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PubMed2024

Virulence-associated variants in Cryptococcus neoformans sequence type 93 are less likely to be associated with population structure compared to independent rare mutations.

Cryptococcus neoformans is a pathogenic yeast that is the causative agent of cryptococcal meningitis. While it is well known that the genotype of C. neoformans impacts patient outcomes, the reason for this association has not been well elucidated. In this study, we examined the relationship between two subpopulations in the sequence type 93 clade of C. neoformans: ST93A and ST93B. We found extensive linkage disequilibrium (LD) among the single nucleotide polymorphisms (SNPs) that differentiate ST93A from ST93B. We also found differences in the extent of linkage among SNPs within each subpopulation; LD was more extensive within ST93B than ST93A. SNPs associated with virulence were in long-range linkage disequilibrium with less frequency than recurrent SNPs not associated with virulence. We investigated the karyotype of ST93A and ST93B using contour-clamped gel electrophoresis and long-read sequencing and found that the extensive long-range linkage was not due to chromosomal rearrangements. Overall, we found that the two subpopulations in ST93 are driven by SNPs in LD. We additionally found that recurrent SNPs associated with virulence were less frequently evolutionarily linked and were two times more likely to be independent, congruent mutations rather than tied to phylogeny.IMPORTANCECryptococcus neoformans is an important pathogen that is widely distributed and ubiquitous in the environment. The majority of the human population has a latent, controlled infection suggesting that C. neoformans is uniquely adapted to cause infection. In spite of this, the reason C. neoformans is a pathogen remains unknown; interestingly, most environmental isolates are avirulent but are genetically very similar to disease-causing virulent isolates. Recent evidence from genome-wide association studies shows that small mutations in key virulence-associated genes are associated with the virulence of specific isolates. The data presented here provide an evolutionary framework for those small mutations. The mutations that impact disease are not being collected over long-term evolution. The mutations may instead occur independently during infection. Identifying these genes that are more likely to be mutated during infection will be fundamental for understanding C. neoformans virulence.

Cryptococcus neoformans↗

PubMed2005

Difficulty in diagnosing chronic meningitis caused by capsule-deficient Cryptococcus neoformans.

We report a case of chronic meningitis due to capsule-deficient Cryptococcus neoformans which could not be diagnosed by routine morphological and immunological cerebrospinal fluid (CSF) examination. Repeated CSF examination and culture did not disclose the fungal body, and the cryptococcal antigen tests were always negative. Culture of ventricular fluid showed non-encapsulated cultured cells which were stained positively by indirect immunofluorescence using diluted sera from another patient diagnosed with cryptococcal meningitis. Inoculation of dispersed colonies into mice resulted in encapsulated C neoformans. It is important to suspect capsule-deficient C neoformans when the conventional diagnostic approaches fail to identify the organism or antigens.

Adult↗

PubMed2001

Encephalomyelitis due to Cryptococcus neoformans var gattii presenting as spinal tumour: case report and review of the literature.

A 24 year old immunocompetent German resident is described who developed multifocal encephalomyelitis due to infection with Cryptococcus neoformans var gatti, commonly considered a disease of tropical regions. In the light of current knowledge on the epidemiology of C neoformans var gatti and the travel history of the patient it is assumed that the infection was acquired outside Europe. As exclusive intramedullary involvement is an outstandingly rare manifestation in spinal cryptococcosis, the particular diagnostic procedure and the therapeutic strategies are discussed

Adult↗

PubMed1976

Beta-glucanases in the yeast Cryptococcus albidus var. aerius. Production and separation of beta-glucanases in asynchronous cultures.

beta-Glucanases were detected in cell-free extracts of the yeast Cryptococcus albidus var. aerius when grown on glucose as the sole carbon source. The production of beta-glucanases was followed in log-phase cells and stationary-phase cells; the maximal production of beta-(1 leads to 3) and beta-(1 leads to 6) glucanases takes place respectively in log-phase and stationary-phase cells. The results show that there are marked differences in the elution profiles on Sephadex G-50 of fractions containing beta-glucanase from cells grown for 12, 24, 48, 72, and 96 h. The possibility either of replacement changes in fractions containing beta-glucanase activity or of a different synthesis of each beta-glucanase during the growth of the yeast is discussed. The results suggest that all fractions containing beta-glucanases hydrolyze both beta-(1 leads to 3) and beta-(1 leads to 6) linkages. Evidence in support of the conclusion that a low molecular form of beta-glucanase has a molecular weight of 2100 +/- 100 is also shown.

