A conceptual framework for nomenclatural stability and validity of medically important fungi: a proposed global consensus guideline for fungal name changes supported by ABP, ASM, CLSI, ECMM, ESCMID-EFISG, EUCAST-AFST, FDLC, IDSA, ISHAM, MMSA, and MSGERC.

The rapid pace of name changes of medically important fungi is creating challenges for clinical laboratories and clinicians involved in patient care. We describe two sources of name change which have different drivers, at the species versus the genus level. Some suggestions are made here to reduce the number of name changes. We urge taxonomists to provide diagnostic markers of taxonomic novelties. Given the instability of phylogenetic trees due to variable taxon sampling, we advocate to maintain genera at the largest possible size. Reporting of identified species in complexes or series should where possible comprise both the name of the overarching species and that of the molecular sibling, often cryptic species. Because the use of different names for the same species will be unavoidable for many years to come, an open access online database of the names of all medically important fungi, with proper nomenclatural designation and synonymy, is essential. We further recommend that while taxonomic discovery continues, the adaptation of new name changes by clinical laboratories and clinicians be reviewed routinely by a standing committee for validation and stability over time, with reference to an open access database, wherein reasons for changes are listed in a transparent way.

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.

Antifungal Susceptibility and Mutations in the Squalene Epoxidase Gene in Dermatophytes of the Trichophyton mentagrophytes Species Complex.

During the past decade, a prolonged and serious outbreak of dermatophytosis due to a terbinafine-resistant novel species in the Trichophyton mentagrophytes-T. interdigitale complex has been ongoing in India, and it has spread to several European countries. The objective of this study was to investigate the molecular background of the squalene epoxidase (SQLE) gene in order to understand the risk of emergence and spread of multiresistance in dermatophytes. Antifungal susceptibility to fluconazole, griseofulvin, itraconazole, ketoconazole, miconazole, naftifine, sertaconazole, and terbinafine was tested in 135 isolates from India, China, Australia, Germany, and The Netherlands. Based on the latest taxonomic insights, strains were identified as three species: T. mentagrophytes sensu stricto (n = 35), T. indotineae (n = 64, representing the Indian clone), and T. interdigitale sensu stricto (n = 36). High MICs of terbinafine (>16 mg/liter) were found in 34 (53%) T. indotineae isolates. These isolates showed an amino acid substitution in the 397th position of the SQLE gene. Elevated MICs of terbinafine (0.5 mg/liter) were noted in 2 (3%) T. indotineae isolates; these isolates lead to Phe415Val and Leu393Ser of the SQLE gene. The stability of the effect of the mutations was proven by serial transfer on drug-free medium. Lys276Asn and Leu419Phe substitutions were found in susceptible T. mentagrophytes strains. The Phe377Leu/Ala448Thr double mutant showed higher MIC values for triazoles. High MICs of terbinafine are as yet limited to T. indotineae and are unlikely to be distributed throughout the T. mentagrophytes species complex by genetic exchange.

[Arthroderma crocatum on human skin]. / Arthroderma crocatum auf menschlicher Haut.

We report on three cases in which Arthroderma (A.) crocatum was isolated from human skin in Germany. The characteristics and epidemiology of this rare geophilic and probably mostly apathogenic dermatophyte are described paying special attention to its gymnothecia. The combination of KOH mount, culture and genetic analysis is the foundation for clinically meaningful conclusions. It is likely that the prevalence of A. crocatum is currently underestimated.

"Indian" strains of Trichophyton mentagrophytes with reduced itraconazole susceptibility in Germany.

We use published reports and three of our own tinea cases as an opportunity to report on "Indian" strains of Trichophyton (T.) mentagrophytes with ITS genotype VIII and reduced susceptibility to itraconazole due to the mutation c.1342G>A in the SQLE gene in Germany. In vitro measurements of resistance revealed normal susceptibility to terbinafine, but markedly reduced susceptibility to itraconazole - although no valid breakpoints are currently defined and minimum inhibitory concentrations (MICs) depend on the methods used. Problems related to the determination and interpretation of MICs are outlined. Our cases show that azole-resistant "Indian" strains of T. mentagrophytes with ITS genotype VIII occurred in Germany as early as 2011, which is earlier than was previously assumed. This variant of the pathogen cannot be phenotypically distinguished from customary strains of T. mentagrophytes; its identification is based on genetics. The taxonomic classification is still under debate. This variant is anthropophilic and causes only mildly inflammatory tinea lesions with many fungal elements. Its further dissemination must therefore be expected. Prerequisites for rapid and valid antimycotic testing against dermatophytes need to be developed.

