Aspects of physiology of Histoplasma capsulatum (A review)

Boguslawski, George; Stetler, Dean A.

doi:10.1007/bf00436235

Cited by 17 publications

(10 citation statements)

References 31 publications

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“…However, the in vivo defect of the C. neoformans met3 auxotrophic mutant stands in contrast to that observed for H. capsulatum, where the pathogenic yeast phase (but not the saprobic mycelial phase) is auxotrophic due to a deficiency in sulfite reductase (Boguslawski & Stetler, 1979 ;Maresca & Kobayashi, 1989), a step in sulfate assimilation which occurs after ATP sulfurylase. The different responses of these two species of pathogenic fungi to deficiencies in the sulfate-assimilation arm of the methionine\cysteine biosynthetic pathway, which could be due to differences in their ecological niches or differences in methionine and\or cysteine uptake systems, are intriguing.…”

Section: Discussionmentioning

confidence: 84%

“…For example, the saprobic mycelial form of the human pathogen Histoplasma capsulatum is prototrophic while the pathogenic yeast form requires cysteine (reviewed by Boguslawski & Stetler, 1979 ;Maresca & Kobayashi, 1989). Similar results are seen in two other dimorphic pathogenic fungi infecting humans, Blastomyces derma- The GenBank accession numbers for the sequences reported in this paper are AY035556 and AF489498.…”

Section: Introductionmentioning

confidence: 99%

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Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant a aThe GenBank accession numbers for the sequences reported in this paper are AY035556 and AF489498.

et al. 2002

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The Cryptococcus neoformans MET3 cDNA (encoding ATP sulfurylase) was cloned by complementation of the corresponding met3 mutation in Saccharomyces cerevisiae. Sequence analysis showed high similarity between the deduced amino acid sequence of the C. neoformans Met3p and other fungal ATP sulfurylases. A C. neoformans met3 mutant was made by targeted insertional mutagenesis, which had the expected auxotrophic phenotype, and reconstituted the met3 mutant to Met M . In vitro, the C. neoformans met3 mutant had a substantial defect in melanin formation, significantly reduced growth rate, and greatly increased thermotolerance. In the murine inhalation infection model, the met3 mutant was avirulent and was deficient in its ability to survive in mice. It is concluded that, in contrast to the yeast form of Histoplasma capsulatum, in C. neoformans the sulfate-assimilation arm of the methionine biosynthetic pathway plays an important role in vitro, even in the presence of abundant exogenous methionine, and is critical for virulence, and indeed for survival, in vivo.

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Section: Discussionmentioning

confidence: 84%

Section: Introductionmentioning

confidence: 99%

Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant a aThe GenBank accession numbers for the sequences reported in this paper are AY035556 and AF489498.

et al. 2002

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“…HMM, a medium optimized for growth of Histoplasma and macrophages, is based on Ham's F-12 cell culture medium and is supplemented with 1.8% glucose and 0.7 mM cystine (30). The increased cystine provides additional organic sulfur, since Histoplasma yeast cells and mycelia differ in their requirements for organic sulfur (31)(32)(33)(34)(35). Although both F-12 and RPMI have cysteine (0.2 mM and 0.4 mM, respectively), we tested if supplementation of F-12 or RPMI with cystine increased the yield of Histoplasma yeasts in microtiter plates.…”

Section: Methodsmentioning

confidence: 99%

Quantitative Microplate-Based Growth Assay for Determination of Antifungal Susceptibility of Histoplasma capsulatum Yeasts

