At the 48th Annual Meeting of The Society for Japanese Medical Mycology, held in October, 2004, we reported our findings from a survey on Trichophyton tonsurans infections in the Hokuriku and Kinki regions of Japan. The survey revealed that a few epidemics had occurred across these regions. In this article, we introduce our subsequent studies relating to 1) molecular epidemiology of isolates taken from people in many parts of Japan and 2) factors affecting adequate sampling of the scalp with hairbrushes, essential for surveying and monitoring the infection.
In total, 198 isolates of Trichophyton tonsurans were analyzed using restriction fragment length polymorphisms of the non-transcribed spacer regions of ribosomal RNA genes. The restriction enzyme Mva I indicated two molecular types of strains, implying that the causative agents of the epidemic had different origins. None of the isolates obtained from the epidemic showed the same restriction profile as that of isolates from aged and sporadic cases.
The published hairbrush method suitable for obtaining samples from the scalp of Judo trainees was reevaluated by changing several factors. We found that sampling should not be done soon after the students' physical training because other fungal elements may give a false positive, samples should not be obtained from students who have recently applied topical antimycotics, and samples should be taken under the guidance of qualified instructors familiar with the sampling method.

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Environments may act as reservoirs for pathogenic fungi, a determinant of the establishment of fungal infection, or an exacerbating factor of disease. In recent years, skin disease caused by geophilic fungi has been decreasing, while case reports of zoonoses from various animals are increasing. Outbreaks of anthropophilic T. tonsurans infection pose a problem to medical mycologists. Tinea pedis is the most common exogenous dermatomycosis in Japan. Although T. rubrum is presumed to be the dominant pathogen of this disease, T. mentagrophytes is detected more frequently from various environments, so far, the reason for this discrepancy has not been fully understood. The latest knowledge about the route of dermatophyte foot infection is as follows: (1) Dermatophyte propagules disseminated from patients may contaminate not only bath-mats but also wood floors, Japanese style mattings, concrete floors, slippers, cushions, etc., and from them adhere to healthy skin. (2) The agar stamping method can easily detect dermatophytes from the skin and the environment. (3) Propagules of T. mentagrophytes can survive for more than three months under certain conditions such as in rubber boots. (4) In order to eliminate dermatophytes gathered in socks and footwear, simple procedures (washing, bathing with hot water, or wiping with a towel) are all effective. (5) Prior application of an antifungal agent promptly eradicates dermatophyte propagules adhering to the skin from the environment. The author also mentioned the possibility of asymptomatic dermatophyte colonization, and the high prevalence of dysgeucia in oral carriers of Candida albicans.

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Recent progress in the epidemiology, diagnosis and treatment of onychomycosis is summarized. The risk factors of this condition were investigated, and the results of analyses of the systemic state of patients, shape of nails, blood circulation around nails, nail growth rate and causative fungi were reported. In making a diagnosis of onychomycosis, the major effect of the quality of collected nail samples on the results was reconfirmed. In addition to the KOH method and culture method, a molecular biological method was introduced. From the therapeutic perspective, 1) prevention of recurrence, 2) identification of patients with high risk of onychomycosis, and 3) attempting new treatments (development of new drugs, improvement of administration and dosage, and evaluation of combined therapy) were discussed. The effectiveness of concomitant use with nail lacquer is anticipated in combined therapy. New studies on accurate diagnosis, early detection and early stage treatment are now being undertaken.

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The genus Malassezia is now divided into eleven species. Different species initiate or aggravate different skin diseases. In seborroheic dermatitis, M. restricta play an important role, while in atopic dermatitis, M. globosa and/or M. restricta are major cutaneous microflora.
M. globosa is a causative species of tinea versicolor, and this species is also a causative species of malassezia folliculitis. We should therefore obtain better knowledge of the ecological and pathogenic roles of malassezia.

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Dermatophytosis, cryptococcosis and sporotrichosis are dermatological zoonosis caused by fungi. Control of dermatophytosis is still difficult, although it is well known as a transmittable disease from animal to human. Exotic animals such as hamster, guinea pig and rabbit, some imported from foreign countries, have become a frequent source of human dermatophytosis, and the emergence of fungal pathogens followed. The nasal cavity of dogs and cats is an important reservoir of pathogens of cryptococcosis, in addition to bird droppings. Though there have been many reports of human sporotrichosis caused by patients' pet, especially cats in Europe and U.S., since there has been no report of zoonotic transmission of sporotrichosis, zoonosis is not believed do be present in Japan.
The interrelation and cooperation of medical and veterinary doctors and all other scientists involved is therefore required to prevent fungal zoonosis, because the condition may be partly due to changes in the environments of human and animal life and the more intimate relationships between humans and animals.

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Five aerobic actinomycete strains isolated from patients in Japan were assigned provisionally to the genus Nocardia based on morphological and physiological characteristics. The five strains, IFM 10481, IFM 0668, IFM 0901, IFM 0583 and IFM 0342, were not classified into any Nocardia species reported as infectious agents in Japan. Therefore, they were studied further to determine their specific taxonomic positions. Detailed chemotaxonomic and physiologic characterization and 16S rDNA sequence data of the five strains showed that they belonged to respective species of Nocardia carnea, N. elegans, N. paucivorans, N. puris and N. takedensis. This is the first isolation report of these five Nocardia species from patients in Japan.

