In vitro study to determine the sexual compatibility of 26 Cryptococcus neoformans isolates was conducted on sunflower (Helianthus annus) seed agar. Among 26 isolates examined, 14 were originated from animals, 6 from men and 6 from environmental substrates; 17 isolates were obtained in Japan and 9 in India. These isolates were crossed with each of the 2 mating types (α and a) of Filobasidiella neoformans var. neoformans or F. neoformans var. bacillispora. All the isolates produced basidiospores, basidia, dikaryotic hyphae with clamp connections when paired with the “a” mating type of F. neoformans var. neoformans. We could not find any isolate showing sexual reproduction with “α” mating type of either variety of F. neoformans. Furthermore, none of the isolates exhibited self-fertile behavior. Our results indicate that the “α” mating type occurs more frequently in both clinical and natural isolates of C. neoformans originated from Japan and India.

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Two cases of keratomycosis, a 35 y.o. male and a 65 y.o. female, were reported. Both developed corneal ulcers and the fungal elements were observed by direct examination. Cultures of the corneal scraping revealed Fusarium solani and Alternaria alternate, respectively. Treatments with topical amphotericin B for the former and oral ketoconazole for the latter were both beneficial.
More than 250 cases of keratomycosis were reported in Japan as of 1988 ; among these the diagnoses were myco logically confirmed in 86 cases. Statistical analysis of these “reliable” cases showed that almost all were over 40 years old. More than 20 species of the fungi were isolated and Fusarium species was the one most frequently found.
MIC (minimal inhibitory concentration) determination was performed using pimaricin, amphotericin B, miconazole and ketoconazole against the two isolates from our cases, and pimaricin and amphotericin B proved much more effective than the other two. Moreover, it was demonstrated that the MIC value was lower (that is, more effective) when the MIC determination was done under a temperature and pH close to that of the physiological human body.

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The clinical usefulness of new serological examinations for candidemia was evaluated. In five cases of candidemia, three cases were positive for fungal index (Toxicolor-Endospecy), two cases were positive for Cand-Tec latex agglutination test, and only one case was positive for Mannan latex agglutination test. No case was positive for measured D-arabinitol, but one case had a positive D-arabinitol/creatinine ratio. All cases were diagnosed as candidemia by fungal culture. Candida albicans were isolated from three cases, Candida tropicalis was isolated from one case, and Candida parapsilosis was isolated from one case. These data demonstrated the usefulness of serological examinations for candidemia.

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The clinically normal skin of 5 patients with tinea capitis and 4 patients with tinea corporis due to Microsporum canis was examined using the finger-press method and the cotton-swab method. Our results demonstrated that M. canis was isolated in high incidences from various parts of the body : from the fingers in 7 out of 9 cases (77.8%), from the dorsa of hands in 3 out of 6 cases (50%), from the neck regions in 4 out of 9 cases (44.4%), and from the backs in 2 out of 5 cases (40%).
Even though patients with tinea capitis were treated with griseofulvin and those with tinea corporis were treated only with topical antimycotics, all cases recovered without developing any new lesions.
Based on these findings, it is suggested that although M. canis was found to be present in great amounts in clinically normal skin, the dermatophyte existed in most cases only as a saprophyte.

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Pathogenic fungi in lesions of nocardiosis or systemic mycoses survived in an airtight container at -80°C. After storage of from 2 weeks to 2 years, the lesion samples were cut into tiny pieces in an antibiotic solution. These tiny pieces and the solution were inoculated on appropriate agar slants/plates and incubated at 27°C. In 4 cases out of 5, the fungi rapidly and abundantly grew upon the media. Nocardia asteroides from nocardiosis, Candida albicans and C. tropicalis from candidiasis, and Aspergillus terreus from aspergillosis were successfully isolated. Before culturing, direct microscopic examination of the pieces from the fungally infected tissues was recommended to confirm the causative agents.

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To explore action mechanism of butenafine hydrochloride (butenafine), a novel antifungal agent, the biochemical effect of the drug on a susceptible dimorphic fungus Sporothrix schenchii in the yeast-phase of growth was studied. The results are summarized as follows :
(1) As monitored by viable counts of fungal cultures, 1.1× 10^-7 M and higher concentrations of butenafine almost completely inhibited growth, with time-dependent decrease of viable counts starting after 24 hours of exposure.
(2) Butenafine at concentrations of 5×10^-8 M or above markedly inhibited biosynthesis of ergosterol and induced intracellular accumulation of large amounts of squalene.
(3) Substantial amounts of K⁺ and inorganic phosphate were rapidly released from the fungal cells after exposure to 1.4×10^-4 M or higher concentrations of butenafine. The rate and extent, of release of these cellular components increased with increasing drug concentrations in the medium and time of exposure. Consistent with this, corresponding concentrations of butenafine also caused a rapid increase in the pH value of the cell suspension.
(4) All the experimental results lead us to the postulations that butenafine has a dual mechanisms of antifungal action. First, like existing allylamines and thiocarbamates, it would primarily inhibit squalene epoxidation and resultant inhibition of biosynthesis of ergosterol could attribute to the antifungal activity of the drug. The second action mechanism of butenafine by which it inhibits fungal growth would be a direct damaging effect on the cell membrane, although this effect was demonstrated at higher drug concentrations. It would be also suggested that, in highly susceptible fungi such as S. schenchii, the former action mechanism plays a major role in the antifungal activity of butenafine, while in less susceptible fungi the latter one does so.

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Using a wild-type strain of dimorphic pathogenic fungus Sporothrix schenckii, with both high susceptibility to butenafine and to an allylamine derivative naftifine, and several tolciclate-resistant mutant strains derived therefrom, the mechanism of action by which butenafine inhibits fungal growth was studied. Some of these mutant strains were revealed to be defective in the activity of squalene epoxidation, while others were not. All of the of tolciclate-resistant mutant strains showed cross-resistance to butenafine and naftifine. On the other hand, no significant difference in the response to a direct membrane-damaging action of butenafine in terms of induction of K⁺-release from the fungal cells was demonstrated between a susceptible wild-type strain and one resistant mutant strain of S. schenckii.
All of these experimental data suggest that in highly susceptible fungi such as S. schenckii, butenafine shares the target of action with thiocarbamates and allylamines, and that inhibition of squalene epoxidation by butenafine is primarily responsible for its antifungal activity.

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A case of chromomycosis on the left thigh of a 65 year-old woman with breast cancer is reported. Clinically, the skin lesion was a light brown, flat surfaced, small nodule that was believed to be the incipient stage of chromomycosis. Histologically, the rete ridges were elongated to engulf the microabscesses where there were many sclerotic cells.
This histological change was considered to be part of the elimination phenomenon toward the typical verrucous-crusted plaques.
Many of the fungal elements recognized in the crusts and tissues were caused by long administration of steroid or by systemic diseases with immunodeficiency.

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The effect of terbinafine, a new allylamine class of antifungal agent, on the ultrastructure of growing hyphae of Trichophyton mentagrophytes cultures in a liquid medium was investigated by scanning and transmission electron microscopy. Hyphal growth was inhibited at terbinafine concentrations above 0.2 ng/ml and several morphological changes such as bending and swelling of hyphae could be observed by scanning electron microscope. Many electron dense granular structures were found in the space between the cell wall and the cytoplasmic membrane, and also in the cytoplasm. Cultures exposed to higher concentrations of terbinafine showed destroyed or collapsed hyphae. In thin sections, degeneration of the cytoplasmic membrane, nuclear membrane and mitochondrial membrane were visible.

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