Mycology an update Part 2: Dermatomycoses: Clinical picture and diagnostics

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1 CME-Article Submitted: Accepted: Conflict of interest None. DOI: /ddg Mycology an update Part 2: Dermatomycoses: Clinical picture and diagnostics Pietro Nenoff 1, Constanze Krüger 1, Jörg Schaller 2, Gabriele Ginter-Hanselmayer 3, Rudolf Schulte-Beerbühl 4, Hans-Jürgen Tietz 5 (1) Laboratory for Medical Microbiology, Mölbis, Germany (2) Dermatopathology Duisburg, Germany (3) Department of Dermatology and Venereology, Medical University Graz, Austria (4) Private Dermatology Practice, Kaiserstraße 94, Dortmund, Germany (5) Institute for Fungal Diseases and Microbiology, Berlin, Germany Section Editor Prof. Dr. Jan C. Simon, Leipzig Summary Most fungal infections of the skin are caused by dermatophytes, both in Germany and globally. Tinea pedis is the most frequent fungal infection in Western industrial countries. Tinea pedis frequently leads to tinea unguium, while in the elderly, both may then spread causing tinea corporis. A variety of body sites may be affected, including tinea glutealis, tinea faciei and tinea capitis. The latter rarely occurs in adults, but is the most frequent fungal infection in childhood. Following antifungal treatment of tinea unguium and also tinea capitis a dermatophytid or hyperergic reaction to dermatophyte antigens may occur. Yeast infections affect the mucous membranes both of the gastro-intestinal system and the genital tract as candidiasis mostly due to Candida albicans. Cutaneous candidiasis affects predominantely the intertriginous regions such as groins and the inframammary area, but also the intertriginous space of fingers and toes. In contrast, pityriasis versicolor is a superficial epidermal fungal infection primarily on the the trunk. Mold infections are rare in dermatology; they play a role nearly exclusively in nondermatophyte-mold (NDM) onychomycosis. The diagnosis of dermatomycoses comprises the microscopic detection of fungi using the potassium hydroxide preparation or alternatively the fluorescence optical Blankophor preparation together with culture. The histological fungal detection with PAS staining possesses a high sensitivity, and it should play a more important role in particular for diagnosis of onychomycosis. Molecular biological methods, based on the amplification of fungal DNA with use of specific primers for the distinct causative agents are on the rise. With PCR, such as dermatophyte-pcr-elisa, fungi can be detected directly in clinical material in a highly specific and sensitive manner without prior culture. Today, molecular methods, such as Matrix Assisted Laser Desorption/Ionization Time-Of-Flight Mass Spectrometry (MALDI TOF MS) as culture confirmation assay, complete the conventional mycological diagnostics. Introduction Dermatophytes are the cause of dermatophytoses or tinea. Tinea pedis is the most common fungal infection in Germany followed by onychomycosis (tinea unguium). Dermatomycoses are infections of the skin, nails, and hair caused by dermatophytes, less commonly yeasts, and only occasionally particularly with respect to toenails also molds. Dermatophytes are the cause of dermatophytoses or tinea. Tinea pedis is the most common fungal infection followed by onychomycosis (tinea unguium) and tinea corporis. As different pathogens, i.e. dermatophyte species, may produce identical clinical pictures, only adequate mycologic workup will eventually shed light on the causal pathogen. Tinea faciei and tinea corporis, as well as 749

2 Tinea capitis has once again become very prevalent among children and adolescents. More frequently than suspected, tinea incognito, sometimes also referred to as tinea atypica, is the cause of an apparently recalcitrant dermatosis. Cutaneous candidiases affect intertriginous areas, not just the large skin folds also interdigital spaces on hands and feet. The microscopic workup using KOH (potassium hydroxide) preparations may nowadays be supplemented, or possibly completely replaced, by the more sensitive fluorescent Calcofluor or Blankophor test. Molecular biology dermatophyte diagnostics using nucleic amplification techniques (polymerase chain reaction) have also become an integral part in dermatomycology. tinea cruris, represent rare clinical conditions. In recent years, particularly tinea capitis has once again become very prevalent among children and adolescents. Not only the frequently purulent, abscess-forming deep hair root infections (kerion) but also recalcitrant hyperkeratotic forms of tinea capitis are part and parcel of dermatologic diagnostics and therapy. The term tinea incognito, sometimes also referred to as tinea atypica, has been increasingly used recently. Experience has shown that initially dermatomycoses, especially dermatophytoses, are frequently not recognized as such. Only once antiinflammatory or antibacterial therapy has proven ineffective, consideration of a mycotic infection leads to the appropriate workup. There are numerous differential diagnoses for dermatomycoses of the skin and nails. Apart from psoriasis vulgaris with its special variants inverse psoriasis and nail psoriasis, a number of other inflammatory skin disorders have to be taken into account. Dermatophytoses of the skin and nails have to be differentiated from yeast infections, primarily by Candida species, which more often affect intertriginous areas, not just the large skin folds also interdigital spaces on hands and feet. Pityriasis versicolor is the most important and most common Malassezia-associated superficial skin infection. Today in addition to classic diagnostic components such as direct microscopy (KOH prep) and fungal cultures, modern mycologic diagnostics also include molecular biology methods. The latter have been increasingly used in laboratories and more recently also in dermatologic office settings. The microscopic workup using KOH (potassium hydroxide) preparations should nowadays be supplemented, or possibly completely replaced, by more sensitive fluorescent Calcofluor or Blankophor tests. Fungal cultures the gold standard in mycologic diagnostics are still time-consuming, with results taking two to four weeks to come back. Only zoophilic dermatophytes show fungal growth within a few days. Nucleic acid amplification techniques have become an integral part of molecular biology diagnostics in dermatomycology. Almost always, polymerase chain reaction (PCR) is used for the direct detection of dermatophyte DNA from skin scrapings, nail clippings, hair roots, and skin swabs. While a disadvantage of molecular biology methods is their cost with respect to personnel and technical devices, they are markedly more sensitive and especially more time-efficient compared to fungal cultures. In some cases, results from molecular biology methods down to the species level may be available as early as 24 hours. Clinical picture of dermatomycoses Dermatophytoses Tinea pedis The clinical picture of dermatophytosis is not pathogen-specific, as various dermatophyte species may be the root cause for tinea pedis, but also for any other tinea variant. Anthropophilic dermatophytes are the pathogenic agents in tinea pedis: Trichophyton rubrum and Trichophyton interdigitale (anthropophilic strains) as well as Epidermophyton floccosum. Tinea pedis frequently starts in the interdigital spaces with hyperkeratotic, dry, scaly, and later on macerated, oozing, and erosive lesions. The clinical picture is not pathogen-specific, as various dermatophyte species may be the root cause for tinea pedis, but also for any other tinea variant. Anthropophilic dermatophytes are the pathogenic agents in tinea pedis, i.e. besides Trichophyton (T.) rubrum also T. interdigitale (anthropophilic strains) and Epidermophyton floccosum. Interdigital tinea pedis clinically resembles the little known psoriasis alba or interdigital psoriasis [1], which, however, may be differentiated on the basis of a negative mycologic workup. Tinea pedis causes moderate pruritus. Formation of rhagades over the course of time leads to burning sensations and pain. Secondary infections with gram-positive (Staphylococcus aureus, A streptococci) and gram-negative 750

