Analysis of bacterial biofilms as a virulence factor for peri-implantitis

我们使用愈合基牙研究了人体种植体沟内初始生物膜的形态形成。在征得7例患者的知情同意后，分别于植入后第10、30、90、150天取出20个愈合基牙，进行扫描电子显微镜观察。第10天，基牙表面已部分形成单层生物膜。观察到了所谓的早期殖民者：球菌和细丝。30d后，生物被膜形成，在人种植体沟内主要观察到长棒和短棒。其次，对人种植体周围炎相关细菌进行了免疫组织化学检查。取4例患者的8颗种植体，制成半薄切片，用4种抗体对牙周病杆菌进行BrownBrenn改良革兰氏染色和链霉亲和素生物素染色。在种植体表面和基牙螺丝内侧均可见革兰氏阳性菌，种植体槽内未检出菌种特异性抗体标记的牙龈卟啉单胞菌和直肠弯曲杆菌。生物膜中ATP含量的增加速度很低，仅为浮游细胞的1/3.0×10~(-5)。葡萄糖酸洗必泰能有效地降低牙龈假单胞菌生物膜的存活率，而盐酸二甲胺四环素和甲硝唑的作用不明显。克隆的PPK基因与大肠杆菌PPK基因的同源性为35.8%，其中PPK基因高度保守

We investigated morphologically initial biofilm formation in the human implant sulcus using healing abutments. After obtaining informed consent from 7 patients, twenty healing abutments were removed carefully on day 10, 30, 90 and 150 after insertion and treated for a scanning electron microscopy. On day 10, mono-layer biofilm have partially formed on the abutment surfaces. So called early colonizers: cocci and filaments, were observed. After 30 days, biofilms developed, and long and short rods were predominantly observed in human implant sulcusNext, human periimplantitis associated bacteria have been examined immunohistochemically. Eight implants obtained from 4 voluntary patients were extracted and processed into semithin sections They were stained BrownBrenn modified Gram stain and streptavidinbiotin method using 4 kinds of antibodies with periodontopathic bactria. Gram positive bacteria were shown not only on the implant surface but also at the inner site of abutment screw, Porphyromonas gingivalis and Campylobacter rectus labeled with speciesspecific antibody were not found in the human implant sulcusBy use of the in vitro model with P. gingivalis biofilm, 1) the biological (ATP) activities, 2) the susceptibility to antimicrobial agents, and 3) the essential gene for development of biofilm (the homologue for polyphosphate kinase (ppk)) were examined. The rate of increase of ATP contents in the biofilm was quite low, and 1/3.0x10^5 compared with that of planktonic cells. Chlorhexidine gluconate was effective at reducing the viability of P. gingivalis biofilm, while minocycline hydrochloride and metronidazole did not have a variable effect. Identity of P. gingivalis ppk gene that was carried out cloning to Esherihia coli ppk gene was 35.8%, including \hat ppk from P. gingivalis was highly conserved