worldwide from common and durum wheat. One isolate from wild wheat relative Aegilops speltoides and two from Ae. cylindrica were also included for comparison. Based on 121,907 informative variants identified relative to the reference Race 1-1 genome, isolates were clustered into 11 major lineages with 100% bootstrap support. The isolates were also grouped based on variation in 1311 predicted secreted protein genes. In gene-coding regions, all groups had high ratios of nonsynonymous to synonymous mutations and nonsense to readthrough mutations. Grouping of isolates based on two main variation principle components for either genome-wide variation or variation just within the secreted protein genes, indicated similar groupings. Variants were distributed across the entire genome, not just within the secreted protein genes. Our results suggest that recurrent mutation and selection play a major role in differentiation within the clonal lineages.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/g3journal/jkab219DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8496273PMC
September 2021

The Relaunch of .

Microbiol Spectr 2021 09 9;9(1):e0039621. Epub 2021 Jun 9.

Infectious Disease and Microbiome Program, Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/Spectrum.00396-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8552684PMC
September 2021

Best practices on the differential expression analysis of multi-species RNA-seq.

Genome Biol 2021 04 29;22(1):121. Epub 2021 Apr 29.

Institute for Genome Sciences, University of Maryland School of Medicine, Baltimore, MD, 21201, USA.

Advances in transcriptome sequencing allow for simultaneous interrogation of differentially expressed genes from multiple species originating from a single RNA sample, termed dual or multi-species transcriptomics. Compared to single-species differential expression analysis, the design of multi-species differential expression experiments must account for the relative abundances of each organism of interest within the sample, often requiring enrichment methods and yielding differences in total read counts across samples. The analysis of multi-species transcriptomics datasets requires modifications to the alignment, quantification, and downstream analysis steps compared to the single-species analysis pipelines. We describe best practices for multi-species transcriptomics and differential gene expression.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13059-021-02337-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8082843PMC
April 2021

Amoeba Predation of Cryptococcus neoformans Results in Pleiotropic Changes to Traits Associated with Virulence.

mBio 2021 04 27;12(2). Epub 2021 Apr 27.

Department of Molecular Microbiology and Immunology, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland, USA

Amoeboid predators, such as amoebae, are proposed to select for survival traits in soil microbes such as ; these traits can also function in animal virulence by defeating phagocytic immune cells, such as macrophages. Consistent with this notion, incubation of various fungal species with amoebae enhanced their virulence, but the mechanisms involved are unknown. In this study, we exposed three strains of (1 clinical and 2 environmental) to predation by for prolonged times and then analyzed surviving colonies phenotypically and genetically. Surviving colonies comprised cells that expressed either pseudohyphal or yeast phenotypes, which demonstrated variable expression of traits associated with virulence, such as capsule size, urease production, and melanization. Phenotypic changes were associated with aneuploidy and DNA sequence mutations in some amoeba-passaged isolates, but not in others. Mutations in the gene encoding the oligopeptide transporter (CNAG_03013; ) were observed among amoeba-passaged isolates from each of the three strains. Isolates derived from environmental strains gained the capacity for enhanced macrophage toxicity after amoeba selection and carried mutations on the CNAG_00570 gene encoding Pkr1 (AMP-dependent protein kinase regulator) but manifested reduced virulence in mice because they elicited more effective fungal-clearing immune responses. Our results indicate that survival under constant amoeba predation involves the generation of strains expressing pleiotropic phenotypic and genetic changes. Given the myriad potential predators in soils, the diversity observed among amoeba-selected strains suggests a bet-hedging strategy whereby variant diversity increases the likelihood that some will survive predation. is a ubiquitous environmental fungus that is also a leading cause of fatal fungal infection in humans, especially among immunocompromised patients. A major question in the field is how an environmental yeast such as becomes a human pathogen when it has no need for an animal host in its life cycle. Previous studies showed that increases its pathogenicity after interacting with its environmental predator amoebae. Amoebae, like macrophages, are phagocytic cells that are considered an environmental training ground for pathogens to resist macrophages, but the mechanism by which changes its virulence through interactions with protozoa is unknown. Our study indicates that fungal survival in the face of amoeba predation is associated with the emergence of pleiotropic phenotypic and genomic changes that increase the chance of fungal survival, with this diversity suggesting a bet-hedging strategy to ensure that some forms survive.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.00567-21DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8092252PMC
April 2021

Editorial: Genomic Characterization of Emerging Human Fungal Pathogens.

