Diagnostic Evaluation of Superficial Mycoses: A Clinico-Pathological and Mycological Study Using KOH Microscopy and Fungal Culture.

Mycoses that are superficial, such as those that affect the skin, hair, and nails, are a significant public health issue that affects the entire world [1-3]. Dermatophytes are the most common cause of these infections; however, yeasts and non-dermatophyte molds (NDMs) are increasingly being implicated in the development of these infections, particularly in highly complicated nail infections [4,5].

In recent years, dermatophytosis in tropical and subtropical regions has been increasingly considered to be a chronic, relapsing, and atypical presentation. As a result, a pure clinical diagnosis is no longer sufficient for modern clinical practice [6-8].Empirical therapy is a common practice; however, accurate laboratory diagnosis is required due to the increasing antifungal resistance and the rapidly changing epidemiology of fungal pathogens in the Indian subcontinent [9-11]. Despite the fact that KOH direct microscopy is capable of providing rapid presumptive evidence of fungal elements, it is not able to identify the specific genus, species, or sensitivity profiles that are responsible for the infection [12].

Therefore, fungal culture continues to be the gold standard for accurate species identification as well as for epidemiological monitoring over an extended period of time [13,14].The current study was conducted with the purpose of examining the clinico-mycological profile of patients who were attending a tertiary care center in Kalaburagi and who were diagnosed with superficial mycoses. Additionally, the study aimed to correlate clinical types with laboratory findings obtained through KOH microscopy and fungal culture.

Study Design and Setting:

This cross-sectional, observational study was conducted in the Department of Dermatology and Microbiology at ESIC Medical College and Hospital Kalaburagi, Karnataka, India.

Study Period:

The study was conducted over a period of one year, from January 2024 to December 2024.

Study Population:

The study included a total of 128 patients presenting with clinically suspected superficial mycoses (including various types of tineae and onychomycosis) during the specified study period.

Specimen Collection and Processing:

After cleansing the affected area with 70% ethanol, clinical specimens (skin scrapings, nail clippings, and hair stumps) were collected using sterile blunt scalpels.

Each specimen was divided into two portions:

Direct Microscopy: The first portion was subjected to KOH preparation (10% for skin/hair, 20% for nails) and examined under a light microscope for the presence of fungal hyphae, spores, or yeast cells.

Fungal Culture: The second portion was inoculated onto Sabouraud Dextrose Agar (SDA) with and without cycloheximide and chloramphenicol. Cultures were incubated at 25°C and 37°C for up to four weeks.

Variables Analyzed:

Patient demographics (age, sex, urban/rural residence), clinical presentation (e.g., Tinea corporis, Tinea cruris, Onychomycosis), KOH status, and specific fungal isolates were recorded and analyzed.

A total of 128 clinically suspected cases of superficial mycosis were evaluated. The demographic distribution is presented in Tables 1, 2, and 3. The peak incidence was observed in the 31–50 age range, comprising 65 out of 128 patients (50.8%). Males (62.5%) were more predominantly affected than females (37.5%). Most patients belonged to an urban demographic (64.8%).

Table 1: Age Distribution of Total and Culture-Positive Cases

Table 2: Sex Distribution

Table 3: Distribution by Residence

Tinea corporis was the most common clinical presentation, accounting for 42.2% (n=54) of the cases, followed by Onychomycosis (25.0%, n=32).

Table 4: Clinical Types of Superficial Mycoses

Comparing the two diagnostic modalities, 74 cases were KOH positive, and 73 were culture positive. 58 cases were positive by both methods. KOH microscopy demonstrated a higher diagnostic yield in culture-negative cases (n=16) compared to culture detecting fungus in KOH-negative cases (n=15).

Table 5: Correlation of KOH Microscopy and Fungal Culture

Among the 73 culture-positive cases, dermatophytes were the predominant pathogens, but non-dermatophyte molds also formed a substantial cohort. Trichophyton mentagrophytes was the single most common isolate (n=21).

Table 6: Profile of Fungal Isolates (n=73)

The correct diagnosis of superficial mycoses is of great importance because of its high prevalence and the ever-increasing problem of recalcitrant infections [15]. In our study in ESIC Medical College and Hospital Kalaburagi maximum number of cases were in third to fifth decades of life (31-50 years). This is in agreement with the results obtained by Singh et al. and other Indian cohorts [16,17] who found that active age group is highly susceptible to fungal infections due to physical exertion, occupational sweating and outdoor activities. We also noted a marked male preponderance (62.5%) which was in agreement with the demographic findings of Bhatia et al. and other regional epidemiological surveys [18,19]. This is generally attributed to increased outdoor labour and use of occlusive footwear or close-fitting clothes.Clinically the most frequent presentation was Tinea corporis (n=54, 42.2%) followed by onychomycosis (n=32, 25.0%). Similar clinical distributions have been previously reported by Kaur et al. and contemporary reports [20,21] in tropical tertiary care settings with high rates of T. corporis attributed to hot, humid climates that promote fungal colonization on glabrous skin.As for diagnostic modalities, KOH was positive in 57.8% of cases and culture in 57%.

Interestingly, 16 cases were KOH positive but culture negative, which can occur secondary to non-viable fungal elements, often because of prior empirical antifungal therapy [22]. Conversely, 15 cases were culture positive with a negative KOH smear which points out that direct microscopy alone may miss low fungal burdens [23].This study describes a major epidemiological shift in the mycological profile. Historically, T. rubrum was reported as the most frequent dermatophyte across India [20]. But our data showed that T. mentagrophytes (n=21) was the predominant species than T. rubrum (n=13).

