Quantitative analysis of Malassezia in the scale of patients with psoriasis using a real-time polymerase chain reaction assay

BACKGROUND: It has been suggested that Malassezia is associated with the development of skin lesions in psoriasis because of the response of the scalp lesions in psoriasis to antifungal agents. Malassezia restricta and M. globosa are the two major members of the cutaneous Malassezia flora in patients with psoriasis, although they have not been analysed quantitatively. OBJECTIVES: This study quantified the two major cutaneous Malassezia species in psoriatic scale from different body sites using a real-time polymerase chain reaction (PCR) assay. METHODS: Scale samples were collected from lesional and nonlesional skin of 20 Japanese patients with psoriasis and fungal DNA was extracted from the samples directly. All the Malassezia species, including the two major species M. globosa and M. restricta, were quantified with high accuracy, using a real-time PCR assay. RESULTS: Colonization by M. restricta was approximately five times higher at all body sites than colonization by M. globosa. Malassezia colonization was significantly lower in patients with hyperlipidaemia than in patients with normolipidaemia. CONCLUSIONS: Malassezia restricta is the predominant species in psoriatic scale.     

Molecular analysis of Malassezia microflora in the lesional skin of psoriasis patients

Systemic and focal infections by microorganisms have been known to induce or exacerbate psoriasis. To investigate the role of Malassezia species in the development of psoriasis, we analyzed the Malassezia microflora in psoriasis patients using a nested polymerase chain reaction (PCR) assay, and compared it with those in atopic dermatitis (AD) patients and healthy subjects. Fungal DNA was directly collected from the lesional and non-lesional skin of the trunk of 22 psoriasis patients by applying a transparent dressing. The extracted DNA was amplified by using specific primers designed for the PCR in the intergenic spacer or internal transcribed spacer area of the ribosomal RNA. All nine of the Malassezia species were detected at different rates from the 22 psoriasis patients. The overall detection rates in lesional and non-lesional skin of M. restricta, M. globosa and M. sympodialis were high (96%, 82% and 64%, respectively), whereas the detection rates of the other species were relatively low. However, there was no difference in the rates between lesional and non-lesional skin areas. The average number of Malassezia species detected in overall sites of the psoriasis patients was 3.7 +/- 1.6 species, although this fact showed no correlation with the severity of the symptoms. The number of Malassezia species detected was 4.1 +/- 1.9 in the AD patients, and 2.8 +/- 0.8 in the healthy subjects, suggesting that the skin microflora of psoriasis patients and AD patients show greater diversity than that of healthy subjects.     

Identification of Malassezia species isolated from scalp skin of patients with psoriasis and healthy subjects

The role of Malassezia species in psoriasis is still undetermined, but several reports have associated these lipophilic yeasts with the development of skin lesions in psoriasis. The aim of our study was to analyze the prevalence of Malassezia species in the scalp lesions of patients with psoriasis and assess the distribution of the species according to patient sex, age, and duration of the disease. Forty psoriatic patients with scalp involvement and the same number of clinically healthy individuals were included in the study. The samples were obtained by scraping the skin surface of the scalp of all subjects and then incubated on modified Dixon agar. The yeasts isolated were identified by their morphological and physiological properties according to Guillot et al method. M.globosa in its yeast phase was a predominant species (55%), followed by M.slooffiae (18%) and M.restricta (10%), the latter being the most common species isolated from healthy scalp skin. We found significant difference in the distribution of Malassezia species between psoriatic and healthy scalp skin and in the distribution of Malassezia species according to the severity of the scalp involvement.     

