基于PCR-LIS-SSCP的白念珠菌菌丝相与酵母相ERG11基因比较研究

目的通过对白念珠菌菌丝相与酵母相ERG11基因PCR-LIS-SSCP图谱的比较,探讨两相细胞在DNA水平的差异。方法分别抽提16株来自同一亲本且对氟康唑敏感性不同的白念珠菌菌丝相与酵母相基因组DNA,根据ERG11基因序列设计7对引物,对其进行分段PCR扩增,扩增产物经单链变性后,以非变性聚丙烯酰胺凝胶进行SSCP分析。结果16株白念珠菌菌丝相与酵母相均能扩增出目的片段,SSCP图谱显示两相细胞的ERG11基因序列存在多位点差异。结论白念珠菌菌丝相与酵母相ERG11,基因存在着差异。     

白念珠菌菌丝相与酵母相ERG11基因PCR-RFLP比较分析

目的通过对白念珠菌菌丝相与酵母相ERG11基因限制性片段长度多态性分析,比较两相细胞在DNA水平的差异。方法分别抽提来自同一亲本、对氟康唑敏感性不同的白念珠菌(CA-1,CA-2,CA-4,CA-6,CA-7,CA-9,CA-11,CA-13~CA-17)菌丝相与酵母相基因组DNA,根据ERG11基因序列设计一对引物,对其进行PCR扩增(包括部分上游和下游非编码区),扩增产物分别用AccⅠ和MunⅠ消化、琼脂糖凝胶电泳后观察。结果各株白念珠菌菌丝相与酵母相均能扩增出约1 734bp大小的目的片段,CA-7,CA-14,CA-16和CA-17株两相细胞ERG11基因RFLP图谱存在差异,其余受试菌株未见差异。结论从DNA全貌来看,白念珠菌菌丝相与酵母相ERG11基因存在差异。     

白念珠菌对特比萘芬、咪康唑和制霉菌素的敏感性测定

目的测定白念珠菌的菌丝相和酵母相对特比萘芬、咪康唑和制霉菌素的敏感性。方法参照美国临床实验室标准化研究所制定的M27-A3方案,测定特比萘芬、咪康唑和制霉菌素对从临床获得的14株白念珠菌的菌丝相和酵母相MIC值。结果 (1)特比萘芬对白念珠菌的菌丝相及酵母相MIC结果差异无统计学意义,且其对白念珠菌敏感株和耐药株间MIC结果差异亦无统计学意义;(2)咪康唑对白念珠菌的菌丝相及酵母相MIC结果差异有统计学意义,后者明显较低;该药对白念珠菌敏感株和耐药株间MIC结果的差异亦有统计学意义,后者明显较高;(3)制霉菌素对白念珠菌敏感株和耐药株间MIC结果差异无统计学意义,但制霉菌素对白念珠菌菌丝相及酵母相MIC结果差异有统计学意义,后者明显较高。结论特比萘芬对白念珠菌菌丝相无抑制效果,且与氟康唑间无交叉耐药。咪康唑与氟康唑对白念珠菌存在交叉耐药,且咪康唑对白念珠菌酵母相抑制效果优于菌丝相。制霉菌素对白念珠菌菌丝相有抑制效果,且与氟康唑间无交叉耐药。     

白念珠菌菌丝相和酵母相ERG11基因部分序列差异性的探讨

目的 研究白念珠菌菌丝相和酵母相细胞ERG11基因碱基序列上的差异，探讨两相细胞之间的差异性。方法 分别抽提从同—HIV阳性患者体内分离到的7株对氟康唑敏感程度不同的白念珠菌菌丝相和酵母相的DNA，此系白念珠菌经染色体水平及DNA水平证实来源于同一亲本。根据ERG11编码序列设计一对引物，对ERG11的近3’端的310bp的碱基序列进行PCR扩增，引物序列为：上游引物5’-GGGAAAGTTTCTAAAGGGG-3’：下游引物5’-TATGrITAATCCAACTAAGTAA-3’。经PCR产物直接测序比较两相细胞ERG11基因碱基序列上的差异。结果 1株氟康唑剂量依赖性敏感和2株氟康唑耐药白念珠菌的菌丝相与酵母相细胞间均出现ERG11基因1547位点、1587位点和1617位点的不一致。结论 白念珠菌的菌丝相和酵母相ERG11基因的部分序列存在差异。     

