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.

In vitro investigation of the principle of action of ammonium bituminosulfonate ointments on a 3D skin model.

Ammonium bituminosulfonate preparations have been used for various dermatological diseases since the 19th century. The dark preparation is known as the so-called "drawing salve" (Ichtholan®) for the treatment of abscesses and furuncles. The underlying activity is in part the loosening of the skin, which facilitates pus extraction and treatment of deep inflammations. For this investigation 3D skin models were incubated with ointments containing different ammonium bituminosulfonate concentrations. Histological and immunohistochemical staining as well as penetration investigation were carried out. The effect of dark ammonium bituminosulfonate ointments on skin loosening was investigated to reveal the underlying mechanism. The skin loosening effect could be proved by HE-staining for ammonium bituminosulfonate treated skin models. This effect was concentration dependent. While treatment with ammonium bituminosulfonate ointment had no influence on keratin expression, high concentrated ointments led to decreased filaggrin and laminin expression. Treatment of skin models with ABS ointments led to an increased skin permeability, which was concentration dependent. For the first time the skin loosening effect of ammonium bituminosulfonate ointment has been demonstrated on 3D skin models. This effect is at least in part caused by the interaction of the substance with structure dependent proteins of the epidermis.

Efficacy of antifungal agents against fungal spores: An in vitro study using microplate laser nephelometry and an artificially infected 3D skin model.

Dermal fungal infections seem to have increased over recent years. There is further a shift from anthropophilic dermatophytes to a growing prevalence of zoophilic species and the emergence of resistant strains. New antifungals are needed to combat these fungi and their resting spores. This study aimed to investigate the sporicidal effects of sertaconazole nitrate using microplate laser nephelometry against the microconidia of Trichophyton, chlamydospores of Epidermophyton, blastospores of Candida, and conidia of the mold Scopulariopsis brevicaulis. The results obtained were compared with those from ciclopirox olamine and terbinafine. The sporicidal activity was further determined using infected three-dimensional full skin models to determine the antifungal effects in the presence of human cells. Sertaconazole nitrate inhibited the growth of dermatophytes, molds, and yeasts. Ciclopirox olamine also had good antifungal activity, although higher concentrations were needed compared to sertaconazole nitrate. Terbinafine was highly effective against most dermatophytes, but higher concentrations were required to kill the resistant strain Trichophyton indotineae. Sertaconazole nitrate, ciclopirox olamine, and terbinafine had no negative effects on full skin models. Sertaconazole nitrate reduced the growth of fungal and yeast spores over 72 h. Ciclopirox olamine and terbinafine also inhibited the growth of dermatophytes and molds but had significantly lower effects on the yeast. Sertaconazole nitrate might have advantages over the commonly used antifungals ciclopirox olamine and terbinafine in combating resting spores, which persist in the tissues, and thus in the therapy of recurring dermatomycoses.

Point mutations in the squalene epoxidase erg1 and sterol 14-α demethylase erg11 gene of T indotineae isolates indicate that the resistant mutant strains evolved independently.

BACKGROUND: The T indotineae population shows a high amount of terbinafine resistant isolates based on different point mutations of squalene epoxidase erg1 (ergosterol) gene. A significant proportion of these isolates also show azole resistance. OBJECTIVES: Elucidation of the molecular mechanism for azole resistance, especially the identification of mutations in the sterol 14-α demethylase Erg11 genes, which encode for enzymes interacting with azoles. METHODS: Sequencing of putative Erg11 genes and analysis of phenotypic resistance pattern using a microplate-laser-nephelometry-based growth assay. RESULTS: Four different types of Erg11B mutants were detected; two double mutants with Ala230Thr/Asp441Gly, respectively, Ala230/Tyr444His and single mutants with Gly443Glu, Tyr444Cys and Tyr444His. All isolates featured the wild type genotype of Erg11A. All strains demonstrated different combinations of Erg1 and Erg11 genotypes. CONCLUSION: Resistance against terbinafine and azoles developed several times independently within the T indotineae population. The challenge for fungal treatment is, therefore, that species identification is not enough for prediction of therapeutic efficacy of antifungals. In the future, it will also become important to analyse genes involved in resistance mechanisms.

Abdominal Symptoms Assessed With the CFAbd-Score are Associated With Intestinal Inflammation in Patients With Cystic Fibrosis.