A Hundred Years of Diagnosing Superficial Fungal Infections: Where Do We Come From, Where Are We Now and Where Would We Like To Go?

Superficial fungal infections have been known for hundreds of years. During the 20th century new diagnostic methods were developed and the taxonomy changed several times, which, unfortunately, resulted in many fungi having several names (synonyms). The taxonomy is important, as species-specific identification guides clinicians when choosing the most appropriate antifungal agent, and provides an indication of the source of infection (anthropophilic, zoophilic or geophilic). Traditional diagnostic tests (direct microscopy, culture and histopathology) are still widely used, but molecular-based methods, such as PCR, have many advantages, and increasingly supplement or replace conventional methods. Molecular-based methods provide detection of different genus/species spectra. This paper describes recent changes in dermatophyte taxonomy, and reviews the currently available diagnostics tools, focusing mainly on commercially available PCR test systems.

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.

Polyphasic Discrimination of Trichophyton tonsurans and T. equinum from Humans and Horses.

The anthropophilic dermatophyte Trichophyton tonsurans and its zoophilic counterpart T. equinum are phylogenetically closely related. The barcoding marker rDNA internal transcribed spacer (ITS) shows limited variation between these two species. In the current study, we combined molecular approaches with phenotypic data to determine the species boundaries between T. tonsurans (n = 52) and T. equinum (n = 15) strains originating from humans (n = 40), horses (n = 26), and a mouse (n = 1). Culture characteristics and physiology on Trichophyton agar media 1 and 5 were evaluated. Multi-locus sequencing involving ITS, partial large rDNA subunit (LSU), ß-tubulin (TUB), 60S ribosomal protein (RPB), and translation elongation factor-3 (TEF3) genes, and the mating-type (MAT) locus was performed. Amplified fragment length polymorphism data were added. None of the test results showed complete mutual correspondence. With the exception of strains from New Zealand, strains of equine origin required niacin for growth, whereas most strains from human origin did not show this dependence. It is concluded that T. tonsurans and T. equinum incompletely diverged from a common lineage relatively recently. MAT1-1 and MAT1-2 are the main distinguishing genes between the two species.

S1 guidelines: Tinea capitis.

Tinea capitis describes a dermatophyte infection of scalp and hair that predominately occurs in children. The diagnostic workup includes microscopic examination, culture and/or molecular tests. Treatment is guided by the specific organism involved and should consist of systemic agents as well as adjuvant topical treatment. The aim of the present update of the interdisciplinary German S1 guidelines is to provide dermatologists, pediatricians and general practitioners with a decision tool for selecting and implementing appropriate diagnostic and therapeutic measures in patients with tinea capitis. The guidelines were developed based on current international guidelines, in particular the 2010 European Society for Pediatric Dermatology guidelines and the 2014 British Association of Dermatologists guidelines, as well as on a review of the literature conducted by the guideline committee. This multidisciplinary committee consists of representatives from the German Society of Dermatology (DDG), the German-Speaking Mycological Society (DMykG), the German Society for Hygiene and Microbiology (DGHM), the German Society of Pediatric and Adolescent Medicine (DGKJ) and the German Society for Pediatric Infectious Diseases (DGPI). The Division of Evidence-based Medicine (dEBM) provided methodological assistance. The guidelines were approved by the participating medical societies following a comprehensive internal and external review.

Species Distinction in the Trichophyton rubrum Complex.

