Dermatophytes and skin dermatophytoses in Southeast Asia-First epidemiological survey from Cambodia.

BACKGROUND: Dermatomycoses count to the most frequent dermatoses in Cambodia. OBJECTIVES: The aim of this survey was to investigate the occurrence of dermatophytes in this Southeast Asian country. METHODS: From June 2017 to July 2018, skin scrapings were taken from 67 patients with superficial dermatophytosis for mycological diagnostics. Identification of dermatophytes was confirmed by sequencing of the 'internal transcribed spacer'-(ITS) region of the rDNA, and the gene of the Translation Elongation Factor (TEF)-1α. RESULTS: Patients were suffering from tinea corporis and tinea inguinalis/cruris 42/67 (63%), tinea capitis/faciei 14/67 (21%), tinea corporis/capitis/faciei 6/67 (9%), tinea manuum/pedis 2/67 (3%), tinea pedis 2/67 (3%) and tinea manuum 1/67 (1%). Both, by culture and/or PCR, a dermatophyte was detected in 52 (78%) out of 67 samples. Culture positive were 42 (81%) of 52, PCR positive were 50 (96%). The following dermatophytes were found: Trichophyton (T.) rubrum, 36/52 strains (69%, 29 by culture), T. mentagrophytes/T. interdigitale (TM/TI) 9/52 (17%, six by culture) and Microsporum (M.) canis 5/52 strains (10%, by culture). One strain of Nannizzia (N.) incurvata 1/52 (2%) and N. nana 1/52 (2%) was isolated. Based on sequencing, we demonstrated that two T. mentagrophytes strains out of the nine TM/TI represented the new ITS genotype XXV Cambodia. We found one T. mentagrophytes strain genotype VIII (now, reclassified as T. indotineae). This isolate was terbinafine resistant, and it exhibited the amino acid substitution Phe397Leu in the squalene epoxidase. Three strains of T. interdigitale genotype II* were isolated. CONCLUSION: This is the first survey on epidemiology of dermatophytes in Cambodia. Currently, T. rubrum represents the most frequent species in Cambodia. One Indian strain genotype VIII T. mentagrophytes was found. A highlight was the first description of the new T. mentagrophytes genotype XXV Cambodia.

Resistance Profile, Terbinafine Resistance Screening and MALDI-TOF MS Identification of the Emerging Pathogen Trichophyton indotineae.

The emerging pathogen Trichophyton indotineae, often resistant to terbinafine (TRB), is known to cause severe dermatophytoses such as tinea corporis and tinea cruris. In order to achieve successful treatment for these infections, insight in the resistance profile of T. indotineae strains and rapid, reliable identification is necessary. In this research, a screening medium was tested on T. indotineae strains (n = 20) as an indication tool of TRB resistance. The obtained results were confirmed by antifungal susceptibility testing (AST) for TRB following the in vitro broth microdilution reference method. Additionally, AST was performed for eight other antifungal drugs: fluconazole, itraconazole, voriconazole, ketoconazole, griseofulvin, ciclopirox olamine, naftifine and amorolfine. Forty-five percent of the strains were confirmed to be resistant to terbinafine. The TRB resistant strains showed elevated minimal inhibitory concentration values for naftifine and amorolfine as well. DNA sequencing of the squalene epoxidase-encoding gene showed that TRB resistance was a consequence of missense point mutations in this gene, which led to amino acid substitutions F397L or L393F. MALDI-TOF MS was used as a quick, accurate identification tool for T. indotineae, as it can be challenging to distinguish it from closely related species such as Trichophyton mentagrophytes or Trichophyton interdigitale using morphological characteristics. While MALDI-TOF MS could reliably identify ≥ 95% of the T. indotineae strains (depending on the spectral library), it could not be used to successfully distinguish TRB susceptible from TRB resistant strains.

S1 Guideline onychomycosis.