OBJECTIVES: This prospective study evaluated the relationship between fecal markers of intestinal inflammation and cystic fibrosis (CF)-associated abdominal symptoms. These were assessed using the CFAbd-Score, a CF-specific patient-related outcome measure developed and validated, following FDA guidelines. METHODS: In feces from patients with CF (n = 41) and healthy volunteers (n = 27), concentrations of fecal calprotectin (FC), M2-pyruvate kinase (M2-PK), interleukins IL-1ß, IL-6, IL-8, and neutrophilic elastase (NE) were measured. Abdominal symptoms during the 2 preceding weeks were recorded using the CFAbd-Score. This patient-reported outcome measure (PROM) for assessment of the multi-organic abdominal involvement in CF includes 28 items in five domains. RESULTS: Inflammatory parameters FC, IL-1ß, M2-PK, and NE in feces, as well as CFAbd-Scores resulted significantly higher in CF patients than in healthy controls (all P < 0.01). Furthermore, significant differences between both groups were found for pain-symptoms, disorders of bowel movement, impaired quality of life, as well as disorders of eating and appetite. With 83% sensitivity and 74% specificity, FC was the most reliable measure for CF-related intestinal inflammation, which, in the CFAbd-Score, was associated to significantly higher rates of abdominal pain, as well as to general quality of life items such as gastrointestinal-related impaired sleep and frustration. CONCLUSION: Using the CFAbd-Score as a CF-specific PROM for identification and quantification of abdominal symptoms revealed that abdominal pain and impaired quality of life are associated with intestinal inflammation in CF.

Brain Derived Neurotrophic Factor Deficiency is Associated with Cognitive Impairment and Elevated Phospholipase A2 Activity in Plasma of Mice.

Decreased levels of Brain-Derived Neurotrophic Factor (BDNF) are a common finding in schizophrenia. Another well-documented protein linked to schizophrenia is intracellular Ca2+-independent Phospholipase (PLA2). However, the potential association between PLA2 and BDNF with regard to schizophrenia has yet to be examined. In the present study, male and female BDNF knockout mice, a possible genetic model of schizophrenia, were exposed to prenatal stress and tested in the nest test, open field test and T-maze. Following behavioral tests, whole brain and plasma samples were harvested to measure the activity of PLA2. BDNF knockout mice showed cognitive deficits in the T-maze. Furthermore, there was a quadratic association of PLA2 with performance in the open field test. Moreover, BDNF deficiency and female sex were associated with elevated plasma PLA2 levels. The cognitive impairment of BDNF heterozygous mice as well as their increased PLA2 activity in plasma is consistent with findings in schizophrenia patients. The particular elevation of PLA2 activity in females may partly explain sex differences of clinical symptoms in schizophrenia (e.g. age of onset, severity of symptoms). Additionally, PLA2 was significantly correlated with body and adrenal weight after weaning, whereby the latter emphasizes the possible connection of PLA2 with steroidogenesis.

Keratinocyte and Fibroblast Wound Healing In Vitro Is Repressed by Non-Optimal Conditions but the Reparative Potential Can Be Improved by Water-Filtered Infrared A.

It is a general goal to improve wound healing, especially of chronic wounds. As light therapy has gained increasing attention, the positive influence on healing progression of water-filtered infrared A (wIRA), a special form of thermal radiation, has been investigated and compared to the detrimental effects of UV-B irradiation on wound closure in vitro. Models of keratinocyte and fibroblast scratches help to elucidate effects on epithelial and dermal healing. This study further used the simulation of non-optimal settings such as S. aureus infection, chronic inflammation, and anti-inflammatory conditions to determine how these affect scratch wound progression and whether wIRA treatment can improve healing. Gene expression analysis for cytokines (IL1A, IL6, CXCL8), growth (TGFB1, PDGFC) and transcription factors (NFKB1, TP53), heat shock proteins (HSP90AA1, HSPA1A, HSPD1), keratinocyte desmogleins (DSG1, DSG3), and fibroblast collagen (COL1A1, COL3A1) was performed. Keratinocyte and fibroblast wound healing under non-optimal conditions was found to be distinctly reduced in vitro. wIRA treatment could counteract the inflammatory response in infected keratinocytes as well as under chronic inflammatory conditions by decreasing pro-inflammatory cytokine gene expression and improve wound healing. In contrast, in the anti-inflammatory setting, wIRA radiation could re-initiate the acute inflammatory response necessary after injury to stimulate the regenerative processes and advance scratch closure.