The Trichophyton rubrum species complex comprises commonly encountered dermatophytic fungi with a worldwide distribution. The members of the complex usually have distinct phenotypes in culture and cause different clinical symptoms, despite high genome similarity. In order to better delimit the species within the complex, molecular, phenotypic, and physiological characteristics were combined to reestablish a natural species concept. Three groups, T. rubrum, T. soudanense, and T. violaceum, could be distinguished based on the sequence of the internal transcribed spacer (ITS) ribosomal DNA barcode gene. On average, strains within each group were similar by colony appearance, microscopy, and physiology, but strains between groups showed significant differences. Trichophyton rubrum strains had higher keratinase activity, whereas T. violaceum strains tended to be more lipophilic; however, none of the phenotypic features were diagnostic. The results of matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and amplified fragment length polymorphism (AFLP) were partially consistent with the ITS data but failed to distinguish the species unambiguously. Despite their close similarity, T. violaceum, T. soudanense, and T. rubrum can be regarded as independent species with distinct geographical distributions and clinical predilections. Trichophyton soudanense is pheno- and genotypically intermediate between T. rubrum and T. violaceum For routine diagnostics, ITS sequencing is recommended.

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.

Arthroderma chiloniense sp. nov. isolated from human stratum corneum: Description of a new Arthroderma species.

A 68-year-old woman was submitted to our hospital because of erythematous and scaly skin lesions. To exclude tinea samples of stratum corneum were collected and used for mycological investigations. In this material, no fungal elements were detected microscopically, but inoculation on Sabouraud agar with cycloheximide yielded a presumptive dermatophyte fungus. Subsequent detailed investigations with conventional morphological and physiological methods and a phylogenetic analysis of the combined LSU rRNA gene (D1/D2 domains) and ITS region sequences suggested that the fungus represents a hitherto undescribed species of the genus Arthroderma. Here, we describe this species as Arthroderma chiloniense sp. nov., EMBL accession no. LT992885. This new species can be distinguished from phylogenetically related Arthroderma species using ribosomal ITS and LSU genes, and 60S L10 protein sequences; specific macroscopic, microscopic and physiological features are lacking. Our attempts to re-isolate this fungus from the patient's skin failed although her skin lesions persisted. Most likely A. chiloniense is a geophilic species that incidentally contaminated or transiently colonised the patient's skin. To avoid diagnostic misinterpretations, it is necessary to distinguish A. chiloniense from truly pathogenic dermatophytes like Trichophyton (T.) rubrum and T. interdigitale which can easily be confused with A. chiloniense based on similar mycelium morphology.

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.

Trichophyton mentagrophytes - a new genotype of zoophilic dermatophyte causes sexually transmitted infections.

BACKGROUND: A new genotype of the zoophilic fungal species Trichophyton (T.) mentagrophytes was recently described in two studies. It was isolated from three patients who had visited Southeast Asia and one patient who had visited Egypt. In contrast to these studies, we have observed a number of cases with the dimensions of an epidemic outbreak. PATIENTS AND METHODS: At the University Hospital Charité Berlin, 43 patients, mostly suffering from highly inflammatory, painful and persistent infections of the pubogenital region were observed between January 2016 and July 2017. Mycological examination was performed with fungal culture and sequencing of the ITS (internal transcribed spacer) region of the ribosomal DNA. Three additional genomic regions were spot-checked. RESULTS: In 37 of the cases, a new genotype of T. mentagrophytes (referred to here as T. mentagrophytes VII) was isolated as the etiological agent, and sequencing revealed identical sequences for all isolates. Most of the infected patients had no history of travel, and only two patients reported contact with animals. CONCLUSIONS: The new genotype clustered phylogenetically among the strains of the zoophilic species T. mentagrophytes with four different DNA markers. While human-to-human transmission of zoophilic dermatophytes is rare, transmission via sexual contact seemed to be quite effective here.

Unequal distribution of the mating type (MAT) locus idiomorphs in dermatophyte species.

