Epidemiology of Dermatophytes and Non Dermatophyte Molds among Patients Attending Rank Higher Specialized Dermatology Clinic, Addis Ababa, Ethiopia

Background: Dermatophytes are keratinophilic fungi that infect keratinized tissues causing diseases known as dermatophytosis. Dermatophytosis is common worldwide and continues to increase. Objective: This study was undertaken to determine the prevalence of dermatophytes and the spectrum of fungal agents in patients attending Rank higher clinic. Methods: a cross sectional study has been conducted, in which 318 Samples were collected from patient’s hair, nail and skin. A portion of each sample was examined microscopically and the remaining portion of each sample was cultured onto plates of Sabouraud’s dextrose agar containing chloramphenicol with and without cycloheximide. Dermatophyte isolates were identified by studying macroscopic and microscopic characteristics of their colonies. Result: Of 318 samples, fungi were detected in 133 (54.4%) by direct wet mount while 148/315(46.5%) of them were culture positive. From these 72/148(46.8 %) were dermatophytes. Among dermatophyte isolates T. tonsurans 29/72(40.2%) was the most common cause of infection. Tinea capitis was the predominant clinical manifestation accounting for 170/315(53.4%) of the cases. Patients with age group 1-14 years were more affected. T. tonsurans was the most common pathogen in tinea capitis, whereas T. mentagrophytes was the most common pathogen in tinea corporis. Conclusion: In this study the prevalence of dermatophytes were higher. Further intensive epidemiological studies of dermatophytes induced dermatophytosis which have public health significance are needed.

by dermatophytes, which are fungi basically molds that require keratin for growth. An increase in the incidence of dermatophytosis has been noted worldwide, especially in developing countries [2,3]. In particular, tinea capitis one of the most common dermatophytosis represents a major public health issue among children in developing countries; mainly in children's of African or Caribbean origin [3].
Geographic location, health care, immigration, climate (temperature, humidity, wind, etc.), overcrowding, environmental hygiene culture, awareness to dermatophytes , age of individuals, hygiene and socioeconomic conditions have been described as major factors for these variations of dermatophyte epidemiology [4,5,6,7]. As human contact among children is more frequently between the ages of 4 and 16 years than in very early childhood; this age group is similarly at greater risk of contracting infectious diseases from different sources [8,9,10].
Dermatophyte fungi have a worldwide distribution, and now days, there are about 40 known species in the genera of dermatophytes. Of these species, about 25 species belong to the three most recognized and prevalent worldwide genera's; Epidermophyton,
Ethiopia is a developing nation located in the tropical region with wet humid climate which makes it to fell into regions with high prevalence of dermatophytosis. So, conducting further studies to know the actual magnitude of dermatophytosis as well as the spectrum of its etiological agents among the general population is of the highest priority [9,10].

Study areas and population
A prospective cross-sectional institution based study was carried out from January 2018 to June 2018 among dermatophytosis suspected patients who visit Rank higher specialized dermatology clinic, Addis Ababa, Ethiopia.

Sample size determination and sampling technique
The required sample size of the study was determined using the formula for single population proportion. A total of 318 clinical samples were collected from patient visiting the Dermatology clinic during the study period were included in the study. Single specimen was taken from each patients based on their clinical manifestation.

