摘要
目的比较咖啡酸玉米琼脂培养基(caffeic acid cornmeal agar medium,CACA)和黑米琼脂培养基(black rice agar medium)对鸽粪中新型隐球菌的分离效果。方法用无菌竹签从鸽舍随机采取鸽粪标本,取0.6g与10ml无菌生理盐水制成悬液,然后按每个平板100μl分别接种咖啡酸玉米琼脂培养基和黑米琼脂培养基,肉眼观察菌落形态、颜色,光学显微镜下观察菌体形态,同时用特异性引物CN4和CN5扩增新型隐球菌URA基因,分别统计两种培养基中出现阳性菌株的平板数目。结果黑米琼脂培养基中新型隐球菌菌落呈棕黄色,外观湿润,状似胶汁。共分离获得23株新型隐球菌,检出率32.86%。CACA中新型隐球菌菌落较黑米琼脂培养基上的菌落小,褐色,干燥。共分离出11株新型隐球菌,检出率为15.71%。黑米琼脂培养基的检出率与CACA比较差异有统计学意义(P<0.05),且丝状真菌覆盖性生长的平板数目少于CACA,污染程度低于CACA;通过墨汁染色,在光学显微镜下观察新型隐球菌菌体呈圆形或卵圆形,外有宽厚荚膜,PCR扩增得到了目的条带产物。结论黑米琼脂培养基作为新型隐球菌的选择性培养基其分离培养效果(检出效果和检出率)优于CACA。
Objective To compare how effective cornmeal agar incorporating caffeic acid （CACA） and black rice agar are at isolating Cryltococcus neoformans from pigeon droppings. Methods Sterile bamboo sticks were used to random ly collect pigeon droppings from a pigeonry. A suspension was created from 0.6 g of pigeon droppings and 10 ml of sterile normal saline, and plates were then separately furnished with 100μl of CACA and black rice agar. The shape and color of colonies were observed and mycelial morphology was examined using an optical microscope. The URA gene was amplified using the specific primers CN4 and CN5. Th.e number of positive strains was counted in the 2 culture media. Results C. neoformans colonies in the black rice agar were tan in color and moist, like glue. Twenty three strains of C. neoforroans were isolated at a rate of 32. 86%. C. neoformans colonies in CACA were smaller in size than the colonies on the black rice agar. The colonies on CACA were brown and dry. Eleven strains of C. neoformans were isolated from the medium at a rate of 15.71%. C. neoformans was isolated from black rice agar at a significantly （P〈0.05） higher rate than from CACA. Fewer plates with CACA were covered with filamentous fungal growth and plates with CACA had less contamination. India ink staining revealed round or oval cells and a thick capsule when C. neoformans was observed under an optical microscope, and PCR amplification yielded products in target bands. Conclusion Black rice agar was better as a selective medium than CACA.
出处
《中国病原生物学杂志》
CSCD
北大核心
2014年第6期492-495,共4页
Journal of Pathogen Biology
基金
国家自然科学基金项目(No.31060019
31100093)
云南省教育厅重大专项资助项目(No.ZD2010010)
云南省中青年学术带头人后备人才资助项目(No.2012HB-042)
云南省高校滇西生物多样性科技创新团队基金项目(云教函〔2011〕93号)
大理学院第一批专业综合改革生物科学专业综合改革建设项目(TDⅠ-05)