Front Genet 2021 25;12:674765. Epub 2021 Mar 25.

National Institute for Communicable Diseases, National Health Laboratory Service, Johannesburg, South Africa.

View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fgene.2021.674765DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8027301PMC
March 2021

Genome-Wide Analysis of Experimentally Evolved Candida auris Reveals Multiple Novel Mechanisms of Multidrug Resistance.

mBio 2021 04 5;12(2). Epub 2021 Apr 5.

VIB Center for Microbiology, Leuven, Belgium

is globally recognized as an opportunistic fungal pathogen of high concern, due to its extensive multidrug resistance (MDR). Still, molecular mechanisms of MDR are largely unexplored. This is the first account of genome-wide evolution of MDR in obtained through serial exposure to azoles, polyenes, and echinocandins. We show the stepwise accumulation of copy number variations and novel mutations in genes both known and unknown in antifungal drug resistance. Echinocandin resistance was accompanied by a codon deletion in hot spot 1 and a substitution in "novel" hot spot 3. Mutations in and further increased the echinocandin MIC. Decreased azole susceptibility was linked to a mutation in transcription factor and overexpression of the drug efflux pump Cdr1, a segmental duplication of chromosome 1 containing , and a whole chromosome 5 duplication, which contains The latter was associated with increased expression of , , and but not The simultaneous emergence of nonsense mutations in and was shown to decrease amphotericin B susceptibility, accompanied with fluconazole cross-resistance. A mutation in , a gene mainly known for its role in DNA damage homeostasis, further increased the polyene MIC. Overall, this study shows the alarming potential for and diversity of MDR development in , even in a clade until now not associated with MDR (clade II), stressing its clinical importance and the urge for future research. is a recently discovered human fungal pathogen and has shown an alarming potential for developing multi- and pan-resistance toward all classes of antifungals most commonly used in the clinic. Currently, has been globally recognized as a nosocomial pathogen of high concern due to this evolutionary potential. So far, this is the first study in which the stepwise progression of multidrug resistance (MDR) in is monitored Multiple novel mutations in known resistance genes and genes previously not or vaguely associated with drug resistance reveal rapid MDR evolution in a clade II isolate. Additionally, this study shows that experimental evolution can be a powerful tool to discover new drug resistance mechanisms, although it has its limitations.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.03333-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8092288PMC
April 2021

Clade-specific chromosomal rearrangements and loss of subtelomeric adhesins in Candida auris.

Genetics 2021 05;218(1)

Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.

Candida auris is an emerging fungal pathogen of rising concern due to global spread, the ability to cause healthcare-associated outbreaks, and antifungal resistance. Genomic analyses revealed that early contemporaneously detected cases of C. auris were geographically stratified into four major clades. While Clades I, III, and IV are responsible for ongoing outbreaks of invasive and multidrug-resistant infections, Clade II, also termed the East Asian clade, consists primarily of cases of ear infection, is often susceptible to all antifungal drugs, and has not been associated with outbreaks. Here, we generate chromosome-level assemblies of twelve isolates representing the phylogenetic breadth of these four clades and the only isolate described to date from Clade V. This Clade V genome is highly syntenic with those of Clades I, III, and IV, although the sequence is highly divergent from the other clades. Clade II genomes appear highly rearranged, with translocations occurring near GC-poor regions, and large subtelomeric deletions in most chromosomes, resulting in a substantially different karyotype. Rearrangements and deletion lengths vary across Clade II isolates, including two from a single patient, supporting ongoing genome instability. Deleted subtelomeric regions are enriched in Hyr/Iff-like cell-surface proteins, novel candidate cell wall proteins, and an ALS-like adhesin. Cell wall proteins from these families and other drug-related genes show clade-specific signatures of selection in Clades I, III, and IV. Subtelomeric dynamics and the conservation of cell surface proteins in the clades responsible for global outbreaks causing invasive infections suggest an explanation for the different phenotypes observed between clades.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/genetics/iyab029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8128392PMC
May 2021

Genomic insights into the host specific adaptation of the Pneumocystis genus.