南通和南京马拉色菌毛囊炎临床调查及致病菌种分析

目的 探讨南通和南京马拉色菌毛囊炎的易感因素及致病菌种在不同地区、不同部位马拉色菌毛囊炎中的菌种分布情况。方法 对符合病例收集纳入标准的患者进行问卷调查,取毛囊内容物进行真菌涂片、培养;并根据形态学和生理生化特征进行菌种鉴定。结果 241例临床诊断为马拉色菌毛囊炎的患者,真菌涂片204例阳性,涂片阳性率84.65%;收集标本259份,共得阳性株213株,其中马拉色菌209株,念珠菌4株（占1.54%）,真菌培养阳性率82.24%。菌种鉴定：209株马拉色菌活化菌种后,可供鉴定的马拉色菌菌株186株,共检测到6个菌种的马拉色菌,其中糠秕马拉色菌111株（59.68%）、斯洛菲马拉色菌43株（23.12%）、合轴马拉色菌17株（9.14%）、球形马拉色菌9株（4.84%）、厚皮马拉色菌4株（2.15%）、钝形马拉色菌2株（1.08%）。不同个体、不同部位的菌种分布：胸部、后背、腹部和面颈部以糠秕马拉色菌为主,上肢、肩部和头顶以斯洛菲马拉色菌为主,下肢均为球形马拉色菌。同一个体、不同发病部位存在不同的菌种,主要为糠秕马拉色菌合并合轴或斯洛菲马拉色菌。 结论 南通和南京马拉色菌毛囊炎存在6种马拉色菌致病菌种,糠秕和斯洛菲马拉色菌是主要的致病菌种。     

Molecular analysis of Malassezia microflora in seborrheic dermatitis patients: comparison with other diseases and healthy subjects

Malassezia species colonize the skin of normal and various pathological conditions including pityriasis versicolor (PV), seborrhoeic dermatitis (SD) and atopic dermatitis (AD). To elucidate the pathogenic role of Malassezia species in SD, Malassezia microflora of 31 Japanese SD patients was analyzed using a PCR-based, culture-independent method. Nested PCR assay using the primers in the rRNA gene indicated that the major Malassezia species in SD were M. globosa and M. restricta, found in 93 and 74% of the patients, respectively. The detection rate and number of each species varied similarly in SD, PV and healthy subjects (HSs), whereas AD showed higher values. Real-time PCR assay showed that the lesional skin harbored approximately three times the population of genus Malassezia found in nonlesional skin (P<0.05), and that M. restricta is a significantly more common species than M. globosa in SD (P<0.005). Genotypic analysis of the rRNA gene showed that the M. globosa and M. restricta from SD patients fell into specific clusters, and could be distinguished from those collected from HSs, but not from those colleted from AD patients. Our results indicate that certain strains of M. restricta occur in the lesional skin of SD patients.     

Cutaneous Malassezia flora in atopic dermatitis differs between adults and children

BACKGROUND: Malassezia species are suspected to be involved in the development of skin lesions in atopic dermatitis (AD) when the response of adult AD to anti-inflammatory treatments is poor. However, a comparative analysis of Malassezia flora between adults and children with AD has not been performed. OBJECTIVES: To compare the cutaneous Malassezia flora between adults and children with AD. METHODS: Scale samples were collected from skin lesions of 58 patients with AD in the head and neck regions (28 males and 30 females; 31 children and 27 adults), and fungal DNA was extracted from the samples directly. The number and identities of the Malassezia species were analysed with high accuracy using a polymerase chain reaction-based culture-independent method. The in vivo level of anti-Malassezia IgE antibody was also assayed. RESULTS: Malassezia restricta was the predominant species in the children with AD, while both M. restricta and M. globosa predominated in the adults. The adults showed increased sensitization in terms of anti-Malassezia-specific IgE responses in the sera to both M. globosa and M. restricta in comparison with the children. CONCLUSIONS: The cutaneous Malassezia flora differs significantly between the two age groups.     

