Showing papers in "Medical Mycology in 1998"

Antifungal drug resistance in pathogenic fungi.

Abstract: Failures of drug treatment in fungal infections combined with improvements in performances and standardization of antifungal susceptibility testing have drawn attention to the problem of antifungal resistance and its underlying mechanisms. Resistance of Candida species and Cryptococcus neoformans to flucytosine (5FC) develops during monotherapy. Acquired resistance results from a failure to metabolize 5FC to 5FUTP and 5FdUMP, or from the loss of feedback control of pyrimidine biosynthesis. A combination of 5FC and amphotericin B (AmB) reduces the appearance of resistant C. albicans isolates. Resistance to AmB is unusual. C. lusitaniae is the most susceptible to AmB resistance. C. neoformans with decreased AmB susceptibility has been isolated from an HIV-infected patient. Acquired resistance to AmB is often associated with alteration of membrane lipids, especially ergosterol. Concomitant with the widespread use of fluconazole there have been increasing reports of fluconazole resistance in Candida species and C. neoformans. Fluconazole resistance was mostly associated with prior use of fluconazole as intermittent therapy or prophylactic continuous treatment for recurrent thrush. In contrast to fluconazole, itraconazole is active against C. krusei. Decreased susceptibility to itraconazole is observed over time in C. albicans isolates becoming resistant to fluconazole. Decreased susceptibility to itraconazole and SCH-56592 was also observed in a few Aspergillus fumigatus isolates. Failure to accumulate azole antifungals has been identified as a cause of resistance in several post-treatment C. albicans, C. glabrata and C. krusei isolates. In azole-resistant C. albicans isolates from AIDS patients with oropharyngeal candidiasis, multidrug efflux transporters of the ATP-binding cassette (ABC) superfamily and of the class of major facilitators (MF) have been shown to be responsible for the low level of accumulation of azole antifungal agents. Two genes for these transporters, the ABC-transporter gene CDR1 and the MF gene, CaMDR1 (BEN) were shown to be overexpressed in resistant C. albicans isolates. Overexpression of BEN in Saccharomyces cerevisiae conferred resistance to fluconazole and terbinafine. CDR1 overexpression in S. cerevisiae conferred cross-resistance to fluconazole, itraconazole, ketoconazole and terbinafine. C. albicans clinical isolates resistant to azole antifungal agents over-expressing the ABC-transporter genes CDR1 and CDR2 were less susceptible to the morpholine derivative amorolfine. In C. glabrata isolates azole resistance is based on over-expression of the CgCDR gene. A reduced susceptibility of ergosterol biosynthesis is another mechanism of resistance described in a number of post-treatment C. albicans, C. neoformans and Histoplasma capsulatum isolates. Mutations have been reported in the CYP51A1 genes of resistant C. albicans isolates. Over-expression of CYP51A1 in C. albicans and C. glabrata may also account for a decreased susceptibility to azole antifungal agents.

The role of malassezia species in the ecology of human skin and as pathogens

Abstract: The new taxonomic structure of the lipophilic genus Malassezia was presented with key characteristics for the seven described species. Among techniques used for epidemiological surveys, the pulsed field gel electrophoresis (PFGE) was found to be of little value in contrast to randomly amplified polymorphic DNA (RAPD). Immunological studies still yielded conflicting results but at least the immunomodulatory capacity of Malassezia yeasts appeared to be related to the cell wall lipids. A review of Malassezia infections together with the present consensus for their prevention and treatment was also made.