Commun Biol 2021 03 8;4(1):305. Epub 2021 Mar 8.

Critical Care Medicine Department, NIH Clinical Center, National Institutes of Health (NIH), Bethesda, MD, USA.

Pneumocystis jirovecii, the fungal agent of human Pneumocystis pneumonia, is closely related to macaque Pneumocystis. Little is known about other Pneumocystis species in distantly related mammals, none of which are capable of establishing infection in humans. The molecular basis of host specificity in Pneumocystis remains unknown as experiments are limited due to an inability to culture any species in vitro. To explore Pneumocystis evolutionary adaptations, we have sequenced the genomes of species infecting macaques, rabbits, dogs and rats and compared them to available genomes of species infecting humans, mice and rats. Complete whole genome sequence data enables analysis and robust phylogeny, identification of important genetic features of the host adaptation, and estimation of speciation timing relative to the rise of their mammalian hosts. Our data reveals insights into the evolution of P. jirovecii, the sole member of the genus able to infect humans.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s42003-021-01799-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7940399PMC
March 2021

Genomic epidemiology of a case cluster in Glasgow, Scotland, 2018.

Microb Genom 2021 03 23;7(3). Epub 2021 Feb 23.

Broad Institute of MIT and Harvard, Cambridge, MA, USA.

In 2018, a cluster of two cases of cryptococcosis occurred at the Queen Elizabeth University Hospital (QEUH) in Glasgow, Scotland (UK). It was postulated that these cases may have been linked to pigeon droppings found on the hospital site, given there have been previous reports of associated with pigeon guano. Although some samples of pigeon guano taken from the site yielded culturable yeast from genera related to , they have since been classified as or spp., and no isolates of were recovered from either the guano or subsequent widespread air sampling. In an attempt to further elucidate any possible shared source of the clinical isolates, we used whole-genome sequencing and phylogenetic analysis to examine the relationship of the two isolates from the QEUH cases, along with two isolates from sporadic cases treated at a different Glasgow hospital earlier in 2018. Our work demonstrated that these four clinical isolates were not clonally related; while all isolates were from the VNI global lineage and of the same mating type (MATα), the genotypes of the two QEUH isolates were separated by 1885 base changes and belonged to different sub-lineages, recently described as the intercontinental sub-clades VNIa-93 and VNIa-5. In contrast, one of the two sporadic 2018 clinical isolates was determined to belong to the VNIb sub-lineage and the other classified as a VNIV/VNI hybrid. Our work demonstrated that the two 2018 QEUH isolates and the two prior clinical isolates were all genetically distinct. It was not possible to determine whether the QEUH genotypes stemmed from independent sources or from the same source, i.e. pigeons carrying different genotypes, but it should be noted that whilst members of allied genera within the were isolated from the hospital environment, there were no environmental isolations of .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1099/mgen.0.000537DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8190611PMC
March 2021

Comparative genomics of white and opaque cell states supports an epigenetic mechanism of phenotypic switching in Candida albicans.

G3 (Bethesda) 2021 02;11(2)

Department of Molecular Microbiology and Immunology, Brown University, Providence, RI 02912, USA.

Several Candida species can undergo a heritable and reversible transition from a 'white' state to a mating proficient 'opaque' state. This ability relies on highly interconnected transcriptional networks that control cell-type-specific gene expression programs over multiple generations. Candida albicans, the most prominent pathogenic Candida species, provides a well-studied paradigm for the white-opaque transition. In this species, a network of at least eight transcriptional regulators controls the balance between white and opaque states that have distinct morphologies, transcriptional profiles, and physiological properties. Given the reversible nature and the high frequency of white-opaque transitions, it is widely assumed that this switch is governed by epigenetic mechanisms that occur independently of any changes in DNA sequence. However, a direct genomic comparison between white and opaque cells has yet to be performed. Here, we present a whole-genome comparative analysis of C. albicans white and opaque cells. This analysis revealed rare genetic changes between cell states, none of which are linked to white-opaque switching. This result is consistent with epigenetic mechanisms controlling cell state differentiation in C. albicans and provides direct evidence against a role for genetic variation in mediating the switch.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/g3journal/jkab001DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8366294PMC
February 2021

Application of an optimized annotation pipeline to the Cryptococcus deuterogattii genome reveals dynamic primary metabolic gene clusters and genomic impact of RNAi loss.