Onychomycosis is a fungal infection of the fingernails and toenails. In Europe, tinea unguium is mainly caused by dermatophytes. The diagnostic workup comprises microscopic examination, culture and/or molecular testing (nail scrapings). Local treatment with antifungal nail polish is recommended for mild or moderate nail infections. In case of moderate to severe onychomycosis, oral treatment is recommended (in the absence of contraindications). Treatment should consist of topical and systemic agents. The aim of this update of the German S1 guideline is to simplify the selection and implementation of appropriate diagnostics and treatment. The guideline was based on current international guidelines and the results of a literature review conducted by the experts of the guideline committee. This multidisciplinary committee consisted of representatives from the German Society of Dermatology (DDG), the German-Speaking Mycological Society (DMykG), the Association of German Dermatologists (BVDD), the German Society for Hygiene and Microbiology (DGHM), the German Society of Pediatric and Adolescent Medicine (DGKJ), the Working Group for Pediatric Dermatology (APD) and the German Society for Pediatric Infectious Diseases (DGPI). The Division of Evidence-based Medicine (dEBM) provided methodological assistance. The guideline was approved by the participating medical societies following a comprehensive internal and external review.

Tinea capitis et barbae caused by Trichophyton tonsurans: A retrospective cohort study of an infection chain after shavings in barber shops.

BACKGROUND: Tinea capitis is a highly contagious infectious disease caused by dermatophytes. In Central Europe, it is mainly caused by zoophilic dermatophytes, as, for example Microsporum (M) canis or Trichophyton (T) mentagrophytes and increasingly by anthropophilic fungi. T tonsurans was commonly related to the Tinea gladiatorum, where transmission occurred between infected persons or via contaminated floors. OBJECTIVE: Reporting the transmission of this highly contagious dermatophyte for the first time via beard shaving and hairdressing in barber shops in Germany. PATIENTS AND METHODS: 18 young male patients developed tinea capitis and/or barbae shortly after shavings of the beard and/or hair in a barber shop. Native, cultural and molecular diagnostics as well as tissue biopsies and resistance tests were performed of skin and hair samples. RESULTS: In all samples, T tonsurans could be identified. The medical history and the clinical picture suggest a transmission through contaminated hairdressing tools. The patients were treated with terbinafine or itraconazole in combination with or exclusively with topical antimycotics. CONCLUSION: The transmission and a resulting increase in the incidence of infections with T tonsurans may be due to shavings with direct skin contact of insufficiently disinfected hairdressing tools. This path of infection has already been observed in Africa and is now being described for the first time in Germany. Knowledge of the pathogen and its transmission ways are essential to interrupt the chain of infection.

Taxonomy of the Trichophyton mentagrophytes/T. interdigitale Species Complex Harboring the Highly Virulent, Multiresistant Genotype T. indotineae.

A severe outbreak of highly virulent and multi-resistant dermatophytosis by species in the Trichophyton mentagrophytes/T. interdigitale complex is ongoing in India. The correct identity of the etiologic agent is a much-debated issue. In order to define species limits, a taxonomic study was undertaken combining molecular, morphological, and physiological characteristics as evidence of classification. Molecular characteristics show that T. mentagrophytes s. str. and T. interdigitale s. str. can be distinguished with difficulty from each other, but are unambiguously different from the Indian genotype, T. indotineae by sequences of the HMG gene. The entities were confirmed by multilocus analysis using tanglegrams. Phenotypic characters of morphology and physiology are not diagnostic, but statistically significant differences are observed between the molecular siblings. These properties may be drivers of separate evolutionary trends. Trichophyton mentagrophytes represents the ancestral, homothallic cloud of genotypes with a probable geophilic lifestyle, while T. indotineae and T. interdigitale behave as anthropophilic, clonal offshoots. The origin of T. indotineae, which currently causes a significant public health problem, is zoonotic, and its emergence is likely due to widespread misuse of antifungals.

[Trichophyton tonsurans-an emerging pathogen in wrestling in Germany]. / Trichophyton tonsurans ­ ein Emerging-Pathogen im Ringsport in Deutschland.

