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On-line version ISSN 1806-4841
An. Bras. Dermatol. vol.86 no.4 Rio de Janeiro July/Aug. 2011
Identification of Malassezia yeast species isolated from patients with pityriasis versicolor*
Vanessa PetryI; Fernanda TanhausenII; Luciana WeissIII; Thais MilanIV; Adelina MezzariV; Magda Blessmann WeberVI
IDermatologist, Master's degree student at the Federal University of Health Sciences of Porto Alegre (UFSPA), Porto Alegre, Rio Grande do Sul, Brazil.
IIUndergraduate medical student, Federal University of Health Sciences of Porto Alegre (UFSPA), Porto Alegre, Rio Grande do Sul, Brazil
IIIDermatologist, Mae de Deus Hospital Group, Porto Alegre, Rio Grande do Sul, Brazil
IVResident Physician in Dermatology, Federal University of Health Sciences of Porto Alegre (UFSPA), Porto Alegre, Rio Grande do Sul, Brazil
VPhD., Professor at the Federal University of Health Sciences of Porto Alegre (UFSPA), and at the School of Pharmacy, Federal University of Rio Grande do Sul (UGRGS), Porto Alegre, Rio Grande do Sul
VIPhD., Adjunct Professor of Dermatology and Professor of the Dermatology Residency Program, School of Medicine, Federal University of Health Sciences of Porto Alegre (UFSPA), Porto Alegre, Rio Grande do Sul, Brazil
Pityriasis versicolor (PV) is a disease with worldwide distribution. Twelve different species of Malassezia yeast have been described. The objective of this study was to determine which species of Malassezia are more prevalent in patients with pityriasis versicolor. Samples were collected by scraping the lesions of 87 patients with a clinical suspicion of pityriasis versicolor. The samples were then submitted to fungal microscopy and culture to identify the species. The species found were: Malassezia sympodialis (30%), Malassezia furfur (25.7%), Malassezia globosa (22.7%), Malassezia restricta (12.1%), Malassezia obtusa (7.6%) and Malassezia slooffiae (1.5%).
Keywords: Dermatology; Dermatomycoses; Fungi; Mycoses.
Pityriasis versicolor (PV) is a disease with worldwide distribution. 1-3 The diagnosis of PV is based on clinical findings and confirmed by direct microscopy. 2 Culture and molecular analysis of Malassezia microflora may be used to identify different Malassezia species. 4 In 1996, Guillot et al. described a method for identifying species of Malassezia have been described; however, not all are clinically relevant in humans. 6 The objective of this study was to determine which species of Malassezia were most prevalent in the population of patients with pityriasis versicolor receiving care at the dermatology outpatient clinic of the Federal University of Health Sciences of Porto Alegre (UFCSPA).
A cross-sectional study was conducted in which patients with PV being seen at the dermatology outpatient clinic over a one-year period were invited to participate. Patients under 16 years of age, those who had used systemic or topical antifungal medication in the preceding month and patients known to have an immunosuppressive disease were excluded from the study. The study was approved by the Internal Review Board of UFCSPA under approval letter #383/07. Data analysis was performed using the SigmaStat software. Student's t-test and the chi-square test were used for continuous and categorical variables, respectively. The Mann-Whitney/Wilcoxon Two-Sample Test (Kruskal-Wallis test for two groups) was used to compare quantitative variables between two independent groups. The diagnosis of pityriasis versicolor was confirmed by clinical observation and direct microscopy. For the primary isolation of the fungi in the material collected, Sabouraud agar and Dixon's medium were used, with the addition of olive oil in both cases. After seeding, the material was incubated at 32-35ºC for 36 days. Glabrous colonies of a creamy-yellow color and a furrowed surface were observed, the reverse of the colony also being creamy-yellow in color. Following primary isolation, final identification of the species was made using the methodology outlined by Guillot et al. (1996), which is based on the phenotypic characteristics of the species: the capacity to grow in the absence of lipids, production of the enzyme catalase, description of the micromorphology and the ability to assimilate different concentrations of Tween (20, 40, 60 and 80) in Sabouraud agar containing 0.05% of chloramphenicol and 0.05% of cycloheximide. 5 The Tween assimilation test was performed using a suspension of colonies inoculated into a plate containing Sabouraud agar supplemented with 0.05% chloramphenicol and 0.05% cycloheximide. Each polysorbate (Tween 20, 40, 60 and 80) was added to fill up equidistant wells made in the inoculated agar. The plates were then incubated at 32ºC for 5-7 days.