Cell-Free System↗

PubMed1976

beta-Xylosidases in the yeast Cryptococcus albidus var. aerius.

beta-xylosidase activity has been detected in cell-free extracts and in culture fluids when Cryptococcus albidus var. aerius was grown on glucose as the sole carbon source. The enzyme appears to be constitutive. Mild acid treatment of whole cells suggested that the total activity is located in the periplasmic space and some experiments indicated that it is partially associated with the cell walls. DEAE-Sephadex A50 chromatography has shown that there are two different forms of beta-xylosidase in the cell-free extracts, but only one form is present in the supernatants of culture.

Cell Wall↗

PubMed1979

A modified method for testing inositol assimilation by Cryptococcus species.

The proposed method for rapid testing of inositol assimilation incorporates shake cultures in an indicator-based broth containing inositol (1%), yeast nitrogen base (0.067%), bromocresol purple, and a heavy inoculum. Of 153 yeast isolates investigated, inositol assimilation was shown with the modified method, as also by the Adams-Cooper procedure, in all the 123 isolates, representing 11 species of Cryptococcus. The results were negative by both the methods in the remaining 30 isolates belonging to Candida, Rhodotorula, Torulopsis, Pichia, Saccharomyces, and Sporobolomyces. The modified method was found to be significantly more effective than the Adams-Cooper procedure; the results could be read within 36 h by the former as against 336 h by the latter method. The superiority of the modified method to the Adams-Cooper procedure is attributed to increased aeration in shake cultures, a heavier inoculum, and reduced concentration of yeast nitrogen base.

Cryptococcus↗

PubMed1980

Catabolism of 3-O-methyl-D-glucopyranose by Cryptococcus laurentii.

When Cryptococcus laurentii is grown on 3-O-methyl-D-glucopyranose (3-O-MeGlc), two metabolic products are methanol and an extracellular mannan. Hydrolysis of the mannan produced during growth on 3-O-MeGlc labelled with 14C in the 1 or 6 position yields mannose with retention of the labelling pattern in the carbon chain. O-Methyl-labelled substrate yields radioactive methanol. Three possible mechanisms of O-demethylation are ether cleavage, peroxidatic attack, and beta-elimination. The latter mechanism was shown to be the most likely through the use of double-labelled (3H and 14C) 3-O-MeGlc.

3-O-Methylglucose↗

PubMed1980

Effect of 3-O-methyl-D-glucose on the production of glycosidases by Cryptococcus laurentii and Saccharomyces cerevisiae.

3-O-Methyl-D-glucose (3-O-MeGlc) or a mixture of 3-O-MeGlc and glucose stimulate invertase, beta-glucosidase, alpha-glucosidase, and alpha-galactosidase production by Cryptococcus laurentii. They also increase invertase and alpha-glucosidase production by Saccharomyces cerevisiae. The stimulatory effect of 3-O-MeGlc is not caused by competition with glucose for transport nor by a direct action on glycosidases. It is proposed that 3-O-MeGlc acts as a structural rather than as a functional analogue of glucose displacing it from regulatory sites to relieve catabolite repression. Evidence is presented suggesting that intracellular cAMP levels may be related to the effect of 3-O-MeGlc.

3-O-Methylglucose↗

PubMed1991

Susceptibility of congenitally immunodeficient mice to a nonencapsulated strain of Cryptococcus neoformans.

The susceptibility of congenitally immunodeficient mice to a nonencapsulated strain of Cryptococcus neoformans (strain M7) was evaluated. Gnotobiotic mice with defined congenital defects in innate immunity (beige) or cell-mediated immunity (athymic) or with combined defects in innate and cellular immunity (beige athymic) were i.v. challenged with C. neoformans M7. The nonencapsulated strain of C. neoformans produced a persistent low-grade infection in the brains of all immunodeficient and immunocompetent mice used in this study. Immunocompetent mice (nu/+;bg/+) and immunodeficient bg/bg mice readily cleared nonencapsulated cryptococci from their kidneys, liver, lungs, and spleen. In contrast to nu/+ mice, nu/nu mice had a reduced capacity to clear nonencapsulated cryptococci from their kidneys and liver after i.v. challenge. Both bg/bg-nu/nu and bg/bg-nu/+ mice developed a low-grade infection in their kidneys, liver, lungs, and spleen, which was maintained throughout the 21-day study. Persistent infections were not due to reversion to an encapsulated state. These data indicate that a capsule may not always be necessary for C. neoformans to survive, in vivo, in tissues of immunodeficient and immunocompetent mice.

