Surface material analysis for human papillomavirus detection in nail Bowen's disease caused by HPV type 58.

Over the past few years, cases of human papillomavirus (HPV) infection in nail Bowen's disease have been reported. This disease presents diagnostic challenges due to its similarity to nail malignant melanoma, particularly with respect to the clinical manifestation of black nail streaks. While skin biopsy is usually employed for diagnosis, it is an invasive procedure. We report the case of a 52-year-old healthy Japanese male with a pigmented streak on the nail of the fourth finger of his right hand, which had extended from the central to the lateral nail fold within 4 months. Dermoscopic examination revealed a dark-brown pigmented band with splinter microhemorrhage. Clinically, nail Bowen's disease was suspected. The lesion was excised in strips under local anesthesia. Histopathological examination revealed hyperkeratosis, parakeratosis, papillomatosis, and dyskeratotic cells with atypical nuclei irregularly arranged. Immunohistochemistry using anti-HPV L1 antibody detected HPV-positive cells in the upper epidermis and stratum corneum of the nail matrix. Mucosal high-risk HPV type 58 DNA was detected from brush cytology of the keratotic surface prior to surgery, which was confirmed in formalin-fixed, paraffin-embedded excised samples using polymerase chain reaction (PCR) and subsequent direct DNA sequencing. Our case highlights HPV type 58 as a potential causative agent of nail Bowen's disease and shows that brush cytology of the surface material prior to excision may be a useful and less invasive way for mucosal high-risk HPV detection. PCR analysis of the nail surface could serve as a supplementary diagnostic tool for nail Bowen's disease.

Vasculitis-like herpes zoster in the course of treatment with tofacitinib in ulcerative colitis: An assessment of local viral distribution by RNA in situ hybridization.

A 67-year-old man had taken the janus kinase (JAK) inhibitor, tofacitinib, for ulcerative colitis. He was referred to our department for a refractory ulcer on his lower leg. We suspected vasculitis and performed skin biopsy. Histopathological examination showed multinucleated giant cells in the epidermis and fibrinoid degeneration of small vessels in the upper dermis. Varicella zoster virus (VZV) DNA was detected by polymerase chain reaction and we diagnosed the patient with atypical vasculitis-like herpes zoster. The patient was treated with oral valacyclovir, but the rash persisted and took 2 months to heal. Immunostaining using anti-VZV antibody was positive mainly in epidermal keratinocytes, but was also observed to be positive in cells in the dermis. We further performed RNA in situ hybridization using a VZV ORF9 mRNA probe and clearly showed that the distribution of VZV mRNA extended into the dermis, including the dermal vessel walls and the eccrine sweat glands as well as the epidermis. The internal administration of JAK inhibitors may induce regional widespread VZV infection including vessels and involved in the formation of prolonged vasculitis-like manifestation. RNA in situ hybridization can be a potent tool for detecting the spread of VZV infection in the skin.

Detection of varicella-zoster virus in two dermatomes of herpes zoster duplex bilateralis in an immunocompetent host.

An 85-year-old woman with no history of herpes zoster (HZ) presented with a primary lesion of erythema and blistering on her left thigh and a secondary similar lesion on her right chest which had appeared at 4 and 3 days before presentation, respectively. Tzanck smears for both lesions were positive, revealing multinucleated giant cells. Immunochromatography to detect varicella-zoster virus (VZV) antigen (DermaQuick®VZV) showed positive on the left thigh at initial onset but negative on the right chest at subsequent onset. The latter repeatedly tested negative for VZV by DermaQuick®VZV. A skin biopsy of the subsequent onset area revealed giant cells, and inclusion bodies were observed in the epidermis. Immunohistochemical staining with anti-VZV antibody and polymerase chain reaction to detect VZV DNA were positive. The patient was diagnosed with HZ duplex bilateralis and treated with acyclovir. The right thoracic region of the posterior part of the lesion became negative for DermaQuick®VZV. It is thought that expression of viral antigens was suppressed in the right thoracic region, i.e., the late-onset area.

A Case of Tinea Corporis Caused by Trichophyton benhamiae var. luteum from a Degu and Evolution of the Pathogen's Taxonomy.

