Wu Li-jie, Wang Ben, Liao Qin-ping, Zhang Rui
Department of Obstetrics and Gynecology, Peking University First Hospital, Beijing 100034, China; Department of Obstetrics and Gynecology,China-Japan Friendship Hospital, Beijing 100029, China.
Department of Obstetrics and Gynecology,Beijing Tsinghua Changgung Hospital, Beijing 102218, China.
Beijing Da Xue Xue Bao Yi Xue Ban. 2015 Dec 18;47(6):933-8.
To investigate the female vaginal Lactobacillus crispatus biofilm by using confocal laser scanning microscopy (CLSM),thus revealing the formation of biofilm.
The cover slide biofilm culture approach in vitro was employed for induction of the vaginal Lactobacillus crispatus biofilm formation. Following the culture for 2, 4, 8, 12, 16, 20, 24, 48, 72, 96 and 120 hours, the cover slide was removed for subsequent staining with the fluoresce in isothiocyanate-conjugated concanavalin A(FITC-ConA) and propidium (PI).This was followed by determination of the formation and characteristics of the vaginal Lactobacillus crispatus biofilm by using CLSM.
The CLSM images of biofilm formation at different time points were captured, suggesting that the vaginal Lactobacillus crispatus adhesion occurred at h 4, which was in reversible attachment, then more and more Lactobacillus crispatus aggregated at h 8 to h 20, which was in irreversible attachment.Lactobacillus crispatus clustered at h 20, with early development of biofilm architecture.Then the biofilm with extracellular matrix around the bacteria was set up at h 24,with gradual matureation at h 24 to h 48.The biofilm dispersed at h 72. The biofilm density of cultivating for 20 hours was 42.7 × 10⁻³ ± 6.8 × 10⁻³ ,and for 24 hours increased to 102.5 × 10⁻³ ± 23.1 × 10⁻³, suggesting a significant difference, P<0.05. This meant that mature biofilm was formed at h 24.
The vaginal Lactobacillus crispatus is able to form typical biofilm with distinct developmental phases and architecture characteristics.Mature biofilm is formed at h 24 to h 48, then the biofilm begins to disperse.
运用共聚焦激光扫描显微镜（CLSM）研究卷曲乳杆菌生物膜，以揭示生物膜的形成过程。
采用体外盖玻片生物膜培养法诱导阴道卷曲乳杆菌生物膜形成。培养2、4、8、12、16、20、24、48、72、96和120小时后，取出盖玻片，随后用异硫氰酸荧光素偶联的伴刀豆球蛋白A（FITC-ConA）和碘化丙啶（PI）进行染色。接着使用CLSM测定阴道卷曲乳杆菌生物膜的形成及特征。
获取了不同时间点生物膜形成的CLSM图像，表明阴道卷曲乳杆菌在4小时时发生黏附，处于可逆性附着阶段，随后在8至20小时有越来越多的卷曲乳杆菌聚集，处于不可逆性附着阶段。卷曲乳杆菌在20小时时聚集，生物膜结构开始初步形成。然后在24小时时形成了围绕细菌的带有细胞外基质的生物膜，并在24至48小时逐渐成熟。生物膜在72小时时分散。培养20小时时生物膜密度为42.7×10⁻³±6.8×10⁻³，24小时时增加至102.5×10⁻³±23.1×10⁻³，差异有统计学意义，P<0.05。这意味着在24小时时形成了成熟生物膜。
阴道卷曲乳杆菌能够形成具有明显发育阶段和结构特征的典型生物膜。成熟生物膜在24至48小时形成，随后生物膜开始分散。
