Abstract:

Objective    To compare the biofilm formation of P. gingivalis  W83 and its sialidase-deficient mutant strain（ΔPG0352） under stressful conditions. Methods    P. gingivalis  W83 and ΔPG0352 were cultured in trypticase soy broth（TSB）and supplemented with 5 μg/mL hemin and 1 μg/mL menadione under stressful conditions，including temperature，oxidatie stress and pH values. Biofilms were formed on the 96-well plates for four days，and stained with crystal violet. Detect the biofilm formation with spectrophotometer. Results    Both P. gingivalis  W83 and ΔPG0352 strains could form biofilms in normal culture conditions. P. gingivalis  W83 formed more biofilm，compared to ?PG0352 （P < 0.05）. The biofilm formations of P. gingivalis  W83 and ΔPG0352 decreased in 34℃ and 41℃ （P < 0.05）. As the concentration of H2O2 increased，biofilm formations of both strains decreased more and more seriously. The biofilm formations of ΔPG0352 were less than those of P. gingivalis  W83，when the ultimate concentration of hydrogen peroxide was 0.1，0.25 and 0.5 mmol/L. Both P. gingivalis  W83 and ?PG0352 formed less biofilm in alkaline condition（pH = 9），ΔPG0352 even less，and were unable to form biofilm in acidic condition（pH = 5）. Conclusion    Sialidase gene deletion can affect P. gingivalis  biofilm formation. Biofilms formations of both P. gingivalis  W83 and ΔPG0352 will decrease under stressful conditions. 

Key words: Porphyromonas gingivalis, sialidase, biofilm

摘要：

目的    探讨在不同培养条件下唾液酸酶基因缺失对牙龈卟啉单胞菌（Porphyromonas gingivalis，P. gingivalis ）生物膜形成的影响。方法    不同温度、过氧化氢（H2O2）浓度及pH值条件下，在96孔板中用含有5%无菌脱纤维羊血、氯化血红素（5 μg/mL）和维生素K（1 μg/mL）的TSB培养基培养P. gingivalis 野生株（W83）和唾液酸酶基因突变株（ΔPG0352）4 d，结晶紫染色、酒精脱色后，采用酶标仪检测生物膜的形成情况。结果       P. gingivalis W83和ΔPG0352在正常条件下均能形成生物膜，与P. gingivalis  W83相比，ΔPG0352形成的生物膜较少。P. gingivalis  W83和ΔPG0352在34℃和41℃条件下生物膜形成均较正常培养条件下（37℃）形成的生物膜少，差异均有统计学意义（均P < 0.05）。随着H2O2浓度的增加，W83和ΔPG0352形成的生物膜均逐渐减少；在H2O2终浓度分别为0.1、0.25和0.5 mmol/L时，ΔPG0352形成的生物膜均比W83少（均P < 0.05）。在碱性条件下（pH = 9），两者的生物膜形成均较正常条件下（pH = 7.2）减少，并且ΔPG0352形成生物膜比W83少；但在酸性条件下（pH = 5），两种菌株均不能形成生物膜。结论    唾液酸酶基因缺失影响P. gingivalis 的生物膜合成。在非正常培养条件下，无论是P. gingivalis  W83还是ΔPG0352，其生物膜的形成能力均有所减弱。

关键词: 牙龈卟啉单胞菌, 唾液酸酶, 生物膜