1株致体股癣的颗粒型红色毛癣菌变种的形态学观察与基因鉴定

目的：探讨1株致体股癣的颗粒型红色毛癣菌变种的形态学特点及核糖体基因序列变化。方法1例体股癣患者皮损中分离出1株红色毛癣菌,观察形态学特点,并用PCR测定核糖体转录间区（ITS）、D1/D2区、非转录区基因序列（NTS区）内串联重复亚单位（TRS）。结果该菌株可产生鹿角形菌丝,大分生孢子有中空现象及单侧出芽产孢特征,可在42℃生长;ITS和D1/D2区基因测序鉴定为红色毛癣菌,但TRS-1和TRS-2基因序列分别有2个、1个碱基差异。结论该菌株可能是颗粒型红色毛癣菌的1个变种。     

我国代表地区须癣毛癣菌复合体的分子鉴定与分型研究

目的对我国代表地区的须癣毛癣菌菌株进行分子再鉴定和分型研究。方法选取我国南北方8个省市地区经表型鉴定的须癣毛癣菌菌株47株,通过再培养形态观察、生理试验;PCR扩增核糖体DNA(rDNA)的内转录间隔区(ITS)和核糖体大亚基(LSU)D1-D2区,测序后利用数据库进行序列比对,对须癣毛癣菌复合体进行再鉴定;PCR扩增rDNA非转录间隔区(NTS)的三个串联重复亚单位S0、S1和S2区,进行种内分型,并比较不同部位来源菌株型别的差异性。结果我国南北方8个省市地区47株须癣毛癣菌中3株鉴定为断发毛癣菌,6株鉴定为无性型苯海姆节皮菌,其余均鉴定为万博节皮菌中的亲人型趾间毛癣菌;三对不同引物扩增38株趾间型毛癣菌和2株苯海姆节皮菌NTS区,共产生28种特征性带型。带型和菌株来源及发生部位无相关性。结论我国分离自人类须癣毛癣菌复合体的主要组成菌种为趾间毛癣菌;ITS区结合LSU D1-D2区测序有助于鉴定须癣毛癣菌复合体至种水平;NTS区的三个串联重复亚单位所产生的特征性指纹图提供了一种快速、稳定的分子生物学种内分型方法,可应用于趾间毛癣菌感染的流行病学研究。     

红色毛癣菌菌株的特异性PCR鉴定方法研究

目的建立一种快速的红色毛癣菌分子生物学鉴定方法。方法根据红色毛癣菌保守区域-真菌核糖体DNA（rDNA）的转录间隔区（ITS）设计特异性引物,采用上游：ITS19865＇GAC ACC AAG AAA AAA TTC TCT GAA GA3＇,下游：ITS24415＇GTC CTG AGG GCG CTG AA3＇为引物对45株红色毛癣菌、5株须癣毛癣菌和1株紫色毛癣菌菌株的DNA进行PCR扩增,观察产物电泳带型的差异。结果 45株红色毛癣菌均能扩增出目的片段,5株须癣毛癣菌和1株紫色毛癣菌均无目的片段扩增出。结论红色毛癣菌可用特异引物PCR方法快速鉴定。     

须癣毛癣菌的形态学及分子生物学鉴定

目的对临床上分离自人(36株)及狐狸(5株)的须癣毛癣菌菌株进行新的分类系统鉴定,并检测传统的分类方法是否能满足临床鉴定需要。方法①观察原鉴定为须癣毛癣菌菌株在沙氏培养基、1%蛋白胨培养基、溴甲酚紫乳固体葡萄糖琼脂培养基(BCP-MSG)和显微镜下形态学及尿素酶、毛发穿孔等生理学试验表现。②通过ITS区段和LSU区段分子生物学序列分析进行新的分类系统的菌种分型,并对传统形态学和生理学鉴定方法进行检测。结果①41株须癣毛癣菌形态学及生理学试验符合须癣毛癣菌(38株)和红色毛癣菌(3株)菌落的特点。②ITS区段序列分析发现ITS区段能将须癣毛癣菌和红色毛癣菌准确的鉴定到种,但无法明确其种内分型;而LSU区段序列分析可对36株(36/38)须癣毛癣菌有性型做出明确的鉴定。结论传统实验室鉴定方法仍具有其有效性及可靠性。通过分子生物学鉴定,临床分离的须癣毛癣菌皆为指(趾)间毛癣菌(38/38),而LSU区段的序列分析鉴定狐狸源性菌株皆属于本海姆节皮菌,有别于大多数人源性菌株有性型为万博节皮菌,对于须癣毛癣菌的菌种鉴定更优于ITS区段,但分子生物学试验还需结合形态学的观察,才能够对菌种做出正确的鉴定。     