3 Figure 1 Moccasin-type tinea caused by Trichophyton rubrum. The most severe variant of tinea pedis presents with dry, erythematous, scaly lesions with prominent borders. Moccasin-type tinea pedis is difficult to clinically differentiate from atopic foot dermatitis or plantar psoriasis. In order to rule out inflammatory dermatoses, e.g. dyshidrotic eczema or contact dermatitis, a mycologic workup should always be performed when tinea pedis is suspected. (Escherichia coli, Klebsiella spp., Pseudomonas aeruginosa, Proteus spp.) bacteria result in maceration and fetor. With respect to differential diagnoses, (interdigital) erythrasma by Corynebacterium minutissimum has to be ruled out; in immunosuppressed patients, also interdigital candidiasis. The hyperkeratotic type of tinea pedis spreads across the sole as well as medially and laterally along the edges and back of the foot. Moccasin-type tinea pedis is clinically barely differentiable from atopic foot dermatitis or plantar psoriasis (Figure 1). Another differential diagnosis is pitted keratolysis (keratoma plantare sulcatum) due to maceration in plantar hyperhidrosis and marked colonization by Corynebacterium spp. and Kytococcus sedentarius as well as Dermophilus congolensis [2]. Another rare differential diagnosis is the autosomal recessive palmoplantar keratoderma of the Gamborg-Nielsen type, which may occasionally also be secondarily infected by T. rubrum [3]. The third variant of tinea pedis is the inflammatory vesiculobullous or dyshidrotic type. It is located either in the arch or at the medial edge of the foot [4] (Figure 2a, b) and typically shows small, superficial, pruritic vesicles on erythema and exfoliations of the stratum corneum with scaling. Dyshidrotic eczema, contact dermatitis, and impetigo all have to be ruled out. Moreover, juvenile plantar dermatosis or dermatitis plantaris sicca (corresponds to plantar atopic dermatitis) should be considered. Here again, mycologic tests will bring certainty. There is an overlap with keratolysis exfoliativa (lamellar dyshidrosis), which as of late is considered a distinct entity [5]. It is most likely neither dyshidrotic eczema nor a fungal infection or an Id reaction in terms of a dermatophytid. Currently, incomplete or immature corneodesmolysis is viewed as the chief pathomechanism in this palmoplantar dermatosis. Tinea corporis The English term for tinea corporis and tinea capitis is ringworm. Dermatophytoses on the skin appear as centrifugally growing, erythemato-squamous, scaly plaques. This explains the English term ringworm for tinea corporis and tinea capitis, which is almost identical to an older German term. Due to its peculiar ring-like morphology, tinea circinata represents a special 751

4 Figure 2 Tinea pedis caused by Epidermophyton floccosum in a 29-year-old female marathon runner. Vesiculobullous tinea pedis presenting with vesicles, hyperkeratosis and desquamation involving the sole and toes (a). Tinea pedis with maceration, erythema, hyperkeratosis, and rhagades (b). Tinea imbricata (tokelau) caused by Trichophyton concentricum is well known in Southeast Asia, but also in Africa and South America. In Great Britain, the anthropophilic species Trichophyton tonsurans is the second most common pathogen of tinea corporis after Trichophyton rubrum. form of tinea corporis, frequently caused by T. tonsurans and sometimes also Microsporum (M.) gypseum [6]. It is difficult to differentiate from tinea imbricata (tokelau), primarily occurring in Southeast Asia (Vietnam, Polynesia), but also in Africa and South America, caused by the slow-growing dermatophyte T. concentricum [7]. In adults, tinea corporis is predominantly caused by T. rubrum, occasionally also T. interdigitale. Differential diagnostic considerations include psoriasis vulgaris, impetigo, atopic dermatitis, contact dermatitis, erythema multiforme, but also granuloma annulare. By contrast, zoophilic dermatophytes are the primary cause for tinea corporis in children and adolescents (Figure 3a, b). Source of infection are small, domesticated, furry animals that either suffer from dermatophytosis or simply represent asymptomatic carriers of zoophilic dermatophytes. In the USA, Latin America (Mexico), but also Great Britain, the anthropophilic species T. tonsurans is the second most common pathogen of tinea corporis after T. rubrum [8]. In Africa, T. violaceum and M. audouinii play a crucial role. Tinea inguinalis and Tinea glutealis The old term eczema marginatum (Hebra) corresponds to tinea inguinalis. In most cases, tinea inguinalis results from autoinoculation in patients with preexisting tinea pedis et unguium. Although T. rubrum is a common pathogen, T. interdigitale and Epidermophyton floccosum have also been isolated in the inguinal region. Apparently, women are far less commonly affected by tinea inguinalis than men. However, in rare cases dermatophytes have also been isolated as causal pathogens in suspected intertrigo of the inguinal region in women. Due to the accentuated borders of the macerated and scaly lesions in tinea inguinalis, the clinical picture corresponds to so-called eczema marginatum (Hebra), first described in 1860 by Ferdinand Ritter von Hebra ( ), 752

5 Figure 3 Tinea corporis on a girl s upper chest caused by a zoophilic dermatophyte. The centrifugally growing lesions show central clearing and typically prominent borders (a). Inflammatory tinea corporis with centrifugally growing, erythemato- squamous plaques on the shoulder of a girl affected by a zoophilic dermatophyte infection (b). Figure 4 Tinea corporis glutealis caused by Trichophyton rubrum in a 70-year- old man. The scaly plaques with accentuated borders developed as a result of autoinoculation from the patient s own tinea pedis and tinea unguium. founder of the Vienna School of Dermatology. Tinea glutealis is frequently misdiagnosed as anal dermatitis, while failing to recognize its mycotic genesis (Figure 4). In some patients with tinea glutealis, inflammatory follicular papulopustules may be found in addition to typical erythemato-squamous plaques with prominent margins. 753

6 Tinea cruris Tinea cruris is a rare form of dermatophytosis. Differential diagnoses include psoriasis vulgaris, nummular dermatitis, stasis dermatitis, and contact dermatitis. A special variant in women is tinea granulomatosa follicularis et nodularis cruris which may occur as a result of shaving the lower extremities. It should be noted that in the Anglo-American literature the term tinea cruris is frequently incorrectly used when referring to tinea inguinalis. Tinea incognito Initially, tinea incognito is often erroneously regarded as infectious and inflammatory dermatosis resulting in antiinflammatory treatment with topical corticosteroids. By definition, tinea incognito is a dermatophyte infection that has lost its typical clinical appearance due to the unjustified use of topical corticosteroids or calcineurin inhibitors. Tinea incognito, recently also termed tinea atypica, displays many overlapping features with the T. rubrum syndrome [9]. In tinea incognito, i.e. unrecognized tinea, cutaneous dermatophytosis is not considered as differential diagnosis. Taking only various other infectious and inflammatory dermatoses into account eventually results in erroneous antiinflammatory treatment with topical corticosteroids. There has been a worldwide increase in tinea incognito. On the one hand, this may be attributed to an actual increase in the number of patients prone to contract extensive tinea corporis. On the other hand, however, it has to be critically noted that abandonment of a targeted mycologic workup mirrors the lack of differential diagnostic consideration of dermatomycosis in the first place. Seitz et al. [10] have recently described four patients with extensive tinea corporis initially not recognized as such. Instead, the patients were diagnosed with cutaneous T cell lymphoma or bullous pemphigoid and received treatment further predisposing them to dermatophyte infections (corticosteroids or azathioprine). By definition, tinea incognito is a dermatophyte infection that has lost its typical clinical appearance due to the unjustified use of topical corticosteroids or calcineurin inhibitors [11]. Of 283 patients with tinea incognito in Korea, more than half (59 %) had previously been treated by non-dermatologists or self-medication. Strikingly, the average duration of tinea incognito was 15 ± 25 months. Clinically, lesions initially resembled eczema-like and psoriasis-like dermatoses as well as lupus erythematosus. Trunk and face were frequently affected. Among 67 isolated fungal strains, T. rubrum was found to be the most common (73 %). In patients with tinea pedis or tinea unguium, recalcitrant eczema-like dermatoses on the trunk or face should always raise the suspicion for tinea incognito. In the case of a 55-year-old female Japanese patient, pruritic erythematous and erosive lesions on the face and nape were clinically suspected as pemphigus erythematosus [12], however, direct and indirect immunofluorescence (anti-desmoglein 1 and 3 antibodies) were negative. Only mycologic evaluation of skin scales eventually revealed the diagnosis tinea faciei et corporis incognito. Here, surprisingly, M. canis was the pathogen. Trichophyton rubrum syndrome In T. rubrum syndrome, at least four body sites are affected: feet (plantar), hands (palmar), nails, as well as one other site. Diagnostic criteria in T. rubrum syndrome include microscopic fungal detection (KOH or Blankophor ) from all four sites as well as cultural detection of T. rubrum from at least three out of four sites. Trichophyton (T.) rubrum syndrome represents a chronic and generalized dermatophytosis. Prior terms such as chronic dermatophytosis syndrome, generalized chronically persistent rubrophytia, tinea corporis generalisata and dry-type T. rubrum infection all refer to the same dermatomycotic entity. In T. rubrum syndrome, at least four body sites are affected: feet (plantar), hands (palmar), nails, as well as one other site. The inguinal region which is a common site of tinea, is explicitly excluded. The second diagnostic criterion in T. rubrum syndrome includes microscopic fungal detection (KOH or Blankophor ) from all four sites. The third criterion is obvious; cultural detection of T. rubrum from at least three out of four sites (Figure 5a, b). 754