从花斑癣患者的皮损区及非皮损区分离和鉴定马拉色菌

目的 研究花斑癣患者皮损区及非皮损区马拉色菌菌种构成;不同解剖部位、皮损颜色及各菌种的分布情况;患者病情和年龄与菌种构成的关系。方法 用无菌胶带粘取113例花斑癣患者皮损区及非皮损区共629个部位的皮屑,分别接种于含菜子油培养基中分离马拉色菌,用生理生化及形态学方法鉴定菌种。结果 皮损区与相对应的非皮损区马拉色菌分离阳性率无差别,非皮损区额部和胸背部分离阳性率高于上、下肢。共分离到565株马拉色菌,鉴定出合轴马拉色菌(44.78%)、糠秕马拉色菌(32.94%)、球形马拉色菌(11.68%)、钝形马拉色菌(5.84%)及限制马拉色菌(4.76%)共5个种,有27处(5.01%)同时分离到两种菌。皮损区与非皮损区菌种构成无明显差别,限制马拉色菌主要从额部分离出。菌种构成与皮损面积无关,但与皮损颜色和患者年龄有关。皮损颜色与病程无关。结论 花斑癣患者皮损区与非皮损区马拉色菌分离阳性率和菌种构成基本一致,与病情无关,而不同解剖部位、皮损类型及年龄患者的菌种构成有一定差异。     

Distribution of Malassezia species in pityriasis versicolor and seborrhoeic dermatitis in Greece. Typing of the major pityriasis versicolor isolate M. globosa

BACKGROUND: The expansion of the genus Malassezia has generated interest in the epidemiological investigation of the distribution of new species in a range of dermatoses, on which variable results have been reported from different geographical regions. No data are thus far available from South-east Europe (Greece). OBJECTIVES: To study the distribution of Malassezia species in pityriasis versicolor (PV) and seborrhoeic dermatitis (SD) and to investigate whether polymorphisms in the internal transcribed spacer (ITS) 1 region facilitate detection of M. globosa and M. sympodialis subtypes. METHODS: In total, 109 patients with PV and SD and positive Malassezia cultures were included in the study. Age, gender, primary/recurrent episode, disease extent and clinical form of PV were recorded. ITS 1 polymorphisms of M. globosa and M. sympodialis type and clinical strains were investigated by polymerase chain reaction (PCR)-single-strand conformational polymorphism (SSCP) analysis. RESULTS: Malassezia globosa was the prevalent species isolated from PV and SD either alone (77% and 39%, respectively) or in combination (13% and 18%, respectively) with other Malassezia species. The pigmented form of PV was strongly correlated with the female gender. PCR-SSCP differentiated five subgroups of M. globosa with one being associated with extensive clinical disease. All M. sympodialis isolates displayed a homogeneous ITS 1 PCR-SSCP profile. CONCLUSIONS: Malassezia species isolation rates were in agreement with those reported from South-west Europe. PCR-SSCP of the ITS 1 is useful for highlighting prospective clinical implications of M. globosa subtypes.     

豚鼠皮肤球形马拉色菌感染模型的建立

目的：建立皮肤球形马拉色菌感染的动物模型。方法：分别采用涂菌1次法、涂菌1次并皮下注射甲泼尼龙法、连续7d涂菌法3种方法诱导豚鼠皮肤感染球形马拉色菌。结果：连续7d涂菌法适合于此模型的建立,得到了满意的皮损及菌落数。结论：皮肤菌落计数能更直观、客观地反映药物的短期疗效,是评价皮肤球形马拉色菌感染模型较科学的指标。     

马拉色菌与人角质形成细胞株共同培养液对培养人黑素细胞生长及酪氨酸酶mRNA表达的影响

目的:观察不同马拉色菌与人角质形成细胞株共同培养液对培养人黑素细胞生长及酪氨酸酶mRNA表达的影响,探讨花斑癣的色素改变是否与马拉色菌的不同菌种有关。方法:将球形、钝形、糠秕、合轴马拉色菌分别与人角质形成细胞株共同培养,24 h后收集上清液,按一定比例加入黑素细胞培养体系中继续培养24 h:①用四甲基偶氮唑兰比色法观察对黑素细胞生长的影响;②用实时荧光定量PCR法测定黑素细胞酪氨酸酶mRNA的表达。结果:各共同培养液对黑素细胞生长无明显影响,但除球形马拉色菌与人角质形成细胞共同培养液可上调黑素细胞酪氨酸酶mRNA的表达(P<0.05)外,未见其它种马拉色菌与人角质形成细胞株共同培养液对黑素细胞酪氨酸酶mRNA的表达有影响。结论:球形马拉色菌与人角质形成细胞株共同培养液刺激黑素细胞后,能提高酪氨酸酶mR-NA的表达,该菌可能与花斑癣的色素沉着有关。     