BACKGROUND: Trichophyton benhamiae, an anthropophilic dermatophyte, can cause dermatophytosis in humans and animals with rising zoonotic infections through pets, primarily in Europe. Dermatophytosis from T. benhamiae is often misdiagnosed due to its inflammatory symptoms. We report a case of tinea corporis from T. benhamiae var. luteum in a Japanese woman, contracted from pet Czech degus. CASE: The 40-year-old patient developed neck papules resembling acne. Initial treatment with topical antibiotics and steroids exacerbated the rash. Fungal elements were not detected by direct potassium hydroxide examination. Skin biopsy confirmed fungal elements in the stratum corneum and hair follicles, and tinea corporis was diagnosed. Oral terbinafine 125 mg was initiated without topical agents. Erythematous papules appeared on her limbs, determined as a trichophytid reaction. After two months, her skin improved significantly. Fungal culture identified T. benhamiae var. luteum colonies with a yellowish hue. Mating tests classified the strain as Americano-European race (-) with MAT1-1 genotype. This was diagnosed as tinea corporis from T. benhamiae var. luteum, likely transmitted from pet Czech degus. CONCLUSIONS: The incidence rate of pet-transmitted cutaneous fungal infections may increase in Japan with the trend to keep exotic pets. Dermatologists must recognize dermatophytosis clinical features from anthropophilic dermatophytes to prevent misdiagnosis and understand evolving nomenclature and pathogenesis.

Possibly the first case of onychomycosis by Fusarium lactis: Case presentation and literature review of onychomycosis by Fusarium species.

Fusarium species (spp.) is frequently found in soil and plant residues and on plant bodies in all climatic zones worldwide. Although there have been few reports of onychomycosis caused by Fusarium spp., it is characterized by drug sensitivity and other characteristics. Here, we report what may be the first case of onychomycosis caused by Fusarium lactis. We analyzed the mycology and characterized previously reported cases of onychomycosis caused by Fusarium spp. A 73-year-old otherwise healthy woman presented with discoloration and thickening of her right thumbnail with paronychia. Direct microscopy revealed unevenly swollen hyphae, and a Grocott-stained nail specimen showed septate hyphae. Based on the morphological features and gene analysis of fungus isolated from the nail, we diagnosed onychomycosis caused by F. lactis belonging to Fusarium fujikuroi species complex. Partial nail removal and topical application of 1% luliconazole solution resolved the condition in 6 months. Minimum inhibitory concentrations for isolated F. lactis showed high sensitivity to luliconazole but not itraconazole or terbinafine. The isolated F. lactis was temperature-sensitive. A search of the literature revealed 57 cases of onychomycosis caused by Fusarium spp. with delineated clinical characteristics. Since those cases were investigated using morphological and/or molecular methods, we analyzed them by species complex as well as species. Onychomycosis caused by Fusarium spp. is predominantly found on the big toe, with Fusarium solani species complex and Fusarium oxysporum species complex accounting for over 70% of cases. Infection of only one digit with paronychia is a characteristic clinical manifestation of onychomycosis caused by Fusarium spp. Since there has been an increase in instances of molecular determination of Fusarium spp., it is deemed necessary to clarify its clinical and fungal nature. Due to its characteristic drug sensitivity and temperature-sensitive nature, new treatments are expected to be developed.

[Trichophyton tonsurans Infection].

Trichophyton tonsurans infection has been prevalent among individuals involved in contact sports in Japan since about 2000. The present review focuses on its diagnosis, molecular epidemiology, drug susceptibility, and infection control. The most commonly observed lesions of T. tonsurans, an anthropogenic dermatophyte, are tinea corporis and tinea capitis. However, the presence of asymptomatic carriers must be considered for infection control. Genotypic epidemiology using restriction fragment length polymorphisms (RFLP) in the non-transcribed spacer (NTS) region of the ribosomal RNA gene showed a lack of diversity of genotypes, and only the NTS I genotype is detected at present. In regard to drug susceptibility, terbinafine drug resistance has not been found to be associated with the RFLP genotypes, and it is assumed that there are no terbinafine-resistant strains in Japan. T. tonsurans coexisted with other fungi and bacteria in the scalp of asymptomatic carriers without affecting species diversity. T. tonsurans is an anthropogenic dermatophyte and may be difficult for the human immune system to eliminate. During an infection outbreak, screening of infection and treatment including asymptomatic carriers are essential to eradicate the infection.