Trichophyton (T.) tonsurans is considered as the main causative agent of tinea gladiatorum (ringworm) in contact and martial arts worldwide and regularly leads to outbreaks. In the national wrestling squad in Leipzig, dermatophytoses occurred frequently and recurrently in children and adolescents for over a 2-year period. The wrestlers came to the dermatologist's office for clinical examination and sampling. Dermal scales and hair roots as well as smears were examined mycologically with fluorescence optical preparation, fungal culture, and polymerase chain reaction (PCR) for dermatophyte DNA. Sequencing of the dermatophyte rDNA served as culture confirmation test. Environmental investigations in the wrestler training center included contact cultures and smears from surfaces, in particular from the mats. T. tonsurans was culturally and/or with PCR detectable in 21 out of 25 children and adolescents plus one trainer. T. tonsurans grew in one of ten contact cultures of mats and floors in the wrestling training center, and T. interdigitale was found in another culture. Smears from the mats resulted in a culture of T. tonsurans detection twice. The PCR was positive for T. tonsurans three times. Within 14 days, T. tonsurans developed small, flat, radiating, granular and white-colored colonies with a mahogany-brown reverse side on the fungal culture media. The sequencing of the internal transcribed spacer (ITS) region of the rDNA and the translation elongation factor 1 α (TEF 1 α) gene confirmed the species T. tonsurans in all cases. T. interdigitale that was found from a mat was also identified by sequencing. Eight T. tonsurans strains were subjected to in vitro susceptibility testing to terbinafine. All isolates were sensitive to terbinafine in vitro with minimal inhibitory concentrations of ≤ 0.1 µg/ml.

[Report of the first case of tinea corporis bullosa by Trichophyton tonsurans]. / Bericht des ersten Falls einer bullösen Tinea corporis durch Trichophyton tonsurans.

Blister formation in tinea corporis is rare. Bullous tinea is usually evoked by zoophilic dermatophytes. A 20-year-old woman presented in our out-patient department with a painful pruritic bullous skin eruption at the left forearm. Clinically, a 4â€¯× 3 cm symmetrical plaque with sharp borders and peripheral versiculation and serous crusts was seen. Histologically there was a marked spongiotic dermatitis with fungal elements in periodic acid stain (PAS). By fungal culture, PCR and gene sequencing Trichophyton (T.) tonsurans was identified. To the best of our knowledge, this is the first published case of T. tonsurans as the causative agent of bullous tinea corporis.

Visualising virulence factors: Trichophyton benhamiaes subtilisins demonstrated in a guinea pig skin ex vivo model.

BACKGROUND: Dermatophytoses rank among the most frequent communicable diseases in humans, and the zoonotic transmission is increasing. The zoophilic dermatophyte Trichophyton (T.) benhamiae is nowadays one of the main causes of tinea faciei et corporis in children. However, scientific data on molecular pathomechanisms and specific virulence factors enabling this ubiquitous occurrence are scarce. OBJECTIVES: To study tissue invasion and the expression of important virulence factors of T. benhamiae, isolates that were recovered from two groups of hosts (humans vs. guinea pigs (GP)) using an ex vivo skin model. METHODS: After confirmation of species identity by ITS sequencing, CFU suspensions of dermatophyte isolates (n = 20) were applied to the skin infection model and cultured. Employing specific immunofluorescence staining techniques, the expression of subtilisin 3 and 6 and metallocarboxypeptidase A was analysed. The general mode of invasion was explored. Results were compared with biopsies of naturally infected GP. RESULTS: All isolates were successfully recovered and proliferated well after application to the infection model. Progressive invasion of hyphae through all skin structures and destruction of explants were observed with early events being comparable to natural infection. An increasing expression of the examined virulence factors towards the end of culture was noticed but no difference between the two groups of isolates. CONCLUSIONS: For the first time, important in vivo markers of dermatophytosis were visualised immunohistochemically in an ex vivo skin infection model and in skin biopsies of GP naturally infected with T. benhamiae. More research on the underlying pathomechanisms of dermatophyte infection is urgently needed.