Biological sample collection
The samples were collected from January 2018 to June 2018 using convenient sampling technique. Before collecting the sample the infected area was cleaned with 70% (v/v) ethanol. Skin, nail, and finger scrapings were collected aseptically using sterile blades and transferred into sterile plastic petri-dishes. In tinea capitis suspects, dull broken hairs from the margin of the scalp lesion with forceps was sampled and transferred to sterile folded papers and transferred in to sterile petri dishes.
Culture and microscopic examination: A portion of each sample from a nail, finger skin and scalp scraping was mounted in a drop of 10% (w/v) potassium hydroxide on a clean microscopic slide. After 5 minutes; the mount preparation was examined under low (×10) and high (×40) power magnification for the presence of any fungal elements. The remaining portion of each clinical sample from the different sites was cultured in to Sabouraud's dextrose agar plates containing chloramphenicol with and without cycloheximide (Oxoid, Basingstoke, England) prepared according to the instruction of the manufacture. All inoculated plates were then incubated at inverted position at 25-30 oC for 4-6 weeks. Culture plates containing the Sabouraud's dextrose agar were examined twice a week for any fungal growth. In the absence of growth during 6th week, the results were considered negative. Those suspected colonies for dermatophytes were sub cultured into potato dextrose agar for the production of spores. Cultures of dermatophytes were identified by examining macroscopic and microscopic characteristics of their colony on Sabouraud's dextrose agar. For the macroscopic identification; rate of growth, texture, topography, and pigmentation of the reverse and front side of the culture were employed.
Mold isolates was identified microscopically by placing pieces of the colony from Sabouraud's dextrose agar plates in to clean microscopic slide and stained using lacto phenol cotton blue stain. Each preparation was observed microscopically after placing a cover slip. In addition, urease test was also used in differentiating between some members of Trichophyton species. For the identification of yeasts, candida CHROME agar was also used.

Statistical analysis
Data was collected, double entered, cleaned and analyzed using SPSS version 20 software according to the study objectives. Frequency and percentage were used for investigation of the outcome.

Result
A total of 318 dermatophytosis suspected patients were included in this study. Among these; fungi were detected in 133(41.8%) samples using potassium hydroxide wet mount while, 148 (46.5%) were culture positive (Table 1).
In the present study a total of 318 clinical samples were collected from suspected cases of dermatophytosis of which 125 (39.3%) were males and 193(60.7 %) females ( Table 2).
The age of the study subject was ranged from 1 to 88 year with a mean age of 16 year.
Among non dermatophyte molds Cladosporium spp was the predominate isolate followed by Neoscytalidim dimidatum and Alternaria spp respectively .
Yeasts were the least common isolates accounting 13(8.7 %) of the total suspects of dermatophytosis (Table 4) (Table 4). Clinical manifestation in relation to age group depicted that patients with age group 1-14 were highly affected accounting for 45.2 % of the cases followed by age group 25-44 years accounting for 30.1 %. Tinea capitis was found to be more in patients of age group 1-14 years and tinea corporis in patients of age group 45-64 years. Tinea unguium was common in patients of age group of 25-44 years ( Table 2).
Tinea capitis was the predominant clinical manifestation accounting for 53.4% of the cases; of which 98 (57.6%) were females and 72 (42.4 %) males. This was followed by tinea corporis and tinea capitis accounting for 30.5% and 16% of the cases, respectively (Table 5). According to species frequency in different areas of involvement, T.tonsurans was the most common pathogen in tinea capitis, whereas T. mentagrophytes was the most common pathogen in tinea corporis (Table 5).

