The parasitic cycle of Coccidioides immitis in murine coccidioidomycosis was investigated using a scanning electron microscope. An arthroconidium suspension containing 3.3×10⁵ arthroconidia in 0.1ml phosphate buffered saline was prepared from a culture of C. immitis IFM 4935 incubated at 37°C for 14 days. As experimental animals, 7 congenitally athymic nude mice and 7 of their heterozygous littermates were used. Each mouse was inoculated intravenously with 0.1ml of the arthroconidium suspension and dessected 3, 5, and 6 days after inoculation.
The liver was severely affected by the fungus, followed by the spleen and lungs in that order. The inoculated arthroconidia swelled and became spherical. Many young spherules with or without cellular responses were observed in the liver of both groups of mice 3 days after inoculation. On that day a few spherules reached the midpoint stae of their parasitic cycle. All the developmental stages, from spherule initials to released endospores, were found in each specimen prepared from the tissue of both groups of mice on the 5th and 6th day after inoculation. These developmental stages were clearly demonstrated by scanning electron microscopy.

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Ultrastructure of Candida albicans during cell division and germ tube formation was studied by serial sections and electron microscopy. 1. Nuclear division was mediated by the formation of a mitotic spindle whose main structural component was a bundle of microtubules developed between the opposite NAOs in the nuclear envelope. Chromosomes were not condensed and nuclear envelope persisted throughout the division. 2. Behavior of mitochondria during cell division was studied by three dimensional reconstruction of serial sections. It was found that giant mitochondria were developed at the time of nuclear migration. 3. The ultrastructure of the germ tube was considered to be very similar to that of a growing hypha of the filamentous fungi, although apical vesicles were sparse and not so aggregated as were in the latter. Three dimensional reconstruction of mitochondria showed the presence of a giant mitochondrion which was produced in the process of germ tube formation.

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Recent information about chemical and ultrastructural architecture of dermatophyte cell walls was summarized. Cell walls of dermatophytes consist of two layers; the inner electron lucent, thick (0.1 to 0.5μm in thickness) and the outer electron dense, thin (20 to 500nm in thickness) layers. The inner layer is composed of microfibrils and an amorphous matrix. The microfibrils are made from the chitinous “bricks” enveloped with glucan, mannan and peptides. The cement-like amorphous matrix is composed of glucan, mannan, galactomannan and peptides. The outer layer, which is biochemically composed of protein, is observed as rodlet fascicles interwoven into a basket-weave-like structure. In the present study, the length of rodlets showed no differences amongst hyphae, micro- and macrocondidia of Trichophyton mentagrophytes, Trichophyton rubrum, Microsporum canis and Microsporum gypseum, and no rodlet sturctures were seen on the cell wall of Epidermophyton floccosum. The orientation of rodlets was approximately perpendicular to the long axis of the hyphae and circular at the base of conidium in young cells. The outer layer of the dermatophyte cell walls is similar to some bacterial cell walls in chemistry and ultrastructure, although the inner layer is apparently different from them.

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On the basis of the biochemical and morphological studies on Paracoccidioides brasiliensis, a hypothesis for the conversion from a yeastlike (Y) form to a mycelial (M) form is proposed as follows. The synthesis of the cell wall components in the outer layer of the Y form cell wall is inhibited by environmental changes, resulting in the formation of a tunnel through which the M form extends. The microfibrils of a linear polysaccharide (chitin) in the inner layer of the weakened Y form cell wall are pressed by turgor pressure between the tunnel and expanding cytoplasm. In this condition, a part of the microfibrils is turned mechanically toward the exit of the tunnel and the synthesis of microfibrils in a more or less longitudinal orientation continues smoothly. However, in the tunnel, the synthesis of microfibrils that lie transversely to the long axis of hypha may be limited or may put a hoop on parallel longitudinal microfibrils. Out of the tunnel, a cylindrical form is maintained by the elongation of the hooped microfibrils in a longitudinal orientation in the apical region. Chitin may be synthesized isotropically in subapical region, resulting in the formation of randomly oriented microfibrils. Branched polysaccharides and proteins are interlaced with chitin and the M form is completed.