Indian Trichophyton mentagrophytes squalene epoxidase erg1 double mutants show high proportion of combined fluconazole and terbinafine resistance.

BACKGROUND: The Indian ITS genotype VIII Trichophyton mentagrophytes population shows a high amount of different erg1 (ergosterol) mutants encoding for squalene epoxidase, which catalyses the first step of ergosterol biosynthesis. OBJECTIVES: Illumination of the implication of point mutations at position Ala448Thr in single and double erg1 T mentagrophytes mutants because mutants of this type were abundantly found within the Indian fungal population. METHODS: Growth in fluconazole or terbinafine containing medium was analysed using a microplate-laser-nephelometry (MLN)-based growth assay. RESULTS: Ala 448 Thr erg1 single mutants were terbinafine sensitive, but about 50% of isolates showed an increased fluconazole resistance, whereas 95% of the double mutants (Phe 397 Leu, Ala 448Thr) demonstrated combined terbinafine and increased fluconazole resistance. CONCLUSION: The new Indian T mentagrophytes populations show several point mutations in erg1. Point mutations at position 397 were previously described and cause terbinafine resistance. A large part of the double mutants exhibit resistance to terbinafine and fluconazole, demonstrating a selective advantage of the combination of both mutations.

Alarming India-wide phenomenon of antifungal resistance in dermatophytes: A multicentre study.

BACKGROUND: An alarming increase in recalcitrant dermatophytosis has been witnessed in India over the past decade. Drug resistance may play a major role in this scenario. OBJECTIVES: The aim of the present study was to determine the prevalence of in vitro resistance to terbinafine, itraconazole and voriconazole in dermatophytes, and to identify underlying mutations in the fungal squalene epoxidase (SQLE) gene. PATIENTS/METHODS: We analysed skin samples from 402 patients originating from eight locations in India. Fungi were identified by microbiological and molecular methods, tested for antifungal susceptibility (terbinafine, itraconazole, voriconazole), and investigated for missense mutations in SQLE. RESULTS: Trichophyton (T.) mentagrophytes internal transcribed spacer (ITS) Type VIII was found in 314 (78%) samples. Eighteen (5%) samples harboured species identified up to the T interdigitale/mentagrophytes complex, and T rubrum was detected in 19 (5%) samples. 71% of isolates were resistant to terbinafine. The amino acid substitution Phe397Leu in the squalene epoxidase of resistant T mentagrophytes was highly prevalent (91%). Two novel substitutions in resistant Trichophyton strains, Ser395Pro and Ser443Pro, were discovered. The substitution Ala448Thr was found in terbinafine-sensitive and terbinafine-resistant isolates but was associated with increased MICs of itraconazole and voriconazole. CONCLUSIONS: The high frequencies of terbinafine resistance in dermatophytes are worrisome and demand monitoring and further research. Squalene epoxidase substitutions between Leu393 and Ser443 could serve as markers of resistance in the future.

Genes Encoding Proteolytic Enzymes Fungalysin and Subtilisin in Dermatophytes of Human and Animal Origin: A Comparative Study.

Dermatophytes are among the most successful fungal pathogens in humans, but their virulence mechanisms have not yet been fully characterized. Dermatophytic fungi secrete proteases in vivo, which are responsible for fungal colonization and degradation of the keratinized tissue during infection. In the present study, we used PCR to investigate the presence of genes encoding fungalysins (MEP) and subtilisins (SUB) in three dermatophyte species whose incidence is increasing in Europe: the anthropophilic Trichophyton rubrum (n = 58), zoophilic Microsporum canis (n = 33), and Trichophyton benhamiae (n = 6). MEP2 and SUB4 genes were significantly correlated with T. rubrum; MEP3 and SUB1 were mostly frequently harbored by M. canis; and MEP1, 2, and 4 and SUB3-7 were most frequently harbored by T. benhamiae isolates (p < 0.05). Furthermore, MEP1-5 and SUB1-3 genes were significantly more prevalent among human clinical isolates of M. canis (n = 17) than among asymptomatic cat isolates of M. canis (n = 16; p < 0.05). Unidentified MEP and/or SUB genes in some isolates in the current study may suggest that other gene repertoires may be involved in the degradation of keratin. The presented analysis of the incidence of MEP and SUB virulence genes in three dermatophyte species of diverse origins provides an insight into the host-fungus interaction and dermatophyte pathogenesis.