Ivacaftor Reduces Inflammatory Mediators in Upper Airway Lining Fluid From Cystic Fibrosis Patients With a G551D Mutation: Serial Non-Invasive Home-Based Collection of Upper Airway Lining Fluid.

In cystic fibrosis (CF) therapy, the recent approval of CF-transmembrane conductance regulator (CFTR) channel modulators is considered to be the major breakthrough. However, the current first-line approach based mainly on pulmonary function to measure effects of the novel therapy, tested by forced expiratory volumes in one second (FEV1), provides restricted sensitivity to detect early structural damages. Accordingly, there is a need for new sensitive surrogate parameters. Most interestingly, these should quantify inflammation that precedes a decline of pulmonary function. We present a novel method assessing inflammatory markers in the upper airways' epithelial lining fluid (ELF) obtained by nasal lavage (NL). In contrast to broncho-alveolar lavage, ELF sampling by NL is an attractive method due to its limited invasiveness which allows repeated analyses, even performed in a home-based setting. In a longitudinal cohort study (ClinicalTrials.gov, Identifier: NCT02311140), we assessed changes of inflammatory mediators in 259 serially obtained nasal lavages taken up to every second day before and during therapy with ivacaftor from ten CF patients carrying a G551D mutation. Patients were trained to sample NL-fluid at home, to immediately freeze and transfer chilled secretions to centers. Neutrophil Elastase, Interleukins IL-1ß, IL-6 and IL-8 in NL were quantified. During 8-12 weeks of ivacaftor-treatment, median values of IL-1ß and IL-6 significantly declined 2.29-fold (2.97→1.30 pg/mL), and 1.13-fold (6.48→5.72 pg/mL), respectively. In parallel, sweat tests and pulmonary function improved considerably. This is the first study assessing changes of airway inflammation on a day-to-day basis in CF patients receiving a newly administered CFTR-modulator therapy. It proves a decline of airway inflammation during ivacaftor-therapy.

Dermatophyte infections in children compared to adults in Germany: a retrospective multicenter study in Germany.

BACKGROUND AND OBJECTIVE: Dermatophyte infections of the skin and nails are common worldwide and vary between geographical areas and over time. The aim of this study was to determine the epidemiological profile of dermatophytes in Germany with a focus on comparing children with adults. PATIENTS AND METHODS: In this retrospective multicenter study, mycological dermatophyte culture results in the period 01/2014 to 12/2016 were analyzed according to identified pathogen, age and gender of patients, and type of disease. RESULTS: Of 1,136 infections (children: n = 88, adults: n = 1,048), 50.8 % were clinically classified as onychomycosis, followed by tinea pedis (34.6 %), tinea corporis (16.2 %), tinea manus (16.2 %), tinea capitis (2.5 %), and tinea faciei (1.2 %). The most frequent pathogen was Trichophyton (T.) rubrum (78.6 %), followed by T. interdigitale (14.3 %), T. benhamiae (3.2 %), T. mentagrophytes (2.1 %), and Microsporum canis (1.7 %). The fungal spectrum differed particularly in tinea corporis and tinea capitis between children and adults with a more diverse pathogen spectrum in children. Trichophyton tonsurans was rarely identified as cause for tinea corporis (2.7 %) or tinea capitis (3.3 %). CONCLUSIONS: Differences in pathogens and frequency of fungal infections between age groups should be considered for optimal selection of the appropriate therapeutic regimen.

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.

[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.

A shell-less hen's egg test as infection model to determine the biocompatibility and antimicrobial efficacy of drugs and drug formulations against Pseudomonas aeruginosa.

A shell-less hen's egg based infection test with Pseudomonas aeruginosa was established to investigate the antimicrobial efficacy of drugs and drug formulations close to the in vivo situation. The test system using preincubated fertilized chicken eggs transferred in petri dishes was optimized with respect to the controlled local application of liquid materials and bacteria as well as the bacterial cultivation conditions. The applicability of the ex ovo infection model was confirmed with antimicrobial susceptibility tests using tobramycin, ciprofloxacin and meropenem. The validity of the ex ovo data was demonstrated by correlation with in vitro data of the CellTiter®-Blue and the microplate laser nephelometry assay. Real-time imaging of the progress of infection and the efficacy of the treatment could be realized by the MolecuLight i:X™ technique. Furthermore, in a proof-of-concept efficacy, biocompatibility and even the presence of irritants were determined side-by-side using commercial ophthalmics. In conclusion, this egg based infection model could bridge the gap between in vitro and in vivo models for the evaluation of antimicrobial susceptibility to reduce animal tests according to the 3R concept.

