红色毛癣菌表型稳定性研究

为探讨红色毛癣菌表型的稳定性,所有菌株采用沙堡琼脂试管培养基(SDA)培养。菌株鉴定及分型依据菌落的特征及色素,大小分生孢子的形态,尿素酶试验和毛发穿孔试验。对近期临床分离的10株红色毛癣菌和1株标准株,间隔4周传代1次,共传4次。207株红色毛癣菌共分离出4个表型,其中绒毛型占首位(45.4%),沟纹型(24.1%),羊毛型(20.8%),粉末型(9.7%),未见颗粒型。保存1年后207株有54株发生形态变异,变异率为26.1%,沟纹型变异最小,相对稳定。11株红色毛癣菌传代后菌落形态或色素发生变异。红色毛癣菌表型不稳定,保存和传代后的菌落形态或产色均易发生变异;沟纹型变异率最低。     

红色毛癣菌基因型稳定性的研究

目的探讨红色毛癣菌基因型的稳定性。方法采用随机引物PCR法和探针与DNA印迹杂交法。以CTAB法提取DNA。随机引物为OPAA11[5-′ACCCGACCTG-3′],印迹法以NS5和ITS4(真菌的特异性引物)为引物,以红色毛癣菌的标准株为模板,扩增出rDNA的部分18S区、ITSⅠ区、5.8S区和ITSⅡ区为探针,并用随机引物法将探针用P32标记,与EcoRⅠ酶切的基因组DNA杂交。结果①AP-PCR法:红色毛癣菌扩增的主要带型是2.2,1.7,1.3,0.9,0.7 kb,所试红色毛癣菌传代前后扩增带型均无变化。②探针与DNA印迹杂交法:原代(1代)与传代后的(2,3,4代)杂交带型一致,11株红色毛癣菌均表现为3条带,分两型(分子量分别为2.4,3.9,5.9 kb和2.4,4.4,6.5 kb)。结论红色毛癣菌基因型稳定。     

须癣毛癣菌表型的研究

目的：采用传统方法对须癣毛癣菌进行表型的分型。方法：采用沙氏琼脂试管培养基（SDA）培养。菌株鉴定及分型依据菌落的形态，菌丝及大小分生孢子的特征，尿素酶试验，葡萄糖米粉试验和体外毛发穿孔试验。结果：36株须癣毛癣菌共分为5个表型，羊毛状、紧密状和绒毛状居多，三型占72．22％，颗粒状最少。螺旋菌丝主要见于绒毛状和粉末状；羊毛和紧密状的镜下结构相似，见大量细长菌丝及少量圆形小分生孢子；颗粒状的大分生孢子最多。结论：须癣毛癣菌分为5个表型，各型镜下结构不同。     

大连地区红色毛癣菌表型与皮肤癣菌病相关性分析

红色毛癣菌从形态学分5型^[1],对表型及其致病性的研究报道不多。本科2002年1月至2003年10月,对在我科门诊就诊的红色毛癣菌病临床分离培养出的274株红色毛癣菌的表型以及与临床所致疾病的关系进行了初步分析。     

红色毛癣菌的基因分型研究

目的 探讨探针与DNA印迹杂交法对红色毛癣菌的基因分型并分析其基因型与来源地区的相关性。方法 用溴化十六烷基三甲铵（CTAB）法提取DNA，以皮肤癣菌的特异性引物NS5[5′-AACTAAAGGAATTGACGGAAG-3′]与ITS4[5′-TCCTCCGCTTATTGATATGC-3′]为引物，以红色毛癣菌的标准菌株为模板，用PCR法扩增出其rDNA部分18S区，ITSI区，5.8S区和ITSⅡ区为探针，用随机引物法将探针标记^32P，用EcoRI酶切基因组DNA，采用DNA印迹的标准流程将酶切的基因组DNA与探针杂交；显示的不同带型，以此作为红色毛癣菌基因分型的依据。结果 所试49株红色毛癣菌（南京21株，大连26株，北京2株）分为20型（A-T型），其中A-C型占48.98%。南京株绝大多数为3条带，大连株大部分为4条带。结论 用ITS区域探针与DNA印迹杂交法对红色毛癣菌基因组分型敏感性强，分辨力高；南京与大连两地区红色毛癣菌DNA分型具有明显差异，DNA分型对红色毛癣菌病的流行病学，临床疗效的判定以及指导用药均具有重要价值。     

