Analysis of bacterial invasive lesion of refractory periapical periodontitis cases by genetic engineering technique

使用标准培养技术已经从根管中分离出了种类相对广泛的细菌。然而，口腔中只有50%的细菌是可培养的。因此，牙髓感染的细菌多样性被低估了。本研究使用基于pcr的16S rRNA或细菌物种特异性基因测定来评估感染根管中存在的细菌多样性。采用以细菌16S rRNA和细菌物种特异性基因为目标的PCR引物，对难治性根尖周炎患者根管中可能存在的细菌病原体进行鉴定。此外，这些微生物与症状和根管治疗史的关系进行了调查。难治性根尖周炎患者的根管微生物样本纳入本研究。根管周围辐射区较大的根管标本中检出最多的是核梭菌（62.5%），最难检出的是密螺旋体（Treponema denticola），较难培养的还有单宁菌（Tannerella forsythesis）和牙龈卟啉单胞菌（Porphyromonas龈）。根管样品有恶臭或渗出液与牙龈假单胞菌的存在有关。在标本中还检出难治性根尖周病相关微生物粪肠球菌。除细菌外，其中一种真菌白色念珠菌被检测到菌丝和酵母两种类型。这些结果表明，在根管治疗过程中，仍有多种细菌在根管内存活。在根管手术中，经化学力学清洁和根管整形后，根管显微镜观察发现根管骨折，尤其是根尖区骨折。这些结果提示，难治性慢性根尖周炎病变是由细菌在根尖破坏区侵袭生长而形成的，并提示具核假单胞菌、连翘假单胞菌和牙龈假单胞菌与难治性根尖周炎有关。少

A relatively wide range of bacteria have been isolated from root canals using standard culture techniques. However, only 50% of the bacteria in the oral cavity are cultivable. Hence, bacterial diversity in endodontic infections is underestimated. This study used a PCR-based 16S rRNA or bacterial species-specific gene assay to assess the diversity of bacteria present in infected root canals. PCR primers that target the bacterial 16S rRNA and bacterial species-specific genes were used to identify putative bacterial pathogens in root canals with refractory periapical periodontitis. In addition, the associations of these microorganisms with symptoms and a history of endodontic treatment were investigated. Microbial samples from root canals with refractory periapical periodontitis were included in this study.Fusobacterial nucleatum (62.5%) were most frequently detected in the samples from root canals with relatively large peripaical radiolucent area, and Treponema denticola, which is diffic … More ult to culture, Tannerella forsythesis and Porphyromonas gingivalis were also detected. Samples from root canal with foul odor or exudates fluid were associated with the presence of P.gingivalis. Enterococcus faecalis, which is suggested as refractory periapical priodontitis-related microorganism,was also detected in samples. In addition to the bacteria, Candida albicans, one of the fungi, was detected as both hypha and yeast types. These results suggested that various bacterial species still survived in root canals during the root canal treatment. After chemomechanically cleaning and shaping of root canal, the fractures of the root, especially apex area, were found by the endodontic microscopy observation at the endodontic surgery.These observations suggest that refractory chronic periapical periodontitis lesion is formed by bacterial invasion and growth in the area of root apex destroyed by some reasons, and indicated that F.nucleatum, T.forsythensis and P.gingivalis are associated with refractory periapical periodontitis. Less