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.

Superficial fungal infections in the department of dermatology, University Hospital Jena: A 7-year retrospective study on 4556 samples from 2007 to 2013.

BACKGROUND: Superficial fungal infections are often seen in day-to-day clinical practice, and their prevalence continues to rise worldwide. Over the years, a change in the pattern of dermatophytoses has been noted. OBJECTIVES: This study aimed to determine the epidemiologic profile of dermatophytes at the Department of Dermatology, University Hospital Jena, from 2007 to 2013. METHODS: The retrospective study was carried out with a total of 4556 samples collected from 3607 patients suspected of superficial fungal infections during the 7-year study period. RESULTS: Among the 3607 suspected patients, 1951 (54.09%) were men and 1656 (45.91%) were women. Of 4556 samples, 703 (15.43%) samples were positive for fungal culture, which included 585 (83.21%) dermatophytes and 118 (16.79%) non-dermatophytes. Trichophyton (T.) rubrum was the most common isolated pathogen in 73.33% of the dermatophyte cases. Among dermatophyte-infected patients, men were most likely to be affected (63.48%) as well as those of higher age (61 to 80). The most commonly affected areas were nails (33.16%) and feet (33%). T rubrum was recurrently isolated in several regions with exception of the scalp where M canis (58.33%) was the most frequently isolated pathogen. About 16.04% of cases had a history of treatment taken prior to sampling. The majority of the affected individuals did not have any prior animal contact (77.26%). T benhamiae was exclusively associated with contact to guinea pigs. CONCLUSION: Trichophyton rubrum was the most frequently isolated pathogen in several regions except the scalp. New dermatophyte species emerged with time especially T benhamiae.

Evaluation of cell and hemocompatibility of Cucurbiturils.

BACKGROUND: Cucurbiturils (CB) are pumpkin-shaped macrocyclic molecules consisting of methylen-bridged glycoluril units. Because of their complexing characteristics, they can be used as drug containers for medical purposes. For future biomedical and dermal application of CB, the investigation of cell compatibility is essential. Little is known about the influence of CB on eukaryotic cells, especially on dermal keratinocytes. The structurally related cyclodextrins are known to induce cell death by apoptosis in HaCaT keratinocytes as well as hemolysis in erythrocytes. OBJECTIVE: To examine cytotoxic effects of different CB. METHODS: Different cytotoxicity tests were performed on HaCaT keratinocytes and erythrocytes incubated with CB[5], CB[6], and CB[7]. RESULTS: CB[5] and CB[6] did not lead to cytotoxic reactions at high concentrations up to 30 mg/mL whereas incubation with CB[7] triggered apoptosis at a concentration of 3.75 mg/mL. None of the investigated CB caused hemolytic effects on erythrocytes. CONCLUSION: These results confirm the high potential of CB as host-complexes for biomedical and dermal applications.

Tryptanthrin promotes keratinocyte and fibroblast responses in vitro after infection with Trichophyton benhamiae DSM6916.

Exceedingly virulent pathogens and growing antimicrobial resistances require new therapeutic approaches. The zoophilic dermatophyte Trichophyton benhamiae causes highly inflammatory, cutaneous fungal infections. Recently, it could be shown that the plant-derived alkaloid tryptanthrin (TRP) exhibits strong anti-microbial activities against yeasts and dermatophytes. The aim of this study was to analyse the bioactivity of TRP under infectious conditions using an in-vitro dermatophytosis model employing fibroblasts and keratinocytes infected with T. benhamiae DSM6916. Analyses comprised determination of cell viability, effects on the innate immune response including expression and secretion of pro-inflammatory cytokines/chemokines as well as expression of various antimicrobial peptides (AMP), toll-like receptor (TLR) 2 and proliferation marker MKI67. T. benhamiae caused severe inflammation in the cutaneous cell models. TRP almost fully prevented T. benhamiae-derived damage of dermal fibroblasts and substantially reduced it in epidermal keratinocytes. A distinct down-regulation of the expression and secretion of pro-inflammatory cytokines was observed. Further, TRP promoted AMP expression, especially of HBD2 and HBD3, in keratinocytes even without fungal presence. This study provides crucial evidence that TRP is not only a strong antifungal agent but also potentially modulates the innate immune response. This makes it interesting as a natural antimycotic drug for adjuvant treatment and prevention of fungal re-infection.

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.