Comparison of six extraction techniques for isolation of DNA from filamentous fungi

Abstract: Filamentous fungi have a sturdy cell wall which is resistant to the usual DNA extraction procedures. We determined the DNA extraction procedure with the greatest yield of high quality fungal DNA and the least predilection for cross-contamination of equipment between specimens. Each of six extraction methods was performed using Aspergillus fumigatus hyphae. The six methods were: (1) glass bead pulverization with vortexing; (2) grinding with mortar and pestle followed by glass bead pulverization; (3) glass bead pulverization using 1% hydroxyacetyl trimethyl ammonium bromide (CTAB) buffer in a water bath sonicator; (4) water bath sonication in CTAB buffer; (5) grinding followed by incubation with CTAB; and (6) lyticase enzymatic cell lysis. Genomic DNA yields were measured by spectrophotometry and by visual reading of 2% agarose gels, with shearing assessed by the migration of the DNA on the gel. Genomic fungal DNA yields were highest for Method 1, followed by Methods 5 approximately = to 2 >3 approximately = to 4 approximately = to 6. Methods 2 and 5, both of which involved grinding with mortar and pestle, led to shearing of the genomic DNA in one of two trials each. We conclude that the use of glass beads with extended vortexing is optimal for extraction of microgramme amounts of DNA from filamentous fungal cultures.

Cryptococcus neoformans var. gattii--evidence for a natural habitat related to decaying wood in a pottery tree hollow.

Abstract: To study hollows of living trees as the natural habitat of Cryptococcus neoformans in an endemic area of cryptococcosis in the northeastern Brazilian region, samples of decaying wood were collected inside the hollows, plated on niger seed agar and inoculated into mice and hamsters. Identification of C. neoformans was based on morphological and physiological tests. Canavanine-glycine-bromothymol medium was used to screen the varieties and Crypto Check Iatron Kit to serotype the isolates. For a period of 29months C. neoformans var. gattii serotype B was isolated repeatedly from the hollow of a pottery tree (Moquilea tomentosa), pointing to the natural occurrence of C. neoformans var. gatti in decaying wood forming hollows in living trees. Evidence for anatural habitat of the variety gattii other than that related to Eucalyptus camaldulensis are discussed.

Importance of Candida species other than Candida albicans as opportunistic pathogens.

Abstract: Candida species other than C. albicans have become a significant cause of infection in humans. Several of the more commonly isolated of these species are less susceptible to commonly used azole antifungal drugs, a factor that poses significant difficulties for effective treatment. The modern mycology laboratory has an important role to play in several aspects relating to these organisms, including therapy, detection, identification and epidemiological analysis. The application of molecular techniques and phylogenetic analysis has led to the identification of a new species of Candida associated with mucosal candidiasis in HIV-infected individuals named Candida dubliniensis, the clinical significance of which is currently under investigation. Molecular techniques are also being applied to the analysis of determinants involved in pathogenicity of species such as Candida glabratta. These approaches should lead to a better understanding of these organisms and there ability to cause disease and should also provide more effective treatment.

Concentrations of airborne Aspergillus compared to the incidence of invasive aspergillosis: lack of correlation

Abstract: Air sampling of the rooms and corridors of the oncology wards of the hospital was carried out over a 54-week period to assess the concentration of viable Aspergillus conidia. A. fumigatus and A. flavus were recovered at a mean of 1.83 cfu m-3 air sampled. Individual samplings yielded concentrations of up to 11.6 cfu m-3. Other Aspergillus spp. were recovered at a mean of 2.38 cfu m-3 (maximum 32.6 cfu m-3). Concentration was not correlated with season or hospital ward. Review of autopsy results showed an average of 6.6 cases of aspergillosis annually over a 22-year period. No seasonal variation in case incidence was found. Six cases of invasive aspergillosis were diagnosed on the three cancer wards during the air-sampling period, but no association was seen linking these cases with changes in recovery of airborne Aspergillus. A seasonal pattern was not observed in the overall incidence of aspergillosis cases nor concentrations of airborne conidia.

Molecular diagnosis and epidemiology of fungal infections.