7 Figure 5 Trichophyton rubrum syndrome in a 34-year-old woman with minor mental retardation. The dermatophytosis affected fingernails, toenails, hands, feet, trunk, face and scalp. Tinea faciei caused by Trichophyton rubrum (a). Tinea unguium and tinea manuum caused by Trichophyton rubrum in the same patient (b). It still remains unclear whether T. rubrum syndrome represents a distinct nosologic entity. Moreover, previously administered topical or systemic corticosteroid therapy also has to be considered a predisposing factor. Piñeiro et al. [13] pointed towards secondary tinea as a result of autoinoculation from primary tinea pedis, as they found T. rubrum syndrome in 7 % of these secondary infections. Four male patients (age: 38 56) with T. rubrum syndrome showed various predisposing factors. They either suffered from Down syndrome, HIV infection, or diabetes mellitus, or they received topical corticosteroid treatment. Endogenous hypercortisolism in the context of Cushing s disease also has to be regarded as predisposing factor for chronic and recalcitrant dermatophytoses by T. rubrum [14]. Tinea manus Dermatophytes rarely affect the hands. Clinically, tinea manus presents with scaly, hyperkeratotic, centrifugally growing, and erythemato-squamous lesions (Figure 6). Differential diagnostic considerations include palmar psoriasis, atopic hand dermatitis, and contact dermatitides of various origins. The most common pathogen is T. rubrum. As mentioned above, the infection results from autoinoculation of infectious skin and nail material in tinea pedis et unguium. Bullous tinea Bullous tinea is rare. It may occasionally be found in the context of tinea manus or tinea corporis. Rarely, tinea occurs in its bullous form, e.g. also in tinea manus [15]. T. rubrum is usually the causal pathogen but in some cases zoophilic dermatophytes have been found, e.g. Trichophyton species of Arthroderma benhamiae. Clinical examination reveals, often peripherally located, blister formation sometimes with an opaque serous content on erythemato-squamous plaques, leading to differential diagnoses such as impetigo, cutaneous herpes simplex, but also bullous dermatoses. Histologic detection of septate fungal hyphae in PAS stained sections quickly yields the correct diagnosis, though. Occasionally, a biopsy may be omitted in bullous tinea, as histologic evaluation of the blister roof usually suffices. Timely mycologic analysis of skin scrapings or a swab would be an even simpler method. 755

8 Figure 6 Tinea manus caused by Trichophyton rubrum in a 33-year-old patient with atopic dermatitis. Two feet-one hand syndrome A fungal infection of the left hand and both feet, frequently involving finger and toenails, is referred to as two feetone hand syndrome (TFOHS). The two feet-one hand syndrome is caused by dermatophyte transmission from tinea pedis or onychomycosis to the (left) hand, for example when scratching. In case of a fungal infection of one usually the left hand (palm) in tandem with corresponding lesions on both feet (plantar and interdigital mycosis), frequently involving fingernails and toenails, the clinical picture alone allows for the suspected diagnosis of two feet-one hand syndrome (TFOHS) [16]. The pathogen is usually T. rubrum in this tinea manus et pedum et unguium, occasionally also T. interdigitale. Differential diagnoses include dyshidrotic hand and foot eczema, irritant or allergic contact dermatitis as well as palmoplantar psoriasis. TFOHS more frequently affects men. It is caused by dermatophyte transmission from tinea pedis or onychomycosis to the (left) hand, for example when scratching. In rare cases, the right hand may be involved as tinea manus or tinea unguium. There is also a correlation with pedicure performed with the right hand holding the scissors while the left hand is in contact with mycotically diseased skin and nails. In a Chinese patient group of 113 patients with bilateral tinea pedis and unilateral tinea manus, there was a significant correlation between tinea manus and the hand used for scratching the feet [17]. Employing PCR amplification in genotyping tandem repeat elements of ribosomal DNA of the isolates nontranscribed spacer region yielded a match between hand and feet in 94.5 % of cases with respect to species (T. rubrum) and an 80 % match regarding genotype. The contralateral hand is not infected or shows no signs of tinea manus, however, subclinical dermatophyte involvement is possible. Predisposing palmar hyperhidrosis with left-right asymmetry has been suggested. Furthermore, the non-dominating (left) hand is mostly affected, whereas the dominating or working hand shows a better-developed protective stratum corneum. 756