Antigenic components of Malassezia species for immunoglobulin E antibodies in sera of patients with atopic dermatitis

Antigenic components of Malassezia furfur, M. globosa, M. restricta, M. slooffiae, and M. sympodialis were studied for immunoglobulin E antibodies in sera of patients with atopic dermatitis (AD). Antigenic components were extracted from Malassezia cells by treatment with beta-mercaptoethanol, referred to as 2-ME extract. CBB staining and lectin blots using Con A, LCA, PHA-E4, PNA or RCA120 showed that the 2-ME extracts contained several species-dependent components that differed quantitatively and qualitatively among the Malassezia species at the protein level. In the Western blot with the 2-ME extracts, of 54 sera of the patients with AD (54 patients), the patients' IgE antibodies most frequently recognized components showing molecular weights of 43-46 kDa for M. slooffiae, 12-22 kDa for M. sympodialis, 35-40 kDa for M. restricta, 45-50 kDa for M. globosa, and of 67-72 kDa for M. furfur, respectively. In the correlative study, in which the total band intensities generated for each extract in Western blot were compared among the Malassezia species, the intensity for M. globosa was well correlated with that for M. sympodialis (r=0.756). In the Western blot inhibition test, the 2-ME extract of M. globosa partially inhibited the reaction of the antigenic components of other Malassezia species with the patient's IgE antibodies. These results indicated that Malassezia species contained both species-specific and common antigenic components at the IgE antibody level.     

Skin colonization by Malassezia species in neonates: a prospective study and relationship with neonatal cephalic pustulosis

OBJECTIVES: To assess skin colonization by Malassezia species in full-term healthy newborns, to investigate factors associated with colonization, and to look at acnelike cephalic pustulosis associated with this carriage. DESIGN: Samples were obtained from neonates and their mothers 0 to 5 days after birth and again 3 weeks later. Clinical patterns of common acnelike pustulosis were reported as mild (<10 papulopustules), moderate (> or =10 papulopustules), or absent. Direct examination and culture of sample. Identification of yeasts was based on microscopic and physiologic criteria. SETTING: A maternity hospital and the pediatric dermatology unit of a university hospital. PARTICIPANTS: Consecutive series of 102 neonates and their mothers. MAIN OUTCOME MEASURES: Incidence of skin colonization and type of Malassezia species found in neonates and correlation with neonatal cephalic pustulosis (neonatal acne). RESULTS: At the first visit, 11 neonates and 36 mothers had cultures positive for Malassezia. Malassezia sympodialis and Malassezia globosa were preferentially cultured. At 3 weeks, 29 (52%) of 56 neonates and 18 (32%) of 56 mothers had cultures positive for only M sympodialis and M globosa. Breastfeeding was not associated with a higher prevalence of Malassezia carriage in neonates. Malassezia colonization was higher when pustulosis was more severe and M sympodialis was found in pustules. CONCLUSIONS: Malassezia colonization begins at birth and increases in the first weeks of life. A high prevalence of M sympodialis in neonates is noted from birth. Its association with neonatal acne is confirmed. Further investigation is needed to study the role of sebum secretion rate and quality in the neonatal period.     