红色毛癣菌和须癣毛癣菌的PCR指纹分析

红色毛癣菌和须癣毛癣菌的传统鉴别主要依据培养形态、生理生化等方法,但红色毛癣菌的菌落产红现象受培养基类型、 pH、培养温度等因素的影响 [1],我们应用 PCR方法,以微小卫星 DNA引物,对这 2种癣菌的基因组 DNA进行扩增,可以在很短时间内把它们区分开来。 泳道 1为 Markerλ DNA/EcoRⅠ＋ HindⅢ,泳道 2、 7、 8、 9为红色毛癣菌,泳道 3～ 6为须癣毛癣菌 图 1 (GACA)4作引物的 PCR扩增结果 一、材料与方法 (一 )实验菌株：本研究采用红色毛癣菌临床分离株 23株 ,须癣毛癣菌临床分离株 11株。红色毛癣菌中 11株来自上海…     

红色毛癣菌基因型与表型的研究

目的 探讨红色毛癣菌基因型与表型、侵犯部位及其来源地区的相关性.方法 基因分型采用ITS区域探针与DNA印迹杂交法,表型分型采用传统方法.结果 所试49株红色毛癣菌(南京21株、大连26株、北京2株)分为20型(A-T型),其中A型9株均为大连株;B型9株中7株为南京株;C型6株均为南京株.5种表型除颗粒型外其他4型均有A型,17株绒毛型和7株沟纹型B型占居首位,6株羊毛型和17株粉末型A型占居首位.分离自甲癣的21株以C型为主;股癣10株和体癣6株以A型为主;足癣8株,B型占居首位;头癣4株G型占居首位.结论 红色毛癣菌基因型与表型、侵犯部位及其来源地区可能具有一定的相关性.     

传代对红色毛癣菌表型和核糖体基因影响的研究

目的通过对传代前后红色毛癣菌表型和核糖体基因的分析,探讨传代对表型变异和核糖体基因的影响。方法采用传统方法鉴定原代及传代后的5株红色毛癣菌临床菌株及1株标准菌株;对原代及20代的菌株核糖体保守区(5.8S和ITS2)和非转录间隔区(NTS)进行PCR扩增和基因测序,比较原代与传代后表型及核糖体基因有无变化。结果红色毛癣菌菌株在传代过程中发生多次表型变异,且变异可逆转。同一菌株的原代与20代PCR扩增指纹图一致,基因序列亦基本一致,其中20代标准株菌株在TRS-1区可见个别碱基的突变。结论红色毛癣菌传代过程中表型极易发生变异和逆转,而核糖体基因相对稳定。     

须癣毛癣菌基因分型的研究

目的探讨ITS区域的探针与DNA印迹杂交法对须癣毛癣菌的基因分型,并分析基因型与来源地区的相关性。方法用溴化十六烷基三甲铵(CTAB)法提取DNA,以皮肤癣菌的特异性引物NS5[5-′AACTTAAAGGAATT-GACGGAAG-3′]与ITS4[5-′TCCTCCGCTTATTGATATGC-3′]为引物,以须癣毛癣菌的标准菌株为模板,用PCR法扩增rD-NA部分18S区、ITSⅠ区、5.8 s区和ITSⅡ区为探针,随机引物法将探针标记,EcoR1酶切基因组DNA,按ECL试剂盒标准流程将酶切的基因组DNA与探针杂交;根据杂交的不同带型而分型。结果所试35株须癣毛癣菌分为14型(A~N型),A,B,C,D四型占62.86%。南方株以A和C型为主,北方株以B和D型为主。结论用ITS区域探针与DNA印迹杂交法对须癣毛癣菌基因组分型敏感性强、分辨力较高;南北方须癣毛癣菌DNA分型存在一定差异;DNA分型对须癣毛癣菌病的流行病学调查和菌学的生物学研究具有重要价值。     

红色毛癣菌随机扩增DNA多态性分型研究

红色毛癣菌(Trichophyton rubrum)是皮肤癣菌中最常见的病原菌,我国大多数地区的检出率在30%以上^[1],传统的表型分型法将其分为五型,但是,由于红色毛癣菌大多数为不典型株,而且传代过程中表型特征可发生转变或丧失,因此,表型分型的不稳定性导致它不能准确反映红色毛癣菌的遗传特征。我们应用随机扩增DNA多态性(RAPD)分析法对46株分别从广州、香港、雅加达(印尼)3个城市浅部真菌感染患者获得的红色毛癣菌进行DNA分型,对三地红色毛癣菌的表型特征与基因型别关系进行了初步探讨。     