G3 (Bethesda) 2021 02;11(2)

Département de Mycologie, Institut Pasteur, Unité Biologie des ARN des Pathogènes Fongiques, F-75015 Paris, France.

Evaluating the quality of a de novo annotation of a complex fungal genome based on RNA-seq data remains a challenge. In this study, we sequentially optimized a Cufflinks-CodingQuary-based bioinformatics pipeline fed with RNA-seq data using the manually annotated model pathogenic yeasts Cryptococcus neoformans and Cryptococcus deneoformans as test cases. Our results show that the quality of the annotation is sensitive to the quantity of RNA-seq data used and that the best quality is obtained with 5-10 million reads per RNA-seq replicate. We also showed that the number of introns predicted is an excellent a priori indicator of the quality of the final de novo annotation. We then used this pipeline to annotate the genome of the RNAi-deficient species Cryptococcus deuterogattii strain R265 using RNA-seq data. Dynamic transcriptome analysis revealed that intron retention is more prominent in C. deuterogattii than in the other RNAi-proficient species C. neoformans and C. deneoformans. In contrast, we observed that antisense transcription was not higher in C. deuterogattii than in the two other Cryptococcus species. Comparative gene content analysis identified 21 clusters enriched in transcription factors and transporters that have been lost. Interestingly, analysis of the subtelomeric regions in these three annotated species identified a similar gene enrichment, reminiscent of the structure of primary metabolic clusters. Our data suggest that there is active exchange between subtelomeric regions, and that other chromosomal regions might participate in adaptive diversification of Cryptococcus metabolite assimilation potential.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/g3journal/jkaa070DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8022950PMC
February 2021

An oxindole efflux inhibitor potentiates azoles and impairs virulence in the fungal pathogen Candida auris.

Nat Commun 2020 12 22;11(1):6429. Epub 2020 Dec 22.

Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada.

Candida auris is an emerging fungal pathogen that exhibits resistance to multiple drugs, including the most commonly prescribed antifungal, fluconazole. Here, we use a combinatorial screening approach to identify a bis-benzodioxolylindolinone (azoffluxin) that synergizes with fluconazole against C. auris. Azoffluxin enhances fluconazole activity through the inhibition of efflux pump Cdr1, thus increasing intracellular fluconazole levels. This activity is conserved across most C. auris clades, with the exception of clade III. Azoffluxin also inhibits efflux in highly azole-resistant strains of Candida albicans, another human fungal pathogen, increasing their susceptibility to fluconazole. Furthermore, azoffluxin enhances fluconazole activity in mice infected with C. auris, reducing fungal burden. Our findings suggest that pharmacologically targeting Cdr1 in combination with azoles may be an effective strategy to control infection caused by azole-resistant isolates of C. auris.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1038/s41467-020-20183-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7755909PMC
December 2020

Phylogenomic Analysis of a 55.1-kb 19-Gene Dataset Resolves a Monophyletic that Includes the Species Complex.

Phytopathology 2021 07 9;111(7):1064-1079. Epub 2021 Sep 9.

Departamento de Genética, Campus de Excelencia Internacional Agroalimentario, Universidad de Córdoba, Córdoba, Spain.

Scientific communication is facilitated by a data-driven, scientifically sound taxonomy that considers the end-user's needs and established successful practice. In 2013, the community voiced near unanimous support for a concept of that represented a clade comprising all agriculturally and clinically important species, including the species complex (FSSC). Subsequently, this concept was challenged in 2015 by one research group who proposed dividing the genus into seven genera, including the FSSC described as members of the genus , with subsequent justification in 2018 based on claims that the 2013 concept of is polyphyletic. Here, we test this claim and provide a phylogeny based on exonic nucleotide sequences of 19 orthologous protein-coding genes that strongly support the monophyly of including the FSSC. We reassert the practical and scientific argument in support of a genus that includes the FSSC and several other basal lineages, consistent with the longstanding use of this name among plant pathologists, medical mycologists, quarantine officials, regulatory agencies, students, and researchers with a stake in its taxonomy. In recognition of this monophyly, 40 species described as genus were recombined in genus , and nine others were renamed Here the global community voices strong support for the inclusion of the FSSC in , as it remains the best scientific, nomenclatural, and practical taxonomic option available.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1094/PHYTO-08-20-0330-LEDOI Listing
July 2021

Experimental Evolution Identifies Adaptive Aneuploidy as a Mechanism of Fluconazole Resistance in Candida auris.