9 Majocchi granuloma (granuloma trichophyticum) Majocchi granuloma (granuloma trichophyticum) is a granulomatous follicle-associated inflammatory reaction caused by a dermatophyte. Majocchi granuloma (granuloma trichophyticum) is a chronic intracutaneous/subcutaneous dermatophytosis first described by the Italian dermatologist Domenico Majocchi ( ) in 1883 [18]. The pruritic and scaly erythemato-squamous plaques display follicular papules and pustules. Superficial dermatophytosis with subsequent deep infiltration of mycelia along hair follicles results in their rupture leading to a granulomatous follicle-associated inflammatory reaction. It particularly affects immunosuppressed patients, e.g. following a cardiac transplant [19]. In most cases of Majocchi granuloma, T. rubrum is the causal pathogen, but other dermatophytes have also been described, e.g. T. tonsurans as pathogen of tinea corporis gladiatorum clinically presenting as Majocchi granuloma [20]. A patient with rheumatoid arthritis on methotrexate and prednisolone developed severely pruritic follicular papules and nodules on the inner aspect of the thigh. Histology revealed mixed periadnexal infiltrates of histiocytes, neutrophils, and lymphocytes in the mid and deep dermis as well as fungal elements in the vicinity of hair shafts in PAS staining (culture: T. rubrum), yielding the diagnosis of nodular granulomatous perifolliculitis caused by T. rubrum (Majocchi granuloma) [21]. Dermatophytid A dermatophytid or mycid occurs in the context of a so-called Id reaction or hyperergic reaction to dermatophyte components as a result of antimycotic treatment of extensive tinea pedum or any other dermatophytosis. A dermatophytid or mycid likely occurs more often than suspected in the context of a so-called Id reaction [22]. It is characterized by a hyperergic reaction to dermatophyte components as a result of antimycotic treatment of extensive tinea pedum. Dermatophytids may basically also occur in any other tinea variant [23], as recently described in an 8-year-old boy with tinea capitis [24]. Id reactions appear as symmetric dyshidrotic eruptions on hands and feet, or as nodular and multiform eruption, predominantly on the extremities. It is frequently falsely misconstrued as adverse drug reaction or intolerance to antimycotic agents, but lesions slowly resolve, even if the drug is continued [25]. The decision to do so may, however, prove difficult in some cases. Tinea faciei Differential diagnoses of tinea faciei include impetigo, atopic dermatitis, contact dermatitis, discoid lupus erythematosus, and herpes zoster. It is characterized by erythematous, centrifugally growing, discretely scaly lesions with prominent borders, frequently on the cheeks, but also on the eyelids and sometimes in the submandibular region (Figure 7a c). Mildly pruritic (or even non-pruritic), scaly facial lesions with accentuated borders should therefore always prompt a mycologic workup to rule out tinea faciei. Among others, differential diagnostic considerations include impetigo, atopic dermatitis, contact dermatitis, discoid lupus erythematosus, and herpes zoster (Table 1). In children, zoophilic dermatophytes zoophilic strains of T. interdigitale as well as M. canis and Trichophyton species of Arthroderma benhamiae are the primary pathogens in tinea faciei. Microsporosis is the term formerly used for infections of the face and hairy scalp by Microsporum species. Nowadays, these infections should preferably be referred to as tinea capitis or tinea corporis, as they may be caused by both Microsporum as well as Trichophyton species. Tinea capitis Tinea (Latin for wood worm or moth) is the historic term for dermatophyte infections of the hairy scalp that result in a moth-eaten clinical appearance. The formerly common term microsporosis referred to infections of the face and hairy scalp by Microsporum species; in the 1950s mostly M. audouinii, but also M. canis and M. gypseum [26]. As early as 1969, Braun-Falco and Meinhof came to 757

10 Figure 7 Tinea faciei of the eyelids with inflammatory erythema and scaling in a child (a). Tinea faciei of the left cheek and right eyebrow in a child caused by a zoophilic strain of Trichophyton interdigitale (formerly Trichophyton mentagrophytes) (b). Tinea faciei of the perioral region in a 7-year-old boy. The inflammatory, slightly raised plaque with prominent margins is covered with yellow scabs. The differential diagnosis primarily included impetigo resulting in unsuccessful treatment with topical antibacterial agents. The lesions eventually resolved, once topical antimycotic treatment with sertaconazole had been initiated (c). Table 1 Differential diagnoses of tinea faciei. 1. (Allergic) contact dermatitis 2. Atopic dermatitis 3. Discoid Lupus erythematosus 4. Granuloma faciale 5. Herpes zoster 6. Impetigo 7. Lyme disease (erythema migrans, lymphocytoma) 8. Lymphocytic infiltration of the skin 9. Perioral dermatitis 10. Polymorphous light eruption 11. Psoriasis vulgaris 12. Rosacea 13. Sarcoidosis of the face 14. Malar rash in systemic lupus erythematosus 15. Seborrheic dermatitis Tinea capitis is the most important and most common dermatophyte infection in children between 3 and 8 years of age, commonly in the 5 th and 6 th year. the conclusion that dermatophytoses should not be named after their respective pathogen as in microsporosis by Microsporum species. The same holds true for the old term trichophytia profunda capillitii. Thus, mere assessment of the clinical morphology in tinea capitis does not offer any indication as to the causal dermatophyte species. Tinea capitis is the most relevant dermatophyte infection in children between 3 and 8 years of age, commonly in the 5 th and 6 th year. While in the past boys were thought to be more frequently affected, particularly with respect to M. canis infections [27], this gender difference seems to have disappeared. Currently, even in the USA, there is an even distribution between girls and boys [28]. 758

11 In the USA unlike Germany the anthropophilic dermatophyte Trichophyton tonsurans is the most common pathogen in tinea capitis. In Germany, tinea capitis is caused by the same zoophilic dermatophytes responsible for tinea corporis and tinea faciei. In the USA, on the other hand, the anthropophilic dermatophyte T. tonsurans is the most common pathogen in tinea capitis. Sub-Saharan-Africa shows a particularly high prevalence of dermatophytoses (tinea corporis and tinea capitis) especially among school-aged children. In Gondar, Ethiopia, 47.5 % of 372 students living in a rural area exhibited clinical signs of tinea capitis [29], predominantly caused by T. violaceum, followed by T. verrucosum in second place. A recent study from Tunisia also points to T. violaceum as most common cause for tinea capitis, followed by M. canis, Trichophyton interdigitale complex, and T. verrucosum [30]. In France, the anthropophilic fungus M. audouinii has been increasingly found, especially in immigrant families from Africa [31]. Recently, Munich, Berlin, Hannover, and Wittlich (Rhineland-Palatinate) have also seen outbreaks of M. audouinii infections among children of African immigrants. Geophilic dermatophytes M. gypseum being the clinically most relevant species rarely cause tinea capitis. Present in soil and dust, children may contract them, e.g. while playing outside, with subsequent infection of the skin and occasionally the scalp. Tinea capitis in adults Unlike children, adults are very rarely affected by tinea capitis. The most likely pathogen of tinea capitis in North and Central America is Trichophyton tonsurans. In Germany, Trichophyton rubrum is the most likely pathogen in adult tinea capitis as a result of autoinoculation. In endothrix infections by T. tonsurans, T. violaceum, T. soudanense, and T. verrucosum, (arthro)spores and mycelia may be microscopically detected inside the hair shaft. In ectothrix infections by M. canis, M. audouinii, and T. interdigitale (zoophilic strains), spores and hyphae aggregate outside the hair shaft. Unlike children, adults are very rarely affected by tinea capitis [32]. Due to the atypical clinical picture, other disorders are often initially considered, resulting in antiinflammatory topical corticosteroid therapy. Potential differential diagnostic considerations are listed in Table 2. There has been an unusual favus case caused by T. schoenleinii in a 34-year-old, Kosovo-born female patient from Würzburg. [33]. Here, nonspecific symptoms like scaling, erythema, and round alopecia with loss of hair follicles were initially misdiagnosed as lichen planus with subsequent topical clobetasol propionate treatment. Histology eventually yielded the correct diagnosis of tinea capitis, once spores and hyphae had been detected. Trichoscopy occasionally shows so-called corkscrew hairs in adult tinea capitis [34]. A review article on tinea capitis from California described a case initially misdiagnosed as dissecting cellulitis (folliculitis et perifolliculitis capitis abscedens et suffodiens (Hoffmann)). Both disorders almost exclusively affect dark-skinned, i.e. Africans and African Americans, in this particular case, a Hispanic man. The most likely pathogen in North and Central America is T. tonsurans. There has been a report of an immunosuppressed male HIV patient from India who suffered from tinea capitis circinata caused by M. audouinii [35]. In Germany, T. rubrum is the most likely pathogen in the majority of cases of adult tinea capitis, similar to tinea incognito as a result of autoinoculation [36]. Pattern of involvement of hair and hair roots in tinea capitis Dermatophytes either show an endothrix or an ectothrix pattern of hair root involvement. In endothrix infections by T. tonsurans, T. violaceum, T. soudanense, and T. verrucosum, (arthro)spores and mycelia may be microscopically detected inside the hair shaft, without any destruction of the cuticle. In ectothrix infections by M. canis, M. audouinii, and T. interdigitale (zoophilic strains), spores and hyphae aggregate in a cuff-like fashion outside the hair shaft. Clinical picture of tinea capitis Six different forms of tinea capitis may be differentiated [37]. 759