Epidemiology of pityriasis versicolor in Adana, Turkey

Pityriasis versicolor is a common superficial mycoses of the skin. It is now recognized that the causative organisms of this infection are different species of Malassezia . The aim of this study was to determine the distribution of Malassezia species in patients with pityriasis versicolor in Adana, Turkey. In total, 97 patients positive for Malassezia elements, namely, yeast cells and short hyphae in microscopic examination, were included in the study. All samples were inoculated in plates containing modified Dixon's medium. However, only 44 of the patients (45.4%) showed Malassezia spp. in culture. Malassezia globosa (47.7%) was the most commonly isolated species followed by Malassezia furfur (36.4%) and Malassezia slooffiae (15.9%). Mixed Malassezia species were not isolated. In conclusion, M. globosa was found to be the predominant PV isolate in Adana, Turkey.     

Malassezia globosa as the causative agent of pityriasis versicolor

BACKGROUND: The taxonomic revision carried out in 1996 on the genus Malassezia, which now comprises seven different species, made necessary a re-evaluation of the data concerning the ecology and pathogenicity of these lipophilic yeasts. Very little has been published since then. OBJECTIVES: The aim of this work was to contribute to the knowledge of the aetiology of pityriasis versicolor (PV) with a mycological study made according to the new species. METHODS: Ninety-six patients with PV completed the study. In all of them, samples were taken from the lesions for direct microscopy with KOH+ Parker ink and culture. Samples were also taken from normal skin of the trunk and the forehead of the same patients for culture. Cultures were made in modified Dixon medium in Petri dishes, incubated at 31 degrees C and the isolates were identified by morphological and physiological characteristics. RESULTS: In the PV lesions, direct microscopy always showed the typical mixture of globose blastoconidia and pseudomycelium. Only one patient, having received previous topical antimycotic treatment, was negative. The spherical yeasts observed in vivo were morphologically identical to the globose yeasts characteristic of M. globosa. In culture, M. globosa was found in 97% of cases, alone in 60% of them and associated with M. sympodialis in 29% and M. slooffiae in 7%. These two species were also found in similar percentages on the clinically uninvolved skin of the trunk, whereas M. globosa was not isolated at these sites. However, on the forehead, a small number of colonies of M. globosa was recovered in 12% of cases. CONCLUSIONS: These results support the data previously reported and strongly suggest that M. globosa in its mycelial phase is the causative agent of PV.     

从夫妻双方的头皮屑中分离和鉴定马拉色菌

目的 探讨夫妻双方的头皮屑中马拉色菌的带菌情况，菌种构成及夫妻间共用梳子与马拉色菌菌种一致性的关系，方法 采用Leeming和Notman培养基，Dixon培养基和含菜子油培养基分别同时培养114对夫妻的头皮屑，观察培养阳性率与头皮屑严重程度的关系。根据生理生化学及形态学特点鉴定菌种，并比较在这3种培养基中菌落初长时间及对菌种有无选择性。结果 从114对夫妻（共228人0中153人的头皮屑中分离到马拉色菌，阳性率为67.11%。培养阳性率与头皮屑的严重程度有正相关关系，3种培养基共分离到马拉色菌，阳性率为67.11%。培养阳性率与头皮屑的严重程度有正相关关系。3种培养基共分离到459株马拉色菌，从中鉴定出合轴马拉色菌（33.33%)，球形马拉色菌（25.05%)，限制马拉色菌（15.47%)，糠秕马拉色菌（13.73%)和钝形马拉色菌（12.42%)共5个种，3种培养基对菌种无选择性，但在含菜子油的培养基中菌落初长时间最短，同一头皮屑在3种培养基上分离到同一菌种有129人（56.58%)，分离到2种菌的有24人（10.53%)，共用梳子的夫妻双方的头皮屑中菌种相同者（35.96%)显著多于菌种不同者（13.16%)；夫妻间共用梳子者头皮屑中菌种相同者（50.67%)显著多于非共用梳子者（7.69%)。结论 头皮屑中的优势菌种主要为合轴和球形马拉色菌，夫妻双方的头皮屑中马拉色菌菌种一致率较高，而夫妻间共用梳子组的菌种一致率更高，提示马拉色菌可在夫妻之间传播，共用梳子可能是重要的传播媒介。     