2015

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bStandardized methodologies for determining the antifungal susceptibility of fungal pathogens is central to the clinical management of invasive fungal disease. Yeast-form fungi can be tested using broth macrodilution and microdilution assays. Reference procedures exist for Candida species and Cryptococcus yeasts; however, no standardized methods have been developed for testing the antifungal susceptibility of yeast forms of the dimorphic systemic fungal pathogens. For the dimorphic fungal pathogen Histoplasma capsulatum, susceptibility to echinocandins differs for the yeast and the filamentous forms, which highlights the need to employ Histoplasma yeasts, not hyphae, in antifungal susceptibility tests. To address this, we developed and optimized methodology for the 96-well microtiter plate-based measurement of Histoplasma yeast growth in vitro. Using optical density, the assay is quantitative for fungal growth with a dynamic range greater than 30-fold. Concentration and assay reaction time parameters were also optimized for colorimetric (MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] reduction) and fluorescent (resazurin reduction) indicators of fungal vitality. We employed this microtiter-based assay to determine the antifungal susceptibility patterns of multiple clinical isolates of Histoplasma representing different phylogenetic groups. This methodology fulfills a critical need for the ability to monitor the effectiveness of antifungals on Histoplasma yeasts, the morphological form present in mammalian hosts and, thus, the form most relevant to disease.T he increasing incidence of fungal disease necessitates adequate and timely assessment of antifungal susceptibility to guide the selection and implementation of antifungal therapies. Consequently, the Clinical and Laboratory Standards Institute (CLSI) and the European Committee on Antimicrobial Susceptibility Testing (EUCAST) have established reference test methods for susceptibility to the major classes of antifungals, which include the polyenes, azoles, and the recently developed echinocandins (1, 2). These standards utilize broth macrodilution or microdilution assays for testing yeasts (M27-A3 [3] and E.DEF 7.1 [4]) and filamentous fungi (M38-A2 [5] and E.DEF 9.1 [6]). Procedures, including inoculum preparation, media, assay duration, and endpoint definitions, have now been established for testing some of the more commonly encountered yeast-form pathogenic fungi (i.e., Candida species and Cryptococcus). However, notably missing from the M27-A3 and E.DEF 7.1 standards for testing of yeasts are methods for testing the yeast forms of the dimorphic fungi.The dimorphic fungi that cause systemic disease are characterized by distinct yeast and filamentous morphological states (7,8). Temperature is the principal morphology-determining factor, with yeast forms characterizing infections in mammals (9). Histoplasma capsulatum is the most common clinically encountered dimorphic fungal pathogen in the United States, with an estimated 10,000 to 20,000...

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“…Early studies of yeast growth in culture indicated that yeasts but not mycelia of most Histoplasma species are auxotrophic for cysteine due to temperature-dependent expression of sulfite reductase and the consequent inability to incorporate inorganic sulfate into cysteine (28)(29)(30)(31)(32). Organic sulfhydryls, such as cysteine, also reduce the redox potential, which contributes to yeast phase differentiation (33)(34)(35).…”

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confidence: 99%

Histoplasma capsulatum Depends on De Novo Vitamin Biosynthesis for Intraphagosomal Proliferation

2014

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During infection of the mammalian host, Histoplasma capsulatum yeasts survive and reside within macrophages of the immune system. Whereas some intracellular pathogens escape into the host cytosol, Histoplasma yeasts remain within the macrophage phagosome. This intracellular Histoplasma-containing compartment imposes nutritional challenges for yeast growth and replication. We identified and annotated vitamin synthesis pathways encoded in the Histoplasma genome and confirmed by growth in minimal medium that Histoplasma yeasts can synthesize all essential vitamins with the exception of thiamine. Riboflavin, pantothenate, and biotin auxotrophs of Histoplasma were generated to probe whether these vitamins are available to intracellular yeasts. Disruption of the RIB2 gene (riboflavin biosynthesis) prevented growth and proliferation of yeasts in macrophages and severely attenuated Histoplasma virulence in a murine model of respiratory histoplasmosis. Rib2-deficient yeasts were not cleared from lung tissue but persisted, consistent with functional survival mechanisms but inability to replicate in vivo. In addition, depletion of Pan6 (pantothenate biosynthesis) but not Bio2 function (biotin synthesis) also impaired Histoplasma virulence. These results indicate that the Histoplasma-containing phagosome is limiting for riboflavin and pantothenate and that Histoplasma virulence requires de novo synthesis of these cofactor precursors. Since mammalian hosts do not rely on vitamin synthesis but instead acquire essential vitamins through diet, vitamin synthesis pathways represent druggable targets for therapeutics.

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Aspects of physiology of Histoplasma capsulatum (A review)

Cited by 17 publications

References 31 publications

Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant a aThe GenBank accession numbers for the sequences reported in this paper are AY035556 and AF489498.

Molecular and genetic analysis of the Cryptococcus neoformans MET3 gene and a met3 mutant a aThe GenBank accession numbers for the sequences reported in this paper are AY035556 and AF489498.

Quantitative Microplate-Based Growth Assay for Determination of Antifungal Susceptibility of Histoplasma capsulatum Yeasts

Histoplasma capsulatum Depends on De Novo Vitamin Biosynthesis for Intraphagosomal Proliferation

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