Antimicrob Agents Chemother 2020 12 16;65(1). Epub 2020 Dec 16.

Department of Infectious Diseases, Huashan Hospital, and State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai, China

is a newly emerging fungal pathogen of humans and has attracted considerable attention from both the clinical and basic research communities. Clinical isolates of are often resistant to one or more antifungal agents. To explore how antifungal resistance develops, we performed experimental evolution assays using a fluconazole-susceptible isolate of (BJCA001). After a series of passages through medium containing increasing concentrations of fluconazole, fungal cells acquired resistance. By sequencing and comparing the genomes of the parental fluconazole-susceptible strain and 26 experimentally evolved strains of , we found that a portion of fluconazole-resistant strains carried one extra copy of chromosome V. In the absence of fluconazole, cells rapidly became susceptible and lost the extra copy of chromosome V. Genomic and transcriptome sequencing (RNA-Seq) analyses indicate that this chromosome carries a number of drug resistance-related genes, which were transcriptionally upregulated in the resistant, aneuploid strains. Moreover, missense mutations were identified in the genes , , and in all experimentally evolved strains. Our findings suggest that the gain of an extra copy of chromosome V is associated with the rapid acquisition of fluconazole resistance and may represent an important evolutionary mechanism of antifungal resistance in .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/AAC.01466-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7927865PMC
December 2020

Blastomycosis in Africa and the Middle East: A Comprehensive Review of Reported Cases and Reanalysis of Historical Isolates Based on Molecular Data.

Clin Infect Dis 2021 10;73(7):e1560-e1569

Agricultural, Life and Environmental Sciences, University of Alberta, Edmonton, Alberta, Canada.

Background: Blastomycosis has been reported from countries in Africa and the Middle East, but a decades-long debate has persisted regarding whether this is the same disease known in North America and caused by Blastomyces dermatitidis and Blastomyces gilchristii.

Methods: We reviewed published cases of human and veterinary blastomycosis from Africa and the Middle East. We abstracted epidemiological and clinical features of cases, including sites of disease, diagnosis, management, outcomes, and, where available, genetic and antigenic typing of case isolates. In addition, we sequenced nucleic acids from 9 clinical isolates from Africa deposited in global collections as B. dermatitidis; for 5, we sequenced the internal transcribed spacer regions, and for the other 4 we sequenced the whole genomes.

Results: We identified 172 unique human patients with blastomycosis, including 159 patients from 25 African countries and 12 patients from 5 Middle Eastern countries, and also identified 7 reports of veterinary blastomycosis. In humans, cutaneous disease predominated (n = 100/137, 73%), followed by pulmonary (n = 73/129, 57%) and osteoarticular involvement (n = 61/128, 48%). Unusual direct microscopy/histopathological presentations included short hyphal fragments in tissues (n = 23/129, 18%). There were 34 genotyped case isolates that comprised 4 species: Blastomyces percursus (n = 22, 65%), from 8 countries throughout all regions; Blastomyces emzantsi (n = 9, 26%), from South Africa; B. dermatitidis (n = 1, 3%), from the Democratic Republic of Congo; and B. gilchristii (n = 2, 6%), from South Africa and Zimbabwe.

Conclusions: Blastomycosis occurs throughout Africa and the Middle East and is caused predominantly by B. percursus and, at least in South Africa, B. emzantsi, resulting in distinct clinical and pathological patterns of disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/cid/ciaa1100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8492124PMC
October 2021

Understanding the Emergence of Multidrug-Resistant : Using Whole-Genome Sequencing to Describe the Population Structure of Species Complex.

Front Genet 2020 10;11:554. Epub 2020 Jun 10.

Mycotic Diseases Branch, Centers for Disease Control and Prevention, Atlanta, GA, United States.