白念珠菌ERG11基因启动子部分序列分析

目的 探讨白念珠菌菌丝相与酵母相ERG11基因启动子部分(-440～-1)碱基序列的差异以及ERG11基因启动子突变与白念珠菌对氟康唑敏感性的关系。方法 分别提取从同一亲本来源、对氟康唑敏感性不同的白念珠菌菌丝相与酵母相基因组DNA,根据ERG11基因启动子序列及其编码序列设计一对引物,对ERG11基因上游启动子部分碱基序列(-503～53)进行PCR扩增,经PCR产物直接测序比较白念珠菌菌丝相与酵母相ERG11基因启动子序列(-440～-1)的差异以及对氟康唑敏感性不同的白念珠菌ERG11基因启动子序列的差异。结果 白念珠菌菌丝相与酵母相ERG11启动子序列未发现差异。氟康唑敏感株白念珠菌ERG11基因启动子的-365,-353,-328,-310,-308,-299,-295,-293,-292,290,-289位点为杂合状态,且在-284位点有一等位基因发生单碱基缺失(-284delT);而在白念珠菌氟康唑剂量依赖性敏感和耐药株上述杂合现象消失。结论 白念珠菌菌丝相与酵母相ERG11基因启动子-440～-1区碱基序列无差异;ERG11基因启动子突变可能与白念珠菌对氟康唑耐药有关。     

白念珠菌菌丝相与酵母相ERG11基因部分序列的比较研究

目的 通过对白念珠菌菌丝相与酵母相ERG11基因部分序列的比较,探讨两相在DNA水平的差异。方法 对5株分离自同一HIV阳性患者的白念珠菌分别进行菌丝相与酵母相培养后,抽提基因组DNA,PCR扩增ERG11基因(包括部分上游和下游非编码区),用下游引物对部分ERG11基因进行测序。结果CA-7菌丝相与酵母相ERG11基因序列存在差异,分别位于第1547,1587和1617位点。结论 白念球菌菌丝相与酵母相在DNA水平存在差异,在进行白念珠菌的病原真菌学研究中菌丝相为适宜的研究对象。     

真菌病

20071738白念珠菌菌丝相与酵母相ERG11基因差异性分析/姜红浩(广州暨南大学附一院皮肤科),张宏,乔建军…∥中华皮肤科杂志.-2007,40(4).-217~220对白色念珠菌菌丝相与酵母相ERG11基因进行PCR-RELP及基因测序研究,结果表明,该菌两相ERG11无论在氟康唑敏感株(CA-7,CA-8),还是剂量     

白念珠菌菌丝相与酵母相ERG11基因差异性分析

目的 探讨白念珠菌菌丝相与酵母相ERG11基因间的差异.方法 将16株同一亲本来源、对氟康唑敏感性不同的白念珠菌(CA-1～CA-9、CA-11～CA-17)进行菌丝相与酵母相培养后,分别抽提基因组DNA,设计引物扩增ERG11基因,扩增产物分别用Acc I、Mun I、Afl Ⅱ消化,电泳后比较两相细胞RFLP图谱,同时用下游引物P₂对ERG11基因扩增产物进行反向测序.结果 Acc I、Mun I、Afl Ⅱ RFLP图谱及基因测序结果均显示菌株CA-7、CA-8、CA-14、CA-16、CA-17的菌丝相与酵母相间存在差异,两相细胞ERG11基因在1547、1587、1617三位点存在差异,位点编号按照基因库中ERG11基因序列编号(登录号X13296).结论 白念珠菌菌丝相与酵母相ERG11基因间存在差异,在进行白念珠菌体外研究时,选用其菌丝相更能反映体内真实情况.     

