Mycologic Evaluations in Chronic Leg Ulcers

      Abstract: Background. Thus far, the role of fungi in superinfection of chronic leg ulcers has been poorly studied. Many articles are based on either a small number of patients or single cases. Furthermore, the study conclusions are conflicting. Objective. The objective of this study was to evaluate the percentage of mycotic superinfections and their clinical importance in chronic leg ulcers. Methods. A group of 149 consecutive patients without diabetes was subjected to mycologic examination of the ulcers. Two specimens were obtained from each ulcer. Results. Mycologic examinations were positive in 11 patients (7.4%). Candida albicans was the most frequently isolated species (7 patients), followed by C. parapsilosis (2 patients), C. krusei (1 patient), C. parapsilosis and C. lipolytica (1 patient). Neither dermatophytes nor molds were isolated. No particular features characterized the clinical appearance of ulcers superinfected by fungi, and no clinical improvement of ulcers with mycotic superinfections was observed with oral antimycotic therapy. Conclusion. Based on the results of this study, mycologic examinations may be considered unnecessary in patients without diabetes who suffer with chronic leg ulcers. The role of fungi in superinfections of chronic ulcers of the skin of lower limbs has been poorly studied thus far. More than 20 articles on this topic have been published over the last 40 years,^1-21 many of which have been based on a small number of patients^5,7,10,13,16 or single cases.^2,11,18 Furthermore, conclusions of these studies are conflicting.      This study presents the results of mycologic examinations performed on a group of 149 patients without diabetes who had chronic leg ulcers.

Patients and Methods

     All patients were observed at the Outpatient Department for Chronic Ulcers of the Skin, Institute of Dermatological Sciences, University of Milan, Italy. None of the patients had diabetes but all had chronic leg ulcer(s) of at least 2 months’ duration. Mycologic examinations were carried out after a 2-week wash out period using topical and/or systemic anti-infectious agents. All ulcers were previously washed with 0.9% NaCl. Two swabs were subsequently applied at the inner border, as well as at the deeper portion of each ulcer bed. Pathologic material was cultured on Sabouraud-dextrose agar medium with an added 0.5% chloramphenicol. Plates were subsequently incubated at 28°C for two weeks.      Results of semiquantitative cultures were evaluated as 1+ (light growth), 2+ (moderate growth), and 3+ (heavy growth). Isolated yeasts were identified by germ tube production and morphology on potato-carrot-ox gall agar. Isolates not producing chlamydospores were identified by means of ID32C® (bioMérieux, Rome, Italy). Results The study group consisted of 149 consecutive patients without diabetes (75 men and 74 women). The age range was 28 to 95 years (mean: 67.2 years). Etiopathogenetic classification of ulcers is presented in Table 1. Mycologic examinations were positive in 11 patients (7.4%). Five of the 11 patients had venous ulcers, 2 had atherosclerotic ulcers, and 1 each had a post-traumatic, pressure, vasculitic, and neuropathic ulcer. The following fungal species were isolated: C. albicans (7 patients), C. parapsilosis (2 patients), C. krusei (1 patient), C. parapsilosis, and C. lipolytica (1 patient). No other fungi (dermatophytes or moulds) were isolated. Seven patients were treated with oral fluconazole (100 mg/day for 2 weeks) and 4 with oral itraconazole (200 mg/day for 2 weeks). No clinical improvement of the ulcers was observed with this therapy.

Discussion

     As previously mentioned, several studies published on mycotic superinfection of chronic lower leg ulcers are based on small numbers of patients or are anecdotal cases. Furthermore, conclusions of these studies were different. In studies based on a large number of patients^{3,9,12,14,20,21} the percentage of ulcers with mycotic superinfection ranged from 1%²¹ to 24%.⁹ However, this percentage reached 100% in some studies based on a small number of patients: 17/17 patients with mycotic superinfections in the study by Smith et al⁷ and 6/6 patients in two Swedish studies^10,13 (Table 2). C. albicans was most frequently isolated;^3,9,10,12,13 however, according to other authors, C. parapsilosis was more common.^5,7,16,18,20 Hansson et al12 studied antibody titers against C. albicans using indirect immunofluorescence. Mean antibody titer was 434 in patients whose ulcers were superinfected by C. albicans, 123 in control patients with negative mycologic cultures, and 258 in control patients with basal cell carcinomas or actinic keratoses. Other yeasts, such as C. famata,²⁰ C. glabrata,^16,20 C. guilliermondii,⁹ C. humicola,¹⁶ C. kefir,²⁰ C. krusei, ²⁰ C. lipolytica, ²⁰ and C. tropicalis,^16,20 were isolated almost exclusively in patients with diabetic foot ulcers^16,20 (Table 2).

     The isolation of molds, such as Fusarium (F.) oxysporum^2,5 and F. solani,⁵ was rare. To the authors’ knowledge, dermatophytes were isolated in 1 ulcer, which was superinfected by Trichophyton rubrum¹² (Table 2). Clinical appearance of skin ulcers superinfected by fungi does not present with any particular features. According to Simonart³, in 12 out of 24 patients in whom antimycotic treatment induced the disappearance of C. albicans, the clinical course of the ulcers was more favorable. Smith et al⁷ treated 17 patients with nystatin ointment, which was applied once per week before the application of compressive bandages for a total of 1 month. Yeasts were eradicated in all cases, but clinical response was not closely related to the disappearance of the yeasts.      According to these authors, topical nystatin did not have any obvious clinical effect on overall healing rates. These results were confirmed by Dereume,⁹ who stated that the disappearance of yeasts after topical treatment did not consistently hasten healing. Conversely, Hansson et al¹⁰ observed that the treatment of 6 patients with topical clotrimazole resulted in pain relief and more rapid healing. As previously mentioned, in the present study none of the patients showed clinical improvement in their ulcers with antimycotic treatment. Particularly, no changes were observed with regard to erythema and edema of periulcerative tissue. Additionally, no changes were observed in the wound borders, exudate features, wound bed, depth, or wound size.

Conclusion

     The results of this mycologic study, which is based on the largest number of patients published in the literature to date, are summarized as follows: 1) mycotic superinfections of chronic ulcers of the legs are rare: in the reported group of patients, positivity was 7.4% (11 out of 149 patients). This percentage is surprisingly low considering the high number of broad-spectrum antibiotics—both topical and systemic—that these patients often use because of bacterial superinfections. It is well known that antibiotics ease the development of fungal superinfections; 2) Mycotic superinfections were exclusively caused by yeasts. C. albicans was the most frequently involved species (7/11 patients); 3) Clinical appearance of skin ulcers superinfected by fungi is not characterized by particular features; 4) Oral antimycotic treatment did not provide clinical improvement for the ulcers with mycotic superinfection. Based on the results of this study, mycologic examinations may be considered unnecessary in patients without diabetes who suffer with chronic leg ulcers.