Animals↗

PubMed1993

Mannosyl transfer in Cryptococcus neoformans.

A particulate enzyme preparation from Cryptococcus neoformans transferred the mannosyl residue from GDP-mannose to an acceptor consisting of a commercial preparation of methyl 3-O-alpha-mannopyranosyl-alpha-mannopyranoside (containing 10% 2-O-alpha-mannopyranosyl-alpha-mannopyranoside). The configuration of the new bond was alpha by its susceptibility to alpha-mannosidase; the amount of product was dependent on the concentration of enzyme, of GDP-mannose, and of acceptor. The optimal temperature and pH were 37 degrees C and 7.0, respectively. Manganous ion was required for activity and acetyl coenzyme A was stimulatory. Studies suggested that dolichyl phosphate intermediates were not involved in this mannose transfer. The fact that none of the several acapsular mutants tested were deficient in this mannosyltransferase suggested that this enzyme was not involved in synthesis of backbone mannan linkages in capsular polysaccharide. NMR analysis of the methylmannotriose product showed only alpha(1-->2) linkages between sugar moieties. This mannosyltransferase evidently extends alpha(1-->2) mannan by adding another alpha(1-->2)-linked mannosyl residue. Its activity is appropriate for a role in synthesis of "high mannose" oligosaccharide moieties of glycoproteins.

Cryptococcus neoformans↗

PubMed1993

Analysis of ploidy and sexual characteristics of natural isolates of Cryptococcus neoformans.

We report here an extensive examination of the ploidy of a basidiomycetous yeast. The ploidy of 34 strains of Cryptococcus neoformans var. neoformans, isolated mostly from Japanese patients, was determined by means of quantitative fluorescence microscopy after DNA staining with propidium iodide. The sexual characteristics of the strains including the two tester strains were as follows. Twenty-four strains were of the heterothallic alpha type, 1 was of the heterothallic a type, and 2 were self fertile; the remaining 7 showed no sexual response. The following strains were found to be haploid: the heterothallic a strain, 20 of the 24 heterothallic alpha strains, and 3 of the 7 asexual strains. Thus, the C. neoformans strains were mostly haploid, as the sexual response of this organism suggests. Both of the self-fertile strains and four of the seven asexual strains were diploid. Unexpectedly, 4 of the 24 heterothallic alpha strains were diploid.

Cryptococcus neoformans↗

PubMed1994

Cryptococcus neoformans infection can elicit protective antibodies in mice.

An IgG1 monoclonal antibody generated from a mouse infected with Cryptococcus neoformans modified the course of intravenous cryptococcal infection in A/J mice by prolonging survival and reducing lung fungal burden, brain mass, and serum polysaccharide levels. The results demonstrate that C. neoformans infection can elicit useful antibodies.

Animals↗

PubMedDate not supplied

Reduced recovery of a Cryptococcus neoformans adherence mutant from a rat model of cryptococcosis.

Stable mutants of Cryptococcus neoformans (strain CSF-1) induced by treatment with ultraviolet light and nitrosoguanidine were isolated that demonstrated reduced adherence to glial cells in culture. Adherence of the mutants, as measured by a radiometric assay, was reduced by 50-70% of that attained for the parent CSF-1 strain. The adherence mutants appeared to be phenotypically similar to the CSF-1 strain. However, all but one mutant (designated as CSF-23) demonstrated slightly slower growth rates than the wild-type strain. The CSF-1 and CSF-23 strains were injected intravenously and intratracheally into normal rats and rats immunosuppressed by cyclophosphamide treatment, and the organ distribution and recovery of viable yeasts determined over 2-96 h. During this relatively short period of observation the majority of the yeasts were localized in the lungs. By either route of injection, the recovery of the CSF-23 adherence mutant was reduced by as much as 90% of that obtained for the wild-type strain. The results indicated that host cell adherence may be important for the persistence of cryptococci in tissue and that further studies with the adherence mutants are warranted.

Animals↗

PubMed2002

Genetics of Cryptococcus neoformans.

