Resistant Dermatophytosis, the Causative Species, and Treatment

Background : Dermatophytosis was a fungal infection caused by molds (dermatophytes). Dermatophytes are a group of closely related filamentous fungi able to damage and utilize keratin found in the skin, hair, and nails. Objective: To to isolate the species which cause resistant dermatophytosis, and clinical types of tinea, as well as to assess the response to different antifungal therapies. Patients and Methods: Ninety-two patients with different clinical types of tinea infections (ringworm) were seen in a private clinic in Baquba city for the period from May 2021 to December 2021, they were (42) females and (50) males, their ages ranged from (7-70) years with a mean age of (27.57±8) years. All patients were diagnosed clinically as ringworm and supported by isolation of the species from samples either by direct examination of samples or cultures on Sabouraud media, and the patients were treated by combination therapy of systemic and topical terbinafine and systemic azole (itraconazole capsules) for (1-3) months. Results: All patients complained of widespread and concomitant tinea. The most common causative dermatophytes species were Epidermopyton floccosum (44%) and Trichophyton rubrum (22%), less commonly Microsporum audouinii (9.8%), and the others as in Table (1). The most common clinical type was tinea pedis (29.3%) then tinea cruris (26.1%), tinea corporis (22.8%), and tinea ungum (20.7%) as in Table (2). All patients were cured on combination therapy of systemic (terbinafine and itraconazole) and topical (terbinafine). Five patients (5.4%) showed relapses of disease after discontinuation of therapy and retreated by the same method. Conclusion: Epidermophyton floccosum and Trichophyton rubrum were the most common cause of resistant tinea. Terbinafine and itraconazole were good therapeutic options for the treatment of resistant tinea.


Introduction
Dermatophytosis is a group of fungal infections caused by molds (dermatophytes) [1]. A range of infections ranging from limited skin, subcutaneous to disseminated and may be fatal infection, variable according to the defense immunity of the host [2]. Dermatophytes are a group of pathogenic, not-opportunistic fungi, which need the ORIGINAL RESEARCH Published: 30 June 2023 Doi: 10.26505/DJM.24027091113 breakdown of keratin of the skin, hair, and nails to supply nutrition for survival and production of lytic enzymes (proteases and lipases)which allows fungal colonization and induction of infection in the individuals [3,4]. Dermatophytes are classified according to the source into Anthropophilic, Zoophilic, and Geophilic. Anathropophilic usually enhances chronic resistant infections with a minimal inflammatory response (e.g. Trichophyton tonsurans) [4], which is opposite to severe inflammation induced by Zoophilic type (e.g. Arthroderma benhamiae M. canis) [5]. The development of resistant dermatophytes against different antifungal therapy may result from defects in cellular immune response and production of pro-inflammatory (IL2, IL6, and IL8) rather than antiinflammatory (IL4, IL10, and IL11) interleukins [6].
The most common dermatophytes which liable to develop resistance against systemic antifungals are Trichophyton rubrum, T. interdigital, and T. mentagrophytes [7,8]. Widespread use of topical therapy, containing a combination of antibacterial, antifungal, and steroids or topical potent steroids (like Clobetasol dipropionate) is responsible for the development of resistance dermatophyte or relapsing typical and widespread infections [9][10][11][12]. The resistant dermatophytosis is diagnosed clinically and supported or confirmed both in vivo and in vitro by either susceptibility testing (high MIC) or through molecular study (resistance mutation) [12][13][14][15][16].
The clinical criteria which are useful as an indicator of multidrug resistance and modified dermatophytosis are 1. widespread infection previously treated with multiple regimens. 2. Presence of concomitant infections such as tinea corporis, cruris, and faciei, or localized tinea not cleared by normal antifungal regimens therapy [17,18]. many studies deal with resistant dermatophytosis, and combination therapies of (Terbinafine, Azole, And Griseofulvin) are used [19].
The present study aimed to isolate and identify the species of dermatophytes that cause resistant dermatophytosis (tinea), the clinical types of tinea, and how to treat these infections.

Patients and Methods
A total number of (92) patients with different clinical types of tinea (ringworm) infections, were seen and examined in a private clinic in Baquba city, they were (42) females and (50) males, and their ages ranged from (7-70) years, for the period from May 2021 to December 2021. They complained of skin lesions of different sizes and different sites of the body, for (1-12) month's duration of lesion appearance, the lesions were diagnosed clinically by a specialist physician as dermatophytosis and supported by isolation of dermatophytes by culture and study of microconidia by using direct KOH examination.
All patients were interrogated regarding their age, sex, occupation, marital status, residence, family history, duration of the lesions and medical therapies. All patients were either misdiagnosed and treated by topical corticosteroids or treated by topical and systemic antifungals of bad quality. Most of the patients had a family history of the same lesions, affecting more than one member, particularly the partners. The clinical diagnosis of tinea was supported and confirmed by isolation of the dermatophytes, either by direct examination or culturing of scrubbing samples, (92) samples were taken from the patients and examined as follows:

Direct examination
Nail clippings and skin scrubs were exposed to direct inspection by putting the sample on a clean glass slide and adding drops of 30% KOH, then covering the specimen with a cover slide and leaving the slide for (30) minutes for the nail clips and directly examined for skin scrubs then for the presence of fungal hyphae [20].