The recent emergence of a multidrug-resistant yeast, , has drawn attention to the closely related species from the complex that include , and the recently identified . Here, we used antifungal susceptibility testing and whole-genome sequencing (WGS) to investigate drug resistance and genetic diversity among isolates of complex from different geographic areas in order to assess population structure and the extent of clonality among strains. Although most isolates of all four species were genetically distinct, we detected evidence of the in-hospital transmission of and in one hospital in Panama, indicating that these species are also capable of causing outbreaks in healthcare settings. We also detected evidence of the rising azole resistance among isolates of and in Colombia, Panama, and Venezuela linked to substitutions in gene as well as amplification of this gene in in isolates in Colombia suggesting the presence of evolutionary pressure for developing azole resistance in this region. Our results demonstrate that these species need to be monitored as possible causes of outbreaks of invasive infection.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3389/fgene.2020.00554DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7298116PMC
June 2020

Mutations in : a Novel Genetic Determinant of Clinical Fluconazole Resistance in Candida auris.

mBio 2020 05 12;11(3). Epub 2020 May 12.

Department of Clinical Pharmacy and Translational Science, University of Tennessee College of Pharmacy, Memphis, Tennessee, USA

has emerged as a multidrug-resistant pathogen of great clinical concern. Approximately 90% of clinical isolates are resistant to fluconazole, the most commonly prescribed antifungal agent, and yet it remains unknown what mechanisms underpin this fluconazole resistance. To identify novel mechanisms contributing to fluconazole resistance in , fluconazole-susceptible clinical isolate AR0387 was passaged in media supplemented with fluconazole to generate derivative strains which had acquired increased fluconazole resistance Comparative analyses of comprehensive sterol profiles, [H]fluconazole uptake, sequencing of genes homologous to genes known to contribute to fluconazole resistance in other species of , and relative expression levels of , , and were performed. All fluconazole-evolved derivative strains were found to have acquired mutations in the zinc-cluster transcription factor-encoding gene and to show a corresponding increase in expression relative to the parental clinical isolate, AR0387. Mutations in were also identified in a set of 304 globally distributed clinical isolates representing each of the four major clades. Introduction of the most common mutation found among fluconazole-resistant clinical isolates of into fluconazole-susceptible isolate AR0387 was confirmed to increase fluconazole resistance by 8-fold, and the correction of the same mutation in a fluconazole-resistant isolate, AR0390, decreased fluconazole MIC by 16-fold. Taken together, these data demonstrate that can rapidly acquire resistance to fluconazole and that mutations in significantly contribute to clinical fluconazole resistance. is an emerging multidrug-resistant pathogen of global concern, known to be responsible for outbreaks on six continents and to be commonly resistant to antifungals. While the vast majority of clinical isolates are highly resistant to fluconazole, an essential part of the available antifungal arsenal, very little is known about the mechanisms contributing to resistance. In this work, we show that mutations in the transcription factor significantly contribute to clinical fluconazole resistance. These studies demonstrated that mutations in can arise rapidly upon exposure to fluconazole and that a multitude of resistance-associated mutations are present among the majority of fluconazole-resistant isolates from a global collection and appear specific to a subset of lineages or clades. Thus, identification of this novel genetic determinant of resistance significantly adds to the understanding of clinical antifungal resistance in .
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.00365-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7218281PMC
May 2020

Threats Posed by the Fungal Kingdom to Humans, Wildlife, and Agriculture.

mBio 2020 05 5;11(3). Epub 2020 May 5.

Department of Molecular Genetics, University of Toronto, Toronto, Ontario, Canada

The fungal kingdom includes at least 6 million eukaryotic species and is remarkable with respect to its profound impact on global health, biodiversity, ecology, agriculture, manufacturing, and biomedical research. Approximately 625 fungal species have been reported to infect vertebrates, 200 of which can be human associated, either as commensals and members of our microbiome or as pathogens that cause infectious diseases. These organisms pose a growing threat to human health with the global increase in the incidence of invasive fungal infections, prevalence of fungal allergy, and the evolution of fungal pathogens resistant to some or all current classes of antifungals. More broadly, there has been an unprecedented and worldwide emergence of fungal pathogens affecting animal and plant biodiversity. Approximately 8,000 species of fungi and Oomycetes are associated with plant disease. Indeed, across agriculture, such fungal diseases of plants include new devastating epidemics of trees and jeopardize food security worldwide by causing epidemics in staple and commodity crops that feed billions. Further, ingestion of mycotoxins contributes to ill health and causes cancer. Coordinated international research efforts, enhanced technology translation, and greater policy outreach by scientists are needed to more fully understand the biology and drivers that underlie the emergence of fungal diseases and to mitigate against their impacts. Here, we focus on poignant examples of emerging fungal threats in each of three areas: human health, wildlife biodiversity, and food security.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.00449-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7403777PMC
May 2020

Tracing the Evolutionary History and Global Expansion of Candida auris Using Population Genomic Analyses.

mBio 2020 04 28;11(2). Epub 2020 Apr 28.

