The echinocandin (candin) class of antifungal drugs inhibit β – 1,3 – glucan synthase and block synthesis of β – 1,3 – glucan, an important polysaccharide in fungal cell walls. Candins are used widely for treatment of systemic infections caused by Candida and Aspergillus because of their high potency and low toxicity to humans. The incidence of candin resistance has been rare compared to that of azole resistance, although candin-resistant clinical isolates of C. albicans, C. glabrata, C. krusei and C. tropicalis have been reported in the USA and Europe in recent years. These isolates possess hundred – fold higher MIC values for candins than sensitive strains, as well as candin-resistant β – 1,3 – glucan synthase activities. Their candin resistance is associated with amino acid substitutions in the echinocandin resistant region (Ech^R) of the FKS gene that encodes a catalytic subunit of the β – 1,3 – glucan synthase. However, the effect of these amino acid substitutions on the drug-protein interaction and the molecular basis for the resistance is unknown. The exposure of fungi to candin drugs induces stress responses that activate networks involving transcriptional regulators and components controlling signal transduction of the pathways responsible for maintenance of fungal cell wall integrity. The fungal cell wall is still an attractive drug target and further investigation into the mechanisms of candin resistance and structural analysis of the β – 1,3 – glucan synthase protein complex will facilitate the development of broad spectrum inhibitors of fungal cell wall synthesis.

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Pneumocystis (Pc) jirovecii causes severe interstitial pneumonia in patients with immunodeficiency, in whom this fungus adheres with type – I alveolar epithelial cells. Therefore, it is important to perform quick diagnosis and treatment for Pc pneumonia (PcP). In general, a combination of two antifolate agents, sulfamethoxazole (inhibition of dihydropteroate synthase (DHPS)) and trimethoprim (inhibition of dihydrofolate reductase), is the first choice for PcP treatment, and pentamidine or atovaquone (inhibition of cytochrome b) are the alternative reagents for the therapy. Amino acid substitutions of drug – binding sites in DHPS shown in genotypic analysis have been reported to be associated with failures of prophylaxis ⁄ treatment or severe mortality for PcP, while there is another article showing a negative relationship between the DHPS mutations and poor prognosis. Drug sensitivity tests using the phenotypes as well as genotypes are necessary, although it is difficult to culture Pc. This review focuses on the relationship between mutations of drug – targeting molecules and treatment failure based on original data and other reports. In addition, trials of phenotypic analyses for Pc are described as promising investigations.

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The present article describes our studies to know the usefulness of in situ hybridization (ISH) to identify various kinds of mold observed in tissue sections and ⁄ or cytological preparations from the lesions of patients with invasive fungal infection. To establish the precise procedure for ISH in formalin – fixed and paraffin-embedded sections, various pretreatments were attempted. The condition finally chosen is written here providing a favorable outcome regarding to both intensity and specificity of signals on outline of molds observed in the tissue sections when specimens were treated with both heat and proteinase K and, solutions were adjusted to higher pH value.
Therefore, usefulness of promising probes, two each DNA and peptide nucleic acid (PNA) were verified with a favorable pretreatment condition, using lungs of mice experimentally infected and ⁄ or those obtained from autopsies with invasive mold infection. As the result, DNA probes targeting alkaline proteinase (ALP) gene and retrotransposon Afut – 1 gene of Aspergillus fumigatus showed specific signal intensity for the Aspergillus species and A. fumigatus, respectively. PNA probes for Candida albicans and the Fusarium species also showed satisfactory specificity. We wish to emphasize that ISH can be a valuable tool to identify medically important molds in formalin – fixed and paraffin – embedded tissue sections or cytological preparations.

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Tinea corporis and the tinea pedis model in guinea pig with Trichophyton mentagrophytes are well established models of dermatophytoses. We attempted to provide animal infection models for T. tonsurans, endemic in Japan, and Malassezia restricta, an important pathogenic factor in seborrhoeic dermatitis, by utilizing the tinea corporis model. An inoculum of the organisms was applied to the back skin of male guinea pigs. T. tonsurans infected animals showed follicular inflammation mimicking those seen in humans. Interestingly, anthropophilic T. tonsurans showed a high infection rate in animal skin. Meanwhile, a single application of M. restricta, as well as consecutive applications to the surface of the skin without any pretreatment, succeeded in producing scales mimicking seborrhoeic dermatitis, but application of the pathogens after the tape stripping of the stratum corneum failed to induce infection. These models using guinea pigs were considered to be useful for studying the pathogenesis of, and evaluating therapies for, T. tonsurans infection and seborrhoeic dermatitis.