Indirect examination (Culture of specimens)
Specimens of nail clipping and skin fragments and hair were cultured on Sabouraud Dextrose Agar (SDA) plates at pH 6.5, the plates (triplicate) were incubated at 28℃ for 7-14 days, with daily examination and observation.

Identification of the dermatophytes
According to [21] the bellow criteria were taken into consideration during identification: -Colony morphology (color and consistency).
-Reverse color (color changes with age). -Microscopic characteristics (microconidia and macroconidia, their size, arrangement, shape, and conidial ontogeny). The presence of growth was seen favorably. Colony morphology on medium, colony microscopic features and other pertinent biochemical tests were carried out for further assessment and dermatophyte species detection. The shape and arrangement of the spores were examined microscopically.

Growth on Potato Dextrose Agar (PDA) Medium
This test was used to differentiate between T. mentagrophytes and T. rubrum, according to the ability to produce red color on the reverse side of the plates with PDA medium. The fungal inoculum was taken from the colony edge and put in the center of the plates, and then the plates were incubated at 28℃ and examined after 14 days for the observation of media color [22]. All patients were treated with topical terbinafine cream three times daily, terbinafine tablets (250 mg) orally once daily and itraconazole capsules (100 mg) twice daily for one to three months according to the response.

Statistical Analysis
The data were analyzed using numbers, and percent. Frequency and also demographic data were analyzed by descriptive statistics, and quantitative data were expressed as mean ± standard deviation of the mean (SD).

Results
Ninety-two patients with different clinical types of dermatophytosis were seen and examined in a private clinic in Baquba City for the period from the 1 st of May 2021 to the 12 th of December 2021. They were (42) females and (50) males with ages ranged from (7-70) years and a mean (27.57 ± 8) years.
All patients had widespread ringworm and concomitant infection (more than one clinical type). Some pateints had tinea corporis, cruris, and faciei , others had tinea corporis, cruris,and pedis, while some pateints had tinea corporis, unguim, and cruris, as shown in Table ( topical corticosteroid therapy or by insufficient antifungal therapy. Also, all patients had more than one member of the family was infected, and the patient's residents from all cities of Diyala Province. The clinical diagnosis was supported and confirmed by the isolation of dermatophytes from scrubbing off the lesions either by microscopical examination or by culturing, and which give the following results as shown in Table ( According to the site of infection as shown in Table( For most patients with concomitant infection, the isolate showed T. rubrum, T. mentagrophytes, and M. audouinii, and for those with tinea pedis, most of them were infected by E. floccosum. Flow up of therapy for one to three months, all patients showed clearance of the ringworm, and five patients (4.6%) showed relapses of the disease after discontinuation of therapy, and another course of therapy was started.

Discussion
Fungal infections were a common disease in Iraq, both dermatophytes and candidiasis. In this study, 100 patients were included, and most of them have different types of dermatophytosis, which was concordant with other studies [5]. In our study, the development of resistant dermatophytosis appeared as a result of either using a topical potent corticosteroid (Clobetazol diprobanol) or using topical azole antifungal of bad quality or the insufficient duration which results in the emergence of resistant dermatophyte, which is also similar to other studies [9][10][11][12]23].
Clinically our study showed that dermatophytosis was a widespread infection and in most of the patients was concomitant, which means the presence of tinea corporis, cruris, faciei, and sometimes tinea pedis or unguium, which also go with other studies [17,18].
Eighty patients (80%) showed a positive family history of more than one member, especially the partners, this indicates that the tinea was contagious and the species of dermatophyte which cause dermatophytosis were anthropophilic in origin and this was supported by isolation of the T. rubrum from (21) patients (21%), M. audouinii from (9) patients (9%), and E. floccosum from (41) patients (41%) of samples these were anthropophilic, which agree with other studies of first groups [1,24] and different from the second and third groups [7,8].
Also regarding the species of dermatophyte which developed resistance in our study were approximately similar to that seen in different countries, (6%) Trichophyton Spp., (29%) Microsporum spp. and (2%) Epidermophyton floccosum but with different percentage [8]. Also, the type of therapy was similar in our study and other different studies [19,24], but the clearance was better in our study.

Conclusions
Epidermophyton floccosum and Trichophyton rubrum were the most common cause of resistance dermatophytosis. Tinea pedis was the most common clinical type of infection. The most common cause of the development of resistance was a rowing use of topical corticosteroids or bad-quality of antifungal therapy. Oral terbinafine and itraconazole were good options for the treatment of resistant cases of dermatophytosis in combination with topical terbinafine cream.

Recommendations
We recommend that you see a specialist doctor if symptoms of the aforementioned disease appear, as soon as possible, to take the necessary treatment in the right way, and to avoid taking steroids randomly.

Source of funding:
The current study was funded by our charges with no any other funding sources elsewhere.
Ethical clearance: Ethical approval was obtained from the College of Medicine / University of Diyala ethical committee for this study.