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PubMedDate not supplied

Enhanced activity of antifungal drugs by lysozyme against Cryptococcus neoformans.

The in vitro susceptibility of 16 isolates of Cryptococcus neoformans to three antifungal drugs and lysozyme in combination was determined using an urea broth microdilution method. The antifungal activities of each drug alone against 16 isolates of Cr. neoformans were determined as mean minimal inhibitory concentrations (MICs). MICs of fluconazole, itraconazole and terbinafine were 2.0 micrograms ml-1, 0.004 microgram ml-1 and 0.25 microgram ml-1, respectively. Lysozyme alone inhibited the growth of Cr. neoformans in a dose-dependent manner, although the lysozyme was unable to kill the cells of Cr. neoformans at the highest concentration of 20 micrograms ml-1. The mean MICs of fluconazole, itraconazole and terbinafine in combination with lysozyme were 0.13 microgram ml-1, 0.004 microgram ml-1 and 0.03 microgram ml-1 respectively. The antifungal activity of fluconazole and terbinafine in combination with lysozyme against Cr. neoformans was greatly enhanced compared with that of each drug alone. Itraconazole was unable to enhance the antifungal activity, as it demonstrated higher activity against Cr. neoformans when alone rather than in combination. Lysozyme was confirmed to enhance the antifungal activity of fluconazole and terbinafine in vitro.

Antifungal Agents↗
PubMedDate not supplied

Fungaemia due to Cryptococcus laurentii and a review of non-neoformans cryptococcaemia.

Cryptococcus laurentii is one of several non-neoformans cryptococci that have rarely been associated with human infection. The spectrum of clinical infection due to non-neoformans species ranges from skin lesions to fungaemia. Most cases of non-neoformans fungaemia have been nosocomially acquired and have been associated with indwelling intravascular catheters and neutropenia. Limited data on in vitro susceptibilities of non-neoformans cryptococci show these species to be more resistant to fluconazole and flucytosine than most Cr. neoformans. Two such cases are presented here.

Adult↗
PubMed1998

Phenotypic and genotypic differentiation of several human and avian isolates of Cryptococcus neoformans.

The Cryptococcus neoformans strains isolated from two human cases could be diagnosed as Cr. neoformans var. neoformans by differentiation on the basis of their characteristics determined by proline, canavanine and EDTA urease tests. The results of the serovar assignment were: for the isolate from the meningoencephalitis patient with lethal outcome, serovar A; for the strain isolated from the osteomyelitis patient with benign course, serovar D. Also, the PCR fingerprinting using primers (GACA)4, (CAC)5 and FM 1 resulted in a clear and reproducible assignment of the Cr. neoformans strains to the varieties neoformans and gattii, respectively, and, in addition, it confirmed the serovar assignment. No statistically confirmed differences in virulence between the osteomyelitis and the meningoencephalitis strain could be established by i.v. testing in mice, nor did the PCR with several primers provide any clues to a genetically determined higher virulence of the meningoencephalitis strain. The different classification as serovars A and D does not allow any conclusions concerning different virulence. It was not possible to retrospectively establish the sources of infection of the two Cr. neoformans infections, but pigeon faeces may well have played a role as a reservoir for one of the illnesses.

Animals↗
PubMedDate not supplied

Isolates of Cryptococcus neoformans serotype A and D developed on canavanine-glycine-bromthymol blue medium.

Two isolates of Cryptococcus neoformans serotype A and one isolate of serotype D from pigeon droppings were found to grow on canavanine-glycine-bromthymol blue (CGB) medium, when the Japanese isolates of Cr. neoformans were examined for their serotype and biochemical characteristics. The susceptibility to canavanine and the activity in assimilation of glycine were analysed on these three isolates. They were resistant to canavanine at the high concentration of 3.6 mmol l-1 and developed by assimilating the glycine even at a concentration of 7 mmol l-1. These isolates were proved to develop well on CGB medium, which contains 0.1 mmol l-1 of canavanine and 133 mmol l-1 of glycine. Three isolates of Cr. neoformans developed on CGB medium were also confirmed to be serotype A or D by the molecular analysis.

Animals↗
PubMedDate not supplied

Susceptibility testing of Cryptococcus neoformans using the urea broth microdilution method.

