The fungi Malassezia are a constituent of the skin microbiota in humans and some animals. The fungi

may cause skin diseases or even organ and/or generalized infections in the presence of appropriate predisposing

factors. To evaluate the occurrence of Malassezia fungi on the skin in patients with psoriasis and search for a relationship between the occurrence of Malassezia and the severity of skin lesions, age and sex of the patients. The materials comprised smears sampled from four sites: scalp, face, chest and back.

Malassezia spp. were isolated in cultures on modified Dixon medium and identified on the basis of morphological

and biochemical features. Malassezia spp. were isolated in 55.1% of the patients. The fungi were most prevalent on the back (33.3%)

and least on the scalp (17.5%) and face (19%). The prevalence on the chest was 30.2%. In the group of patients with

AD localized to the head and neck Malassezia spp. were cultured in 71.4% of patients. No statistically significant differences were found between Malassezia prevalence in males vs. females.

No relationship with the patients' age was found. Higher values of the atopic dermatitis severity index (SCORAD)

were found in the patients in whom Malassezia spp. were isolated. Malassezia spp. were particularly common in

the group of patients with AD localized to the head and neck

Krzyściak, Paweł

Macura, Anna

Rup, Elżbieta

Skóra, Magdalena

Advances in Dermatology and Allergology

English

The fungi Malassezia are a constituent of the skin microbiota in humans and some animals. The fungi
may cause skin diseases or even organ and/or generalized infections in the presence of appropriate predisposing
factors. To evaluate the occurrence of Malassezia fungi on the skin in patients with psoriasis and search for a relationship between the occurrence of Malassezia and the severity of skin lesions, age and sex of the patients. The materials comprised smears sampled from four sites: scalp, face, chest and back.
Malassezia spp. were isolated in cultures on modified Dixon medium and identified on the basis of morphological
and biochemical features. Malassezia spp. were isolated in 55.1% of the patients. The fungi were most prevalent on the back (33.3%)
and least on the scalp (17.5%) and face (19%). The prevalence on the chest was 30.2%. In the group of patients with
AD localized to the head and neck Malassezia spp. were cultured in 71.4% of patients. No statistically significant differences were found between Malassezia prevalence in males vs. females.
No relationship with the patients' age was found. Higher values of the atopic dermatitis severity index (SCORAD)
were found in the patients in whom Malassezia spp. were isolated. Malassezia spp. were particularly common in
the group of patients with AD localized to the head and neck