Multilocus Microsatellite Analysis of the Molecular Epidemiology of Microsporum canis Isolated in Japan.

Microsporum canis, a major causative agent of zoonotic dermatophytosis, has become prevalent in Japan. Molecular epidemiological surveys using multilocus microsatellite typing (MLMT), a sensitive genotyping tool for fungi, have been conducted to reveal intraspecies polymorphisms of M. canis.The present study utilized MLMT optimized for Japanese strains of M. canis to analyze epidemic trends of fungal infection. Six individual loci were targeted; namely, MS1, 2, 4, 5, 6, and 7. Analysis of data from 1974 through 2022 identified 416 strains, which were sorted into 60 genotypes by MLMT.The major genotypes showed changes in dominance during this period-changes that may reflect historical increases and decreases in the numbers of patients infected with M. canis patients. The main origins of infection included animal breeders and pet stores, as well as stray cat communities. Forty-nine episodes of familial outbreaks and cohabitant animal infections were recorded, and genotypes responsible for each episode were determined. MLMT analysis is not only a robust tool to understand population structures, but likely the most suitable method for tracking M. canis infections.

Skin surface material for detecting human papillomavirus infection of skin warts.

Warts, caused by human papillomavirus (HPV) infection, have various clinical presentations, making them difficult to differentiate from clavus, callus, and sometimes, squamous cell carcinoma. Although skin biopsies are the gold standard, a less-invasive method of examining these lesions is desired. Ninety patients with warts and related diseases, such as clavus and callus, were recruited to explore new differentiation methods using the surface of the warts. DNA was extracted from three types of specimens in each case: surface swab, shaved hyperkeratotic scale, and post-shaved surface swab. Total DNA was successfully extracted from these three specimens and was sufficient for subsequent HPV DNA detection. We analyzed samples for the HPV type and HPV viral load using polymerase chain reaction (PCR). Fifty-five cases were PCR-positive, and HPV1a, 2a, 4, 27, 57, and 65 were detected. The amount of HPV1a DNA produced was significantly greater than that of other HPV types. Regarding the correlation between the clinical diagnosis and HPV detection, the positive agreement rate was 90.9%, the negative agreement rate was 40.0%, and the overall agreement rate was 71.1%. Ten of the 21 cases clinically diagnosed as plantar warts were PCR-negative, especially in elderly patients. This suggests that it is difficult to distinguish plantar warts from clavus and callus in clinical practice. Although the amount of HPV DNA in the removed keratinization scale was highest for all HPV types, HPV detection by swabbing before and after shaving is also useful for follow-up as well as for differential diagnosis.

MALDI-TOF MS identification of Exophiala species isolated in Japan: Library enrichment and faster sample preparation.

Exophiala species cause chromoblastomycosis, mycetoma, and phaeohyphomycosis, which are occasionally fatally in immunocompromised patients. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) provides rapid and accurate examination of isolated bacteria and some fungal isolates, but the preparation method for filamentous fungi is complicated. In this study, 31 clinical isolates of Exophiala spp. in Japan were identified by MALDI-TOF MS with a library enriched by adding data. To simplify the sample preparation method, two modified methods were compared with the standard method for filamentous fungi. The agar cultivation sample preparation method reduced the time required for liquid culture and was considered suitable for clinical use. In 30 of 31 clinical isolates of Exophiala spp., the species identified by MALDI-TOF MS with the highest score matched the species identified by sequencing the internal transcribed spacer region. Exophiala dermatitidis, E. lecanii-corni, and E. oligosperma were identified above the genus level, while E. jeanselmei and E. xenobiotica were often not identified at the species level. The identification scores tended to be lower for less-registered strains in the in-house library. It is suggested that library enrichment and the modified preparation method may facilitate early diagnosis of rare fungal infections by Exophiala spp. in clinical laboratories using MALDI-TOF MS.

A Case of Oral Candidiasis Caused by Candida dubliniensis During Treatment of Humanized Monoclonal Antibody to Interleukin-17A.