This switch is well supported by recent literature. Dogra et al. and regional studies [8,24] have also reported a similar changing epidemiological trend, with T. mentagrophytes being heavily implicated in the rise of chronic, steroid-modified dermatophytosis (including Tinea incognito cases, of which we recorded 5).

The isolation of non-dermatophyte molds (NDMs) was also very high, especially Aspergillus species (n=18). The high percentage of onychomycosis cases (n=32) in our cohort is in good agreement with the high rate of NDMs. Aspergillus and Scopulariopsis are now increasingly recognized as the primary pathogens of nail infections and not laboratory contaminants and require specific culture identification to direct appropriate systemic therapy [5,25]. This has been comprehensively established by Gupta et al. and subsequent mycological studies.

Our study reveals that Tinea corporis remains the most prevalent clinical form of superficial mycoses, mainly affecting active adult males. The dominance of Trichophyton mentagrophytes and the high isolation rate of Aspergillus species suggest a continuing shift in the fungal epidemiological scenario in India. Both modalities were successful in identifying cases that were missed by the other and the combined use of KOH microscopy and fungal culture continues to be the gold standard for accurate and reliable diagnosis and targeted therapeutic intervention.

1.       Hay RJ, Johns NE, Williams HC, et al. The global burden of skin disease in 2010: an analysis of the prevalence and impact of skin conditions. J Invest Dermatol. 2014;134(6):1527-34.

2.       Havlickova B, Czaika VA, Friedrich M. Epidemiological trends in dermatophytosis worldwide. Mycoses. 2008;51 Suppl 4:2-15.

3.       Ameen M. Epidemiology of superficial fungal infections. Clin Dermatol. 2010;28(2):197-201.

4.       Welsh O, Vera-Cabrera L, Welsh E. Onychomycosis. Clin Dermatol. 2010;28(2):151-9.

5.       Gupta M, Sharma NL, Kanga AK, Mahajan VK, Tegta GR. Onychomycosis: Clinico-mycologic study of 130 patients from Himachal Pradesh, India. Indian J Dermatol Venereol Leprol. 2007;73(6):389-92.

6.       Verma SB, Panda S, Nenoff P, et al. The unprecedented epidemic-like scenario of dermatophytosis in India: II. Clinicomycological and therapeutic aspects. Indian J Dermatol Venereol Leprol. 2021;87(3):326-32.

7.       Rajagopalan M, Inamadar A, Mittal A, et al. Expert Consensus on The Management of Dermatophytosis in India (ECTODERM India). BMC Dermatol. 2018;18(1):6.

8.       Dogra S, Uprety S. The menace of chronic and recurrent dermatophytosis in India: Is the problem deeper than we perceive? Indian Dermatol Online J. 2016;7(2):73-6.

9.       Sahoo AK, Mahajan R. Management of tinea corporis, tinea cruris, and tinea pedis: A comprehensive review. Indian Dermatol Online J. 2016;7(2):77-86.

10.    Nenoff P, Verma SB, Vasani R, et al. The current Indian epidemic of superficial dermatophytosis due to Trichophyton mentagrophytes-A molecular study. Mycoses. 2019;62(4):336-56.

11.    Khurana A, Sardana K, Chowdhary A. Antifungal resistance in dermatophytes: Recent trends and therapeutic implications. Fungal Genet Biol. 2019;132:103255.

12.    Panhetiya PN, Patel K. Direct microscopy versus culture for the diagnosis of superficial mycoses. J Clin Diagn Res. 2017;11(9):WC01-WC04.

13.    Weitzman I, Summerbell RC. The dermatophytes. Clin Microbiol Rev. 1995;8(2):240-59.

14.    Hainer B. Dermatophyte infections. Am Fam Physician. 2003;67(1):101-8.

15.    Pires CAA, Cruz AM, Del Rosso JQ. Epidemiology of dermatophytosis in a tertiary care dermatology center. J Clin Aesthet Dermatol. 2014;7(2):30-5.

16.    Singh S, Beena PM. Profile of dermatophyte infections in Baroda. Indian J Dermatol Venereol Leprol. 2003;69(4):281-3.

17.    Agarwal US, Saran J, Agarwal P, et al. Clinico-mycological study of dermatophytes in a tertiary care centre in northwest India. Indian J Dermatol Venereol Leprol. 2014;80(2):194.

18.    Bhatia VK, Sharma PC. Epidemiological and mycological profile of dermatophytosis in a tertiary care center in North India. Indian J Dermatol. 2014;59(3):314-15.

19.    Lakshmanan A, Ganeshkumar P, Mohan SR, et al. Epidemiological and clinical pattern of dermatomycoses in rural India. Indian J Med Microbiol. 2015;33 Suppl:134-6.

20.    Kaur R, Kashyap B, Bhalla P. A five-year mycological study of dermatophytosis in a tertiary care hospital in Delhi. Indian J Med Res. 2015;141(3):323-26.

21.    Rengasamy M, Shenoy MM, Majumdar B, et al. Clinico-mycological profile of dermatophytosis in a tertiary care centre. Indian J Dermatol. 2011;56(4):460.

22.    Peerapur BV, Inamdar AC, Pushpa PK, et al. Clinicomycological study of dermatophytosis in Bijapur. Indian J Med Microbiol. 2004;22(4):273-4.

23.    Sharma R, Adhikari L, Sharma RL. Incidence of dermatophytosis in Sikkim. Indian J Med Microbiol. 2012;30(1):123.

24.    Vineetha M, Girish PN, et al. Clinicomycological study of dermatophytosis in Kerala. Indian J Dermatol. 2018;63(4):300-304.

25.    Sen SS, Rasul ES. Dermatophytosis in Assam. Indian J Med Microbiol. 2006;24(1):77-8.