Viability of pathogenic dermatophytes during a 4-week treatment with 1% topical luliconazole for tinea pedis.

The viability of pathogenic fungi in the scale was investigated during topical administration of 1% luliconazole (LLCZ). Thirteen tinea pedis patients found to be positive on KOH examination were assessed by mycological examinations and quantitative real-time polymerase chain reaction (PCR) targeted internal transcribed spacer (ITS) in ribosomal RNA gene at the initial visit and after 2 and 4 weeks of treatment. Assays showed that the average copy number of ITS DNA had significantly decreased to 22.9% at 2 weeks and 4.8% at 4 weeks compared with the initial visit. LLCZ topical treatment could defeat almost pathogenic dermatophytes in the scales within 4 weeks.

Variations in ribosomal DNA copy numbers in a genome of Trichophyton interdigitale.

BACKGROUND: Ribosomal DNA (rDNA) reportedly has multiple copies in the fungal genome. The internal transcribed spacer (ITS) region in rDNA is useful for investigating relationships between close taxonomic relatives. Thus, ITS has been widely used as a target gene in medical mycology for the detection of pathogenic fungi and identification of fungal species. However, the rDNA copy number in a genome of Trichophyton interdigitale, the pathogen causing dermatophytosis, currently remains unknown. OBJECTIVE: Clarification of the rDNA copy number in a genome of T. interdigitale. METHODS: rDNA copy numbers in 64 clinical isolates of T. interdigitale were examined using quantitative real-time PCR (qPCR) with the absolute quantitative method targeting TruMDR2, a single-copy control gene and the ITS region in rDNA. RESULTS: The copy numbers of the rDNA subunit varied among the 64 strains tested, from 24 to 116 copies per genome. The average rDNA copy number ± standard deviation was 59 ± 16. No correlations were observed between the rDNA copy number and colony colour, colony morphology or molecular type of the non-transcribed spacer region in rDNA. Experiments on rDNA copy numbers obtained from independent colonies of each strain in single-spore cultures revealed that the copy number was homogeneous within each strain. CONCLUSION: This is the first study to estimate copy numbers of the rDNA subunit in a genome of T. interdigitale. The rDNA copy number of T. interdigitale varied among the strains tested and was homogeneous within each strain.

Pathogenic Dermatophytes Survive in Nail Lesions During Oral Terbinafine Treatment for Tinea Unguium.

Tinea unguium caused by dermatophyte species are usually treated with oral antimycotic, terbinafine (TBF). To understand the mechanisms of improvement and recalcitrance of tinea unguium by oral TBF treatment, a method of quantifying dermatophyte viability in the nail was developed, and the viability of dermatophytes was analyzed in toenail lesions of 14 patients with KOH-positive tinea unguium treated with oral TBF 125 mg/day for up to 16 weeks. Mycological tests, including KOH examination and fungal culture, and targeted quantitative real-time PCR for internal transcribed spacer (ITS) region, including rRNA, were demonstrated at the initial visit and after 8 and 16 weeks of treatment. Assays in eight patients showed that average ITS DNA amount significantly decreased, to 44% at 8 weeks and 36% at 16 weeks compared with 100% at initial visit. No significant difference was observed between at 8 and 16 weeks, despite the TBF concentration in the nail supposedly more than 10-fold higher than the minimum fungicidal concentration for dermatophytes. This finding suggests the pathogenic dermatophytes in nail lesions could survive in a dormant form, such as arthroconidia, during oral TBF treatment. Both antimycotic activity and nail growth are important factors in treatment of tinea unguium.

Quantification of dermatophyte viability for evaluation of antifungal effect by quantitative PCR.

Dermatophytosis is a common disease caused by dermatophyte fungi such as Trichophyton rubrum and Trichophyton mentagrophytes. A method of quantifying fungal viability in the lesions of dermatophytosis is indispensable for understanding the therapeutic process and outcome; however, no such method has yet been developed. The aim of this study was to develop a method for quantifying dermatophyte viability by quantitative polymerase chain reaction (qPCR). The internal transcribed spacer (ITS) and D1/D2 regions, including each of rRNA and rDNA, were chosen as the targets, and dermatophyte-specific primer pairs were designed corresponding to ITS and D1/D2 regions. The amounts of target RNA and DNA after heat or antifungal treatment were measured by qPCR and compared with colony-forming unit (CFU) counts. RNA and DNA could extract from dermatophytes by mechanical pulverization of conidia using a Multi-Beads Shocker cell disruptor. Our method was sufficiently sensitive to detect 10 copies by qPCR using both ITS and D1/D2 primer pairs. The most sensitive target was ITS-cDNA after heat or antifungal treatment, and essentially consistent with CFU counts. On the other hands, ITS-DNA and D1/D2-DNA were not decreased soon after heat or antifungal treatment, but those were decreased significantly and reflected the CFU counts after 48 h of antifungal treatment. We conclude that ITS-cDNA is useful mainly for quantifying dermatophyte viability at early responses, but ITS-DNA and D1/D2-DNA are also available for evaluation, which does not need an early response.