Jagiellonian Univeristy Repository

Postępy Dermatologii i Alergologii XXVIII; 2011/3 187Address for correspondence: Prof. Anna B. Macura MD, PhD, Department of Mycology, Chair of Microbiology, Jagiellonian University, Collegium Medicum, 18 Czysta, 31-121 Krakow, Poland, tel. +48 12 633 08 77, ext. 231, e-mail: mbmacura@cyf-kr.edu.plDistribution of Malassezia species in patients with atopic dermatitis – quality assessmentElżbieta Rup1, Magdalena Skóra2, Paweł Krzyściak2, Anna B. Macura21Department of Dermatology, Jagiellonian University, Collegium Medicum, Krakow, PolandHead: Prof. Anna Wojas-Pelc MD, PhD2Department of Mycology, Chair of Microbiology, Jagiellonian University, Collegium Medicum, Krakow, Poland Head: Prof. Anna B. Macura MD, PhDPost Dermatol Alergol 2011; XXVIII, 3: 187–190Original paperAbstractIntroduction: The fungi Malassezia are a constituent of the skin microbiota in humans and some animals. The fungimay cause skin diseases or even organ and/or generalized infections in the presence of appropriate predisposingfactors.Aim: To evaluate the occurrence of Malassezia fungi on the skin in patients with psoriasis and search for a rela-tionship between the occurrence of Malassezia and the severity of skin lesions, age and sex of the patients.Material and methods: The materials comprised smears sampled from four sites: scalp, face, chest and back.Malassezia spp. were isolated in cultures on modified Dixon medium and identified on the basis of morphologicaland biochemical features. Results: Malassezia spp. were isolated in 55.1% of the patients. The fungi were most prevalent on the back (33.3%)and least on the scalp (17.5%) and face (19%). The prevalence on the chest was 30.2%. In the group of patients withAD localized to the head and neck Malassezia spp. were cultured in 71.4% of patients.Conclusions: No statistically significant differences were found between Malassezia prevalence in males vs. females.No relationship with the patients’ age was found. Higher values of the atopic dermatitis severity index (SCORAD)were found in the patients in whom Malassezia spp. were isolated. Malassezia spp. were particularly common inthe group of patients with AD localized to the head and neck.Key words: Malassezia spp., atopic dermatitis, skin diseases.IntroductionThe fungi Malassezia are opportunists of significantclinical importance in humans and in animals. Malasseziaspp. may be isolated from healthy human and animal skin.These fungi are involved in certain dermatological dis-eases and, in the case of considerable immunologicalinsufficiency, they can cause systemic infections [1-6]. Cur-rently, by techniques of molecular biology, 13 Malasseziaspecies were identified, with 12 lipid-dependent species(M. furfur, M. globosa, M. obtusa, M. sympodialis, M. sloof-fiae, M. nana, M. dermatis, M. restricta, M. equina,M. japonica, M. yamatoensis and M. caprae) and one lipid-independent species (M. pachydermatis) among them [1,6-9]. Frequency of the superficial fungal infections causedby species of the Malassezia genus depends on many dif-ferent factors, such as socio-economic, climatic or geo-graphical factors [10].Atopic dermatitis (AD) is a chronic recurrent pruriticdermatosis, affecting mainly children. Atopic dermatitismay occur as an isolated disease or as a manifestation ofa systemic allergic disorder, together with hay fever,bronchial asthma or food allergies. Atopic dermatitis isalso observed in 1-3% of adults. The disease may occur forthe first time in adulthood, persist from childhood or recurafter a long period of remission [11-13]. Malassezia spp.seem to be an important factor in the pathogenesis of ADin adults, particularly when the skin lesions are localizedto the head and neck. Since the 1980s it has been observedthat the skin lesions in patients with AD disappear afterthe administration of local and systemic antifungal agents[14]. The prevalence of Malassezia spp. on the skin of adultPostępy Dermatologii i Alergologii XXVIII; 2011/3188patients with AD compared to the healthy population issimilar and is between 50% and 80%, depending on thetesting method. However, it was noticed that in patientswith AD, positive results of skin tests with Malassezia spp.allergens and high plasma levels of specific IgE antibodiesagainst Malassezia spp. are found significantly more oftenthan in the control groups [15-17]. Therefore, currently,Malassezia spp. are considered to be an allergenic ratherthan an infectious agent, in the case of patients with AD.Exacerbation of the skin lesions in patients with AD mayoccur also as a result of activation of inflammatory reac-tions by the release of hydrolytic enzymes by Malasseziaspp. [18, 19].AimThe objective of the study was