[Microsporum ferrugineum-an anthropophilic dermatophyte in Germany : Case report and review of the literature]. / Microsporum ferrugineum ­ ein anthropophiler Dermatophyt in Deutschland : Patientenbeschreibung und Übersicht über die Literatur.

Three boys from the same city, treated by the same dermatologist, developed tinea capitis. Two of them, 4 and 8 years old, underwent mycological diagnostic workup. However, no pathogens familiar in this country, such as Microsporum (M.) canis or Trichophyton (T.) tonsurans, were isolated, but instead that of a dermatophyte that has not been found in Germany for decades. Both dermatophyte isolates showed white-beige-brownish colonies with a flat, radiating edge and a central, verrucous curvature. The sequencing of the internal transcribed spacer (ITS) region of the rDNA confirmed the suspicion of M. ferrugineum already expressed based on the morphological picture. The anthropophilic dermatophyte occurs in the Middle East, Asia, Eastern Europe and Africa and is considered to be the cause of tinea capitis or tinea corporis in children and adolescents. In 2016, M. ferrugineum has again been isolated in Germany, probably as a result of migration movements. The fungus is strikingly isolated to martial arts, especially wrestlers. It mainly affects children and adolescents, some with a Russian-German background. The anthropophilic dermatophyte is transmitted directly from person to person, especially in the case of tinea capitis. An indirect transmission, for example, via mats in martial arts is likely.

[Occupational Trichophyton verrucosum infection in a cattle farmer]. / Berufliche Infektion mit Trichophyton verrucosum bei einem Rinderzüchter.

Ringworm in cattle may cause an occupational skin disease in humans. Trichophyton verrucosum lead to a highly inflammatory fungal skin infection that is often misdiagnosed as bacterial disease and consequently mistreated with antibiotics. To establish the correct diagnosis, it is necessary to collect skin scales; in addition, a skin biopsy can be helpful. Deep dermatophyte infections by Trichophyton verrucosum can be treated effectively with oral terbinafine. In addition, it is necessary to pay careful attention to use suitable hygiene measures in the stables to protect against reinfection.

Database establishment for the secondary fungal DNA barcode translational elongation factor 1α (TEF1α) 1.

With new or emerging fungal infections, human and animal fungal pathogens are a growing threat worldwide. Current diagnostic tools are slow, non-specific at the species and subspecies levels, and require specific morphological expertise to accurately identify pathogens from pure cultures. DNA barcodes are easily amplified, universal, short species-specific DNA sequences, which enable rapid identification by comparison with a well-curated reference sequence collection. The primary fungal DNA barcode, ITS region, was introduced in 2012 and is now routinely used in diagnostic laboratories. However, the ITS region only accurately identifies around 75% of all medically relevant fungal species, which has prompted the development of a secondary barcode to increase the resolution power and suitability of DNA barcoding for fungal disease diagnostics. The translational elongation factor 1α (TEF1α) was selected in 2015 as a secondary fungal DNA barcode, but it has not been implemented into practice, due to the absence of a reference database. Here, we have established a quality-controlled reference database for the secondary barcode that together with the ISHAM-ITS database, forms the ISHAM barcode database, available online at http://its.mycologylab.org/ . We encourage the mycology community for active contributions.

A clarion call for preventing taxonomical errors of dermatophytes using the example of the novel Trichophyton mentagrophytes genotype VIII uniformly isolated in the Indian epidemic of superficial dermatophytosis.

