Phagocytosis, oxidative metabolism and phagolysosomal pH of rat alveolar macrophages (AM) were studied at different points of time after challenge with Cryptococcus neoformans. Phagocytosis was evaluated using a fluorescent quenching technique which distinguishes between attached and ingested organisms. The nitroblue tetrazolium (NBT) test was used as an indirect measurement of the oxidative metabolism of the phagocytes. The pH of the phagolysosomes was measured using a cytofluorometric technique. Both the attachment and ingestion of serum opsonized C. neoformans by AM were slow during the first hours of incubation, but were considerable after 24 h. The oxidative metabolism of Am challenged with the yeast was insignificant during the first hour, but reached high levels after 24 h. Most phagolysosomes in AM with ingested cryptococci had a pH < 5.5. Our results indicate that these AM defence mechanisms, although poor during the first hours after exposure to the yeast, are of significance after 24 h. Thus, in the immunocompetent host the AM should prevent the dissemination of C. neoformans from the lungs.
(1 --> 3)-beta-D-glucan, a characteristic fungal molecule, is known to increase in blood in invasive candidiasis, aspergillosis and cryptococcosis. This report shows that the plasma glucan concentration was also elevated in four patients infected with Fusarium, Trichosporon beigelii, Saccharomyces cerevisiae and Acremonium. In three of the patients, the elevation preceded positive blood cultures by 5-17 days, and in one of them it even preceded the onset of fever by 6 days. In a fourth patient, the glucan level decreased with clinical improvement. Plasma (1 --> 3)-beta-D-glucan determination appears to be useful also for diagnosis and follow-up of these unusual deep mycoses.
Mice treated with cyclophosphamide were infected intravenously with titrated doses of Trichosporon asahii TIMM3140 or TIMM3144. Mortality and survival time correlated with inoculum size and fungal growth was found in the lung, heart, liver, spleen, kidney and brain. Fluconazole and amphotericin B prolonged survival time and reduced the CFU in the kidney, with fluconazole being superior to amphotericin B. The serum level of (1-->3)-beta-D-glucan was reduced by antifungal therapy and correlated well with the CFU/organ in kidney, but not CFU/ml in blood.
A retrospective study was conducted in France to investigate Fusarium infections which are now recognized as emerging opportunistic infections. The clinical and mycological findings for 31 cases diagnosed between 1984 and 1993 by members of the French Groupe d'Etudes des Mycoses Opportunistes were analysed. All suffered from haematological disease, most often acute leucaemia (n = 19). Twenty-two had received cytostatic chemotherapy and ten had undergone bone marrow transplantation. Prolonged aplasia and pancytopenia were present in 18 and 11 patients, respectively. Skin (61%) and blood (42%) were the sites most frequently involved. Fusarium solani (n = 7), Fusarium oxysporum (n = 7), Fusarium verticilloides (n = 7) were the species most frequently isolated. Nine antifungal treatments were used, associated with colony-stimulating factors in five cases. None was unambiguously superior to all the others. The overall mortality was 51.6% with a specific mortality greater than or equal to 25.8%. The disseminated form of the infection was associated with poor prognosis (P < 0.02) whereas improving granulocyte count improved prognosis (P < 0.001). More aggressive cytostatic regimens used for patients with haematological malignancies have favoured the emergence of Fusarium infections. As prognosis is closely correlated with neutrophil recovery, the promising results obtained with the use of colony-stimulating factors should be further evaluated.
Paranasal sinus mycoses are endemic in rural populations of northern India. To study host-parasite interactions, we developed an animal model of paranasal sinus mycoses. After failure in small animals such as mice and rats, we used New Zealand white rabbits weighing 2.5-3 kg. Inoculum sizes consisted of 0.75-1.0 x 10(8), 0.75-1.0 x 10(7), 0.75-1.0 x 10(6) conidia of a clinical isolate of Aspergillus flavus. The inoculum was injected at a spot 0.5 cm in front of the alveolar process of the maxilla and 0.5 cm below the maxillary process of frontal bone and vertically to a depth of 0.5 cm across the bone directly into the nasal sinus. Paranasal sinus mycoses proven by culture and histopathology developed in 67% of animals injected with 0.75-1.0 x 10(8) conidia and 17% of animals with 0.75 x 10(7) conidia. No lesions were found in the group injected with 0.75-1.0 x 10(6) conidia. Precipitating antibody against culture filtrate antigen was found in rabbits with paranasal sinus mycoses. Therefore, rabbits can be used as an animal model to study paranasal sinus mycoses.