汉防己甲素对氟康唑抗白念珠菌活性的增效作用

目的 探讨体外汉防己甲素对氟康唑抗白念珠菌活性是否有增效作用.方法 参照微量稀释法确定汉防己甲素对白念珠菌的非细胞毒性剂量,并测定氟康唑单独及联合汉防己甲素时对16株白念珠菌的MIC.结果 在终质量浓度≤40μg/ml汉防己甲素的作用下,白念珠菌的存活率>95%.其中汉防己甲素(30～40μg/mL)可抑制白念珠菌菌丝相形成.氟康唑单独作用时,对16株白念珠菌的MIC值为0.250～64μg/mL;与汉防己甲素(40μg/mL)联合时,氟康唑对受试菌的MIC值降至0.125～16μg/mL,且终点清晰,“拖尾现象”消火.结论 汉防己甲素在体外对氟康唑的抗白念珠菌活性有增效作用.     

白念珠菌菌丝相随机扩增多态性DNA分析

目的　以白念珠菌菌丝相为研究对象 ,比较对氟康唑敏感程度不同的白念珠菌之间随机扩增多态性DNA(RAPD)图谱的差异。方法　用随机引物 5′ TCACCACGGT 3′对同一亲本来源的 16株白念珠菌菌丝相基因组DNA进行扩增 ,扩增产物进行变性聚丙烯酰胺凝胶电泳并拍照。结果　菌株CA 1～CA 13的电泳带型一致 ,均有 6条电泳带 ;CA 14～CA 17的电泳带型一致 ,均有 7条电泳带 ;CA 14～CA 17比CA 1～CA 13多 1条约 45 0bp的电泳带。 结论　氟康唑耐药株白念珠菌菌丝相基因组DNA的RAPD图谱与氟康唑敏感菌株的有差异     

Verrucous form of chilblain lupus erythematosus

A 45-year-old woman had symmetrical livid plaques with yellowish hyperkeratoses for 5 years, which progressed on to the fingers and toes and on the soles of the feet. Two years later creamy, whitish areas and maceration appeared on the buccal mucosa and the lips. A skin biopsy revealed massive collagen hyaline degeneration in the perivascular area, hyperkeratosis and hypergranulosis, small lymphocyte infiltrates with several melanophages and extravasates of erythrocytes in the upper corium in perivascular areas and hydropic degeneration of basal keratinocytes. The findings using direct immunofluorescence were compatible with lupus erythematosus (LE). Laboratory investigation showed a slight leucopenia and thrombopenia, a slightly elevated erythrocyte sedimentation rate, hypocomplementaemia C3 and C4, a high titre of rheumatoid factor and antinuclear antibodies positivity of extractable nuclear antigen. The results reflected probably the development of a systemic form of the disease. The patient was successfully managed by methylprednisolone and hydroxychloroquine. After 1 year of therapy, a new skin biopsy revealed a substantial reduction of hyperkeratosis and hyaline degeneration of collagen tissue in the perivascular areas. The combination of the extensive hyperkeratosis and hyalinization thus seems to be features of the long-lasting, untreated lesions in chilblain LE.     

A pattern of association between clinical form of vitiligo and disease-related variables in a Brazilian population

Background

Vitiligo classification systems are often based exclusively on the number and distribution of the white patches. To what extend these classification protocols reflect possible different pathophysiological basis for vitiligo or carry any prognostic value is currently unknown.

Objective

To investigate patterns of association between type of vitiligo and common disease-related variables, in order to advance on the understanding of the exact nature of different clinical forms of disease, as well as to identify features with prognostic value for clinical progression of early diagnosed vitiligo.

Materials and methods

This is a cross-sectional study of a population sample from south of Brazil composed of 586 independent vitiligo-affected individuals. Different strategies of case-control analysis were employed to test for association between the three most common vitiligo clinical types and age of onset, family history of vitiligo, occurrence of Köebner phenomenon (KP) and presence of autoimmune co-morbidity.