Abstract: A variety of methods are utilized for DNA strain subtyping of Candida spp. because no 'gold standard' exists. Random amplified polymorphic DNA (RAPD) or restriction enzyme analysis (REA) are useful to determine the source of an outbreak, but more reproducible and discriminatory methods such as Southern hybridization and pulsed field gel electrophoresis (PFGE) may be required. When applied to some nosocomial Candida infections, multiple strains and species have been identified. Microevolution of yeast species occurs and epidemiologically related isolates may show minor pattern differences, creating uncertainty as to whether they are distinct strains. Approximately 1000 isolates of Aspergillus fumigatus from environmental and clinical sources were typed by REA probed with an A. fumigatus-specific retrotransposon-like sequence. Patients with no symptom of aspergillosis may carry several strains, whereas patients with pulmonary aspergillosis may carry one or two strains; nocosomial transmission of aspergillosis was proven in 39% of the patients studied; any given environmental strain can be infectious; the environmental population of A. fumigatus is extremely diverse and no specific niche was found in the hospital. A PCR assay was designed to target conserved 18S-ribosomal DNA (rDNA) sequences shared by most fungi and a 687 bp product was amplified from 25 medically important fungal species. Studies with blood, cerebrospinal fluid and sputum specimens from patients with mycoses indicated that the PCR assay is more sensitive in diagnosing invasive fungal infections than blood culture methods. More specific identification is obtainable with genus/species-specif c probes designed from within the PCR-amplified sequences for C. albicans, C. krusei, C. lusitaniae, Pneumocystis carinii, Cryptococcus neoformans, Aspergillus/Penicillium spp. and C. glabrata/Saccharomyces cerevisiae. A. fumigatus and A. niger were differentiated by denaturing gradient gel electrophoresis. In situ hybridization (ISH) detected a 648 bp fragment of the 18S rDNA of C. neoformans and a 568 bp fragment of the alkaline proteinase gene of A. fumigatus in tissues from experimentally infected animals. In ISH, the entire process can be automated, making this procedure rapid and easy. The difficulty in establishing a diagnosis of invasive candidiasis has prompted the quest for a clinically useful PCR test for candidaemia. The universal fungal oligonucleotide primer pair, ITS3 and ITS4, amplifies portions of the 5.8S ad 28S rDNA subunits, and the ITS2 region. Although rRNA genes are highly conserved, the ITS regions are distinctive. DNA probes were designed from ITS2 that were specific for 16 different Candida species. Simple, rapid sample preparation was suitable for PCR analysis of BacT/Alert blood culture bottles. Sample preparation, PCR, and EIA detection of the amplicon from five different Candida species was accomplished in 7 h, 2.5 days sooner than by conventional culture methods. As well as saving time, minor yeast species among a major species, or among bacteria, were simultaneously detected. PCR-EIA using a microtitration plate format had sensitivity 10-times greater than that obtained with ethidium bromide-stained agarose gels. Taqman combines in one step PCR, probe hybridization, and fluorescent signal generation. Taqman PCR had sensitivity equivalent to PCR-EIA and required only 5 h, including sample preparation.

First isolation of Cryptococcus neoformans var. gattii, serotype C, from the environment in Colombia.

Abstract: The natural habitat of Cryptococcus neoformans var. gattii, serotype B in the environment was established by Australian investigators who demonstrated its association with species of Eucalyptus. The aim of the present study was to search for the habitat of this variety in a city of Colombia, where clinical cases due to this variety occur with great frequency. For a period of 5 months detritus, vegetable material and air samples in and around 68 almond trees (Terminalia catappa) located in the city were studied. C. neoformans var. gattii serotype C was the only variety isolated from two of the 68 trees sampled. These treeswere positive for 4 of the 5 months during which they were studied. From the first positive sample kept under refrigeration, it was possible to isolate the fungus up to 3 months later. This is the first report of the isolation of serotype C from the environment. More studies are required in order to establish the ecological significance of this finding.

Detection of PCR-amplified fungal DNA by using a PCR-ELISA system.

Abstract: In order to speed up and standardize polymerase chain reaction (PCR)-based detection of medically important fungi in clinical samples we established a combination of commercially available kits for DNA extraction, PCR amplification and detection of the amplicons. The PCR plate assay proved to be as sensitive and specific as our previously published assay (5 cfu ml(-1) blood). Moreover, in a selected group of patients, all patients with proven and probable invasive fungal infection were found to be PCR-positive. Thus the PCR plate assay was found to be a sensitive, technically simplified and standardized method with potential for adaptation to automation.