In vitro susceptibility of the seven Malassezia species to ketoconazole, voriconazole, itraconazole and terbinafine

Fifty-five strains, either authentic or ex-type, of seven Malassezia species were investigated for in vitro susceptibility to various concentrations (0.03-64.0 microg/mL) of three azole drugs, ketoconazole, voriconazole and itraconazole, as well as the allylamine terbinafine, using the agar dilution method. All strains of the seven Malassezia species were susceptible to the three azole drugs at low concentrations. M. furfur, M. sympodialis, M. slooffiae, M. pachydermatis, M. globosa, M. obtusa and M. restricta were most sensitive to ketoconazole and itraconazole, with minimum inhibitory concentrations (MICs) ranging from < or = 0.03 to 0.125 microg/mL. The recently introduced antifungal, voriconazole, was also very effective, with MIC80 values < or = 0.03 microg/mL for 80% of strains. MICs of terbinafine against the seven Malassezia species ranged from 相似文献   

Malassezia globosa and restricta: breakthrough understanding of the etiology and treatment of dandruff and seborrheic dermatitis through whole-genome analysis

Dandruff and seborrheic dermatitis (D/SD) share an etiology dependent upon three factors: sebum, microbial metabolism (specifically, Malassezia yeasts), and individual susceptibility. Advances in microbiological and analytical techniques permit a more detailed understanding of these etiologic factors, especially the role of Malassezia. Malassezia are lipid-dependent and demonstrate adaptation allowing them to exploit a narrow niche on sebum-rich skin. Work in our and our collaborators' laboratories has focused on understanding these adaptations by detailed analysis of biochemistry and gene expression. We have shown that Malassezia globosa and M. restricta predominate on dandruff scalp, that oleic acid alone can initiate dandruff-like desquamation, that M. globosa is the most likely initiating organism by virtue of its high lipase activity, and that an M. globosa lipase is expressed on human scalp. Considering the importance of M. globosa in D/SD (and the overall importance of commensal fungi), we have sequenced the M. globosa and M. restricta genomes. Genomic analysis indicates key adaptations to the skin environment, several of which yield important clues to the role Malassezia play in human disease. This work offers the promise of defining new treatments to D/SD that are targeted at changing the level or activities of Malassezia genes.     

Molecular analysis of Malassezia species isolated from hospitalized neonates

Abstract:  Malassezia species are a part of the skin microflora of neonates. Under certain circumstances, they can cause diseases ranging from simple pustulosis to lifethreatening fungemia in newborn infants. Little information is available about the epidemiology of Malassezia species in neonates. In the present study, we successfully isolated Malassezia yeasts from 68.7% of hospitalized neonates. Using the polymerase chain reaction-restriction fragment length polymorphism method (PCR-RFPL), M. furfur (88.06%) was identified as the most isolated species, followed in frequency by M. globosa (10.48%), M. obtusa (0.73%), and M. slooffiae (0.73%). Among the variables studied, only a longer stay in the ward resulted in a higher colonization rate. Using multiple logistic regression, only the type of hospital and ward had some effects on the colonization rate. Our results supported the hypothesis that neonates acquire Malassezia flora through direct contact with their mothers or hospital personnel.     

Effects of topical vehicles on growth of the lipophilic Malassezia species

In the present study, the abilities of major Malassezia species, M. sympodialis, M. globosa and M. furfur, to assimilate topical agents, which have been widely used as a material of ointment for skin diseases, were tested. Obvious growth of M. furfur on GYEP agar plate was noted in the presence of white petrolatum, purified white petrolatum, hydrophilic ointment and heparinoid in hydrophilic ointment, and also M. sympodialis showed similar growth when they were cultured with hydrophilic or heparinoid in hydrophilic ointment. In contrast, M. globosa did not grow on GYEP in the presence of the any topical agents tested. These results suggest that Malassezia species, especially M. furfur and M. sympodialis, assimilate several topical agents and showed the drug-depended cell growth.