Revival of favus in Japan caused by Trichophyton schoenleinii.

Favus is a type of dermatophytosis known to produce yellow scutula around hair follicles. Most cases of this disease worldwide are infections of Trichophyton schoenleinii. Favus has rarely been reported in Japan throughout the last four decades, and T. schoenleinii has not been clinically isolated in any case during the period. Here, we report a case of favus of vellus hair observed in a 63-year-old Japanese woman. Fungal culture showed negative; however, we detected fungal elements in the crust and hair bulbs by Grocott staining. Pathogenic fungi were identified as T. schoenleinii by polymerase chain reaction-based DNA sequencing, targeting the internal transcribed spacer regions of the rRNA gene using the formalin-fixed, paraffin-embedded tissue sample. She was successfully treated with p.o. administration of terbinafine and topical application of luliconazole cream.

Unique Temporal Expression of Triplicated Long-Wavelength Opsins in Developing Butterfly Eyes.

Following gene duplication events, the expression patterns of the resulting gene copies can often diverge both spatially and temporally. Here we report on gene duplicates that are expressed in distinct but overlapping patterns, and which exhibit temporally divergent expression. Butterflies have sophisticated color vision and spectrally complex eyes, typically with three types of heterogeneous ommatidia. The eyes of the butterfly Papilio xuthus express two green- and one red-absorbing visual pigment, which came about via gene duplication events, in addition to one ultraviolet (UV)- and one blue-absorbing visual pigment. We localized mRNAs encoding opsins of these visual pigments in developing eye disks throughout the pupal stage. The mRNAs of the UV and blue opsin are expressed early in pupal development (pd), specifying the type of the ommatidium in which they appear. Red sensitive photoreceptors first express a green opsin mRNA, which is replaced later by the red opsin mRNA. Broadband photoreceptors (that coexpress the green and red opsins) first express the green opsin mRNA, later change to red opsin mRNA and finally re-express the green opsin mRNA in addition to the red mRNA. Such a unique temporal and spatial expression pattern of opsin mRNAs may reflect the evolution of visual pigments and provide clues toward understanding how the spectrally complex eyes of butterflies evolved.

Comparison of Characteristics of Two Topical Therapeutic Agents for Onychomycosis.

Two topical therapeutic agents were approved in Japan from 2015 to 2016, adding new options for onychomycosis therapy in the clinical field. In order to confirm the differences of formulation properties and nail pharmacokinetics between 5% luliconazole solution and 10% efinaconazole solution, drug concentration and antifungal activity in the nail were measured after topical treatment using human nail plates. In the in vitro permeation study, concentration of each drug was measured in the transversely sliced nail after single treatment with the two topical therapeutic agents. The results showed that concentration of luliconazole is higher than that of efinaconazole at all nail layers, differing by 1.7-8.4 times at each measurement point. Next, we examined antifungal activities of each drug in sliced nail after 14-day topical treatment. Mean rates of formation of inhibition zones for 5% luliconazole solution and 10% efinaconazole solution were 71.0% and 12.6%, respectively, and were statistically different. These results show that the two topical therapeutic agents have different properties, and suggest that 5% luliconazole solution has good nail permeation and retention characteristics. Moreover, luliconazole was found to retain enough antifungal activity in the nail plate against Trichophyton spp. after treatment with the topical agent.

Protective effect against Parkinson's disease-related insults through the activation of XBP1.

The accumulation of misfolded and unfolded proteins in endoplasmic reticulum (ER) induces ER stress, activating the unfolded protein response (UPR). Recent evidence has suggested the relationship between UPR and dopaminergic neuronal cell death in Parkinson's disease (PD); however, it remains unclear whether it makes sense to modulate UPR, to mitigate the progression of PD. In this study, we investigated a role of the IRE1 alpha-XBP1 pathway in the survival of dopaminergic cells, under stress induced by PD-related insults. The exogenous expression of the active-form XBP1 (XBP1s) protein had protective effects against cell death induced by 1-methyl-4-phenylpyridinium (MPP+) and proteasome inhibitors. Moreover, adenoviral XBP1s expression significantly suppressed the degeneration of dopaminergic neurons in the mouse model of PD, as induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). These results demonstrate that the enhancement of XBP1 could be a novel PD therapeutic strategy.