Broad Institute of MIT and Harvard, Cambridge, Massachusetts, USA

has emerged globally as a multidrug-resistant yeast that can spread via nosocomial transmission. An initial phylogenetic study of isolates from Japan, India, Pakistan, South Africa, and Venezuela revealed four populations (clades I, II, III, and IV) corresponding to these geographic regions. Since this description, has been reported in more than 30 additional countries. To trace this global emergence, we compared the genomes of 304 isolates from 19 countries on six continents. We found that four predominant clades persist across wide geographic locations. We observed phylogeographic mixing in most clades; clade IV, with isolates mainly from South America, demonstrated the strongest phylogeographic substructure. isolates from two clades with opposite mating types were detected contemporaneously in a single health care facility in Kenya. We estimated a Bayesian molecular clock phylogeny and dated the origin of each clade within the last 360 years; outbreak-causing clusters from clades I, III, and IV originated 36 to 38 years ago. We observed high rates of antifungal resistance in clade I, including four isolates resistant to all three major classes of antifungals. Mutations that contribute to resistance varied between the clades, with Y132F in as the most widespread mutation associated with azole resistance and S639P in for echinocandin resistance. Copy number variants in predominantly appeared in clade III and were associated with fluconazole resistance. These results provide a global context for the phylogeography, population structure, and mechanisms associated with antifungal resistance in In less than a decade, has emerged in health care settings worldwide; this species is capable of colonizing skin and causing outbreaks of invasive candidiasis. In contrast to other species, is unique in its ability to spread via nosocomial transmission and its high rates of drug resistance. As part of the public health response, whole-genome sequencing has played a major role in characterizing transmission dynamics and detecting new introductions. Through a global collaboration, we assessed genome evolution of isolates of from 19 countries. Here, we described estimated timing of the expansion of each clade and of fluconazole resistance, characterized discrete phylogeographic population structure of each clade, and compared genome data to sensitivity measurements to describe how antifungal resistance mechanisms vary across the population. These efforts are critical for a sustained, robust public health response that effectively utilizes molecular epidemiology.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.03364-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7188998PMC
April 2020

Genomic diversity of the human pathogen Paracoccidioides across the South American continent.

Fungal Genet Biol 2020 07 20;140:103395. Epub 2020 Apr 20.

Universidad Nacional del Nordeste, Resistencia, Chaco, Argentina. Electronic address:

Paracoccidioidomycosis (PCM) is a life-threatening systemic mycosis widely reported in the Gran Chaco ecosystem. The disease is caused by different species from the genus Paracoccidioides, which are all endemic to South and Central America. Here, we sequenced and analyzed 31 isolates of Paracoccidioides across South America, with particular focus on isolates from Argentina and Paraguay. The de novo sequenced isolates were compared with publicly available genomes. Phylogenetics and population genomics revealed that PCM in Argentina and Paraguay is caused by three distinct Paracoccidioides genotypes, P. brasiliensis (S1a and S1b) and P. restrepiensis (PS3). P. brasiliensis S1a isolates from Argentina are frequently associated with chronic forms of the disease. Our results suggest the existence of extensive molecular polymorphism among Paracoccidioides species, and provide a framework to begin to dissect the connection between genotypic differences in the pathogen and the clinical outcomes of the disease.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.fgb.2020.103395DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7385733PMC
July 2020

Tracking a Global Threat: a New Genotyping Method for Candida auris.

mBio 2020 03 10;11(2). Epub 2020 Mar 10.

Institut Pasteur, Molecular Mycology Unit, CNRS UMR2000, National Reference Center for Invasive Mycoses and Antifungals (NRCMA), Paris, France.