An urea broth microdilution method to assay the susceptibility of Cryptococcus neoformans to antifungal drugs was newly developed. Using this method, urease activity of the fungus was measured instead of the viability by checking colony development. The urease activities were indicated by colour changes in optical density at 545 nm. The end point in this assay was considered as 99% inhibitory concentration. When we measured antifungal activities of the three drugs against 16 isolates of Cr. neoformans using this assay method, mean minimum-inhibitory concentrations (MICs) of fluconazole, itraconazole and terbinafine were 2.0 micrograms ml-1, 0.008 microgram ml-1 and 0.25 microgram ml-1 respectively. This assay method resulted in higher sensitivity in MICs of the three antifungal drugs than the broth microdilution method recommended by the Committee for Laboratory Standards of the Japanese Society for Medical Mycology. The results obtained using this assay method support the more effective evaluation of antifungal substances in susceptibility testing of Cr. neoformans.

Antifungal Agents↗
PubMed2004

Isolation of Cryptococcus neoformans in Antwerp Zoo's nocturnal house.

Cryptococcosis was diagnosed postmortem in a striped grass mouse (Lemniscomys barbarus) housed in the nocturnal department of Antwerp Zoo. Eight of the remaining mice in the cage were captured. Cryptococcus neoformans was isolated from the lung of one animal. Two mice had an elevated serum cryptococcal antigen titre. On examination of the pooled faecal samples collected from 17 animal species housed in 23 cages of the nocturnal department, the pathogenic yeast was isolated from the faeces of the striped grass mice and a degu (Octodon degus). Numerous Cr. neoformans colonies were isolated from a tree-trunk, tree-stumps, and decaying wood collected from a hollow tree used to decorate the animals cage. Subsequent examination in four other cages of the nocturnal department revealed that all the sampled tree-trunks were colonized by Cr. neoformans. The fungus was isolated from the air sampled in the cage of the degu. Air samples collected in the public and service corridors remained negative. All the isolated strains were identified as Cr. neoformans var. neoformans serotype A.

Animals↗
PubMed2004

Prevalence, serotypes and mating patterns of Cryptococcus neoformans in the pellets of different avifauna in Madras, India.

A total of 887 pellets of different avifauna were screened for the presence of Cryptococcus neoformans. One hundred and six of 887 samples (12%) yielded Cr. neoformans in culture. The report on the isolation of Cr. neoformans from the pellets of the crow appears to be new and of greater significance because of the ubiquitous prevalence of this bird in India. The prevalence of both MAT a and MAT alpha mating types were recorded. The serotype D was predominant over serotype A. The findings of the present study reveal the growing diverse ecological niche of Cr. neoformans in a the pellets of various avifauna in India.

Animals↗
PubMed2004

Effects of several antifungal drug combinations against clinical and environmental isolates of Cryptococcus neoformans from China.

The in vitro interactions of caspofungin (CSP) with terbinafine (TRB) and ravuconazole (RVC) with 5-fluorocytosine (5-FC) were tested against 82 clinical and environmental isolates of Cryptococcus neoformans from China. The interaction of CSP with TRB proved synergistic against those isolates with a CSP MIC < or =2 microg ml-1 (5% of the isolates), additive against 42% of the isolates and indifferent against 53%. The effects of RVC with 5-FC were synergistic, additive or indifferent against 8%, 26% and 67% of the isolates, respectively. No antagonistic effects were found among any of the drugs. The combinations of CSP with TRB and RVC with 5-FC may display beneficial effects in a strain-dependent manner, while in no case showed antagonistic effects. These data might be of use to design safer and more efficient treatments for patients with cryptococcosis and warrant further evaluation.

Animals↗
PubMed2005

Proven pulmonary cryptococcosis due to capsule-deficient Cryptococcus neoformans does not differ clinically from proven pulmonary cryptococcosis due to capsule-intact Cr. neoformans.

We compared the clinical manifestations, laboratory and radiologic findings, and outcomes of eight patients with proven pulmonary cryptococcosis (PC) caused by capsule-deficient Cryptococcus neoformans with those of six patients with PC caused by capsule-intact Cr. neoformans. The presentations and outcomes did not differ significantly between the groups.

Adult↗
PubMed2005

In vitro susceptibility of Cryptococcus neoformans clinical isolates from Egypt to seven antifungal drugs.

The in vitro susceptibility of 29 clinical isolates of Cryptococcus neoformans to fluconazole, miconazole, itraconazole, ketoconazole, flucytosine, nystatin and amphotericin B was tested by broth and colorimetric microdilution methods. Most of the isolates showed uniform patterns of susceptibility to the used antifungal agents. Only three isolates exhibited resistance [fourfold or greater rise in the minimum inhibitory concentrations (MICs)] to the tested antifungal drugs. The MIC50 and MIC90 were 0.5-8 mg l(-1) for 5-flucytosine, 0.2-8.25 mg l(-1) for nystatin, 0.5-16 mg l(-1) for fluconazole and 0.2-12.5 mg l(-1) for miconazole. However, MIC50 and MIC90 were in narrow range for the clinical yeast isolates in both methods used and showed 0.5-1 mg l(-1) for amphotericin B and 0.016-0.25 mg l(-1) for both ketoconazole and itraconazole. The combination of fluconazole plus flucytosine showed greater synergistic and fungicidal activity compared with that of fluconazole plus amphotericin B or the use of individual drugs.