to evaluate the occur-rence of Malassezia fungi on the skin in patients with pso-riasis and search for a relationship between the occur-rence of Malassezia and the severity of skin lesions, ageand sex of the patients.Material and methodsExperimental groupThe experimental group consisted of adult patientsof both sexes, who were diagnosed with atopic dermati-tis according to the criteria of Hanifin and Rajka. Clinicalcondition of the patients was assessed using SCORAD(Severity Scoring of Atopic Dermatitis). Isolation and iden-tification of Malassezia spp.Material for the mycological examination wasobtained from each patient using sterile swabs moist-ened with saline. Smears were sampled from four sitesof the body: 1) scalp, 2) face (nasolabial grooves and fore-head), 3) chest (sternal region), 4) back (interscapularregion). The material was inoculated on modified Dixonmedium, which was composed of: malt extract 36 g, pep-tone 6 g, ox bile 20 g, Tween 40 10 ml, glycerol 2 ml, oleic acid 2 ml, chloramphenicol 250 mg, agar 12 g, dis-tilled water 1000 ml. Cultures were incubated at 30°C for10 days. Fungi were identified to the level of species onthe basis of the colony morphology, microscopic appear-ance of the fungal cells obtained in the culture, by exam-ining the ability of the fungi to grow without addition oflipids and by performing the urease test on Christensen’ssolid medium (with Tween 40 and Tween 80).ResultsThe obtained data were analysed using the program-ming language and software environment for statisticalcomputing R (version 2.9.1, 2009).The study included 49 patients (29 women and 20 men) with atopic dermatitis, aged between 18 and 69.The results are presented in the Table 1 and in the Figure 1.Malassezia spp. were isolated from 27 patients (55.1%),including 16 women (55.2%) and 11 men (55%). In total,63 isolates of Malassezia spp. were obtained from differ-ent body locations (37 from women and 26 from men).No statistically significant difference was observed in thefrequency of isolation of Malassezia spp. between womenand men (χ2 test, p = 0.9905).Elżbieta Rup, Magdalena Skóra, Paweł Krzyściak, Anna B. MacuraTab. 1. Characteristics of patients included in the studySex Number Age, x ± SD Number of Malassezia spp. isolates from particular skin regions SCORAD, x ± SD (Me)Scalp Face Chest BackF 29 34.3 ±13.8 7 8 11 11 42.1 ±16.7 (42.5)M 20 31.9 ±12.2 4 4 7 10 38.8 ±16.5 (36.5)Σ 49 33.3 ±13.1 11 12 18 21 40.8 ±16.5 (38.7)F – female, M – male, x ± SD – mean ± standard deviation, Me – medianFig. 1. Comparison of the SCORAD index values in patientswith and without Malassezia skin colonization. M(–)/F(–) –male/female without Malassezia skin colonization, M(+)/F(+)– male/female with Malassezia skin colonization. The boxrepresents 1-3 quartiles, the median is marked by a hori-zontal line, the range value by whiskers, the outliers bya circle8070605040302010SCORAD valueF (–) F (+) M (–) M (+)Presence of MalasseziaPostępy Dermatologii i Alergologii XXVIII; 2011/3 189Predominantly, Malassezia spp. were isolated from theinterscapular region. Overall, 21 isolates were obtainedfrom this site (33.3% of all isolates of Malassezia spp.): 11 isolates from women and 10 isolates from men. Leastfrequently, Malassezia spp. were isolated from the scalp(11 isolates – 17.5%) and only slightly more often from theseborrhoeic regions of the face (12 isolates – 19.0%). Thistendency was seen in both women and men. However, inpatients with particularly severe skin lesions on the skinof the head and neck, in the course of AD, Malassezia spp.were isolated from these sites in 71.4% of cases. From theskin of the chest, Malassezia spp. were isolated in 30.2%of the patients (11 isolates from women and 8 isolatesfrom men). In 74.1% of the included patients in whomMalassezia spp. were found (68.7% women and 81.8%men), these fungi could be isolated from at least 2 dif-ferent sites of the body. In 18.5% of the patients (25%women and 9% men), Malassezia spp. were found in allexamined locations, at the same time. In the study, no difference in the frequency of occur-rence of Malassezia spp. depending on the age of thepatients could be seen (Student's t-test, p = 0.3656).Severity of skin lesions and subjective symptoms (pru-ritus, sleep disorder) in the course of AD expressed by theSCORAD index was between 10.2 and 80 (mean 40.8).Higher values of the SCORAD index were seen in thegroup in which Malassezia spp. could not be found (Mann-Whitney test, p = 0.004347). In patients with severe AD(SCORAD index above 40) Malassezia spp. were isolatedin 43.5% of the patients, whereas in the group with theSCORAD index below 40, the fungi were isolated from65.4% of the patients.DiscussionThe role of