Results

Individuals affected by segmental vitiligo showed lower average age of onset (16 years) when compared with vulgaris (23.9 years) and acrofacial cases (29 years) (p < 0.001). The distribution of occurrence of KP, family history of vitiligo and co-occurrence of autoimmune disease followed a gradient pattern, with high, intermediate and low chance of occurrence of all three variables observed for vulgaris, acrofacial and segmental vitiligo, respectively (p < 0.001 for overall distribution).

Conclusion

Results indicate a uniform pattern of association between vitiligo clinical forms and KP, positive vitiligo family history and occurrence of co-morbidity autoimmune. The impact of the observed pattern of association over disease prognosis and classification is discussed.     

Development of atopic dermatitis-specific communication tools: Interview form and question and answer brochure

At first consultation, it is sometimes difficult for patients to decide which questions they want to ask most. We investigated whether an improvement in interview forms would identify the questions that patients want to ask doctors and help patients express their needs. First, we developed a two-part interview form specifically for atopic dermatitis (AD) patients. The first part was related to diagnosis. In the second part, we determined the most frequently asked questions by patients in daily AD clinics and included these in a prompt interview form, which we called "Questions You May Want to Ask". We compared this new interview form with the standard interview one used in our hospital. Then we made a brochure with answers to those questions. Finally, we evaluated the usefulness of these communication tools. The usefulness of the AD-specific interview form and the answer brochure was validated by patients and/or their surrogates. The majority of them recognized the necessity for and usefulness of these tools to communicate appropriately with their doctors. The answer brochure significantly increased their understanding of AD. The AD-specific interview form and the answer brochure are useful communication tools to improve doctor-patient relationships.     

Atypical clinical form of cutaneous leishmaniasis: erysipeloid form

A 60‐year‐old woman was admitted to our clinic with a gradually enlarging red papule on her face. Her history revealed that, 9 months previously, a painless, red papule of 1–2 cm in diameter had occurred in the middle of her face and, with time, had enlarged to cover her nose, both cheeks, and eyelids. It was diagnosed as a superficial skin infection, and topical and systemic antibiotics were prescribed; however, no response was obtained. In the last 2 months, a sore had formed in the middle of her nose. The patient lives in the east of Eastern Anatolia, where no case of cutaneous leishmaniasis has been reported in the last 20 years. On dermatologic examination of the patient, an erythematous, indurated, slightly squamous, 10 cm × 5 cm, butterfly‐shaped plaque with sharply defined borders was seen on both cheeks, lower eyelids, and the whole nose ( Fig. 1 ). In smears taken from the lesion, a number of amastigotes belonging to Leishmania were determined in the intracellular and extracellular area ( Fig. 2 ). Histopathologic examination of the cutaneous lesion showed scattered infiltration composed of mononuclear cells, histiocytes, plasma cells, and small epithelioid granulomas. Gram smear and anaerobe–aerobe culture prepared from the lesion were negative. The total blood count and sedimentation rate of the patient were within normal limits. Routine biochemical tests, urine analysis, chest radiography, and intradermal purified protein derivative (PPD) skin test were all normal. Antinuclear antibody and antistreptolysin antibody examinations were negative.

Figure 1 Open in figure viewer PowerPoint Butterfly‐shaped infiltrated erythematous plaque on the face     

Complete form of pachydermoperiostosis

Pachydermoperiostosis (PDP) or primary hypertrophic osteoarthropathy (PHO) is a rare hereditary disease characterized by digital clubbing, pachydermia, and periostosis. Its pathogenesis is uncertain and the diagnosis is based on clinical and radiological data. A complete form of the syndrome is reported in a male patient with disease onset in adolescence, with compatible clinical and radiological findings, presenting the three cardinal findings as well as other associated manifestations, such as hyperhidrosis and acne.