After this period, the growth around each well, indicating assimilation of the substrate and a positive result, was observed. The set of positive results permitted differentiation between the species.
Samples were collected from 87 patients, 51 (58.6%) female and 36 (41.4%) male. The mean age of the patients was 31 ± 15 years. With respect to the site of the lesions, 90% of the patients had lesions on more than one part of the body, the most common being the lower back (53 patients) and the upper back (36 patients). Sixty-seven patients (77%) in whom the condition was clinically suspected tested positive at direct mycological examination, while 66 (75%) had a positive mycological culture. With respect to the cultures, 30% consisted of Malassezia sympodialis, 25.7% Malassezia furfur, 22.7% Malassezia globosa, 12.1% Malassezia restricta, 7.6% Malassezia obtusa and 1.5% Malassezia slooffiae. Figure 1 shows the site of lesions in patients in relation to the species of Malassezia isolated in culture. The difference between the duration of the lesions and recurrences of the disease, according to the species of Malassezia, is shown in Table 1.
The prevalence of pityriasis versicolor was greater in female patients and in young patients (mean age 31 years). These data are in agreement with reports in the literature that emphasize the higher frequency of this infection in young people due to the lipophilic characteristics of this type of fungus.2 The most prevalent species of Malassezia in this sample population was M. sympodialis followed by M. furfur and M. globosa. These findings differ from those reported in a study carried out in the state of Goias in Brazil in 2006 in which a prevalence of M. furfur of 77.8% was found. However, they are in agreement with the findings of Framil et al., who also reported M. sympodialis as being the most prevalent species. 7 Relapses are often reported by patients and cases of recurrence were found for almost all the species isolated; however, the number of relapses was not significantly different between the species of fungus. Framil et al. reported a predominance of M. sympodialis in patients who had experienced more relapses; nevertheless, this finding was not confirmed in the present study.8 The most prevalent sites of infection were the back, chest and abdomen in the case of most of the species and this finding is in agreement with the results published by Miranda et al. (2006), who also reported the back and chest as being the most common sites of the lesions. 9 No statistically significant difference was found between the species as a function of gender, age or the duration of the lesions. To the best of our knowledge, there are no reports in the literature in which statistically significant differences were found between the clinical and demographic data and the species of Malassezia. Further studies with larger sample sizes may be required to clarify this issue.
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8. Framil VMS, Melhem M, Zaitz C. Pitiríase versicolor: estudo conceitual, etiológico, imunológico e perfil de sensibilidade in vitro a derivados azólicos [Tese]. São Paulo (SP): Faculdade de Ciências Medicas da Santa Casa de São Paulo; 2006. [ Links ]
9. Miranda KC, Araújo CR, Soares AJ, Lemos JA, Souza LKH, Silva MRR. Identificação de espécies de Malassezia em pacientes com pitiríase versicolor em Goiânia-GO. Rev da Soc Bras de Med Tropical. 2006;39:582-3. [ Links ]
Mailing address: Received on 18.12.2009. * Study conducted at the Dermatology Department, Federal University of Health Sciences of Porto Alegre (UFSPA), Porto Alegre, Rio Grande do Sul, Brazil.
Rua Artur Rocha 67/301 -Auxiliadora
90450-171 Porto Alegre -RS, Brazil
Approved by the Advisory Board and accepted for publication on 14.06.2010.
Conflict of interest: None
Financial funding: None
Received on 18.12.2009.
* Study conducted at the Dermatology Department, Federal University of Health Sciences of Porto Alegre (UFSPA), Porto Alegre, Rio Grande do Sul, Brazil.