应用RAPD技术对我国不同地域红色毛癣菌分离株的遗传多样性研究

目的探讨我国不同地域红色毛癣菌分离株的遗传多样性。方法采用随机扩增DNA多态性(RAPD)方法对来源于我国不同地域(江苏南京,山东济南,广东广州)的32株红色毛癣菌临床分离株进行DNA多态性分析。结果红色毛癣菌种内差异明显,根据遗传相似性分成三大聚类群,与地域差异及取材部位无明显相关性,而与表型具有一定相关性。结论随机扩增DNA多态性方法可用于红色毛癣菌的DNA分型,其DNA带型具有一定的遗传变异性,与菌株表型有一定关系,与地域差异、侵犯部位无明显相关性。     

皮肤癣菌体外蛋白水解酶活性测定

目的观察皮肤癣菌的体外蛋白水解酶活性;比较分离自不同感染部位的红色毛癣菌的体外蛋白水解酶活性。方法实验菌株包括来自不同感染部位的红色毛癣菌22株、须癣毛癣菌3株、犬小孢子菌5株,进行体外培养,并利用9-羟基乙酚噻唑标识的酪蛋白和酶标仪检测真菌细胞外蛋白水解酶的活性。结果须癣毛癣菌的体外蛋白水解酶活性高于红色毛癣菌和犬小孢子菌（P〈0.05）,而红色毛癣菌和犬小孢子菌之间无差异（P〉0.05）。红色毛癣菌的细胞外蛋白水解酶活性在分离自浅部感染部位的菌株之间无差异（P〉0.05）,但高于引起毛癣菌肉芽肿的菌株（P〈0.05）。结论不同的皮肤癣菌体外蛋白水解酶活性可能不同;分离自不同感染部位的同一菌种的体外蛋白水解酶活性也有可能不同。     

106株红色毛癣菌基因分型与药敏分析CSCD

目的探索海南地区的红色毛癣菌基因型与感染部位、药敏的关系。方法基于红色毛癣菌核糖体rDNA非转录区(NTS)的基因分型进行种内分型,依据“CLSI-M38-A2”方案进行药敏实验。结果106株红色毛癣菌TRS-1基因型有5种带型,其中TypeⅠ52株(49.06%),TypeⅡ14株(13.21%),TypeⅢ5株(4.72%),TypeⅣ型3株(2.83%),其他带型32株(30.18%)。TRS-2基因型有3种带型,其中TypeⅠ68株(64.15%),TypeⅡ6株(5.66%),其他带型32株(30.19%)。药敏结果MIC几何均数由低至高分别为特比萘芬(0.0092μg/mL)、伏立康唑(0.0181μg/mL)、伊曲康唑(0.1491μg/mL)、酮康唑(0.1630μg/mL)、氟康唑(2.3164μg/mL)。有42株菌表现出对抗真菌药物不敏感,不敏感菌株TRS-1和TRS-2分型均以TypeⅠ为主。结论海南地区流行的红色毛癣菌TRS-1和TRS-2基因型均以TypeⅠ为主。以对特比萘芬(0.0092μg/mL)、伏立康唑(0.0181μg/mL)MIC几何均数最低。本结果发现本地区的红色毛癣菌不敏感菌株与基因带型关系不大,也与感染部位无关,可能与来源有关。     