First report of Cryptococcus laurentii meningitis and a fatal case of Cryptococcus albidus cryptococcaemia in AIDS patients

Abstract: We report the first case of Cryptococcus laurentii meningitis and a rare case of Cryptococcus albidus cryptococcaemia in AIDS patients. Both infections were treated with amphotericin B and flucytosine. The C. laurentii meningitis was controlled after 2 weeks of treatment with no evidence of infection 20 months later. The patient with C. albidus cryptococcaemia, despite the amphotericin B/flucytosine combination therapy, died on the 14th day of treatment. The minimum inhibitory concentrations (MICs) for C. laurentii, as determined by Etest on RPMI 1640 agar, were 0.25 microg ml(-1) of amphotericin B, 1.25 microg ml(-1) flucytosine, 4 microg ml(-1) fluconazole, 0.50 microg ml(-1) itraconazole and 1.0 microg ml(-1) of ketoconazole. The MIC of amphotericin B for C. albidus was 0.5 microg ml(-1), flucytosine 1.25 microg ml(-1), fluzonazole 4 microg ml(-1), itraconazole 0.5 microg ml(-1) and ketonazole 0.25 microg ml(-1). The agreement of the amphotericin B MIC values obtained in antibiotic medium 3 by the broth microdilution method, with those obtained on casitone medium by Etest, was within a two-dilution range for both isolates. C. laurentii may cause meningitis and may also involve the lungs in AIDS patients.

Molecular typing of Malassezia species with PFGE and RAPD.

Abstract: The currently recognized seven species of Malassezia all have different karyotypes which do not vary intraspecifically, except in M. furfur whichdisplayed two different karyotypes. In contrast, random amplified polymorphic DNA (RAPD) typing showed the presence of genetic variation in all species. It is concluded that karyotype analysis is useful for species identification, and RAPD typing can be used in epidemiological investigations.

Purification and Characterization of a 315 Kda Keratinolytic Subtilisin-Like Serine Protease from Microsporum Canis and Evidence of Its Secretion in Naturally Infected Cats

Abstract: A keratinolytic protease, secreted as the major component by a feline clinical isolate of Microsporum canis cultivated in a minimal medium containing cat keratin, was purified by affinity chromatography on bacitracin-agarose and gel filtration. The apparent molecular mass of the enzyme was 31·5 kDa and the pI was 11·8. The enzyme was not glycosylated and its first 15 N-terminal amino acids showed numerous similarities with other fungal subtilisins. The optimum pH was around 9 whileinactivation of the enzyme was reversible at pH4, but not at pH 11. The enzyme was stable at 37 °C with an apparent optimum temperature around 55 °C. PMSF, soybean trypsin inhibitor (SBTI) and chymostatin strongly inhibited the proteinase. The highest affinity (Km of 0·37 mm) and physiological efficiency (kcat/Km) were obtained for the synthetic substrate N-Suc-Ala-Ala-Pro-Phe-p-nitroanilide. These results indicate that the keratinase belongs to the subtilisin-like serine protease family. Purified rabbit immunoglobulins G prepar...

In vitro activity of voriconazole against selected fungi

Abstract: Fifty-nine isolates consisting of 14 genera and 33 species of ascomycetes, basidiomycetes, and zygomycetes were tested against amphotericin B, fluconazole, itraconazole and voriconazole using an in vitro modified macrobroth dilution procedure based upon the NCCLS M27-A standard method for yeasts. The triazoles voriconazole and itraconazole had similar MIC values, except for Acremonium alabamensis, A. strictum, Fusarium oxysporum, F. solani and Wangiella dermatitidis, which had substantially lower voriconazole MIC values. Voriconazole MIC values were lower than those for itraconazole for the 17 species of Trichosporon tested. Fluconazole had high MIC values, often greater than 128 microg ml-1.

Pathogenicity of basidiospores of Filobasidiella neoformans var. neoformans.