12 Table 2 Differential diagnoses of tinea capitis. 1. Alopecia areata 2. Bacterial abscess of the hairy scalp 3. Discoid lupus erythematosus 4. Erosive pustular dermatitis of the scalp 5. Folliculitis decalvans 6. Dissecting cellulitis (folliculitis et perifolliculitis capitis abscedens et suffodiens (Hoffmann)) 7. Hyperkeratotic actinic keratoses 8. Acne keloidalis nuchae 9. Impetigo 10. Lichen planopilaris (Lassueur Graham Little syndrome) 11. Pyodermas: furuncle/carbuncle 12. Seborrheic dermatitis of the scalp 13. Pityriasis amiantacea (formerly tinea amiantacea) 14. Pityriasis capillitii 15. Psoriasis capitis 16. Sterile eosinophilic pustulosis (Ofuji) 17. Syphilis II (alopecia syphilitica as DD of moth eaten tinea) 18. Trichotillomania 19. Scarring alopecia Zoophilic dermatophytes like M. canis and T. species of Arthroderma benhamiae cause round, erythematous, hyperkeratotic, scabbed, scaly, centrifugally growing, and alopecic tinea capitis lesions. Microsporum canis infections show a greenish-yellow fluorescence under Wood light. The black dot variant of tinea capitis is the result of hair shafts broken off at skin level in infections by Trichophyton species. Kerion represents the most severe form of tinea capitis characterized by an abscess-like deep infection of the scalp. 1. Gray patch tinea capitis: It exhibits typical disc-like alopecic lesions covered by a gray scaly layer. Hairs break off directly above the skin surface resulting in a stubble field -like clinical picture. In some cases, an inflammatory component with erythema may be completely missing. Zoophilic dermatophytes like M. canis and T. species of Arthroderma benhamiae frequently cause these round erythematous, hyperkeratotic, scabbed, scaly, centrifugally growing, and alopecic lesions (Figure 8). When examined under Wood light, a UV lamp emitting 365 nm UVA radiation, M. canis infections show a green-yellow fluorescence. Favus with scutulum formation caused by T. schoenleinii may be misinterpreted as Pityriasis amiantacea [38]. 2. Moth-eaten tinea capitis: In adults, this variant of tinea capitis should not be confused with alopecia syphilitica [39], a rare form of stage 2 syphilis clinically also referred to as moth-eaten alopecia. 3. Black dot tinea capitis: This scarcely inflammatory tinea variant is the result of hair shafts broken off at skin level in infections by Trichophyton species, such as T. tonsurans, T. interdigitale, T. soudanense, or T. violaceum, but also by M. audouinii (Figure 9). 4. Pityriasis capillitii-type tinea capitis: There is diffuse scaling of the scalp without signs of inflammation. Potential pathogens are e.g. T. tonsurans and M. audouinii. 5. Pustular tinea capitis: Characterized by yellow pustules. 6. Kerion: This represents the most severe form of tinea capitis characterized by an abscess-like deep infection of the scalp. A hypersensitivity reaction may lead 760

13 Figure 8 Tinea capitis caused by Trichophyton species of Arthroderma benhamiae in a child from Berlin. The alopecic scalp area shows erythema, inflammatory infiltration, and hyperkeratotic scabs. Figure 9 Tinea capitis caused by Microsporum audouinii in an African boy. Black dot tinea capitis. Alopecic areas display hair shafts broken off at skin level. Tinea capitis profunda occurs in infections caused by zoophilic dermatophytes, such as Trichophyton interdigitale or Trichophyton verrucosum, but also Trichophyton species of Arthroderma benhamiae. to granuloma formation in kerion [40]. Patients typically experience symptoms like pain and fatigue as well as nuchal and cervical lymphadenopathy. Fever is uncommon. Tinea capitis profunda occurs in infections caused by zoophilic dermatophytes, such as T. interdigitale or T. verrucosum, but also Trichophyton species 761

14 of Arthroderma benhamiae. Peripheral hairs may be epilated without difficulty. In case of T. verrucosum infections, there is no fluorescence under Wood light. Infections of the eyebrows Infections of the eyebrows by dermatophytes are also subsumed under tinea capitis. Due to their slow growth, such infections have proven to be recalcitrant and require prolonged treatment. Dermatophytes on the scalp: tinea capitis or merely carrier status? Children may also merely be dermatophyte carriers. Especially Trichophyton tonsurans and Microsporum audouinii may be asymptomatically present on the scalp or cause only discrete pityriasis capillitii. In superficial tinea capitis caused by anthropophilic dermatophytes, the alopecia areata-like lesions on the scalp are mostly dry, more or less erythematous, hyperkeratotic, and scaly. Some children are merely pathogen carriers. Thus, especially T. tonsurans, but also M. audouinii, frequently give rise to discrete pityriasis capillitii. 5.7 % of 194 Turkish wrestlers were found to merely be asymptomatic carriers of T. tonsurans, mostly on the hairy scalp [41]. If left untreated, these individuals play a significant role in the transmission of tinea capitis among the population. In some cases, T. tonsurans causes recalcitrant and protracted dermatomycoses. There has been the case of a 12-year-old Japanese girl who, for ten years, suffered from recurring tinea corporis and tinea faciei caused by T. tonsurans as a result of asymptomatic tinea capitis [42]. Secondary bacterial infection in tinea capitis The presence of bacteria in tinea capitis is usually merely a sign of secondary infection. Secondary bacterial infections may potentially occur in tinea capitis, occasionally leading to misdiagnosis and subsequent topical and systemic antibiotic therapy. Some cases even result in unnecessary surgical procedures like incision and abscess drainage. As conventional mycologic diagnostics in slow-growing dermatophytes, e.g. T. verrucosum, are initially negative, other causal pathogens, first and foremost Staphylococcus aureus, come to the fore. However, these bacteria are simply a sign of secondary infection in tinea capitis (profunda). Secondary infections by LA-MRSA (livestock-associated methicillin-resistant Staphylococcus aureus) may give rise to considerable diagnostic and therapeutic problems in cases of purulent, abscess-forming tinea capitis caused by T. verrucosum[43]. Tinea gladiatorum corporis et capitis caused by Trichophyton tonsurans Trichophyton tonsurans is frequently indirectly transmitted via contaminated mats in martial arts, e.g. wrestling, and causes tinea gladiatorum. T. tonsurans is frequently indirectly transmitted via contaminated wrestling mats, hence the term tinea corporis or tinea capitis gladiatorum. Microtraumata caused by skin abrasions on the mat facilitate the dermatophyte s entry into the stratum corneum of the epidermis. Direct transmission from skin to skin while wrestling is less likely, though. Clinical picture of onychomycosis Onychomycosis The percentage of onychomycoses among all nail disorders is very high (50 %). It is estimated that roughly 2 3 % of the worldwide population suffer from fungal nail diseases. According to the Foot Check Study, the prevalence of onychomycosis 762