We report the case of a 77-year-old woman who had been diagnosed with psoriasis vulgaris at another hospital at age 33 and treated with various therapies since then. At 61 years old, she was diagnosed with psoriatic arthritis with phalangitis, swelling of both hands, and sacroiliac arthritis, and was treated with oral immunotherapy. At age 76, treatment with a humanized monoclonal antibody to interleukin-17A (ixekizumab) was started due to increased pain associated with her hand inflammation. After ten months of this treatment, she visited our hospital with the complaint of tongue pain and white fungi on the tongue surface. Direct KOH examination of the fungi showed pseudomycelia and spores, and fungal culture (CHROMagar®Candida) showed green and dark green wet colonies. Sequencing of the D1D2 region of ribosomal RNA gene of the nuclear DNA of fungi from the colony identified Candida dubliniensis. She was treated with amphotericin B gargle (Fungizone®syrup) for two weeks, and the lesion improved. Since then, the patient has been treated with ixekizumab with no recurrence of oral candidiasis.

First Isolation of Neoscytalidium dimidiatum from Human Dermatomycosis in Japan.

Neoscytalidium dimidiatum is a common fungus that causes non-dermatophyte dermatomycosis in tropical regions, but there have been no reports of infection with N. dimidiatum in Japan. Here, we report the first isolation of N. dimidiatum from human dermatomycosis in Japan. A 62-year-old healthy Japanese male had been treated with oral terbinafine for tinea pedis diagnosed from a microscopic examination in 2003 with a lesion that was intractable. In 2020, re-identification by sequencing the internal transcribed spacer regions and the D1/D2 domain of the large-subunit (LSU) ribosomal RNA gene revealed that the pathogen was N. dimidiatum. Antifungal susceptibility tests showed that the minimum inhibitory concentration of the drug luliconazole (LLCZ) against the pathogen was 0.00049 µg/mL. The patient's lesions were cured by topical LLCZ. The clinical course and drug susceptibility suggest that LLCZ is a suitable first-line drug for treatment.

Change in dominant genotype of Microsporum canis, a causative fungus of zoonotic dermatophytosis, in Japan over the past 40 years.

Multilocus microsatellite typing was performed on 124 strains of Microsporum canis, which is the most prevalent causative fungus of zoonotic dermatophytosis, isolated in Japan between 1974 and 1981. The strains had been enclosed in glass ampoules by freeze dried process. Genotypes were detected by capillary electrophoresis targeted on six microsatellite regions, and 19 genotypes were found among these 124 strains. The most frequent genotype, which had been reported as genotype L in a previous study, comprised 56 of the 124 strains (45.2%), although genotype L was reported to comprise only six of 165 strains (3.6%) isolated between 2010 and 2017. A decrease in the prevalence of genotype L may have led to the retreat of M. canis infection in 1996 to 2006. On the other hand, genotype A, the second most predominant genotype in the aforementioned studies of strains isolated between 2010 and 2017, comprised just four of 124 strains (3.2%) in the present study. Thus, these studies reveal that prevalences of some major genotypes have changed over the last 40 years. Genotype consistency of strains was proven in all of 12 familial cases, each of which was infected with a single genotype. We emphasize the importance of fungal culture collection for further studies with new techniques in the future.

Genotyping of Trichophyton tonsurans strains isolated between 2016 and 2020, and terbinafine susceptibility of the species in Japan.

Trichophyton tonsurans is the most prevalent fungus which causes dermatophytosis among contact sports players in Japan. We previously surveyed the epidemic of T. tonsurans in Japan from early 2000, and determined the genotypes of isolates by analysis of restriction enzyme fragment length polymorphisms in the non-transcription spacer (NTS) region of ribosomal RNA gene, which enabled discrimination of eight genotypes, namely NTS I to NTS VIII. In the present study, we performed genotyping of T. tonsurans isolated between 2016 and 2020, and investigated the trend of the epidemic and resistance of the pathogen to antibiotic terbinafine (TBF). Regardless of which contact sport they played, the genotype of all 123 strains of T. tonsurans isolated from athletes was NTS I. Genotypes NTS II and III, which were isolated in considerable numbers mainly from wrestlers between 2000 and 2015, were conspicuously absent. TBF susceptibility was screened in 237 T. tonsurans strains isolated between 2000 and 2020 with 28 of these further assessed for minimum inhibitory concentration of TBF and squalene epoxidase gene sequences. None of the strains showed TBF resistance. TBF may still be effective to control the epidemic of T. tonsurans.

