人生殖道源乳酸杆菌的筛选及在小鼠阴道假丝酵母病模型中的应用评价

Lactobacillus: screening from human reproductive tract and therapeutic effect on a mouse model of vulvovaginal candidiasis

摘要:

【目的】分离筛选人阴道环境中具有益生特性的乳酸杆菌，探索外阴阴道假丝酵母病的益生菌疗法。【方法】利用含1%碳酸钙的de Man，Rogosa and Sharpe (MRS)培养基从无症状育龄女性阴道分泌物中分离乳酸杆菌，采用共培养方法评价其对白色念珠菌(Candida albicans)的抑制作用，通过对乳酸杆菌的耐酸性能、体外聚集特性和黏附能力测试考察其益生特性，并进行乳酸杆菌株功能化组合。通过构建小鼠外阴阴道假丝酵母病模型，初步探索乳酸杆菌株组合对C. albicans的抑制作用。【结果】从53个样品中分离得到19株乳酸杆菌，筛选获得4株乳酸杆菌(Lactobacillus crispatus ZH08、L. fermentum ZH09、L. fermentum ZH11和L. crispatus ZH17)具有较强抑制 C. albicans生长的能力。4株乳酸杆菌均能耐受低pH环境，能快速降低培养液pH。其中2株L. fermentum具有更强的抑制活性，能在24 h内快速抑制C. albicans生长，抑制率可达到95%以上；另2株L. crispatus具有更强的聚集特性和对上皮细胞有较强黏附性。乳酸杆菌ZH17和ZH11组合应用小鼠阴道假丝酵母病模型治疗，能显著抑制C. albicans生长和菌丝相转化，促进粘膜修复和缓解炎症。【结论】本研究筛选的乳酸杆菌具有生殖道益生菌的特性，乳酸杆菌组合应用具有潜在的临床前景。

Abstract:

[Objective] To isolate probiotic Lactobacillus strains from the human reproductive tract and explore the therapeutic effect of the strains on vulvovaginal candidiasis. [Methods] We used the de Man, Rogosa and Sharpe agar containing 1% calcium carbonate to isolate the Lactobacillus strains from the vaginal secretions of asymptomatic women in the childbearing age, screened the strains by using the co-culture method with Candida albicans, and then examined their physiological properties. The inhibitory effect of Lactobacillus was further evaluated in a mouse model of vulvovaginal candidiasis. [Results] Nineteen strains of Lactobacillus sp. were isolated from 53 samples. Among of them, 4 strains, i.e., L. crispatus ZH08, L. fermentum ZH09, L. fermentum ZH11, and L. crispatus ZH17, showed strong inhibitory effect against C. albicans, and grew well in a low-pH environment. L. fermentum ZH09 and ZH11 inhibited the growth of C. albicans, with the inhibition effect over 95% within 24 h. L. crispatus ZH08 and ZH17 had strong aggregation property and demonstrated strong adhesion to epithelial cells. Furthermore, the combination of L. fermentum ZH11 and L. crispatus ZH17 significantly inhibited the growth and the switch from budding to hyphal growth of C. albicans, promoted mucosal repair, and relieved inflammation in a mouse model of vulvovaginal candidiasis. [Conclusion] The screened Lactobacillus strains have probiotic properties and the potential in clinical application.