Asthma induced by allergy to Trichophyton rubrum

The worldwide incidence of asthma and of allergic respiratory diseases is increasing (Akiyama K. 'Environmental allergens and allergic diseases.' Rinsho Byori 1997;45(1):13. D'Amato G, Liccardi G, D'Amato M. Environment and development of respiratory allergy. II. Indoors. Monaldi Arch Chest Dis 1994;49(5):412. Weeke AR. Epidemiology of allergic diseases in children. Rhinol Suppl 1992;13:5. Ulrik CS, Backer V, Hesse B, Dirksen A. Risk factors for development of asthma in children and adolescents: findings from a longitudinal population study. Respir Med 1996;90(10):623.) This has been attributed to several factors, including lifestyle changes and an expanding variety of potential causative allergens. Management of asthma entails preventive and acute medications, immunologic therapies, and removal of the identified allergen(s) from the patient's environment. Without the latter, patients may not experience full symptomatic relief. This case report describes a patient who developed bronchial asthma subsequent to an infection of tinea pedis and pedal onychomycosis; antifungal management resulted in full resolution of his tinea pedis, onychomycosis and asthma.     

HaCaT细胞对红色毛癣菌生长的抑制作用

目的 探讨角质形成细胞对红色毛癣菌生长的抑制作用.方法 将HaCaT细胞采用含10%小牛血清的DMEM培养液,5%CO2环境下,37 ℃孵育至80%融合时,弃去培养液,加入用无血清DMEM配制的菌丝段悬液共孵育(浓度0.1×105/mL～1.0×105/mL),并以菌丝段悬液作为阴性对照.分别于4 h、8 h、16 h和24 h制备共孵育后的菌丝段悬液,进行台盼蓝染色计数和菌落计数,对真菌生长状况进行评价并计算真菌生长抑制率.结果 在细胞和菌丝段共孵育4 h、8 h、16 h和24 h,台盼蓝染色法测得的菌生长抑制百分数分别为46.8%±8.0%、64.0%±5.5%、76.3%±7.7%和39.5%±4.9%;菌落计数法测得菌生长抑制百分数分别为39.7%±4.9%、58.0%±8.9%、68.3%±7.4%和39.9%±5.5%.结论 HaCaT细胞对体外培养的红色毛癣菌有一定的抑制作用.     

Molecular strain typing of Trichophyton rubrum indicates multiple strain involvement in onychomycosis

BACKGROUND: Trichophyton rubrum is an important cause of onychomycosis. Molecular strain typing methods have recently been developed to address questions of epidemiology and source of relapse following treatment. OBJECTIVES: To determine whether T. rubrum nail infections are caused by one or more strains of this fungus. METHODS: Nail specimens from 10 patients with onychomycosis due to T. rubrum were cultured and five colonies per culture plate were selected for molecular strain typing. DNA was extracted from these isolates and subjected to a polymerase chain reaction-based typing method that analyses variations in numbers of repetitive elements in the non-transcribed spacer region of the ribosomal RNA gene repeats. RESULTS: In six of 10 specimens, there were two or more T. rubrum strain types present. CONCLUSIONS: This preliminary study suggests that in many cases of fungal nail infection by T. rubrum, multiple strains are involved. This has important implications for epidemiological studies and possibly for therapy.     

Widespread and invasive Trichophyton rubrum infection mimicking Kaposi's sarcoma in a patient with AIDS

Opportunistic fungal infections are commonly encountered in AIDS patients. Candidiasis, tinea pedis, onychomycosis, and deep mycotic infections have been the fungal infections most frequently reported in these patients. Dermatophyte infections can appear to be atypical and aggressive in these patients and may lead to a misdiagnosis. We report a Trichophyton rubrum infection in a 44-year-old man with AIDS that presented as a widespread and multiple tumor-like appearance. After the patient was treated with terbinafine for 21 weeks, the lesions cleared completely. We think that this type of dermatophyte infection is very unusual in patients with AIDS and could lead to inappropriate diagnostic processes and treatments.     

多发性红色毛癣菌病患者不同部位菌株的分子鉴定

目的 用形态学、生物化学和分子生物学方法鉴定和分析从一例多发性皮肤癣菌病患者6个患病部位分离的菌种和菌株相关性。方法 6个部位的分离株先用形态学和生化方法鉴定,再用PCR扩增ITS（内转录间隔区）经序列分析证实全为红色毛癣菌。利用红色毛癣菌特异的随机扩增多态性（RAPD）和串联重复亚单位1和2（Trs-1/Trs-2）的PCR扩增产物判断6菌株基因结构的异同。结果 RAPD可将6株红色毛癣菌分为5个带型。Trs-1扩增产物可将6株菌分为3个带型,并能区分RAPD未能区分的菌株。结论 用RAPD、Trs-1/Trs-2的PCR扩增发现此多发性红色毛癣菌病患者6个感染部位的菌株均不相同,提示同一患者感染的多源性和多克隆性。     