Malassezia spp. in AD has been studiedsince the beginning of the 1980s, when it was observedthat the skin lesions in this dermatosis disappear after theadministration of antifungal agents [14]. It was found thatfungi of the Malassezia genus, on the one hand, can bea component of the normal flora in healthy people, and onthe other hand can act as an allergen and be a direct infec-tious factor causing superficial and systemic fungal infec-tions [20, 21]. Clemmensen et al., examining patients withskin lesions on the face and neck and with concomitantpruritus, observed positive reactions in the skin prick testswith M. furfur antigen [14]. Many clinical trials confirm thepresence of specific IgE antibodies against Malassezia spp.antigens in the plasma in patients with AD [15-17]. The fre-quency of occurrence of Malassezia spp. on the skin inpatients with AD and in the healthy population is similarand varies between 50% and 80%. These differences aredue to the testing method. In our study Malassezia spp.were isolated from 55.1% of the patients with atopic der-matitis, most frequently from the interscapular region(33.3%). Similar results were obtained by Nakabayashi et al.and Yim et al. [22, 23]. In the first of the cited studies, pos-itive culture results were obtained in 54% of the samplesfrom patients with AD and in 50% of the samples fromhealthy people [22]. In the latter one, Yim et al. obtainedpositive culture results in the case of 51.7% of the patientswith AD (the 26S rDNA PCR-RFLP method was used) [23].The species most frequently isolated was M. sympodialis(16.3%). There were significant differences in the occur-rence of particular species, depending on the site of sam-pling. In samples obtained from the scalp and the face,M. restricta was isolated most often (30% and 16.7% respec-tively), whereas in samples from the skin of the chest,M. sympodialis was found most frequently (28.3%). In thestudy conducted by Nakabayashi, predominantly isolatedspecies were M. globosa (22%) and M. furfur (17%). Ma las -sezia furfur was isolated more often from the sites whereskin lesions were observed (21%) than from the unchangedskin (11%) [22]. In our study, in the group with particularlysevere skin lesions on the head and neck, positive cultureresults for Malassezia spp. were obtained in samples fromthe mentioned sites in the case of 71.4% of the patients. In the study, positive culture results for Malasseziaspp. were more frequent in the case of patients with lesssevere AD (SCORAD index below 40) compared to thepatients with severe AD. A possible reason for that couldbe a significantly decreased level of skin lipids in the lat-ter group of patients. In the quantitative studies, it wasfound that in patients with severe AD, the number ofcolonies of Malassezia spp. was lower in the case of iso-lates from skin lesions compared to isolates from theunchanged skin [24]. Another explanation for this phe-nomenon could be intensified migration of the inflam-matory cells to the skin and high concentrations ofreleased mediators having antifungal properties inpatients with severe AD [23].The results of the study suggest that Malassezia is anallergenic rather than an infectious agent in patients withAD. It is worth stressing the role of particular species ofthe Malassezia genus in patients with skin lesions in different locations. Studies on the pathogenicity ofMalassezia spp. have been conducted in the Departmentof Mycology since the 1980s [25-30]. In order to specifythe exact role of Malassezia spp. in patients with AD, appli-cation of molecular biology methods is necessary. Thecharacteristics of particular strains will be a subject offurther studies in our department.The study was done at the Department of Mycology,Chair of Microbiology, Jagiellonian University, CollegiumMedicum, Krakow.References1. Dworecka-Kaszak B, Adamski Z. Zakażenia grzybami z rodza-ju Malassezia. Wydawnictwo SGGW, Warszawa 2005; 61-114.2. Gupta AK, Batra R, Bluhm R, et al. Skin diseases associatedwith Malassezia species. J Am Acad Dermatol 2004; 51: 785-98.Distribution of Malassezia species in patients with atopic dermatitis – quality assessmentPostępy Dermatologii i Alergologii XXVIII; 2011/3190Elżbieta Rup, Magdalena Skóra, Paweł Krzyściak, Anna B. Macura3. Diaz MR, Boekhout T, Theelen B, et al. Microcoding and flowcytometry as a high-throughput fungal identification systemfor Malassezia species. J Med Microbiol 2006; 55: 1197-209.4. Batra R, Boekhout T, Guého E, et al. Malassezia Baillon, emer-ging clinical yeasts. FEMS Yeast Res 2005; 5: 1101-13.5. Ashbee HR. Recent developments in the immunology andbiology of Malassezia species. FEMS Immunol Med Micro-biol 2006; 47: 14-23.6. Cabanes FJ, Theelen