Discussion
Accurate diagnosis based on the clinical symptoms alone is often difficult. Currently, the diagnosis of dermatophytosis is confirmed by clinical examination and screening of the collected clinical specimen by direct microscopy and fungal culture [13]. An accurate diagnosis of dermatophytosis is important for its successful treatment. The risk of developing adverse drug reactions, the cost and long duration of the therapy, and possible interactions with concomitant medications all affects the importance of accurate diagnosis of the condition before commencing therapy [13,14]. In the present study, direct microscopy positivity rate is 41.8% and culture positivity rate is 46.5 % (Table 1). This was in line with study by Tekleberhan et al, in Ethiopia, found that 31.1% and 42.6% using potassium hydroxide direct wet mount and culture results respectively [9]. This high prevalence could be due to that; Ethiopia is a tropical country with wet humid climate, large population size, and low socioeconomic status and this is supported by other studies conducted on dermatophytosis etiologies and risk factors [15,16].
The most susceptible persons to tinea capitis were among patients 1-14 years (45.2%) this could be because of the lack of protective fatty acids in their scalp. This infection was rarely reported in persons above fifty years of age. Earlier, several authors have supported this finding [17,18]. Many cases occurring in adults is involved with hormonal disorders resulting in carryover of tinea capitis from childhood or in patients with severe immunodepression due to leukemia, lymphoma, or treatment with immunosuppressant drugs [19,20].  4.3%) and Trichophyton rubrum (4.3%) being the least [25]. Even though it is not in line with the current finding; a similar study in Ethiopia showed that the prevalence of T.tonsurans in the rate 18.4% [9].
However, in a recent study conducted by Bitew A, 2018 in Addis Ababa, Ethiopia T.violaceum was the dominant causative agent of tinea capitis [21] and this study shows dissimilarity results with the current study. But other studies showed that species of T .tonsurans are circulating in the population. In a study conducted by Raccurt  tonsurans is highly contagiousness and the role of children scalp for tinea capitis [22,23].
In developing countries such as Kenya and other sub-Saharan countries, the most common agent is M. canis followed by T. tonsurans. Trichophyton species had the highest prevalence of 61.3%, with T. tonsurans being the most predominant dermatophyte due its ubiquitous in nature and abundance among human carriers [20,26,27]. This could show that T.tonsurans is spreading in Ethiopia.
The anthropophilic Microsporum species cause a contagious disease and they are endemic in many countries. In the current study; Tinea capitis is the most common clinical disease followed by tinea pedis and tinea corporis. The zoophilic Trichophyton and Microsporum species are seldom responsible for more than minor outbreaks of human infections. T. mentagrophytes, T. verrucosum, T. tonsurans, T. violaceum, and M. audouinii species are causal agents of tinea capitis [24,25].
Tinea corporis was the second most common infection among the enrolled patients with a significant incidence among age groups of 25-44. The site of infection was mostly restricted to face and neck. T. mentagrophytes, T.rubrum and M.audionii were the main causative agents. In a study conducted by Teklebirhan et al 2015; tinea unguium was the dominant clinical manifestation involving 51.1% of the total cases of dermatophytosis. [9]. But in our study Tinea corporis was the second common clinical presentation accounting for 33 (10.8%) next to tinea capitis and this is in line with study conducted in Harari regional state, Ethiopia [28].
Tinea unguium was also observed mainly on the age of 15-24 (Table 4).Since onychomycosis infections in children is not common due to many reasons such as; rapid growth of the nail, have less exposure to fungal infection risk factors than adults such as pedicure and manicure repeated aggressiveness, frequent housework and cosmetic reasons [16,29]. So, the high occurrence of tinea unguium in this study among these age groups could be due to such factors. But in other study more males and elderly patients were highly affected than the adults [30].
Non dermatophytic molds were isolated from 63 cases (42.56%) with Cladosporium spp. as a major isolate accounting 30% of the total non dermatophyte mold isolates. Similar recent study done in Ethiopia supports this finding [21]. Cladosporium species are dematicious fungi, ubiquitous and they are infrequently associated with human and animal opportunistic infection. It is also the most widely spread fungi in the world. It is true also; most of the time appears as a contaminant. But some studies showed that they are associated as opportunistic infection in subcutaneous and disseminated form, especially among immune depressed individuals [32]. Neoscytalidum dimidatum was the second most common isolated fungi from the non dermatophytes. They were isolated from skin and nail scrapings predominantly of toenails. Neoscytalidium dimidiatum and Scytalidium hyalinum are common causative agents of human superficial infections in different parts of the world especially in tropical and subtropical region [21]. Similarly, yeasts were isolated from 13 cases with C. albicans as a major isolate accounting 38.4% of the total yeast isolated and this study was similar with study conducted in Saudi Arabia [33]. Non dermatophyte fungi were isolated as a cause dermatophytosis in many studies [33,34].
Conclusion: This study showed that the prevalence of dermatophytosis was 46.5% which is more or less similar to study conducted in Ethiopia [9] but in developed countries showed that less than 5% [35] which indicates that dermatophytosis is still a common problem in developing countries .Tinea capitis was identified as the most prevalent clinical presentation and children's are the most vulnerable group. This study found that T.

Consent for publication
Not applicable as details like; videos or images related to study subjects were not recorded for this study.

Funding
The study was supported by Addis Ababa University, College of health science, department of medical laboratory science. The funder had no role in data collection, study design, data analysis and interpretation.

Availability of data and material
The data sets used or analyzed during the current study are available from the corresponding author on reasonable request.

Competing of interest
The author's listed above declare that there is no competing of interest.