地塞米松干预下红色毛癣菌对特比萘芬敏感性的影响

目的:探讨地塞米松干预下分离自不同部位的红色毛癣菌对特比萘芬的敏感性。方法:收集不同部位红色毛癣菌共20株,其中体癣、股癣、手足癣和甲癣各5株,采用(NCCLS)M38-A方案微量稀释法,测定不含地塞米松与含不同浓度地塞米松影响下特比萘芬对红色毛癣菌的最小抑菌浓度(MIC)。结果:当地塞米松的浓度为0.05%、0.1%、0.2%时,红色毛癣菌的MIC几何均值分别为0.081μg/mL、0.092μg/mL和0.109μg/mL,与空白组比较无显著性差异(P>0.05);当其浓度为0.4%、0.8%时,红色毛癣菌的MIC几何均值分别为0.196μg/mL、0.319μg/mL,与空白组比较有显著性差异(P<0.05)。分离自四个不同部位的红色毛癣菌MIC几何均值间无显著性差异(P>0.05)。结论:当地塞米松浓度为0.4%、0.8%时可显著降低红色毛癣菌对特比萘芬的敏感性;不同部位红色毛癣菌对特比萘芬敏感性无显著性差异。     

红色毛癣菌致股癣合并龟头癣1例

报道1例红色毛癣茵引起的股癣合并龟头癣。患者男,16岁。因双侧腹股沟及龟头红斑、鳞屑伴痒2周就诊。经真菌培养确诊为红色毛癣茵感染所致的股癣合并龟头癣,予伊曲康唑口服及联苯苄唑乳膏外用治疗2周后痊愈。     

First report on autochthonous urease-positive Trichophyton rubrum (T. raubitschekii) from South-east Europe

BACKGROUND: Trichophyton raubitschekii is a dermatophyte belonging to the T. rubrum complex and is differentiated principally by its positive urease activity and production of profuse macroconidia and microconidia in culture. It is classically isolated from African, South-east Asian and Australian aboriginal patients with tinea corporis or tinea cruris. OBJECTIVES: This study was undertaken to screen Greek and Bulgarian clinical isolates identified as T. rubrum for T. raubitschekii and to delineate these strains by two molecular methods used for the first time in T. rubrum epidemiological studies. METHODS: Ninety-five Greek and 10 Bulgarian strains, originating from various body sites, initially identified as T. rubrum, were screened for urease activity. The biochemical properties and morphology of the urease-positive strains were determined. Strains were delineated with polymerase chain reaction (PCR)-ribotyping amplifying repeat elements of the intergenic spacer region and by PCR fingerprinting. RESULTS: Five Greek and one Bulgarian T. raubitschekii strains were identified comprising isolates from patients with tinea manuum (one), tinea corporis (one), tinea cruris (one) and tinea unguium (three). Only one strain had the classical T. raubitschekii microscopic morphology, whereas the remaining five presented a dominant arthroconidial phenotype. Both typing methods clustered all T. raubitschekii and T. rubrum isolates together in the same group, indicating strain homogeneity in the genetic regions examined. CONCLUSIONS: The reported isolation of T. raubitschekii in the Balkan and South-eastern Mediterranean regions extends the geographical distribution of this species. As the more primitive T. raubitschekii probably represents the parental population of T. rubrum, the Greek and Bulgarian T. raubitschekii strains could represent a remnant of the T. rubrum spread that took place after the First World War, rather than being a recent epidemiological event.     

Epidemiological aspects of Trichophyton rubrum var. raubitschekii in Japan

Trichophyton rubrum var. raubitschekii is a rare anthropophilic dermatophyte isolated around the world from tinea corporis, tinea cruris, tinea pedis and tinea unguium. In this study, the isolation rate of T. rubrum var. raubitschekii was studied in 200 cases of tinea pedis and tinea unguium in Japan. The 200 clinical isolates were shown to be of downy type as their colonies on Sabouraud's dextrose agar were white to cream, suede‐like to downy, with a yellow‐brown to wine‐red reverse, and they produced few macroconidia. The type strain of T. rubrum var. raubitschekii (CBS 100084) and one clinical isolate (KMU 8337; isolated at Kanazawa) of downy type tested positive for urease, but the reference strain of T. rubrum (CBS 392.58) and the remaining 199 clinical isolates tested negative. Further epidemiological investigations are required to study human cases of infection with the granular type of T. rubrum and T. rubrum var. raubitschekii in Japan.