An alarming pan Indian increase in the incidence of superficial dermatophytosis has been noticed over the past 5-6 years. Recent studies have demonstrated emerging predominance of Trichophyton (T.) mentagrophytes as the causative organism in such cases. Interestingly, a distinct Indian genotype of T. mentagrophytes has been identified and recognised with the help of sequencing of the ITS region of the rDNA. That has, however, led to a basic confusion owing to the newly introduced taxonomy of dermatophytes in 2017. According to this most recently suggested classification and new taxonomy of dermatophytes, the former "T. mentagrophytes complex" is differentiated into T. mentagrophytes (zoophilic strains) and T. interdigitale (anthropophilic strains). We have noticed that in some recent studies the causative agent of the chronic, relapsing dermatophytosis outbreak in India has been described as T. interdigitale. In our opinion, it is very likely that these T. interdigitale strains isolated in Delhi and Chennai in India are indeed strains more closely related to the neotype of T. mentagrophytes and not strains of T. interdigitale. We therefore want to underscore the importance of a common nomenclature of species in accordance with the new taxonomy of dermatophytes. This would most likely facilitate better understanding of the issue amongst dermatologists and microbiologists in general. Mistaken identification of Trichophyton isolates not limited to India is very likely to occur due to the lack of appropriate molecular diagnosis which in turn is based on the already published data that presumably wrongly identify one species instead of the other.

The current Indian epidemic of superficial dermatophytosis due to Trichophyton mentagrophytes-A molecular study.

The disease burden of chronic-relapsing and therapy-refractory superficial dermatophytosis dramatically increased in India within the past 5-6 years. In order to evaluate the prevalence of this trend, 201 skin scrapings were collected from patients from all parts of India and were tested for dermatophytes using both fungal culture and a PCR-ELISA directly performed with native skin scrapings. Fungal culture material was identified by genomic Sanger sequencing of the internal transcribed spacer (ITS) region and the translation elongation factor (TEF)-1α gene. In total, 149 (74.13%) out of the 201 samples showed a dermatophyte-positive culture result. Out of this, 138 (92.62%) samples were identified as Trichophyton (T.) mentagrophytes and 11 (7.38%) as Trichophyton rubrum. The PCR-ELISA revealed similar results: 162 out of 201 (80.56%) samples were dermatophyte-positive showing 151 (93.21%) T mentagrophytes- and 11 (6.79%) T rubrum-positive samples. In this study, we show for the first time a dramatic Indian-wide switch from T rubrum to T mentagrophytes. Additionally, sequencing revealed a solely occurring T mentagrophytes "Indian ITS genotype" that might be disseminated Indian-wide due to the widespread abuse of topical clobetasol and other steroid molecules mixed with antifungal and antibacterial agents.

[Extensive tinea corporis due to a terbinafine-resistant Trichophyton mentagrophytes isolate of the Indian genotype in a young infant from Bahrain in Germany]. / Ausgeprägte Tinea corporis durch ein Terbinafin-resistentes Trichophyton-mentagrophytes-Isolat vom indischen Genotyp bei einem Säugling aus Bahrain in Deutschland.

A 6 month-old-female infant from Bahrain visiting Germany with her family for a holiday was seen by us for extensive dermatophytosis of the back, buttocks, chest and groins. Topical treatment by terbinafine for over 2 months was not successful. Other family members including adults and children were treated in Bahrain with topical antifungals and oral voriconazole which was not helpful. Mycological examination performed in Germany revealed the detection of the zoophilic dermatophyte Trichophyton (T.) mentagrophytes. The newly described genotype VIII within the species T. mentagrophytes was identified by sequencing of the "internal transcribed spacer" (ITS) region of the fungal rDNA. This genotype of T. mentagrophytes is the main causative agent of the current epidemic of chronic recalcitrant dermatophytoses in India. Transmission of this Indian genotype of T. mentagrophytes to other countries due to globalization is a serious issue to be considered. Moreover, a significant percentage of these Indian T. mentagrophytes strains are resistant to terbinafine both in vitro and by the way of genetic point mutations in the squalene epoxidase (SQLE) gene. Some are also found to be partially resistant against itraconazole and voriconazole. The point mutation TTC/TTA was found by SQLE mutation analysis in this particular T. mentagrophyte isolate from Bahrain. This point mutation is closely associated with F397L amino acid substitution of the enzyme indicative of in vitro resistance of the dermatophyte against terbinafine. The girl was successfully treated by topical miconazole and later by ciclopirox olamine. This is the first report on an infection due to a terbinafine-resistant T. mentagrophytes strain of the ITS genotype VIII from India in Germany.