15 Figure 10 Onychomycosis caused by Trichophyton rubrum in a 70-year-old woman. Yellow streaks indicate mycotic involvement of the nail matrix. Tinea unguium is an onychomycosis caused by a dermatophyte, most commonly by Trichophyton rubrum. Onychomycosis is a fungal nail infection, which may be caused by dermatophytes, yeasts, or molds. among Germans is 12.4 %. The incidence of onychomycosis is inevitably going to rise, as industrialized societies are getting older. Tinea unguium refers to onychomycosis caused by a dermatophyte most commonly by far T. rubrum, but not by yeasts or molds. The term onychomycosis on the other hand signifies a fungal nail infection which may be caused by dermatophytes, yeasts, or molds or a combination of two (or more) fungi from all three groups. Tinea unguium Distal and lateral subungual onychomycoses start at the distal free edge of the toenails. Yellow streaks correspond to dermatophytomas and point towards fungal matrix involvement. Proximal subungual onychomycosis may be a sign of immunosuppression in affected patients. In Africa, but also in Germany, it occurs in HIV positive and AIDS patients. White superficial onychomycosis refers to a superficial infection of the nail plate by Trichophyton rubrum or Trichophyton interdigitale. As there is no subungual hyperkeratosis in endonychial onychomycosis, onycholysis does not occur. Hyperkeratotically thickened nails with white discoloration are typical of endonychial onychomycosis. Fungal nail infections predominantly start at the distal free edge of the toenails as distal subungual onychomycosis (DSO). The pathogen slowly migrates from the hyponychium at the bottom side of the nail plate proximally towards the matrix, in time resulting in a lateral and distolateral subungual onychomycosis (DLSO). The nail appears thickened and hyperkeratotic with yellowish-brown discoloration. As time progresses, onycholysis sets in. The yellow streaks represent dermatophytomas and point towards fungal matrix involvement (Figure 10). Proximal subungual onychomycosis (PSO) is quite rare. The pathogen progresses from the proximal nail wall (affected by tinea pedis) onto the cuticle and later on onto the eponychium (epithelium at the bottom side of the proximal nail wall). PSO occurs more frequently in immunosuppressed individuals, primarily HIV positive and AIDS patients. The association between PSO and HIV is particularly striking in countries with high HIV prevalence, e.g. Sub-Saharan Africa. Leukonychia trichophytica or white superficial onychomycosis (WSO) refers to a superficial dermatophyte infection of the nail plate, mostly by T. rubrum, but also T. interdigitale. Here, a flat, bright white, plaque-like layer covers the nail plate, sometimes affecting the entire nail surface. Proximal white subungual onychomycosis (PWOM) represents a special variant with white discoloration underneath the proximal part of the nail plate, apart from T. rubrum also caused by T. schoenleinii (not in Germany) and Epidermophyton floccosum. Black superficial onychomycosis caused by the mold Hendersonula toruloidea (current taxonomy: Nattrassia mangiferae) represents another special variant. As there is no subungual hyperkeratosis in endonychial onychomycosis, onycholysis does not occur. In this variant of onychomycosis, the nails are hyperkeratotically thickened and show white discoloration. This form of onychomycosis caused by T. soudanense is likely to be encountered in Africa. 763

16 Figure 11 Total dystrophic onychomycosis and tinea pedis caused by Trichophyton rubrum in an immunosuppressed patient with larynx carcinoma. Total dystrophic onychomycosis (TDO) represents the most severe variant of onychomycosis. Onychia et paronychia candidosa is caused by yeasts and frequently arises as a consequence of bacterial paronychia. Total dystrophic onychomycosis (TDO) represents the most severe variant of onychomycosis (Figure 11). H. Grimmer coined the term glacier nail in the 1960s. At the founding meeting of the German-speaking Mycological Society in Essen on November 15, 1961, he gave a lecture on Histologic studies in nail mycoses (detection of vegetative fungal elements and their significance for griseofulvin therapy). Grimmer then wrote that, apart from exceptions, in a strict anatomical sense, the site of a fungal disease by hyphomycetes primarily represents subungual mycosis and not onychomycosis. The nail bed epithelium reacts to pathogenic effects caused by hyphomycetes with potentially marked acanthosis as well as varying degrees of hyperkeratosis, clinically manifesting itself as elevation of the nail plate. The healthy, growing nail plate pushes itself over the hyperkeratotic nail bed in a glacier-like fashion [44]. They correspond to subungual dermatophytomas, i.e. conglomerates of fungal elements arthrospores, fungal hyphae, and keratin masses underneath the thickened yellowish-brown nail plate. Yellow streaks longitudinal streaks medially or laterally frequently reaching the nail matrix are characteristic for this type of onychomycosis. The entire nail is mycotic and subsequently pushed upward by subungual hyperkeratoses, resulting in onycholysis. In chronic mucocutaneous candidiasis, fingernails become yeast-infected through autoinoculation from mucosal candidiasis of the mouth and tongue, and clinically appear as TDO [45]. By contrast, onychia et paronychia candidosa is caused by yeasts. Yeast infections of the nails are preceded by chronic infections of the proximal and/or lateral nail wall, frequently also by bacterial paronychia. Damage to the matrix then induces structural impairment of the nail plate with horizontal ridges. This may give rise to Candida onycholysis (without nail plate involvement) as distinct clinical entity. Onychomycosis in children Childhood onychomycosis seems to be on the rise. There has been a recent increase in childhood onychomycosis The prevalence of childhood onychomycosis is between 0 % and 2,6 % [46]. Primary cause for 764

17 childhood onychomycosis has been an increase in domestic fungal infections of the feet and nails in parents and grandparents, possibly also siblings. Most cases of onychomycosis show preexisting tinea pedis. Physical activities like soccer are facilitative. Onychomycosis in children with trisomy 21 is less well known, but very common. Obesity and diabetes mellitus represent other significant predisposing factors. From 2002 to 2007, the department of dermatology at the University of Bologna in Italy saw 15 children with onychomycosis, 53 % of whom displayed the same predisposing factors as adults, i.e. domestic onychomycosis, dermatophyte-contaminated surroundings as well as physical activity [47]. Candida onychomycoses have been seen in premature newborns with incompletely developed immune systems, children with chronic mucocutaneous candidiasis, and in iatrogenic immunosuppression. In a retrospective 5-year study at Children s Hospital Colorado and Denver Health Medical Center, 66 (46,8 %) of a total of 141 children with nail disorders suffered from onychomycosis. Among those, the age group between 6 and 10 showed the highest prevalence [48]. Of a total of 1,586 patients from 2008 to 2009, the Institute for Fungal Diseases and Microbiology Berlin, registered 24 children (age: 4 14) with onychomycosis, corresponding to a percentage of 1.5 %. Yeast infections by Candida species Oral candidiasis Candidiases of the oral mucosa (thrush) usually affect immunosuppressed patients. Local factors for example dentures in the elderly also facilitate mucosal infections by Candida albicans. In chronic atrophic oral candidiasis, the tongue and oral mucosa are markedly erythematous. The hyperplastic mucosal infection corresponds to median rhomboid glossitis (midline glossitis). Pseudomembranous candidiasis of the tongue corresponds to thrush. Perlèche or angular cheilitis typically occurs in people with dentures. Candidiases of the oral mucosa (thrush) usually affect immunosuppressed patients. Local factors also facilitate mucosal infections by Candida (C.) albicans, e.g. dentures in the elderly, but also braces in children and adolescents. In chronic atrophic oral candidiasis, the tongue and oral mucosa are markedly erythematous, e.g. in so-called denture stomatitis. The hyperplastic mucosal infection is another variant of candidiasis (median rhomboid glossitis or midline glossitis). Pseudomembranous candidiasis of the tongue corresponds to the common thrush. Symptoms of mucosal candidiasis include burning sensations and occasional pain. Candida leukoplakia affects the tongue and buccal mucosa. Differential diagnoses include oral leukoplakia, mucosal lichen planus, premalignant lesions, squamous cell carcinoma of the oral mucosa, bullous dermatoses like pemphigus vulgaris and Stevens-Johnson syndrome. Perlèche or angular cheilitis typically occurs in people with dentures. In countries with high AIDS prevalence rates, perlèche may even be the first clinical sign of this immunosuppressive disease. Candida cheilitis is a very rare condition. In HIV/ AIDS patients, candidal esophagitis has to be considered in case of retrosternal burning sensations and massive dysphagia. Skin infections by Candida species Cutaneous candidiases predominantly affect intertriginous areas (groins, inframammary skin, interdigital spaces). Cutaneous candidiases predominantly affect intertriginous areas, i.e. groins, abdominal skin folds, inframammary skin, but also interdigital spaces (interdigital candidiasis) (Figure 12a, b). Similar to thrush and perlèche, interdigital mycoses by C. albicans may also be indicative of full-blown AIDS in areas with high HIV prevalence, e.g. Uganda (Sub-Saharan Africa) (Figure 13). Cutaneous candidiasis is typically characterized by erythematous (bright red to purple-red), erosive, dry, scaly, but sometimes also oozing as well as 765