85株皮肤癣菌感染的病原菌分析及标准化体外药物敏感性研究

目的分析深圳地区教学医院门诊皮肤癣菌感染的病原菌特点,测定7种抗真菌药物对85株皮肤癣菌的体外抗真菌活性。方法收集疑诊为皮肤癣菌感染的患者的皮屑(或甲板刮屑、毛发)进行KOH直接镜检和培养。并通过测定皮肤癣菌临床分离菌株的核糖体DNA内转录间隔区(ITS)区序列鉴定菌种。参考CLSI M38-A2方案,测定7种抗真菌药物对85株鉴定为皮肤癣菌的临床分离菌株的体外抗真菌活性。结果 161例疑诊为皮肤癣菌感染的患者中KOH直接镜检皮肤癣菌的阳性率为67.7%,真菌培养皮肤癣菌的阳性率为52.8%。菌种鉴定结果为红色毛癣菌68例、犬小孢子菌7例、石膏样小孢子菌3例、趾间毛癣菌5例、紫色毛癣菌1例、断发毛癣菌1例。体外药敏试验显示特比萘芬(GM MIC,0.032μg·mL~(-1), MIC范围, 0.001～0.125μg·mL~(-1))对6种皮肤癣菌表现出良好的体外抗真菌活性,而伏立康唑(GM MIC, 0.041μg·mL~(-1), MIC范围, 0.032～0.125μg·mL~(-1))在唑类药物中的体外MIC值相对较低。结论本研究中皮肤癣菌感染的致病菌以红色毛癣菌为主。标准化药敏试验证实特比萘芬和伏立康唑对分离的皮肤癣菌具有较强体外抗菌活力,适用于本地区皮肤癣菌感染的治疗。     

参照CLSIM38-A方案测定皮肤癣菌临床株对4种抗真菌药物的敏感性

目的测定并比较临床分离的皮肤癣菌对4种常用抗真菌药物的敏感性,探讨CLSIM38-A方案用于皮肤癣菌药敏试验的可行性。方法实验菌株为31株临床近期分离的皮肤癣菌。其中红色毛癣菌14株,须癣毛癣菌14株,犬小孢子菌1株,铁锈色小孢子菌1株,絮状表皮癣菌1株。4种抗真菌药物为益康唑、伊曲康唑、特比萘芬、伏立康唑。采用M38-A方案微量稀释法,并适当调整试验参数进行体外抗真菌药物敏感性试验。结果红色毛癣菌和须癣毛癣菌对以上4种药物的敏感性无明显差异。犬小孢子菌、铁锈色小孢子菌、絮状表皮癣菌对所有4种抗真菌药物的敏感性都低于红色毛癣菌和须癣毛癣菌。结论M38-A方案经过调整适用于皮肤癣菌药敏试验。     

EZAL-MY96乳酸菌种菌株分离鉴定

目的：对引进法国罗地亚EZAL-MY96直投式酸奶发酵剂(DVS)进行菌株分离鉴定。方法：利用半选择性培养基进行发酵剂菌株分离,并采用糖发酵实验和RAPD指纹图谱进行鉴定。结果：EZAL-MY96中球菌的平板菌落与光镜显微观察、糖发酵实验和RAPD指纹图谱均只有1种情况,而杆菌均有2种情况。结论：从EZAL-MY96菌种中共分离到3株菌,其中1株鉴定为嗜热链球菌,1株为德氏乳杆菌保加利亚亚种,另1株为未知的乳杆菌。     

Keratinolysis by poultry farm soil fungi

Two hundred and eleven dogs (including strictly house and stray dogs) and 170 cattle in and around the city of Madras, India were screened for the presence of dermatophytosis. 106 strains of dermatophytes (89 strains from dogs and 17 strains from bovines) were isolated. 57/106 strains were Trichophyton mentagrophytes var. mentagrophytes and 42/106 strains were of the Microsporum gypseum complex. 5 strains of T. rubrum and 2 strains of T. simii were also obtained in culture. A predominance of M. gypseum complex isolates was recorded in stray dogs and cattle and T. mentagrophytes var. mentagrophytes and T. rubrum in strictly house dogs. The family history of the owners of the most of the dogs had clear records of dermatophytosis. Further, the owners of the 11 dogs that yielded T. mentagrophytes var. mentagrophytes had either tinea corporis or tinea pedis. The etiological agent of all the 11 human cases was T. mentagrophytes var. interdigitale. Similarly the owners of 4 of the 5 dogs that yielded T. rubrum were known T. rubrum patients. All these patients responded to oral griseofulvin or ketaconozole, but the recurrence of lesions was noted with the cessation of treatment. None of the patients had onychomycosis and the family history of all the patients revealed no reports of T. rubrum infections. The pet dogs were presumed to be the source of re-infection. Reversed transmission of dermatophytes from humans to animals may be the reason for the selective predominance of these organisms in strictly house dogs. They also may act as sources of reinfection. Most of the animals had small, occult, scattered lesions. These lesions may either go unnoticed or are ignored by the owners of the animals. The taxonomic status of T. mentagrophytes var. mentagrophytes and T. mentagrophytes var. interdigitale was aligned to their teleomorph Arthroderma vanbreuseghemii. Our study suggests that the periodic screening and medication of all live-stock are essential for the prevention and management of the public health problem caused by dermatophytes. This revised version was published online in August 2006 with corrections to the Cover Date.     