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Animal mycosis, particularly deep mycosis, is one of the most challenging conditions encountered by veterinarians. Pathogens causing mycotic infections in animals include fungi such as Cryptococcus neoformans, Candida spp., and Aspergillus spp. The antifungal drugs used for the treatment of deep mycoses in animals as well as humans are polyenes and azoles. However, the sensitivity of clinical isolates obtained from animals toward these drugs has rarely been assayed. In this study, the antifungal activities of itraconazole and voriconazole against clinical isolates of C. neoformans, Candida spp., and A. fumigatus isolated from animals with mycoses were examined using the broth microdilution method performed according to the guidelines provided by the Clinical and Laboratory Standards Institute. The minimum inhibitory concentrations (MICs) of itraconazole toward the C. neoformans, Candida spp., and A. fumigatus isolates were 0.125 – 1, 0.125 – 2, and 0.25 – 2μg⁄ml, respectively, and those of voriconazole were 0.0625 – 0.5, ≤0.0313 – 0.0625, and 0.0625 – 1μg⁄ml, respectively. The results of the MIC analyses implied that the fungal isolates obtained from infected animals exhibit an equivalent degree of susceptibility to itraconazole and voriconazole, as is observed in the case of isolates obtained from humans. The appropriate antifungal therapeutic strategy for the treatment of mycoses in animals must be selected taking into consideration the host immune status and organ function as well as the in vitro sensitivity of the pathogens to antifungal drugs.

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Seven cases of cryptococcus meningitis in a tertiary care hospital from 2004 – 2007 were reviewed. 85.7% of the patients had headache as their predominant clinical feature. The spectrum of CT ⁄ MR findings ranged from no abnormality, basal ganglion lesion, to intracerebral and intraventricular granulomas. Findings of cerebrospinal fluid(CSF)cytology and biochemistry analysis were inconclusive. Patients were diagnosed by India ink(100%), CSF cryptococcal antigen detection(100%), and CSF culture in 6(85.7%). With the exception of two patients, co-morbidities associated were HIV, diabetes mellitus, and idiopathic CD4 + lymphocytopenia. Six patients were successfully treated with amphotericin B and discharged. A high index of clinical suspicion and laboratory diagnosis achieved early can reduce the overall morbidity and mortality among patients with cryptococcosis.

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A total of 155 sporotrichosis cases examined in Nagasaki prefecture, including 138 cases which had been previously reported and 16 examined from 2002 to 2007, were surveyed and compared. No significant differences were found between 143 cases from 1951 to 2001 and 12 of these from 2002 to 2007 in sex or the affected regions of the body. Males and females were equally affected. The lesions were frequently seen on the face(28.2%, 25.0%)and upper limbs(62.1%, 66.7%). Fixed type(62.1%)was much more frequent than the lymphocutaneous type(37.9%)from 1951 to 2001, but in recent year there was an equal number of two types. The rate of patients over 50 years of age increased from 72.1% to 91.7%, and of note was that there were no patients under 10 years and only 2 was noted over 90 examined after 2002. A remarkable increase in the number of cases in Shimabara area(26.8% → 33.3%). Prior to 1994, potassium hydroxide(KI)was used as therapy in most cases(99.1%), but after 1995, itraconazole was used in over 50% of cases and those treated with terbinafine also increased. KI was used in about 20% of case after 1995 decreasing from then to 2000(8.3%), recently, its use has again increased(25.0%). The period of treatment until cure was achieved for itraconazole was 17.0 weeks and for KI was 10.9 weeks.

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Malassezia-positive smears can be recognized from otitis externa, however, there are few references in the literature to the relation between Malassezia and otitis externa. Therefore, the bacterial and clinical characteristics of 72 cases (63 patients) with otitis externa were investigated at the Department of Otorhinolaryngology, Takinomiya General Hospital to analyze this. Thirty-seven cases were bacterial otitis externa, 20 cases were fungal otitis externa, and 15 cases were etiological agents unknown in this study. The causative organisms in fungal otitis externa were the genera Aspergillus (10 cases), Malassezia (5) and Candida (5), respectively. We suspected that 5 cases were caused by Malassezia because Malassezia cell counts were greater than 10 per field (× 400), and a large number of Malassezia were isolated from all cases. In these cases, many squamous epithelial cells were observed by direct examination, and cells from the middle or basal layer of the ear canal were also recognized in three cases. Therefore, accelerated turnover of epidermal cells of the ear canal was suggested. The main symptoms were itching and fullness in the ear, with observations of redness and erosion in objective deterioration, and we felt that these conditions were similar to seborrheic dermatitis (SD). In addition, these five cases were confirmed as fungus-related otitis externa by their improvement with antifungal agents.

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[in Japanese]

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