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An ultrastructural study was performed on the ascospore formation in Arthroderma vanbreuseghemii. The subcellular events of ascosporogenesis in this fungus were essentially similar to those described for other ascomycetous fungi. Ascospores were delimited by the progressive invagination of the double membrane system, composed of two paired unit membranes. Ascospore maturation began with the separation of the tracks of the double membrane system, which were followed by deposition of cell wall materials in the intercisternal space. Subsequently, the inner track developed into the ascospore plasma membrane and the outer track developed into the ascospore cell wall. At ascospore maturity, the asci underwent lysis, thereby releasing their ascospores.
Cytochemical localization of carbohydrate and thiamine pyrophosphatase was demonstrated to reside in the concentric membrane system, which was concomitantly with the double membrane system. These observations suggest that the concentric membrane system might be involved in cell wall synthesis and thus behave in a manner similar to the Golgi apparatus in higher eukaryotic cells.

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The hemolytic activity of Asp-hemolysin was remarkably inhibited by the addition of chemical reagents such as HgCl₂ and iodine. Injection of Asp-hemolysin-Hg²⁺ by the intravenous route showed strong lethality to mice, but only a poor lethal toxicity by the i. v injection of Asp-hemolysin-Hg²⁺ and by the i. v and i. p injection of Asp-hemolysin-I₂. The hemolytic activity of Asp-hemolysin inhibited by HgCl₂ was excellently restored by the addition of each homogenate-extract prepared from the liver and kidney of mice.
Egg albumin, polypeptone, and proteose peptone showed little restration of hemolytic activity of Asp-hemolysin-Hg²⁺ against hen erythrocytes. Under the same conditions, mouse serum had little effect to Asp-hemolysin-I₂. On the other hand, the hemolytic activity of Asp-hemolysin inhibited by iodine could not be restored after the addition of homogenate-extract, protein preparation, and serum, respectively. The enzymatic hydrolysis of native Asp-hemolysin was not found by the treatment with several proteolytic enzymes, while Asp-hemolysin-Hg²⁺ could be hydrolyzed after treatment with proteolytic enzyme such as α-chymotrypsin, trypsin, pronase P, and A. fumigatus protease. In addition, the hemolytic activity of Asp-hemolysin-Hg²⁺ was not effected by the treatment with trypsin for 150 minutes, but lethal toxicity to mice had yet remained in the toxin. When Asp-hemolysin was treated with thermolysin at 50°C, both activities of hemolysis and lethality were completely inactivated. These results suggest that the hemolytic activity of Asp-hemolysin can not be related to its lethal activity.

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Combination therapy by flucytosine (5-FC) and amphotericin B (AMPH) is very common for the treatment of the patient with deep-seated mycosis. We attempted to investigate the combination effect of 5-FC and AMPH, especially its mechanism, in vitro and in vivo.
Studies were focused on susceptibility of Candida albicans grown on Yeast Morphology Agar to 5-FC with or without AMPH and on cumulative mortality rate and body weight change in experimental C. albicans infection of mice after administration of 5-FC and AMPH in combination or alone.
In addition, we developed a sensitive method to separate and determine 5-FC by high-performance liquid chromatography in the presence of AMPH. By this method, intracellular levels of 5-FC in C. albicans were determined in the presence or absence of AMPH.
Synergistic effect of 5-FC and AMPH was found not only in vitro but also in vivo experiments.
No significant difference in pharmacokinetics of 5-FC by single and combined administration was observed and intracellular levels of 5-FC increased in parallel with the concentration of AMPH added.
These results indicated that the synergistic effect of 5-FC and AMPH in combination is caused by stimulation of the uptake of 5-FC by AMPH into C. albicans cells.

