Molecular biological analysis of oral bacterial population dynamics

1.采用非培养分子生物学方法对龋性牙本质和牙菌斑细菌群落进行分析。16 SrDNA的PCR扩增和DNA测序结果显示，存在多种细菌，包括从口腔中分离较少的细菌或与数据库中登记的细菌没有序列同源性的细菌。此外，细菌种群之间的初始阶段和进行性阶段的龋的差异。在浅龋牙本质中检测到链球菌、放线菌等与菌斑样品相似的细菌，而在深龋牙本质中以乳杆菌为主，表明随着龋的进展，细菌种群发生了明显的变化。在这项研究中，我们分析了3个不同的患者龋牙本质样本。在所有样本中，强龋致病菌如突变链球菌或远缘链球菌的检出率极低或为零，这可能表明各种口腔细菌与龋齿的进展有关。2为了检测污染样本中的特定细菌，我们尝试了牙周病原体的PCR检测，从60名健康青年志愿者牙菌斑中分离放线杆菌和牙龈卟啉单胞菌。通过3年的调查，在不同人群中检出20%~ 30%的伴放线杆菌，而选择性培养基培养法不能分离到伴放线杆菌。

1 Carious dentine and dental plaque bacterial communities were analyzed by the culture-independent molecular biological method. PCR amplification and DNA sequencing of 16SrDNA showed variety of bacterial existence, including organisms less isolated from oral cavity or ones with no sequence homology to the databank registered bacteria. Additionally, bacterial population difference between initial and progressed stage of caries was shown. In the shallow carious dentine bacteria represented Streptococcus, Actinomyces, similar to the plaque sample were detected, but in the deeply progressed carious dentine Lactobacillus was dominant, suggesting that there is a significant change of bacterial population accompanied with the dental caries progression. In this study, we analyzed carious dentine samples from 3 different patients. Of all samples, detection rates of strong carious causative agents such as S.mutant or S.sobrinus are extremely low or zero, which may suggest that various oral bacteria are related to the progression of dental caries.2 In order to detect the specific bacteria from contaminated specimens, we attempted the PCR detection of periodontal pathogens, Actinobacillus actionomycetemcomitans and Porphromonas gingivalis from dental plaque sample of 60 young healthy volunteers. Through 3 years of investigation, against the distinct groups of volunteers, Actinobacillus actionomycetemcomitans, which was not able to be isolated on the selectively media culture, was detected in ratio 20 to 30%.As the above mentioned, analysis using molecular biological culture-independent method brought a new kind of knowledge about the oral bacterial communities.