荧光原位杂交观察口腔原位菌斑中链球菌和具核梭杆菌的动态变化的研究

摘要] 目的 观察口腔原位菌斑形成过程中链球菌和具核梭杆菌的动态变化。方法 专业洁治后,7个志愿者佩戴可形成口内菌斑生物膜以供原位菌斑研究的上颌装置,分别佩戴12 h、1 d、2 d、3 d、5 d后,取下样本,与链球菌和具核梭杆菌的特异性寡核苷酸探针进行荧光原位杂交,于激光共聚焦显微镜下观察菌斑生物膜的形成过程及2种细菌在菌斑形成发育中的分布及比例。结果菌斑生物膜厚度由5.26μm逐渐发育增加为46.93μm。12 h细菌仅形成平铺片状,链球菌占绝大多数,未见具核梭杆菌;此后细菌排列趋于密集,菌斑厚度逐渐增加,最后形成团块状菌斑。链球菌在2、3 d菌斑中比例下降;具核梭杆菌在5 d菌斑中比例比3 d上升107%。结论 链球菌在早期菌斑中为优势菌,具核梭杆菌在菌斑发育中期数量逐渐增加。2种细菌在菌斑形成中起到重要作用。

In vivo dynamics of Streptococcus spp.and Fusobacterium nucleatum in dental plaque biofilm analysed by fluorescence in situ hybridization

Objective To quantify formation and composition of Streptococcus spp.and Fusobacterium nucleatum in dental plaque using fluorescence in situ hybridization（FISH） and confocal laser scanning microscopy.Methods After professional toothcleaning,seven volunteers had to wear a special acrylic appliance,in which six specimens were inserted to allow the build-up of intra-oral biofilms.After （12 h/1 d/2 d/3 d/5 d） wearing,the specimens with the adhering biofilms were removed from the splints and stained with three oligonucleotide probes.Under the confocal laser scanning microscope,biofilm thickness（BT） was evaluated.Formation and composition of Streptococcus spp.and Fusobacterium nucleatum in dental plaque were examined the biofilms were scanned and digital images were made.Results The thickness of formed biofilm increased from 5.26μm after 12 hours after wearing to 46.93μm 5 days after wearing.Streptococcus spp.were predominant in 1-day-old dental plaque and decreased 2 days later（P0.05）.Compared to the first day,Fusobacterium nucleatum increased significantly 5 days later（P0.01）.Conclusions Streptococcus spp.were predominant in early dental plaque biofilm and Fusobacterium nucleatum increased gradually later.These two bacterials were important in dental plaque biofilm formation.