Molecular Epidemiology of Microsporum canis Isolated from Japanese Cats and Dogs, and from Pet Owners by Multilocus Microsatellite Typing Fragment Analysis.

Multilocus microsatellite typing (MLMT) was performed on 93 strains of Microsporum canis (M. canis) isolated between 2012 and 2017 from 75 cats, 8 dogs, and 10 pet owners. These strains were derived from 2 major reservoirs: commercial breeding facilities and pet shops (PS), and stray cats and pet cats that went outdoors and came in contact with stray cats (Outdoor). Six microsatellite markers were used for genotyping. These 93 strains included 22 genotypes; 11 had been previously detected in Japan while the other 11 were new. Strains belonging to the previously reported genotypes, P and A, were distributed widely throughout Japan. Genotype P was the most frequent, accounting for 37 (39.8%) of the 93 strains. Most were derived from Outdoor sources. Genotype A was the second most frequent (11 of 93 strains, 11.8%). Most of the genotype A strains were derived from the PS reservoir. All new genotypes were detected in isolates from cats. Many of these were derived from the Outdoor reservoir. Consistency of infection was observed in 18 of the 19 familial cases. These findings indicate that genotypes differ in strains derived from PS and Outdoor reservoirs. MLMT genotyping is useful for tracking the routes of spread and transmission of M. canis in Japan.

Genotyping of intraspecies polymorphisms of Sporothrix globosa using partial sequence of mitochondrial DNA.

Restriction fragment length polymorphism (RFLP) of mitochondrial DNA (mtDNA) had been used for molecular identification of Sporothrix spp., which is the causative fungi of sporotrichosis and the most prevalent deep-seated dermatomycosis. Also, mtDNA-RFLP had been used to investigate the molecular epidemiology of sporotrichosis. While the current standard for molecular diagnosis is performed by sequence analysis of the calmodulin gene (CAL), correspondence between the results from CAL and mtDNA is of diagnostic and epidemiological interest. Here, we investigated the correspondence between CAL and mtDNA used for molecular identification of Sporothrix globosa and S. schenckii, which are two major species. We also investigated and propose molecular markers suitable to describe the epidemiology of S. globosa, which is considered as a species with few intraspecific polymorphisms. Eighty-seven strains morphologically identified as S. schenckii sensu lato were investigated. They were identified as group A (17 types, 17 strains) or B (14 types, 70 strains) by mtDNA-RFLP. Partial sequences of CAL, internal transcribed spacer, and spacer between atp9 and cox2 genes of mtDNA of these strains were determined. All group A strains corresponded to S. schenckii, and group B to S. globosa. The sequences of the amplicons targeted on the spacer region in mtDNA of S. globosa ranged 510-515 bp in length and exhibited 10 molecular variations, whereas CAL indicated seven molecular variations. In conclusion, most of the S. schenckii sensu lato strains isolated from Japanese sporotrichosis patients were confirmed as S. globosa, because group B, which comprised the majority of strains, matched perfectly with S. globosa by the CAL sequencing study. We proposed sequence variations in the spacer between atp9 and cox2 genes of mtDNA as a suitable molecular epidemiological marker for S. globosa.

A Case of Recalcitrant Phaeohyphomycosis of the Face Caused by Exophiala lecanii-corni.

We describe a case of recalcitrant phaeohyphomycosis caused by Exophiala lecanii-corni, which was previously reported as Exophiala jeanselmei, infection. A 63-year-old Japanese woman presented with a 15-year history of multiple pruritic erythematous patches and plaques on the face. Histopathological examination and fungal culture revealed phaeohyphomycosis by E. jeanselmei. The attempted treatments included 6 g/day 5-flucytosine (5-FC), 100 mg/day itraconazole (ITCZ), and local hyperthermia. 5-FC was effective initially, but the patient deteriorated after discontinuation. Subsequently, she was referred to our hospital. Histopathological examination showed granuloma with multinucleated giant cells with infiltrating fungal hyphae in the dermis. The causative fungus was finally identified as E. lecanii-corni by ribosomal RNA gene analysis. The patient improved after receiving 200 mg/day ITCZ orally for 15 months with local hyperthermia. In the present case, we confirmed the identification of E. lecanii-corni as the causative agent by molecular methods. We also emphasize the importance of combination therapy with antimycotic agents and local hyperthermia in phaeohyphomycosis.