The mating type (MAT) locus is the key regulator of sexual reproduction in fungi. In the dermatophytes and other Ascomycetes this genomic region exists in two distinct forms (idiomorphs) and their balanced presence is a precondition for successful mating in heterothallic fungi. But the MAT locus not only drives sexual reproduction, it has also been shown to influence pathogenicity, virulence, and/or morphological changes in pathogenic fungi of the genera Candida, Histoplasma, and Cryptococcus. In order to find out whether there are similar trends in dermatophytes, we investigated the MAT locus of 19 anthropophilic and zoophilic species via Sanger sequencing and primer walking. We identified for the first time the MAT locus idiomorphs of the dermatophyte species Microsporum audouinii (MAT1-2), M. ferrugineum (MAT1-2), Trichophyton schoenleinii (MAT1-2), T. bullosum (MAT1-1), T. quinckeanum (MAT1-1), T. concentricum (MAT1-1), T. eriotrephon (MAT1-1), and T. erinacei (MAT1-2). In addition, we determined the MAT locus sequence for dermatophyte species whose mating type idiomorphs had been described on the basis of results of classical confrontation experiments (e.g. M. canis, MAT1-2) and we confirmed recently published molecular data (e.g. T. rubrum, MAT1-2). Our results corroborate that MAT locus idiomorphs are unequally distributed in the majority of the analyzed species and the ability to mate with a partner of the opposite sex is limited to a few zoophilic species. Clonal spreads are identified that are connected to one of the idiomorphs and a higher virulence and/or a higher transmission rate to humans (T. benhamiae and T. mentagrophytes). For the imbalanced idiomorph distribution pattern we hypothesize that either: (I) one of the mating type idiomorphs may be extinct due to clonal reproduction (e.g., T. rubrum and M. canis), (II) mating partners of one species adapted to different hosts followed by speciation in the new niche (e.g., T. equinum and T. tonsurans) or (III) unisexual reproduction is the next evolutionary stage of propagation in dermatophytes which involves the extinction of one mating idiomorph.

New insights in dermatophyte research.

Dermatophyte research has renewed interest because of changing human floras with changing socioeconomic conditions, and because of severe chronic infections in patients with congenital immune disorders. Main taxonomic traits at the generic level have changed considerably, and now fine-tuning at the species level with state-of-the-art technology has become urgent. Research on virulence factors focuses on secreted proteases now has support in genome data. It is speculated that most protease families are used for degrading hard keratin during nitrogen recycling in the environment, while others, such as Sub6 may have emerged as a result of ancestral gene duplication, and are likely to have specific roles during infection. Virulence may differ between mating partners of the same species and concepts of zoo- and anthropophily may require revision in some recently redefined species. Many of these questions benefit from international cooperation and exchange of materials. The aim of the ISHAM Working Group Dermatophytes aims to stimulate and coordinate international networking on these fungi.

Molecular epidemiology of Trichophyton quinckeanum - a zoophilic dermatophyte on the rise.

BACKGROUND: Formerly only referred to as a subspecies (T. mentagrophytes var. quinckeanum), T. quinckeanum once again constitutes a distinct species according to the updated taxonomy of dermatophytes. PATIENTS AND METHODS: During routine diagnostic tests conducted at the Mycology Laboratory, Mölbis, Germany, between 11/2013 to 1/2017 (three years and three months), all specimens sent in were examined for T. quinckeanum. Molecular biology methods employed included: 1) DNA hybridization (PCR ELISA), 2) gene sequencing of the ITS region and TEF-1α, and 3) in some cases, MALDI-TOF mass spectrometry. RESULTS: Overall, 62 strains of T. quinckeanum were found. Sixty-eight percent of patients were female; 43 % were children and adolescents (≤ 19 years of age). Cats were a frequent source of infection. Sequencing of all 62 strains revealed a concordance of 100 % with T. quinckeanum sequences contained in the NCBI database. Isolates analyzed by MALDI-TOF mass spectrometry showed specific spectra. CONCLUSIONS: In Germany, the zoophilic dermatophyte T. quinckeanum currently appears to be more prevalent than expected. T. quinckeanum strains were isolated from children and adults with dermatomycosis and tinea capitis. Sources of infection with T. quinckeanum include small rodents (mice), horses, and - remarkably commonly -  cats. Given that unequivocal morphological identification of this dermatophyte is not always possible, molecular methods have to be employed in the diagnosis.