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A new genus, Hortaea Nishimura et Miyaji, is proposed to accommodate Cladosporium werneckii, the etiological agent of a superficial mycotic infection, tinea nigra, and Hortaea werneckii (Horta) Nishimura et Miyaji comb. nov. is provided.
The fungus is characterized by a unique sympodial conidiogenesis accompanying an annellidic one, which has been observed firstly using a scanning electron microscope as follows.
Conidiogenous loci arise from ampullaceous cells formed laterally or terminally on hyphae, lateral branches and projections on hyphae, or directly from the lateral walls of hyphae. These loci elongate and swell in accordance with the production of solitary, terminal conidia, and become thick cylindrical, obclavate or truncate rachises. Bud scars are found on the surface of the rachises, but denticles are absent. Round scars are observed in a few conidiogenous cells on the rachises, but most of the conidiogenous cells show remnants of scars shaped like sickles, half moons or scales. Regardless of the bud scar shapes, a spiral can be drawn on a rachis by tracing the centers of the scars. Namely, the conidiogenous cells are sympodulae because the bud scars are arranged on them sympodially. Such sympodial conidiogenesis is also found in most of the yeast-like conidiogenous cells.
Furthermore, there are a few conidiogenous cells with annellatons, which are irregular in comparison with those of E. dermatitidis or E. jeanselmei.
These results indicate that H. werneckii is definitely different from the other species of the genus Exophiala and other sympodial dematiaceous fungi.

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Following the previous report on in vitro antifungal activities of sulconazole nitrate, a novel imidazole antimycotic, it was reconfirmed that this compound posessed a broad antifungal spectrum with more potent fungistasis against filamentous fungi including dermatophytes and dimorphic fungi than against yeast-like fungi including Candida albicans and C. pseudotropicalis, and that Cryptococcus neoformans, in particular, was extremely sensitive to this drug.
This in vitro antifungal activity of sulconazole nitrate was not much affected by changes of inoculum size or pH of the medium and by the addition of human blood. This activity was exhibited more potently on Sabouraud dextrose agar plates than in Sabouraud dextrose broth.
The fungistatic activity of sulconazole nitrate was almost the same as that of clotrimazole in terms of MIC values against a variety of pathogenic fungi except Cr. neoformans against which the former was found to be more effective than the latter.
The growing cells of C. albicans and C. pseudotropicalis were more sensitive to the killing effect of this drug than the resting cells.

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The consecutive topical administration of 1% or 2% sulconazole nitrate suspended in polyethylene glycol for two weeks resulted in a marked therapeutic effect on cutaneous infection of guinea pigs with a T. mentagrophytes strain in terms of improvement of lesions and of negative culture of inoculated organism, to a degree equivalent to clotrimazole.

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Twenty-eight strains of Sporothrix schenckii, isolated from different clinical types of cutaneous sporotrichosis, nonlesional skin and soil, were examined to know the morphology of yeast-form cells and the temperature sensitivity. Each strain was found to have slight differences in the morphology and the temperature sensitivity. However, these slight differences on above points were not correlated with each other. Nine out of twenty-eight strains were selected for determing GC content of DNA. The DNA was extracted and purified by modified Marmur's method from each strain and the base compositions of DNA were determined by thermal melting method. They were found to be almost constant (52.5-53.7mol% G+C), which was not correlated with heterogeneity in the clinical type of lesion, the morphology of yeast-form cell and the growth temperature. It was suggested that these minor differences observed in the different clinical type, the morphology and growth temperature may be regarded as the variations in the same species.

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A 54-year-old man with fungal keratitis which did not respond to oral ketoconazole was successfully treated with intravenously infused miconazole, a synthetic imidazole derivative. In this case, healing and regression of the corneal ulcer began on the fourth day after the initiation of miconazole monotherapy (from 400mg to 1, 200mg per day). The clinical evidence of corneal infection disappeared completely after three weeks and visual acuity improved from 0.05 to 0.7. Aside from a slight increase in serum triglycerides and β-liporoteins, the drug appears to be well tolerated.

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Aspergillosis with cutaneous and pulmonary involvements in an immature infant is presented. The patient died on the 9th day after delivery. Aspergillus fumigatus was isolated from the bronchial exudate and cutaneous lesions. Autopsy revealed numerous, dichotomously branching hyphae in the pulmonary and cutaneous lesions. There were no cellular reactions in the lesions.

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