Abstract: Basidiospores of Filobasidiella neoformans var. neoformans (progeny of Cryptococcus neoformans MT 100.1 x VR 45980) were able to induce cryptococcosis in Swiss albino mice if inoculated by intraperitoneal injection, nasal instillation or nasal spraying. The latter method, with the aid of a jet nebulizer, was first adopted to imitate the natural entrance of infectious particles. Using this method the small number of basidiospores (7 x 10(3)) could induce cryptococcosis in mice, while the higher number of the parental laboratory-grown yeast cells (1.5 x 10(6)) did not produce infections. By nasal instillation Cyclophosphamide (Cy)-treated mice were more susceptible to the basidiospores, showing 80% cryptococcosis (eight of 10). Seven of the eight infected mice had disseminated cryptococcosis. Immunocompetent mice were more resistant to basidiospore infection than Cy-treated mice, as 40% of that group developed only pulmonary cryptococcosis; none had disseminated infection. Thus, we propose that basidiospores are one form of the infectious propagules of F. neoformans var. neoformans which can cause cryptococcosis, particularly in immunocompromised people.

Standardization of antifungal susceptibility testing and clinical relevance.

Abstract: Standardization of antifungal susceptibility testing became essential to overcome the problem of interlaboratory variability and to determine the clinical relevance of in vitro data. This evolving process began for the yeasts and consequently broth macrodilution and microdilution methods (NCCLS M27 document) have been developed. These tests may not be useful for testing all organism-drug combinations or be the most convenient techniques for routine use in the clinical laboratory. Different alternatives to the NCCLS methods are currently under investigation. The identification of standard guidelines for antifungal susceptibility testing has reduced interlaboratory variability and further progress has been achieved with the determination of tentative interpretive breakpoints for certain drug-yeast combinations. However, these breakpoints are not adequate for interpretations of MICs for fungi-drug combinations beyond the setting for which they were determined. The NCCLS Subcommittee has also generated data for proposed testing guidelines for the moulds.

Antifungal activity of a new triazole, voriconazole (UK-109,496), compared with three other antifungal agents tested against clinical isolates of filamentous fungi.

Abstract: Voriconazole is a new triazole antifungal agent with potent activity against yeast and moulds. We investigated the in vitro activity of voriconazole, itraconazole, amphotericin B and 5-flucytosine against 51 clinical isolates of filamentous fungi. Overall, voriconazole was active (MIC50, 0.5 mg l(-1) and MIC90, 8 mg l(-1)) against these mould isolates. Voriconazole was most active against P. boydii (MIC50, 0.12 mg l(-1)) and Aspergillus spp. (MIC90, 0.5 mg l(-1)) and least active against Fusarium spp. (MIC90, 8 mg l(-1)) and Rhizopus spp. (MIC50, 8 mg l(-1)). Voriconazole was more active than amphotericin B against Aspergillus spp. and P. boydii. By comparison with itraconazole, voriconazole was more active against all isolates except Rhizopus spp. Based on these results, voriconazole has promising activity against commonly encountered isolates of filamentous fungi and its clinical usefulness should be established by further studies.

Isolation of a Paracoccidioides brasiliensis strain from the soil of a coffee plantation in Ibiá, State of Minas Gerais, Brazil

Abstract: Paracoccidioides brasiliensis has rarely been isolated from its habitat in rural areas. In orderto investigate the hypothesis that human infection with this fungus is linked to coffee plantations (Coffea arabica), material was collected monthly over a period of 1 year from farms in the town of Ibia, State of Minas Gerais, Brazil. A total of 760 samples of soil, coffee leaves and fruits was cultured and inoculated into mice. A fungus isolated from the liver of a mouse inoculated with soil showed temperature-dependent dimorphism and in vitro mycelium and yeast phases characteristic of P. brasiliensis. Yeast cells of this fungus caused disseminated infection after intraperitoneal inoculation in Wistar rats from which the fungus was re-isolated. An antigen reacting with sera from patients with paracoccidioidomycosis was obtained from this P. brasiliensis strain; antigenic identity with strain 339 and with four other P. brasiliensis strains was detected by gel immunodiffusion. However, when the exo-antigen was...