Antifungal Agents↗
PubMed2006

Genotyping of Turkish environmental Cryptococcus neoformans var. neoformans isolates by pulsed field gel electrophoresis and mating type.

A total of 26 environmental Cryptococcus neoformans var. neoformans strains isolated from 634 samples of pigeon droppings collected from 54 different provinces of Turkey in 1996 and 1997 were included in this study. The results of pulsed-field gel electrophoresis (PFGE) showed that the 26 strains could be separated into 24 different PFGE patterns. In a mating-type study, of 26 strains, 20 were MATalpha, four were MATa, one was MATa/alpha and one was non-typable by STE20 specific primers. By the polymerase chain reaction typing, all the isolates were serotype A. The extensive heterogeneity among these isolates suggests that a single clonal population may not be present in Turkey. Additionally, the presence of an AMATa/DMATalpha hybrid may indicate the existence of strains that are AMATa mating type in Turkish environment.

Animals↗
PubMed1993

Cryptococcus neoformans in tropical northern Australia: predominantly variant gattii with good outcomes.

BACKGROUND: Infection with Cryptococcus neoformans is common in the Northern Territory of Australia. Disease is life threatening and treatment is prolonged and often complicated by the need for surgery and difficulties with medical therapy. AIMS: To document incidence, demography, risk factors, clinical features and outcomes of infection and to determine differences between gattii and neoformans varieties. METHODS: Case records of all patients (n = 35) diagnosed with cryptococcal infection at the Royal Darwin Hospital between 1976 and 1992 were reviewed retrospectively. Current status of patients was ascertained. Variety identification of isolates was determined by growth in canavanine-glycine-bromthymol blue agar. RESULTS: Of the 35 patients, 23 had meningitis, ten had pneumonia, one had a dermal infection and one had fungaemia with no obvious focus. Twelve (52%) meningitis cases and two (20%) pneumonia cases had no predisposing disease. Thirteen (57%) meningitis cases had concomitant pulmonary cryptococcosis. Twenty-nine patients with Aboriginal and six were Caucasian, with a relative risk for Aboriginals compared with non-Aboriginals of 20.6 (95% CI 8.6-49.5). Arnhemland was the commonest location of infection, with an annual incidence in Aboriginals of 0.14/1000. Fourteen (78%) of 18 isolates tested were C. neoformans var. gattii. Management was characterised by the frequent need for adjunctive surgery and prolonged or repeat courses of systemic antifungal therapy. Despite this, long-term outcomes are encouraging with a mortality of 14% overall and 9% in meningitis patients. The river red gum (Eucalyptus camaldulensis) has a limited distribution in Arnhemland and ongoing studies are seeking alternative environmental sources of C. neoformans var. gattii.

Adolescent↗
PubMed2006

Cryptococcus neoformans capsular polysaccharide component galactoxylomannan induces apoptosis of human T-cells through activation of caspase-8.

The major virulence factor of Cryptococcus neoformans is its polysaccharide capsule composed of glucuronoxylomannan (GXM), galactoxylomannan (GalXM) and mannoproteins. A variety of immunomodulating activities have been described for GXM and mannoproteins but little is known about possible interactions of GalXM with the immune system. In the present article, we investigate the effect of purified soluble GalXM on human T lymphocytes. The results indicate that, GalXM (i) can affect selected immune responses; (ii) causes significant impairment of T cell proliferation and increases interferon-gamma and interleukin-10 production; and (iii) induces apoptosis of T lymphocytes through activation of caspase-8 that terminates with fragmentation of DNA. These results are the first to suggest a role for GalXM in C. neoformans virulence by demonstrating that it can target human T cells, and that it may impair the development of an effective specific T cell response.

Apoptosis↗
PubMed2006

Characterization of a flocculation-like phenotype in Cryptococcus neoformans and its effects on pathogenesis.