Over the past decade, has emerged as an urgent threat to public health. Initially reported from cases of ear infections in Japan and Korea, has since been detected around the world. While whole-genome sequencing has been extensively used to trace the genetic relationships of the global emergence and local outbreaks, a recent report in describes a targeted genotyping method as a rapid and inexpensive method for classifying isolates (T. de Groot, Y. Puts, I. Berrio, A. Chowdhary, and J. F. Meis, mBio 11:e02971-19, https://doi.org/10.1128/mBio.02971-19, 2020).
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.00259-20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7064761PMC
March 2020

Diversity and Complexity of the Large Surface Protein Family in the Compacted Genomes of Multiple Species.

mBio 2020 03 3;11(2). Epub 2020 Mar 3.

Leidos BioMedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, Maryland, USA.

, a major opportunistic pathogen in patients with a broad range of immunodeficiencies, contains abundant surface proteins encoded by a multicopy gene family, termed the major surface glycoprotein (Msg) gene superfamily. This superfamily has been identified in all species characterized to date, highlighting its important role in biology. In this report, through a comprehensive and in-depth characterization of 459 genes from 7 species, we demonstrate, for the first time, the phylogeny and evolution of conserved domains in Msg proteins and provide a detailed description of the classification, unique characteristics, and phylogenetic relatedness of five Msg families. We further describe, for the first time, the relative expression levels of individual families in two rodent species, the substantial variability of the repertoires in from laboratory and wild rats, and the distinct features of the expression site for the classic genes in from 8 mammalian host species. Our analysis suggests multiple functions for this superfamily rather than just conferring antigenic variation to allow immune evasion as previously believed. This study provides a rich source of information that lays the foundation for the continued experimental exploration of the functions of the Msg superfamily in biology. continues to be a major cause of disease in humans with immunodeficiency, especially those with HIV/AIDS and organ transplants, and is being seen with increasing frequency worldwide in patients treated with immunodepleting monoclonal antibodies. Annual health care associated with pneumonia costs ∼$475 million dollars in the United States alone. In addition to causing overt disease in immunodeficient individuals, can cause subclinical infection or colonization in healthy individuals, which may play an important role in species preservation and disease transmission. Our work sheds new light on the diversity and complexity of the superfamily and strongly suggests that the versatility of this superfamily reflects multiple functions, including antigenic variation to allow immune evasion and optimal adaptation to host environmental conditions to promote efficient infection and transmission. These findings are essential to consider in developing new diagnostic and therapeutic strategies.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/mBio.02878-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7064768PMC
March 2020

Evolutionary Persistence of DNA Methylation for Millions of Years after Ancient Loss of a De Novo Methyltransferase.

Cell 2020 01 16;180(2):263-277.e20. Epub 2020 Jan 16.

Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA; Chan-Zuckerberg Biohub, San Francisco, CA 94158, USA. Electronic address:

Cytosine methylation of DNA is a widespread modification of DNA that plays numerous critical roles. In the yeast Cryptococcus neoformans, CG methylation occurs in transposon-rich repeats and requires the DNA methyltransferase Dnmt5. We show that Dnmt5 displays exquisite maintenance-type specificity in vitro and in vivo and utilizes similar in vivo cofactors as the metazoan maintenance methylase Dnmt1. Remarkably, phylogenetic and functional analysis revealed that the ancestral species lost the gene for a de novo methylase, DnmtX, between 50-150 mya. We examined how methylation has persisted since the ancient loss of DnmtX. Experimental and comparative studies reveal efficient replication of methylation patterns in C. neoformans, rare stochastic methylation loss and gain events, and the action of natural selection. We propose that an epigenome has been propagated for >50 million years through a process analogous to Darwinian evolution of the genome.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1016/j.cell.2019.12.012DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7197499PMC
January 2020

Complete Genome Sequences for Two Clinical Isolates from Northern and Southern Vietnam.

Microbiol Resour Announc 2020 Jan 9;9(2). Epub 2020 Jan 9.

Division of Infectious Diseases, Duke University School of Medicine, Durham, North Carolina, USA.

is a thermally dimorphic fungus endemic in China and Southeast Asia that causes fatal infections in immunocompromised individuals, particularly in patients with advanced HIV disease. Here, we report the complete genome sequences of two clinical isolates from northern and southern Vietnam.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1128/MRA.01367-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6952663PMC
January 2020
-->