Mycological control and surveillance of biological waste and compost

Abstract: This paper presents some recent developments regarding current work on hygienic aspects, in particular the presence and dispersion of fungi (e.g. Aspergillus fumigatus), of biological waste and compost.

Fungicidal and binding properties of the natural peptides cecropin B and dermaseptin.

Abstract: In vitro fungicidal properties of cecropin B and dermaseptin were explored using non-germinating and germinating conidia from Aspergillus flavus, A. fumigatus, A. niger, Fusarium moniliforme and F. oxysporum. Cecropin B produced LD50 values for germinating A. flavus, A. fumigatus and A. niger conidia of 3·0, 0·5 and 2·0 μ m, respectively, while dermaseptin gave LD 50 values of 4·0, 0·05 and 2·0 μ m, respectively. Cecropin B produced an LD 50 value of 0·2 μm for non-germinating F. moniliforme and F. oxysporum conidia, while dermaseptin did not reduce either as much as 50% at any level tested. LD50 levels for CB were 0·2 and 0·1 μm, respectively, for germinating F. moniliforme and F. oxysporum conidia. Dermaseptin was less effective, giving LD50 values for germinating F. moniliforme and F. oxysporum conidia of 0·3 and 0·8 μ m, respectively. Neither peptide reduced conidial viabilities of non-germinating Aspergillus spp. Physicochemical studies indicated cecropin B and dermaseptin bound to ergosterol and cho...

Pathogenicity of basidiospores ofFilobasidiella neoformansvar.neoformans

Abstract: Basidiospores of Filobasidiella neoformans var. neoformans (progeny of Cryptococcus neoformans MT 100.1 x VR 45980) were able to induce cryptococcosis in Swiss albino mice if inoculated by intraperitoneal injection, nasal instillation or nasal spraying. The latter method, with the aid of a jet nebulizer, was first adopted to imitate the natural entrance of infectious particles. Using this method the small number of basidiospores (7 x 10(3)) could induce cryptococcosis in mice, while the higher number of the parental laboratory-grown yeast cells (1.5 x 10(6)) did not produce infections. By nasal instillation Cyclophosphamide (Cy)-treated mice were more susceptible to the basidiospores, showing 80% cryptococcosis (eight of 10). Seven of the eight infected mice had disseminated cryptococcosis. Immunocompetent mice were more resistant to basidiospore infection than Cy-treated mice, as 40% of that group developed only pulmonary cryptococcosis; none had disseminated infection. Thus, we propose that basidiospores are one form of the infectious propagules of F. neoformans var. neoformans which can cause cryptococcosis, particularly in immunocompromised people.

Increased incidence of Trichophyton tonsurans tinea capitis in Ontario, Canada between 1985 and 1996

Abstract: In Canada, since the 1990s, Trichophyton tonsurans has emerged as the main cause of tinea capitis. Prior to this the more common agents were T. verrucosum, Microsporum canis and M. audouinii. Over the past few years the incidence of T. tonsurans has increased such that in 1985 and 1996 the cases of mycologically confirmed tinea capitis due to T. tonsurans were 9% and 76%, respectively. The epidemic of T. tonsurans has reduced the role of Wood's lamp in diagnosis of tinea capitis. The age distribution of tinea capitis due to T. tonsurans closely matches that of mycologically confirmed tinea capitis, being most common in children under 14 years of age. There is no significant sex difference in children who develop T. tonsurans tinea capitis; however, subjects are significantly more likely to live in urban than rural areas. This should provide guidance regarding where to concentrate health resources and deliver patient/parent education to combat this epidemic of tinea capitis.

Dermatophytes and host defence in cutaneous mycoses.

Abstract: Dermatophytosis is the infection of keratinized tissues such as hair, nails and the stratum corneum of the skin by dermatophyte fungi. These fungi are onygenalean anamorphs and anamorphic species belonging to the genera Trichophyton, Microsporum and Epidermophyton. An important characteristic of the dermatophytes as parasites is their restriction to the dead keratinized tissues, except in rare cases where the patient is immunosuppressed. In contrast to many fungi, including normally non-pathogenic species, which can invade systemically in severely immunocompromised (e.g. neutropenic) patients, dermatophytes appear to be unable to cause systemic infection in this population. Thus, these fungi appear to have an unique interaction with the immune system. A better understanding of this interaction will contribute significantly to our knowledge of mammalian host defences.

