红色毛癣菌感染豚鼠模型的构建

发布时间：2018-06-03 03:22

本文选题：红色 + 毛癣菌　；参考：《中国协和医科大学》2008年博士论文

【摘要】： 1.鲁比切克毛癣菌,也称作红色毛癣菌鲁比切克变种,1981年由Kane首先描述为一种新的皮肤癣菌,后经证实是红色毛癣菌的变种之一。我们从一名男性股癣患者的皮损处分离并鉴定一株鲁比切克毛癣菌。方法皮屑直接镜检证实真菌感染,培养后进行菌落及镜下形态学观察,生化实验、扩增真菌rDNA的ITS区并进行序列测定的方法鉴定菌株。结果菌株在生理学和形态学上有以下几个方面特征符合鲁比切克毛癣菌的特征:(1)SDA上为绒毛状菌落,PDA上产生红色色素,菌落中央有紫色色素颗粒形成;(2)梨形或棒状小分生孢子,铅笔状大分生孢子,正常条件下培养可产生丰富的节孢子;(3)尿素酶试验阳性,毛发穿孔试验阴性;(4)扩增菌株rDNA的ITS1-4区,测序后与已知红色毛癣菌和鲁比切克毛癣菌序列进行blast比对,显示完全一致。结论该致病菌株为中国首次分离并鉴定的皮肤癣菌-鲁比切克毛癣菌。 2.通过糖皮质激素干预改变豚鼠的免疫状态,使其不能有效地清除接种的红色毛癣菌,从而建立红色毛癣菌感染豚鼠的动物模型。方法自临床标本选取3个不同的红色毛癣菌菌株,分别为短绒毛状菌落、粉末状菌落和鲁比切克毛癣菌(短绒毛状菌落,尿素酶实验阳性)。接种前3天开始,每日1次给予曲安奈德注射液20mg/kg·d股部肌肉注射预处理,接种后给予地塞米松注射液7.5mg/kg腹腔注射,后每隔日重复注射1次,共4次追加应用的方法,促进红色毛癣菌感染动物模型的建立;采用直接镜检、真菌培养和组织病理的方法验证感染结果。结果在应用适当剂量和途径的糖皮质激素的干预后,红色毛癣菌动物接种后6天开始出现直接镜检阳性,至第8天,16只红色毛癣菌感染的实验动物直接镜检均为阳性。感染后第10天,直接镜检持续阳性,毛发内亦见菌丝或孢子,真菌培养均可见与接种菌相同的菌落。组织病理显示:毛囊漏斗部见菌丝和孢子,毛根内见菌丝和孢子。结论糖皮质激素干预下红色毛癣菌可感染豚鼠建立动物模型。 3.研究不同的红色毛癣菌对动物毛发的降解作用。方法:采用体外液体培养红色毛癣菌和鲁比切克毛癣菌,加入数根小鼠或豚鼠毛发,以低浓度的酵母浸膏提供菌株生存所必需的营养,菌株生长所需的碳源和氮源由菌株分解动物毛发获得,培养2周后观察菌落的生长情况及动物毛发结构的改变,须癣毛癣菌作为阳性对照。结果:2周后,红色毛癣菌、鲁比切克毛癣菌和须癣毛癣菌在豚鼠和小鼠毛发上均能生长,菌落生长后,光镜下可见动物毛发正常结构破坏,横纹变模糊消失。结论:红色毛癣菌和须癣毛癣菌一样能够在体外分解豚鼠或小鼠的毛发作为营养,维持菌落的生长。
[Abstract]:1. Trichophyton rubychii, also known as rubesic variety, was first described as a new dermatophytes by Kane in 1981, and later proved to be one of the varieties of Trichophyton rubrum. A strain of Trichophyton rubyceticus was isolated and identified from the lesions of a male patient with tinea femoris. Methods the fungal infection was confirmed by direct microscopical examination of dander. The colony and morphology were observed after culture. The ITS region of fungal rDNA was amplified by biochemical experiments and the strains were identified by sequencing. Results the physiological and morphological characteristics of the strain were in accordance with the characteristics of Trichophyton rubychii. The SDA of Tinea rubestris was a fluffy colony PDA with red pigment, and purple pigment granules formed in the center of the colony to form a Pyriform or rod-shaped microconidium. The pencil-shaped macroconidium, cultured under normal conditions, produced abundant sarcosporum 3) urease positive, hair perforation test negative nil 4) to amplify the ITS1-4 region of the strain rDNA. The sequence was compared with the known sequences of Trichophyton rubrum and Trichophyton rubyceticus by blast. Conclusion this strain is the first dermatophytes isolated and identified in China. 2. The immune state of guinea pigs was changed by glucocorticoid intervention, which could not effectively eliminate the inoculated Trichophyton rubrum, thus establishing the animal model of guinea pigs infected with Trichophyton rubrum. Methods three different strains of Trichophyton rubrum were selected from clinical specimens. They were short villous colony, powdery colony and Trichophyton rubyceticus (short villous colony, positive for urease test). Three days before inoculation, triamcinolone acetonide injection 20mg/kg d was injected intramuscularly once a day. After inoculation, dexamethasone injection 7.5mg/kg was injected intraperitoneally and repeated every other day for 4 times. To promote the establishment of animal model of Trichophyton rubrum infection, direct microscopic examination, fungal culture and histopathology were used to verify the results of infection. Results after the appropriate dose and route of glucocorticoid intervention, the direct microscopic examination of Trichophyton rubrum began to appear 6 days after inoculation, and 16 experimental animals infected by Trichophyton rubrum were all positive under direct microscope on the 8th day. On the 10th day after infection, direct microscopic examination continued to be positive, mycelium or spores were also found in hair, and the same colony was found in fungal culture. Histopathology showed hypha and spores in the funnel of hair follicles and hyphae and spores in hairy roots. Conclusion the animal model of guinea pigs infected with Trichophyton rubrum can be established by glucocorticoid intervention. 3. To study the degradation effect of different Trichophyton rubrum on animal hair. Methods: Trichophyton rubrum and Trichophyton rubychii were cultured in liquid medium in vitro, and several mouse or guinea pig hairs were added to provide the necessary nutrition for the survival of the strain with a low concentration of yeast extract. The carbon and nitrogen sources needed for the growth of the strain were obtained from the decomposition of animal hair by the strain. After 2 weeks of culture, the growth of the colony and the changes of the hair structure of the animal were observed. Results after 2 weeks, Trichophyton rubestris, Trichophyton rubychii and Trichophyton tubuloides could all grow in the hair of guinea pigs and mice. After colony growth, the normal structure of animal hair was destroyed and the cross striation disappeared under light microscope. Conclusion: both Trichophyton rubrum and Trichophyton rubrum can decompose guinea pig or mouse hair as nutrition and maintain colony growth in vitro.
【学位授予单位】：中国协和医科大学
【学位级别】：博士
【学位授予年份】：2008
【分类号】：R-332;R756

【引证文献】