We investigated the phenomenon of cell-cell aggregation (flocculation) in a serotype D strain of Cryptococcus neoformans (ATCC 24067, isolate RC-2). Cell aggregation into clumps of 5-40 cells (clump+ cells) occurred during the early log phase and disappeared in the beginning of the stationary phase (clump- cells). The cell aggregation phenomenon was medium dependent. Clump+ cells could be dispersed by either vortexing or proteinase K digestion. Most importantly, the transient change in cellular phenotype changed several important host-pathogen interactions. Adherence of clump+ cells to murine macrophage-like cells J774.16 was significantly (P < 0.001) enhanced compared with adherence of clump- cells. Furthermore, complement-mediated phagocytosis efficacy of dispersed clump+ cells was significantly higher (P < 0.001) compared with clump- cells. Similar findings were documented with an in vivo phagocytosis assay. Infection of mice with a low inoculum (10(4)) of clump+ cells resulted in lower fungal burden when compared with mice infected with clump- cells. Accordingly, mice infected with clump+ cells survived significantly longer than mice infected with clump- cells. These results indicate that the cellular phenotype undergoes significant changes that result in a transient flocculation-like phenotype. We hypothesize that this cell-cell aggregation is the result of changes in protein content in the polysaccharide capsule. We conclude from our data that the change in cellular phenotype has a dramatic effect on cell adherence, and on complement-mediated phagocytosis, both of which can affect the pathogenesis of the disease in the host. Our results underscore the complexity of studies that investigate host pathogen interactions and may explain differences and inconsistencies observed in in vitro and in vivo assays.

Animals↗
PubMed2006

Monoclonal antibodies can affect complement deposition on the capsule of the pathogenic fungus Cryptococcus neoformans by both classical pathway activation and steric hindrance.

The capsule of the human pathogenic fungus Cryptococcus neoformans presents the immune system with a formidable problem for phagocytosis. Capsule-mediated activation of the alternative complement (C) pathway results in component 3 (particularly, C3) binding to the capsule near the cell wall surface. Hence, for cells with large capsule, C3 cannot interact with the complement receptor (CR) and is not opsonic. However, C activation in either immune serum or in the presence of monoclonal antibody (mAb) to capsular polysaccharide localizes C3 to the capsular edge. When C. neoformans cells were coated with both C and antibody (Ab) opsonins, Ab bound first and promoted C3 deposition at the edge of the capsule. The mechanism for the Ab-mediated change in C3 localization to the capsule edge involved both classical C pathway activation and steric hindrance preventing C3 penetration into the capsule. The change in C3 localization changed the mode of phagocytosis in macrophages, such that localizing C3 at the edge of the capsule allowed phagocytosis through C3-CR3 and C3-CR4 interactions, which did not occur in serum without Ab. These findings reveal a new mechanism of Ab action whereby Abs affect the location of C3 and its interaction with its receptor in macrophages depending on the immunoglobulin concentration.

Antibodies, Fungal↗
PubMed2004

Determination of Cryptococcus neoformans var. neoformans mating type by multiplex PCR.

Mating type plays an important role in the epidemiology and virulence of Cryptococcus neoformans. The present study designed a multiplex PCR method to distinguish the six mating type patterns (Aa, Da, Aalpha, Dalpha, Aa/Dalpha, and Aalpha/Da) of C. neoformans var. neoformans. PCR amplification identified one fragment for Aa (860 bp), Dalpha (413 bp) and Da (645 bp) strains, two fragments for Aalpha (320 and 400 bp) and Aa/Dalpha (860 and 413 bp) strains, and three fragments (645, 400, 320 bp) for an Aalpha/Da strain. The method appears to be a valid, simple and relatively inexpensive tool for epidemiological and virulence studies.

Cryptococcus neoformans↗
PubMed1996

Characterization of Cryptococcus neoformans by random DNA amplification.

Random amplified polymorphic DNA (RAPD) was optimized and used to distinguish between the varieties and serotypes of Cryptococcus neoformans. The RAPD technique distinguished between serotypes A, D or AD within C. neoformans var. neoformans, and revealed further differentiation within each serotype. Four RAPD profiles were clearly recognizable within C. neoformans var. gattii, although its two serotypes, B and C, were only differentiated with one primer combination out of seven.

Cryptococcus neoformans↗
PubMed2001

Genetic diversity in the red yeast Cryptococcus hungaricus and its phylogenetic relationship to some related basidiomycetous yeasts.

Cryptococcus hungaricus is a basidiomycetous yeast with the abilities to synthesize carotenoid pigments and to grow under psychrophile conditions. Six C. hungaricus strains have been isolated so far from different habitats. In this study we wished to clarify the relationships amongst them. Morphological and physiological characters, mitochondrial DNA restriction profiles, and the presence of mycoviruses were examined. Internal transcribed spacers together with the 5.8S rDNA, the D1/D2 region of 26S rDNA, and partial sequences of the 18S rRNA gene were also analysed. On the basis of the phylogenetic analyses the type strain CBS 4214(T) together with four other C. hungaricus isolates were closely related to Bullera armeniaca and Bullera crocea, while strain CBS 6569 was much more similar to Cystofilobasidium than to the other C. hungaricus isolates.

Basidiomycota↗