Cryptococcus neoformans: virulence and host defences.

Abstract: Cryptococcus neoformans represents a model organism for the study of virulence and the host response. In this discussion, there is a focus on the genetic, molecular, and biochemical aspects of C. neoformans as it interacts with the host. Investigations into direct and indirect virulence phenotypes are now possible. The molecular aspects of two major virulence factors, capsule and melanin, are characterized. Yeast polyol metabolism through mannitol is examined as a potential biochemical pathway for virulence. The concept of C. neoformans differentially expressed genes within the host or in response to certain environmental cues can be used indirectly to identify potential virulence genes. However, despite significant progress in molecular pathogenesis with C. neoformans, the future of research in this area will require a certain critical mass of investigators to help share in the developmental costs which continue to occur.

Antibody and/or cell-mediated immunity, protective mechanisms in fungal disease: an ongoing dilemma or an unnecessary dispute?

Abstract: Historically there has been controversy on the relative importance of antibody- and cell-mediated immune responses in the protection against fungal pathogens The controversy was fuelled by the difficulties encountered in obtaining consistent results with polyclonal antibody experiments and I inducing long-lasting immune protection by vaccinations with induce stron cell-mediated responses Recent studies indicate that both antibody- and cell-mediated immune responses can contribute to host protection against Candida albicans and C neoformans At the present time the major issue is not the relative importance of antibody- and cell-mediated immune responses but rather, the mechanisms by which the two arms of the immune system function and cooperate

Cryptococcus neoformans isolated from human dwellings in Rio de Janeiro, Brazil: an analysis of the domestic environment of AIDS patients with and without cryptococcosis.

Abstract: One hundred and fifty-four human dwellings in the metropolitan area of Rio de Janeiro, Brazil were studied. A total of 824 samples of indoor dust, outdoor soil and avian droppings were collected. Cryptococcus neoformans var. neoformans was isolated from 20 (13%) dwellings, comprising five (15.6%) of 32 dwellings of patients with AIDS-associated cryptococcosis; four (8.9%) of 45 dwellings of patients with AIDS but without cryptococcosis; and 11 (14.3%) of 77 dwellings of apparently healthy individuals (P>0.05). The principal factor associated with domiciliar contamination by C. neoformans var. neoformans was the presence of avians in the domestic environment or nearby the home. Cryptococcosis was more frequent among AIDS patients residing in dwellings from which C. neoformans var. neoformans was isolated than among AIDS patients from whose domestic environment the fungus was not demonstrated by the methods used (odds ratio (OR)=2.05). These findings suggest that the distribution of C. neoformans var. neoformans in Rio de Janeiro is not restricted to the classically known biotopes as well as reinforce the possibility of exogenous infection in opportunistic cryptococcosis, including exogenous infection acquired in the domestic environment.

Development of vaccines and their use in the prevention of fungal infections

Abstract: Vaccine approaches to infectious diseases are widely applied and appreciated. Disciplines such as bacteriology and virology have a rich history of successful vaccine development. The complexity of eukaryotic systems presents additional challenges to the development of vaccines against them. These challenges are being met in the fields of parasitology, and are being revisited for application in oncology. Vaccine opportunities exist in medical mycology. The National Institute of Allergy and Infectious Diseases has held a series of workshops in medical mycology where the need to develop vaccines for fungal diseases was noted and where important opportunities were discussed. Major advances in vaccinology and the technology of antigen preparation and delivery have increased feasibility and heightened interest. The recent epidemic of coccidioidomycosis in the American Southwest has demonstrated the need for developing a vaccine as an effective preventive measure for those living in and for those who subsequently move into regions with the endemic mycoses. The XIIth Congress of the International Society for Human and Animal Mycology included a symposium that summarized new vaccination strategies for selected fungi: Candida albicans, Coccidioides immitis, and Trichophyton verrucosum. The goal of the present summary is to provide representative examples of continuing efforts relating to vaccine development within the medical mycological community highlighting Blastomyces dermatidis, Cryptococcus neoformans, Histoplasma capsulatum, Paracoccidioides brasiliensis, and Pythiumn insidiosum.