18 Figure 12 Cutaneous candidiasis caused by Candida albicans in an 81-year-old female diabetic. Inframammary candidiasis with erythematous, macerated, scaly, papular, and pustular lesions and accentuated borders (a). Inguinal candidiasis with violaceous, macerated, and markedly inflamed skin (b). Figure 13 Interdigital mycosis caused by Candida spp. in a 23-year-old HIV positive female patient in Mbarara, Uganda. Pustules and collarette scales scaling are signs of cutaneous candidiasis. 766 acerated especially in intertriginous candidiasis lesions with prominent borm ders. Pustules are also typical of cutaneous candidiasis. Non-intertriginous areas characteristically display erythemato-squamous lesions with collaret-like scaling.

19 Figure 14 Pityriasis (tinea) versicolor in a 23-year-old female student. Hypopigmentated lesions on the trunk, shoulders and upper arms (a). Typical bran-like desquamation on the trunk. The so-called shaving phenomenon in pityriasis versicolor can be evoked by using a wooden spatula (b). A special form of cutaneous candidiasis is diaper dermatitis marked by maceration due to wet and warm surroundings, erosion, and desquamation. Candidal folliculitis barbae should be microbiologically differentiated from bacterial folliculitis or tinea barbae caused by dermatophytes. A special form of cutaneous candidiasis is diaper dermatitis marked by maceration and erosion, yet also whitish patches and desquamation. Wet and warm surroundings are a predisposing factor, further aggravated by urine acting as irritant. Similar lesions can be found in elderly, bedridden, and incontinent patients. Differential diagnoses include contact dermatitis, other types of dermatitis, and inverse psoriasis, which may all be secondarily colonized and sometimes subsequently infected by Candida spp. Candidal folliculitis barbae, which occasionally occurs in men, can only be differentiated from bacterial folliculitis or tinea barbae caused by dermatophytes, such as T. verrucosum or T. rubrum, on the basis of microbiologic evaluation. Only rarely does C. albicans cause deep skin/soft tissue infections. Pityriasis versicolor clinical picture Predilection sites for pityriasis versicolor are the upper trunk, chest, shoulders, and upper arms. Pityriasis versicolor is characterized by macular, hypo and hyperpigmented lesions, in which discrete bran-like scaling may be evoked. In Europeans, pityriasis versicolor never affects the face. Predilection sites for pityriasis versicolor are the upper trunk, back, chest, shoulders, and upper arms. In rare cases, the groins and thighs are involved. The disease typically starts with brownish-red (pityriasis versicolor rubra), macular, oval and round, coalescing, flat, apparently non-scaly lesions. This hyperpigmented and macular form of pityriasis versicolor is usually not pruritic (Figure 14 a). Lesions are light brown in color (café au lait) and spread in a map-like fashion without sharp demarcation [49]. Tangential scraping (wooden spatula, scalpel) evokes bran-like scaling, the so-called shaving phenomenon (Figure 14 b). In Europeans, there is a striking contrast between tanned skin (UV rays, sun, tanning salon) and pale, hypopigmented lesions. Pityriasis versicolor alba, the white pseudochromatic variant of pityriasis versicolor, is usually encountered in the tropics. The perifollicular variant of pityriasis versicolor is confined to follicles. Areolar and periareolar pityriasis versicolor is another rare disease manifestation with light brown macules around the nipples [50]. Pityriasis versicolor never affects the face. Dark-skinned people in tropical countries are the exception. Here, typically hypo and hyperpigmented lesions may occasionally also occur on the face. 767

20 Table 3 Diagnostic sampling in mycology. Type of dermatomycosis Tinea corporis Dry, scaly plaques with prominent borders Tinea pedis interdigitalis et plantaris Tinea unguium (onychomycosis) Tinea capitis Instrument used for sampling Scalpel, curette Sample swab for skin scales Curette, scalpel Scalpel, curette, (scissors), fraise (possibly involving a podiatrist) Epilation tweezers, scalpel Procedure Skin scrapings from (inflammatory) lesional borders Vigorous rubbing with a swab or taking skin scrapings using a curette. Sample swabs may also be directly inoculated onto agar. Scalpel: skin scraping from dry, hyperkeratotic areas of the sole. Disinfection with ethanol (70 %) or some other cutaneous disinfectant. Nowadays, disinfection is regarded as optional, as selective agars are used, largely preventing contamination. Shortening nails with scissors; disposing of whole nail pieces; thereafter, tangential removal of the nail plate. Small and medium-sized, crumbly, partially thready subungual nail clippings at the transition between diseased and healthy tissue represent the optimal material for mycologic evaluation. Alternative: (high-speed) electric nail fraise (podiatrist) 1. Shortening hairs with scissors down to roughly 3 5 mm in length; disposing of cut hairs; harvesting approximately hair roots using epilation tweezers. 2. Skin scrapings using a scalpel 3. Taking a swab from pustules or abscesses 4. Brush method: repeated combing of hair with a sterile head massage brush (alternatively tooth brush), followed by direct inoculation onto culture medium (agar plate) A rule states that topical antimycotic treatment may be started if microscopic findings are positive. Systemic antimycotic therapy may only be initiated after fungal cultures have come back positive. Mycologic lab diagnostics Diagnostics in dermatomycology are based on microscopic and cultural pathogen detection. An old rule states that topical antimycotic treatment may be started if microscopic findings are positive. However, systemic antimycotic treatment, if indicated, should only be initiated after fungal cultures have come back positive. Diagnostic sampling In tinea capitis caused by anthropophilic dermatophytes (Trichophyton tonsurans), the hairbrush method is significantly superior to scalp scraping. Apart from skin scrapings, samples include hair roots in case of suspected tinea capitis and nail clippings in case of suspected onychomycosis (Table 3). Particularly in tinea capitis caused by anthropophilic dermatophytes, primarily T. tonsurans, the hairbrush method is significantly superior to scalp scraping (Table 3) [51]. Immediately after brushing scalp and hair, the brush is gently pressed directly onto the agar plate. This method only allows for cultural fungal detection [52]. As scaling is a dominant feature in tinea capitis caused by anthropophilic dermatophytes (T. tonsurans and M. audouinii), the hairbrush technique makes sense. By contrast, in purulent infections, epilation of hairs and possibly simple swabs are more practicable and advantageous. The hairbrush method has proven useful for screening exams among family members and children at school and kindergarten. However, 768