The glycoprotein gp43 from Paracoccidioides brasiliensis is the main antigenic component in paracoccidioidomycosis (PCM) because it is recognized by 100% of PCM patients. It has also been shown that different fungal strains produce gp43 with at least four isoform profiles. In this study, different isoform profiles from gp43, with pIs ranging from 5.8 to 8.5, were affinity purified from various P. brasiliensis (B-339, S.S., 1925 and I9) exoantigens. Because of the isoform heterogeneity, we questioned whether those isoform profiles could be similarly recognized by acute or chronic PCM patients. By using a specific and sensitive method for detection of human IgG anti-gp43 antibodies, the monoclonal antibody capture immunoassay, we report that not all gp43 isoform profiles are equally recognized in PCM sera when anti-gp43 MAb 17c was employed as capturing antibody. Our results showed that recognition of pI8.5 gp43 isoform was significantly lower for both acute (56%) and chronic patients (71%), compared with gp43 isoforms from the standard strain B-339. On the other hand, when anti-gp43 MAb 8a, which recognizes a different antigenic epitope was used to capture the different gp43 isoform profiles, all patient's sera reacted similarly. The results described suggest that not all the antigenic epitopes expressed by gp43 are equally present in all P. brasiliensis strains.
In 1991, 1208 cases of coccidioidomycosis were reported to the California Department of Health Services, compared with an annual average of 450 during 1986-90. We conducted a study in Tulare County to define the epidemiology of the disease and identify risk factors for severe disease, focusing on the epidemic period September 1991-December 1991. To identify cases, we used data from the Coccidioidomycosis Serology Laboratory at the University of California, Davis, other laboratories, and the Tulare County Health Department's coccidioidomycosis reporting system. We compared patients who were hospitalized with those who were not to determine risk factors for severe disease. We identified 128 cases of acute coccidioidomycosis diagnosed between 1 September and 31 December 1991 (attack rate 41/100,000); south central Tulare County had the highest attack rate. Thirty-five (27%) case-patients were hospitalized. Male sex (relative risk (RR) 2.5, 95% confidence interval (CI) 1.2-5.0), black people and Asian races (RR 4.8, 95% CI 2.4-9.6), and age > or = 20 years (RR 8.3, 95% CI 1.2-57.4) were univariately significant and remained independently associated with hospitalization in multivariate analysis. The 1991 Tulare County outbreak of coccidioidomycosis was part of a much larger outbreak that began in California during 1991 and continued through 1993. The outbreak was preceded by an unusually rainy spring. Although dust reduction measures during times of increased coccidioidomycosis incidence can help reduce exposure, definitive control awaits the development of a safe, effective vaccine.
(1997). Comments on Cladophialophora bantiana. Journal of Medical and Veterinary Mycology: Vol. 35, No. 2, pp. 151-151.
The basidiomycete Coprinus sp. was isolated repeatedly from bronchial secretions and bronchoalveolar lavage of a 40-year-old woman suffering from a relapse of acute lymphoblastic leukemia 5 years after she underwent autologous bone marrow transplantation. Post-mortem microbiological investigation of lung tissue revealed simultaneously growing Coprinus sp. and Aspergillus flavus. Histopathological examination of the lung demonstrated septate hyphae characteristic of both Aspergillus and Coprinus. The basidiomycete Coprinus sp. should be considered as a potential opportunistic pathogen because of its excellent growth at 37 degrees C.
The specificity of random primer R143 for Aspergillus fumigatus DNA was determined in order to test its usefulness in establishing the presence of A. fumigatus DNA in fungal cultures. When PCR reaction products of these cultures were compared with those of 21 other bacterial and fungal DNA samples, R143 proved to produce a 1346 bp band with only A. fumigatus. This band has been sequenced completely and the EcoRI restriction site was used for subsequent confirmation of PCR products. The specificity for A. fumigatus DNA was also confirmed by Southern blotting. Comparison of morphological typing of Aspergillus species in cultures with PCR using R143 on DNA isolated from these cultures showed concordance in 22 of 24 cases. In two cases there was discordance: both times PCR results showed correctly the presence of A. fumigatus, initially not detected by culture. R143 is an A. fumigatus specific random primer, with potential for use in detection of A. fumigatus DNA in clinical specimens.
Albendazole has in vitro activity against Cryptococcus neoformans and reduced in vitro activity for albendazole when compared with Candida albicans. The major metabolite of albendazole, albendazole sulphoxide showed no in vitro activity against isolates of either fungus. Immunocompetent mice infected intravenously (i.v.) with C. albicans were treated with albendazole doses of 20-600 mg kg-1 per day in noble agar or sesame oil for per oral (PO) administration, or 80 mg kg-1 per day in DMSO for intraperitoneal (i.p.) and i.v. administration for 10 days, and were observed for survival. Mice infected with C. neoformans intracranially received albendazole in daily doses of 600 mg kg-1 prepared in DMSO (i.p.) or peanut butter/rat chow (PO) for 10 days and were observed for survival. Mortality was not different between the treated and control animals in any study. Plasma samples from uninfected mice dosed with similar formulations and doses of albendazole were analysed by HPLC for albendazole and albendazole sulphoxide. No albendazole could be detected in any sample, while concentrations of albendazole sulphoxide (286-8697 ng ml-1) were observed in all samples. These data suggest that the absence of in vivo activity for albendazole is due to rapid conversion to the inactive albendazole sulphoxide metabolite.
This paper deals with the discovery of the teleomorph of a Fonsecaea-like pathogen causing chromoblastomycosis in India. A new genus, Ascosubramania, is proposed to accommodate it.