The effects of Malassezia on pro-inflammatory cytokine production by human peripheral blood mononuclear cells in vitro

Abstract: Malassezia spp., the causative agents of pityriasis versicolor, are members of the normal human cutaneous microflora. Utilizing a combination of both enzyme-linked immunosorbent assay (ELISA) and bioassay, we have investigated the ability of both formalin-preserved and viable Malassezia (serovars A, B and C) to modulate pro-inflammatory cytokine (IL-6, IL-1beta and TNF-alpha) release by human peripheral blood mononuclear cells (PBMNC) in vitro, over a 48-h co-incubation period. The results demonstrated that formalin-preserved Malassezia (serovars A, B and C) at mid-exponential phase were generally able to induce a pro-inflammatory cytokine response at a yeast cell to PBMNC ratio of 1:1. In addition, the results consistently demonstrated that at a yeast cell to PBMNC ratio of 20:1, formalin-preserved Malassezia, irrespective of serovar, growth phase or PBMNC donor, were capable of significantly (P<0.05) decreasing the release of both immunochemical IL-6 and IL-1beta plus bioactive IL-1beta and TNF-alpha below that of unstimulated culture medium control values. This was apparent following 24- and 48-h co-incubation times, where maximal cytokine production was detected after 24 h. Similar results were obtained for the effect of viable Malassezia on pro-inflammatory cytokine release by PBMNC. Our results suggest that a possible inhibitory component, present perhaps within the cell wall of Malassezia, was responsible for this depressive effect on pro-inflammatory cytokine production.

PCR-based identification of clinically relevant Pseudallescheria/Scedosporium strains

Abstract: Pseudallescheria boydii (anamorph: Scedosporium apiospermum) and S. prolificans can cause severe infections in both the immunocompromized host and accidentally injured people. A species-specific polymerase chain reaction (PCR) enabling detection and discrimination of these fungi has not been available to date. In view of the difficult treatment especially of S. prolificans infections, a PCR-based detection system has been developed. Based on results of partial sequencing of ribosomal DNA, Scedosporium DNA could be amplified, either at the genus or at the species level. Hybridization probes for the identification of the PCR-products were established.

In vitro evaluation of terbinafine and itraconazole against dematiaceous fungi

Abstract: Two hundred and three isolates representing 15 species of filamentous ascomycetes were evaluated against terbinafine and itraconazole using a modification of the NCCLS M27-A standard reference method for yeasts. The MIC ranges and geometric means were similar, although terbinafine tended to have the lowest values. The loculo-ascomycete clade tested had consistently low MIC geometric mean values for its members, ranging from 0·03 to 0·17 μg ml-1 for terbinafine and 0·03–0·37 μg ml-1 for itraconazole.

Serodiagnosis of histoplasmosis, paracoccidioidomycosis and penicilliosis marneffei; current status and future trends.

Abstract: Effective serodiagnosis of systemic fungal infections is of increasing importance, particularly with regard to the identification of infection with Histoplasma capsulatum, Paracoccidioides brasiliensis and Penicillium marneffei. Methodology has been based either around antibody or antigen detection, although there is clear overlap between the two. Antibody-based detection systems for the diagnosis of histoplasmosis, paracoccidioidomycosis and penicilliosis marneffei have now begun to incorporate a range of highly purified and well-characterized antigens, in contrast to the situation of a few years ago when relatively crudepreparations derived from either whole cells or culture filtrate were used. The application of such antigens offers improvements in reproducibility and specificity, although the detection of meaningful antibody responses in immunosuppressed individuals remains a problem. Partly as a consequence of this a great deal of attention has focused on the development of antigen detection assays, ...