B, Castellá G, Boekhout T. Two new lipid-dependent Malassezia species from domestic animals. FEMSYeast Res 2007; 7: 1064-76.7. Hirai A, Kano R, Makimura K, et al. Malassezia nana sp.,a novel lipid-dependent yeast species isolated from animals.Int J Syst Evol Microbiol 2004; 54: 623-7.8. Sugita T, Takashima M, Kodama M, et al. Description ofa new yeast species, Malassezia japonica, and its detectionin patients with atopic dermatitis and healthy subjects. J ClinMicrobiol 2003; 41: 4695-9.9. Sugita T, Tajima M, Takashima M, et al. A new yeast, Malas-sezia yamatoensis, isolated from a patient with seborrhoeicdermatitis, and its distribution in patients and healthy sub-jects. Microbiol Immunol 2004; 48: 579-83.10. Gupta AK, Boekhout T, Theelen B, et al. Identification andtyping of Malassezia species by amplified fragment lengthpolymorphism and sequence analyses of the internal trans-cribed spacer and large-subunit regions of ribosomal DNA. J Clin Microbiol 2004; 42: 4253-60.11. Katsarou A, Armenaka MC. Atopic dermatitis in older patients:particular points. JEADV 2010.12. Wanat-Krzak M, Kurzawa R. Atopowe zapalenie skóry – etio-patogeneza, diagnostyka i leczenie. Alergologia i pulmono-logia wieku dziecięcego. Klin Pediatr 2002; 10: 237-44.13. Silny W, Czarnecka-Operacz M, Gliński W, et al. Atopic der-matitis – contemporary view on pathomechanism and mana-gement. Position statement of the Polish DermatologicalSociety specialists. Post Dermatol Alergol 2010; 27: 365-83.14. Clemmensen O, Hjorth N. Treatment of dermatitis of the headand neck with ketoconazole in patients with type I sensiti-vity to Pityrosporum orbiculare. Sem Dermatol 1983; 2: 26-9.15. Back O, Scheynius A, Johansson SG. Ketoconazole in atopicdermatitis: therapeutic response is correlated with decreasein serum IgE. Arch Dermatol Res 1995; 287: 448-51.16. Savolainen J, Lintu P, Kosonen J, et al. Pityrosporum and Can-dida specific and non-specific humoral, cellular and cytoki-ne responses in atopic dermatitis patients. Clin Exp Allergy2001; 31: 125-34.17. Lindgren L, Wahlgren CF, Johansson SG, et al. Occurrence andclinical features of sensitization to Pityrosporum orbiculareand other allergens in children with atopic dermatitis. ActaDerm Venereol 1995; 75: 300-4.18. Silny W, Czarnecka-Operacz M. Atopowe zapalenie skóry –udział limfocytów T i komórek Langerhansa w rozwoju zmianskórnych. Alerg Astm Immunol 2000; 5: 15-20.19. Adamski Z. Badania nad rolą drożdżaków lipofilnych Malas-sezia furfur (Pityrosporum ovale, Pityrosporum orbiculare)w różnych dermatozach. Rozprawa habilitacyjna, AMw Poznaniu 1995.20. Borberg A, Faergemann J, Johansson SGO, et al. Pityrospo-rum ovale and atopic dermatitis in children and young adults.Acta Derm Venerol 1992; 72: 187-92. 21. Kieffer M, Bergbrant IM, Faergemann J. Immune reactions toPityrosporum ovale in adult patients with atopic dermatitisand seborrhoeic dermatitis. J Am Acad Dermatol 1990; 22:739-42.22. Nakabayashi A, Sel Y, Guillot J. Identification of Malasseziaspecies isolated from patients with seborrhoeic dermatitis,atopic dermatitis, pityriasis versicolor and normal subjects.Med Mycol 2000; 38: 337-41.23. Yim SM, Kim JY, Ko JH, et al. Molecular analysis of Malasse-zia mikroflora on the skin of the patients with atopic der-matitis. Ann Dermatol 2010; 22: 41-7.24. Schafer L, Kragballe K. Abnormalities in epidermal lipid meta-bolism in patients with atopic dermatitis. J Invest Dermatol1991; 96: 10-5.25. Budak A, Macura AB, Wnuk B, Laskownicka Z. Ocena wrażli-wości grzybów lipoilnych z rodzaju Pityrosporum na leki prze-ciwgrzybicze. Med Dośw Mikrobiol 1985; 37: 51-5. 26. Macura AB, Budak A, Wnuk B, Laskownicka Z. Występowa-nie grzybów lipofilnych z rodzaju Pityrosporum u pacjentówz łupieżem pstrym i ich chorobotwórczość dla zwierzątdoświadczalnych. Przeg Dermatol 1985; 72: 71-6.27. Basta M, Macura AB, Heczko PB. Wpływ wyciągów z lipidówskóry i kwasów tłuszczowych na lipofilny Pityrosporum sp.in vitro. Med Dośw Mikrobiol 1985; 37: 215-8.28. Macura AB, Budak A, Laskownicka Z. Recurrent Pityriasis ver-sicolor. Ann Acad Med Lodz 1985; 27: 25-9.29. Macura AB, Budak A, Wnuk B, Laskownicka Z. Występowa-nie grzybów lipofilnych z rodzaju Pityrosporumu pracowni-ków Huty im. Lenina. Post Dermatol 1986; 3: 355-60.30. Rup E, Skóra M, Krzyściak P, Macura AB. Ocena jakościowawystępowania grzybów z rodzaju Malassezia na skórzeu pacjentów z łuszczycą zwyczajną. Post Dermatol Alergol2010; 4: 264-8.

Distribution of Malassezia species in patients with atopic dermatitis : quality assessment

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Distribution of Malassezia species in patients with atopic dermatitis : quality assessment

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