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By the combined use of agar diffusion, agar vapor and agar vapor-inhibitory assays, contribution of the vapor activity of essential oils was quantitatively estimated. The test organisms were Trichophyton mentagrophytes and Aspergillus fumigatus. Agar vapor assay was used to confirm the vapor activity of the oils. The parameter delta defined as a contribution index of the vapor activity was calculated by (1- b-c/a-c) × 100, where a is inhibitory diameter in the diffusion assay, b is inhibitory diameter in the vapor-inhibitory assay and c is diameter of the sealed ring in the vapor-inhibitory assay (21 mm). Many of the essential oils examined showed a delta value near 100, thus providing the major contribution of the vapor activity to the inhibitory diameter. Essential oils containing aldehyde as major constituent showed low delta value, indicating the major inhibition was due to agar diffusion. Major essential oil components behaved similarly; the delta value was increased in the following order: aldehyde<phenol<alcohol<ester, oxide, hydrocarbon, indicating the enhanced contribution of the vapor activity in that order.

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The patient was a 57-year-old woman who initially consulted our department on May 31, 2002 with a chief complaint of other dermatoses. The initial examination demonstrated deformation of approximately 1/3 of the inner part of the nail plate in the right hallux, in addition to brown discoloration and keratin hypertrophy under the deformed nail plate. KOH-prepared direct microscopy revealed the presence of large round spores and hyphae. Since colonies similar to those of Trichophyton rubrum were obtained by culture, daily administration of terbinafine (125 mg/day) was initiated from June 17, 2002 based on a suspicion of tinea unguium induced by T. rubrum. However, the morphology of the colonies began to change toward a brown powder-like configuration after 2-3 weeks of culture, and band forms were also detected by slide culture, resulting in the diagnosis of onychomycosis induced by Scopulariopsis brevicaulis. From August 6, 2002, the dose of terbinafine was increased to 250 mg/day, which was maintained until November 22, 2002. When the patient visited our department approximately 2 months later, KOH-prepared direct microscopy revealed that she was still positive for S. brevicaulis. Therefore, terbinafine (250 mg/day) was administered again for 1 month from April 25, 2003, followed by a judgment of complete healing on July 4, 2003. Thereafter, there was no recurrence of onychomycosis up until January 29, 2005.

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An epidemiological survey of dermatomycoses and the causative fungus flora of dermatophytoses in Japan for 2002 was made on a total number of 72,660 outpatients who visited 14 dermatological clinics throughout Japan. The results were as follows:
1) Dermatophytosis was the most prevalent cutaneous fungal infection (7,994 cases) seen in these clinics, followed by candidiasis (755 cases) and then Malassezia infections (220 cases).
2) Among dermatophytoses, tinea pedis was the most frequent (4,813 cases: male 2,439, female 2,374), then in decreasing order, tinea unguium (2,123 cases: male 1,093, female 1,030), tinea corporis (497 cases: male 281, female 216), tinea cruris (299 cases: male 249, female 50), tinea manuum (248 cases: male 144, female 104) and tinea capitis including kerion (14 cases, male 6, female 8).
3) Tinea pedis and tinea unguium are seen to increase in summer season, among the aged population and among males in each clinic. When compared to the previous surveys (1992 and 1997) by clinical form, t. unguium patients increased from 1.9% of total outpatients in 1992, to 2.0% in 1997, then to 2.9% in 2002.
4) As the causative dermatophyte species, Trichophyton rubrum was the most frequently isolated among all dermatophyte infections except tinea capitis.
5) T. rubrum was isolated from 63.3% (1,431/2,262) of tinea pedis lesions, followed by Trichophyton mentagrophytes (36.6%, 829/2,262), and also 88.8% (325/366) of t. corporis, 95.4% (185/194) of t. cruris and 85.6% (462/540) of t. unguium.
6) Cutaneous candidiasis was seen in 755 cases (1.0%) of 72,660 outpatients. Intertrigo (347 cases) was the most frequent clinical form, followed by erosio interdigitalis (103 cases) and diaper candidiasis (102 cases). It has a tendency to affect the aged being complicated with topical predisposing factors.
7) Cutaneous Malassezia infections and other superficial fungal infections are seen in 220 cases, without any characteristic features by gender or clinical form.

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The Research Center for Pathogenic Fungi and Microbial Toxicoses, Chiba University is the only organization in Japan to possess a series isolates of Coccidioides spp., which are the most virulent pathogenic fungi and which are treated as biosafety level 3 microorganisms. Recently, the genus Coccidioides has been classified into two species, C. immitis and C. posadasii, based on their endemic areas and genotyping; the former species is endemic to the state of California, and the latter is endemic to other parts of North and South America. We reevaluated 19 isolates of Coccidioides immitis stored in our center using a multiple gene analysis. Five isolates were identified as C. immitis and 14 as C. posadasii. Their sequence information in GenBank will help to identify the two genospecies of Coccidioides spp.

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