Dermatophytosis due to Trichophyton rubrum. Ten-year period (1996-2006) data collection in a Dermatology Department in Mexico City

Dermatophytosis is the most common mycosis in the world up to 80% caused by Trichophyton rubrum. The aim of the present study was to describe the clinical characteristics of the dermatophytosis caused by T. rubrum in a dermatological outpatient clinic during a ten years period, from 1996 to 2005. We collected the data from patients with a dermatophytosis from which we have isolated T. rubrum. A total of 776 patients with dermatophytosis caused by T. rubrum were found. A slight predominance of female patients (56.2%) was observed. The most commonly affected age group was those in the third to the fifth decade of life, and house working women (33.5%) were predominant. Onychomycosis was found in 63% of the cases, tinea pedis in 22.7%, tinea corporis in 5.2% and tinea cruris in 2.8%. In onychomycosis, the first toe nail was the most commonly affected (58.9%) and the dystrophic type was seen in 50.7% of them.     

Contribution to the study of dermatomycosis in Catalonia

We report the results of a study which aim was the mycological identification of specimens coming from patients included in a clinical trial. A total of 445 specimens from patients with clinical diagnosis of dermatomicosis were processed during 8 months (138 pityriasis versicolor, 28 cutaneous candidosis and 279 dermatophytosis). A 48% of pityriasis versicolor cultures were positive for Malassezia furfur, 50% of candidosis cultures were positive for yeasts and 67% of dermatophytosis cultures were positive for dermatophytes. According to our results Candida albicans was the principal causative agent for cutaneous candidosis and Trichophyton mentagrophytes and Trichophyton rubrum were the most frequent isolated species causing dermatophytosis.     

Dermatophytes isolated in Hospital Universitario 12 de Octubre (Madrid, Spain).

Over a 10 year period (January 1988 - December 1997), 3,241 dermatophyte strains were isolated from 18,465 specimens from patients in whom dermatophytosis was suspected clinically. This represents a 17.5% rate of isolation. Trichophyton rubrum (38.44%), Microsporum canis (28.75%), Epidermophyton floccosum (14.5%) and Trichophyton mentagrophytes (13.5%) were the dominant species, and Trichophyton tonsurans (2.09%) has emerged, whilst in the previous decade it had virtually disappeared. Our study is basically based on an out-patient selected population, and tinea corporis (30.79%), followed by tinea cruris (16.69%) and tinea unguium (16.69%) were the most prevalent clinical forms.     

Etiologic and epidemiologic study of dermatomycoses in Navarra (Spain)

We review the etiology of the dermatophytosis in Navarra (Spain) over a 5-year period and it is compared with previous studies. We have isolated 312 strains of dermatophyte fungi in 285 patients (188 men and 97 women). Trichophyton rubrum was the most frequently isolated species (58.6%) followed by Trichophyton mentagrophytes (26.2%) and Microsporum canis (10.5%). Concerning the location of the lesions, tinea pedis was the clinical pattern found in the greatest number of patients, followed by tinea corporis, tinea unguium and tinea capitis. Twenty eight percent of the isolations were accomplished in October and November. More than half of those patients questioned had had epidemiological contact with animals or practiced sports. The rise of tinea pedis in our region is emphasised. The possible causes of this increment are analyzed and some recommendations for its control are made.     