(1-->3)-beta-D-glucan, a characteristic fungal molecule, is known to increase in blood in invasive candidiasis, aspergillosis and cryptococcosis. This report shows that the plasma glucan concentration was also elevated in four patients infected with Fusarium, Trichosporon beigelii, Saccharomyces cerevisiae and Acremonium. In three of the patients, the elevation preceded positive blood cultures by 5-17 days, and in one of them it even preceded the onset of fever by 6 days. In a fourth patient, the glucan level decreased with clinical improvement. Plasma (1-->3)-beta-D-glucan determination appears to be useful also for diagnosis and follow-up of these unusual deep mycoses.
Metarhizium anisopliae var. anisopliae (Metschnikov) Sorokin 1883 to our knowledge has never been reported as an agent of human or animal mycosis. This fungus has great importance as an agent of biological control of different pests and mosquito larvae in Colombia. It has been isolated as the aetiological agent of keratomycosis for the first time from the eye of a Colombian male.
Seventy-three Cryptococcus neoformans isolates and eight other yeast strains were studied. Fingerprints produced by priming with (GACA)4 differentiated C. neoformans from all other yeasts tested and identified the five C. neoformans serotypes. Four major bands of molecular size 800, 540, 475 and 410 bp were recognized for serotypes A, AD and D. Two of them were specific for serotype A and the other two for serotype D isolates. Serotype AD strains were identified by five different genotypic patterns in which at least one of the two bands specific for serotype A and D were present in different combinations. On repeated and simultaneously performed genotype and serotype testing of nine strains, the genotypic pattern did not change, whereas serotyping was unstable in three cases. PCR-fingerprinting using (GACA)4 as a primer proved more stable than serology in discriminating among C. neoformans serotypes A, D and AD and was able to distinguish among serotype AD strains.
Alternaria alternata, Aspergillus spp., Bipolaris spicifera, Curvularia lunata, Epicoccum nigrum and Fusarium solani were isolated repeatedly from groups of patients among 96 diagnosed with allergic fungal sinusitis (AFS). Epicoccum nigrum was obtained consistently from four patients, one of whom yielded mycelial masses consistent in morphology with E. nigrum. Fifteen of the predominant fungi recovered from air samples from selected patients' residences included the same species isolated from the mucin of its inhabitants. Air samples from other buildings, whose occupants (non-AFS individuals) complained of poor indoor air quality or of symptoms of the sick building syndrome (SBS), yielded some of the same species involved in AFS. An association of SBS with AFS was not established. Eight of the species implicated in AFS were found to colonize the surfaces of indoor construction and finishing materials at sites other than the residence of the patient. To our knowledge, this is the first report that E. nigrum can colonize nasal sinuses and cause AFS.
An expression plasmid carrying a heterologous gene fusion between the Cryptococcus neoformans actin promoter and the Escherichia coli reporter gene, LACZ, was constructed to study actin regulation in C. neoformans. Two randomly stable transformants, designated 20.6 and 20.9, were selected for further examination. Both ectopic and homologous recombination with vector insertion in tandem repeats occurred in these transformants. Transformant 20.9 carried more copies of ACTp::LACZ in its genome than 20.6 and this was reflected in expressing higher levels of beta-galactosidase activity. In vitro, these transformants showed higher levels of beta-galactosidase activity expressed when the transformants were propagated at higher temperatures (37 degrees C vs 30 degrees C). However, beta-galactosidase expression in the transformants was variable during logarithmic and stationary growth phases and this differential expression was temperature dependent. This report shows that the constitutive actin gene in C. neoformans is regulated by temperature and growth and this fact should be taken into consideration when actin expression is used as a standard to compare the expression of other regulated genes. Also, a more sensitive reporter construct will be needed for in vivo gene analysis of regulation.
Several investigators have developed monoclonal antibodies against the capsular polysaccharide of Cryptococcus neoformans which have potential therapeutic applications. Using a rat model of C. neoformans meningitis, we studied the biodistribution and pharmacokinetics of a murine anticryptococcal capsular monoclonal antibody (mAb 2H1) after intravenous and intracisternal administration. After intravenous administration of 125I-labelled 2H1 to infected rats, there was no detectable localization of 125I in the brain or cerebrospinal fluid by either gamma-camera imaging of the whole animal or organ scintillation counting. In contrast, direct intracisternal instillation of 2H1 to infected rats resulted in persistent intracranial activity. In addition, the whole body half-life of intravenously administered radio labelled mAb 2H1 was significantly reduced in infected rats compared with uninfected rats. Our observations suggest that if high central nervous system (CNS) levels of mAb are needed to achieve a therapeutic effect in human C. neoformans meningoencephalitis, direct administration of mAb into the cerebrospinal fluid or modification of the mAb to increase penetration into the CNS may be required. Furthermore, higher or more frequent dosing of mAb may be required to maintain therapeutic levels in the presence of infection. This study demonstrates the usefulness of the rat as an experimental system for studying issues related to cryptococcosis.
Onychocola canadensis is a recently recognized cause of human nail infection. We present the first four cases of onychomycosis caused by this organism among patients resident in the UK.