21 scalp scraping and sampling of hair roots using scalpel and tweezers are preferable for individual patients in a pediatric or dermatologic office setting. Microscopic preparations The conventional workup using KOH preparations shows insufficient diagnostic sensitivity, especially in onychomycosis. The fluorescent preparations Blankophor or Calcofluor are the most sensitive microscopic tests for the detection of dermatomycoses. The KOH (potassium hydroxide) examination using 20 % KOH represents the simplest method to microscopically detect fungi in skin scales, nails, and hair roots. Tetraethylammonium hydroxide (TEAH) may be used alternatively. However, the conventional workup using KOH preparations shows insufficient diagnostic sensitivity, especially in onychomycosis (40 68 %). Fluorescent staining with optical brighteners (diaminostilbene) is the most sensitive method to microscopically detect fungi in skin scales, nail clippings, hair roots, hair as well as Scotch tape preparations. These substances bind to chitin, the main cell wall component of fungi. Currently available stains are Blankophor or Calcofluor. Test solutions are prepared with 20 % KOH. Using fluorescent microscopy, spores, yeast cells even tiny Malassezia cells as well as hyphal fragments and arthrospores (disintegrating mycelium) may be differentiated. Cultural dermatophyte detection With respect to cultural fungal detection, every sample should be inoculated onto two culture media, one of them containing cycloheximide (Actidion ) to suppress mold growth. Cultures are incubated at C, optimally at 28 C, for three (to four) weeks. Quite frequently, cultural fungal detection fails, usually because of pretreatment with topical or systemic antimycotics. As fungi are heterotrophic microorganisms, culture media contain organic nutrients required for growth and reproduction, among them a carbon source (glucose), a nitrogen source (peptone, meat extract), water, vitamins, and antibiotics. Every sample should be inoculated onto two culture media, one of them containing cycloheximide (Actidion ) to suppress mold growth. Cultures are incubated at a temperature of C, optimally at 28 C, for three (or four) weeks and visually checked for fungal growth twice weekly. Most dermatophytes also grow at 37 C, yet, because of their adaptation to human skin surface temperature, they do so slightly better at lower temperatures. If a slow-growing dermatophyte is suspected, e.g. T. verrucosum or T. violaceum, the incubation period should be extended to at least four, better 5 6 weeks. The selective dermatophyte agar according to Taplin deserves mention here, as it is a selective but at the same time also differentiation medium. The addition of cycloheximide allows for selective growth of dermatophytes on this cultural medium. One has to bear in mind, though, that, despite the inhibitor, some molds are still capable of growing on this agar. Dermatophytes produce alkaline metabolites that alkalize the initially acidic cultural medium, causing the indicator phenol red to turn from yellow to red, thus signaling dermatophyte growth. T. interdigitale (T. mentagrophytes) does not develop any pigment reminiscent of T. rubrum. This important differential feature of the two most common dermatophytes cannot be assessed on a dye-containing agar. For this, Sabouraud 2 % or 4 % glucose agar is still the best option. By contrast, culture media exhibiting a chemically induced color change (Taplin agar) are inadequate, as they mask the pathogens natural pigment, which also holds true for other pigment-forming dermatophytes. The differentiation of dermatophytes, yeasts, and molds is predicated on macroscopic (upper and bottom side of colonies as well as pigmentation) and microscopic characteristics (formation of macro and microconidia, respectively other growth forms) as well as biochemical properties (for example urea hydrolysis on Christensen urea agar) [53]. Quite frequently, cultural fungal detection fails (sensitivity roughly 70 % in onychomycosis) because of pretreatment with topical or systemic antimycotics. Thus, vital fungi are already inhibited in vivo, preventing them from growing in vitro. 769

22 Figure 15 Histologic detection of dermatophytes. Tinea corporis caused by Trichophyton rubrum. Histology shows hyphae within the stratum corneum, especially adjacent to vital epidermal layers (PAS staining, magnification x 200) (a). Besides septate hyphae within the stratum corneum, the epidermis displays marked spongiosis with neutrophils and incipient intracorneal pustule formation (PAS staining, magnification x 200) (b). Follicle-associated fungal hyphae in dermatophytosis. The epidermis shows hyperkeratosis and parakeratosis with serum lakes, pustule-like accumulation of neutrophils, and a cystically dilated follicular ostium. There is also psoriasiform epithelial hyperplasia with moderate spongiosis. Upper dermal and perifollicular lymphocytes and neutrophils with exocytosis into the epidermis and the follicular ostium (PAS staining, magnification x 100) (c). Dermatophyte hyphae within the stratum corneum labeled with a monoclonal antibody (anti Dermatophyte Mouse Monoclonal Antibody, Xceltis LLC, Mannheim, Germany) allowing for immunohistochemical differentiation of dermatophytes and yeasts. There is, however, some degree of cross-reactivity with various molds (immunohistochemistry magnification x 400. (d). Histologic fungal detection The histologic detection of fungi in fungal nail infections displays a very high sensitivity. In tinea corporis, fungal hyphae are found in the stratum corneum, respectively in follicular epidermal structures (Figure 15 a d). The histologic detection of fungi in fungal nail infections displays a high sensitivity. Specificity, however, is low because it does not offer any indication as to fungal genus or species. While the diagnosis of PSO or PSWO usually requires a punch biopsy from the nail plate, simple nail clippings 770

23 Figure 16 Histologic dermatophyte detection in onychomycosis. Tinea unguium caused by Trichophyton rubrum. Histologic detection of dermatophytic hyphae (Trichophyton rubrum) in pieces of a toenail in tinea unguium. In PAS stained sections, septate hyphae are visible within all layers of the nail (PAS staining, magnification x 200) (a). Higher magnification shows distinctly structured septate fungal hyphae within the amorphous keratin of the nail (PAS staining, magnification x 400) (b). The specificity of histology, however, is low because the morphology of hyphae and spores detected inside the tissue does not offer any indication as to fungal genus or species. To what extent histologically PAS positive fungal elements truly represent evidence for onychomycosis has to be critically checked for plausibility by dermatologists. The possibility of secondary mycotic growth has to be considered. generally suffice in all other forms of onychomycosis [54]. It is imperative that the histologic evaluation of nails be performed by a histopathologist experienced in this field, in order to rule out various differential diagnoses, e.g. yellow nail syndrome, lichen planus of the nail, nail psoriasis, twenty nail dystrophy, congenital and acquired onychodystrophies (brachyonychia), subungual tumors, and melanonychia [55]. In a study conducted at New York University over the course of eight months, all nail samples suspected of onychomycosis were histologically evaluated. A total of 66 patients were thus diagnosed with onychomycosis [54], with 97 % showing subungual hyphae. The remaining 3 % only revealed hyphae in the nail plate but not in subungual tissue. The histologic workup of onychomycosis is a very sensitive method. Herbst et al. [56] employed histologic fungal detection as additional test in patients, in whom repeated microscopic and cultural tests had failed to corroborate the clinical suspicion of onychomycosis. Of 32 histopathologically proven onychomycosis cases, only 14 of 29 (48 %) showed positive cultures. Ten patients suspected of having mycotic leukonychia were mycologically evaluated by culture and nail biopsy [57]. Mycotic leukonychia was subsequently confirmed in nine patients, only four of whom also showed positive cultures. However, H&E and PAS stains revealed fungal elements in all ten patients, eventually yielding a 100 % correlation between onychomycosis and histologic findings. One drawback is the fact that histologic evaluation does not enable any inference as to the causal fungal genus or species. Even the differentiation between dermatophytes, yeasts, or molds is not feasible. In clinical practice, histologic examination has overall proven to be a fast and nearly twice as sensitive method as fungal cultures [58]. Particularly in those cases where neither microscopy nor cultures come back positive for fungi, histologic analysis of nail material facilitates the correct diagnosis (Figure 16a, b). To what extent histologically PAS positive fungal elements truly represent evidence for onychomycosis has to be critically checked for plausibility by dermatologists on a case-by-case basis. This is crucial in order to not false positively interpret secondary mycotic growth, e.g. in nail psoriasis, as fungal nail infection. 771