Cryptococcus neoformans is a pathogenic fungus that primarily afflicts immunocompromised patients, infecting the central nervous system to cause meningoencephalitis that is uniformly fatal if untreated. C. neoformans is a basidiomycetous fungus with a defined sexual cycle that has been linked to differentiation and virulence. Recent advances in classical and molecular genetic approaches have allowed molecular descriptions of the pathways that control cell type and virulence. An ongoing genome sequencing project promises to reveal much about the evolution of this human fungal pathogen into three distinct varieties or species. C. neoformans shares features with both model ascomycetous yeasts (Saccharomyces cerevisiae, Schizosaccharomyces pombe) and basidiomycetous pathogens and mushrooms (Ustilago maydis, Coprinus cinereus, Schizophyllum commune), yet ongoing studies reveal unique features associated with virulence and the arrangement of the mating type locus. These advances have catapulted C. neoformans to center stage as a model of both fungal pathogenesis and the interesting approaches to life that the kingdom of fungi has adopted.

Animals↗

PubMed2006

The biology of the Cryptococcus neoformans species complex.

Cryptococcus neoformans is a major cause of fungal meningoencephalitis in immunocompromised patients. Despite recent advances in the genetics and molecular biology of C. neoformans, and improved techniques for molecular epidemiology, aspects of the ecology, population structure, and mode of reproduction of this environmental pathogen remain to be established. Application of recent insights into the life cycle of C. neoformans and its different ways of engaging in sexual reproduction under laboratory conditions has just begun to affect research on the ecology and epidemiology of this human pathogenic fungus. The melding of these disparate disciplines should yield rich dividends in our understanding of the evolution of microbial pathogens, providing insights relevant to diagnosis, treatment, and prevention.

Animals↗

PubMedDate not supplied

Pneumonia and pleural effusion due to Cryptococcus laurentii in a clinically proven case of AIDS.

Non-neoformans cryptococci were previously considered to be saprophytes and nonpathogenic to humans. Cryptococcus laurentii is frequently used as a biological means to control fruit rot. Interestingly, C laurentii has recently been reported to be a rare cause of infection in humans. The authors report a case of pulmonary cryptococcosis caused by C laurentii in a diabetic AIDS patient who was on antituberculosis and antiretroviral treatments. The sputum smear revealed capsulated yeast cells that were identified as C laurentii. Repeated pleural fluid culture revealed growth of C laurentii. Both respiratory samples were negative for acid-fast bacilli. Moraxella catarrhalis and Klebsiella pneumoniae were also found in the sputum, but not in the pleural fluid. The patient had a good response to oral fluconazole therapy at 600 mg/day for five weeks and was then discharged. The present article is the first to report on the rare pulmonary involvement of C laurentii in the Indian HIV population. These unusual forms of cryptococci create a diagnostic predicament in the rapid diagnosis of pulmonary cryptococcosis. A high degree of suspicion and improvement of techniques for culture and identification will contribute to the early diagnosis and treatment of unusual fungal infections.

AIDS-Related Opportunistic Infections↗

PubMed2005

Pulmonary infection with Cryptococcus neoformans in the face of underlying sarcoidosis.

We present a case of limited pulmonary cryptococcal infection following exposure to pigeon excreta in a patient with sarcoidosis. The pathogenicity of Cryptococcus neoformans depends on the interplay between the immune status of the host and the virulence of the infecting strain. It can range from asymptomatic lung colonization in the immunocompetent host to rapidly progressive meningitis in immunocompromised patients. Immunological models of respiratory disease emphasize a distinction between infections associated with immune suppression on the one hand and diseases such as sarcoidosis believed to involve an excessive Th1-mediated immune response on the other. This case exemplifies the complex nature of immunological responses in the lung and highlights the importance of considering the possibility of co-existent fungal infection in individuals with sarcoidosis. Novel immunotherapeutic options for cryptococcal infection are discussed.

Adult↗

PubMed1978

Effect of a tetracycline antibiotic on the experimental pathogenicity of Cryptococcus neoformans.

The pathogenicity of three stains of Cryptococcus neoformans was experimentally tested by intradermal inoculations into albino rabbits and intraperitoneally into mice. A relationship was found between the number of inoculated cells and the diameter of dermal lesion; moreover, a typical kinetics of lesion evolution and healing has been shown. Treatment of rabbits with deoxytetracycline did not dramatically influence the behavior of dermal lesions. However, in experiments dealing with strain Vi selected for its enhanced dermotropism, the antibiotic did significantly provoke a diminition of the inflammatory area. In mice, the antibiotic caused a marked increase in mortality (as evaluated by both LD50 values and rate of mortality). Neither in rabbits nor in mice, however, were we able to detect a significant effect of the drug on the dissemination of C. neoformans cells in internal organs. There is no simple explanation for the reported observations but it is possible that local factors in the derma or an aspecific antiinflammatory action of deoxytetracycline are responsible for the unusual response of dermal experimental infection to antibiotic treatment.

Animals↗