Characterization of a cDNA clone, encoding a 70 kDa heat shock protein from the dermatophyte pathogen Trichophyton rubrum

Trichophyton rubrum is an anthropophilic fungus causing up to 90% of chronic cases of dermatophytosis. To characterize T. rubrum proteins at the molecular level, we established a cDNA library of this pathogen. Here we describe a recombinant cDNA clone identical to eukaryotic 70kDa heat-shock proteins (HSPs). Western blot analysis using an anti HSP70 monoclonal antibody detected a recombinant fusion protein in Escherichia coli transformed with the expression vector containing the cloned cDNA insert. Southern blot analysis of T. rubrum genomic DNA detected no other members of the HSP70 gene family. Further analysis revealed the presence of two introns within the ORF of the HSP70 gene. In Northern blot analysis, the cDNA clone was hybridized to a RNA species of about 3.5kb which was constitutively expressed by cells cultured at 27 degrees C and was strongly up-regulated after culture at 37 degrees C. In summary, we have cloned the first member of the HSP family of dermatophytes and characterized it as a member of the Dnak subfamily of 70kDa HSPs.     

15例人畜共患体癣的临床分析

目的通过对生活在同一家庭患有体癣的患者、家庭成员和所养宠物进行观察和分析,探讨人畜共患癣病的常见致病菌种以及治疗方案。方法收集2005年3月～2009年1月就诊于我院因接触宠物而致病的体癣患者,对其宠物的带菌情况进行调查,确诊后给予外用特比萘芬乳膏或联苯苄唑凝胶,未愈者给予口服伊曲康唑治疗。结果共收集15例患者,真菌培养结果显示8例为石膏样小孢子菌,4例为须癣毛癣菌,3例为犬小孢子菌。患者皮损真菌培养结果和所养宠物皮损真菌培养结果一致。按上述方法对患者及其宠物进行治疗,所有患者和10只(3只被丢弃)宠物全部治愈。结论 15例人畜共患体癣中,石膏样小孢子菌感染所致的比例最高,其次是须癣毛癣菌和犬小孢子菌,外用特比萘芬乳膏或联苯苄唑凝胶以及口服伊曲康唑胶囊可以有效治疗此类人畜共患皮肤癣菌病。     

Stimulation, inhibition and death of macrophages infected with Trichophyton rubrum

Trichophyton rubrum is the most common pathogen causing dermatophytosis, accounting for approximately 80% of the reported cases of onychomycosis. Since 90% of the chronic dermatophyte infections are caused by T. rubrum, it is likely that this pathogen must have evolved mechanisms that evade or suppress cell-mediated immunity. Several reports have highlighted the participation of phagocytes in the immune defense against fungi; however, few studies have addressed the role of these cells in dermatophytosis. In this study, we investigated the interactions of resident and peritoneal macrophages with T. rubrum. We show here that the interaction of T. rubrum conidia with resident macrophages results in the production of TNF-alpha and IL-10 but not IL-12 and nitric oxide. Infected macrophages down-regulated the expression of co-stimulatory molecules (CD80 and CD54). We also show that phagocytosis of T. rubrum conidia is inhibited by the addition of fungal exoantigens or mannan. Cytotoxicity assays indicated that after 8 h of conidia ingestion macrophage viability decreased drastically. Electron microscopy revealed that the ingested conidia grow and differentiate into hyphae inside macrophages leading to rupture of the macrophage membrane.     

Protein profiles of Malassezia pachydermatis isolated from dogs

Chronic dermatophytosis was observed in 2276 (10.02%) of 22 692 patients with dermatophytosis during a period of 5.5 years. Males were affected at least 3 times more frequently than females. The age group most commonly affected was between 20 and 40 years of age. Females were affected more between the ages of 30 to 40 years. Tinea cruris and tinea corporis were the most common clinical types and tinea pedis was the least common type observed. The most frequent isolate was Trichophyton rubrum followed by T. mentagrophytes and T. violaceum. Ichthyosis vulgaris was the most common cutaneous association whereas atopy and diabetes mellitus were the most common systemic associations. This revised version was published online in August 2006 with corrections to the Cover Date.