MALDI-TOF MS analysis of bovine and zoonotic Trichophyton verrucosum isolates reveals a distinct peak and cluster formation of a subgroup with Trichophyton benhamiae.

The zoophilic dermatophyte Trichophyton verrucosum is the most important causative agent of bovine dermatophytosis. Additionally, it causes profound and poorly healing skin infections in humans indicating the high zoonotic potential. The objective of this study was to establish differentiation of T. verrucosum from other dermatophytes by mass spectrometry and to identify distinct features of the mass spectra. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was successful for identification of this pathogen only after extension of the database of the manufacturer with spectra from T. verrucosum strains, which were identified as such by sequencing of the internal transcribed spacer (ITS) region. MALDI-TOF MS analysis was conducted with 46 field isolates from cattle, two live vaccine strains, and 10 isolates from humans identified as T. verrucosum by sequence analysis of the ITS region. The results suggest a very good agreement of both methods. Comparison with the mass spectra of 68 strains of other keratinophilic fungi revealed that most T. verrucosum wild-type isolates showed a characteristic peak at 7950-7954 m/z, which was missing in the spectra of other keratinophilic fungi and the live vaccine strains. The spectra of T. verrucosum were most similar to the spectra of T. benhamiae, an emerging zoophilic dermatophyte. In summary, MALDI-TOF MS is a powerful and reliable tool to identify T. verrucosum.

Deep facial mycosis due to Trichophyton verrucosum-molecular genetic identification of the dermatophyte in paraffin-embedded tissue-case report and review of the literature.

Deep trichophytosis is relatively uncommon. The infection of the bearded area is also known as sycosis barbae or tinea barbae and can be caused by various fungal species, most often zoophilic fungi. We report on an 80-year-old male patient with severe sycosis barbae who had no animal contact and was treated with systemic antibiosis without improvement. Microbial and mycological investigations using swabs from oozing lesions revealed Staphylococcus haemolyticus and Candida parapsilosis. Histology demonstrated fungal elements in hair follicles. Paraffin-embedded material was subjected to further mycological analysis. For molecular diagnostics DNA was prepared from paraffin sections for real-time polymerase chain reaction (RT-PCR). For sequencing, DNA was isolated from paraffin-embedded skin tissue and the ITS region of the rDNA was selected. Sequencing of the ITS2 region of rRNA revealed a 100% accordance with Trichophyton (T.) verrucosum. Treatment with oral terbinafine achieved a complete remission. Sycosis barbae is an important differential diagnosis for infections of the bearded area. Nucleic acid amplification techniques (NAAT) are more and more used for direct examination of dermatophytes in clinical samples, eg T. verrucosum. NAAT are also used as culture confirmation tests for identification of rare dermatophytes like T. verrucosum. Today, singleplex and multiplex quantitative real-time PCR (qRT-PCR) assays for the detection of the most common dermatophytes including T. verrucosum in clinical specimens are available. Recently, an ITS2 PCR assay has been successfully used for direct detection of T. verrucosum in paraffin-embedded formalin-fixed skin tissue. The PCR is fast and highly specific. The sensitivity of direct molecular detection of the dermatophytes both in native clinical material, and in paraffin-embedded skin tissue can been increased.