根管充填失败后根管内微生物的分子生物学分析

摘要

AIM:To assess microbial diversity in root-well-filled teeth associated with failed endodontic treatment.METHODS:DNA was extracted from 20 root-canal-treated teeth with post-treatment apical periodontitis, the 16S rRNA of all present bacteria was amplified by PCR,followed by cloning and sequencing of the 16S rRNA ampl-icons.RESULTS:Bacteria were present in all cases.15 phylotypes belonging to 4 phyla were found in 423 clones;a mean of 2.7 phylotypes were identified in each sample (range 1 ~6).Enterococcus faecali (60%,12/20)and pep-tostreptoccus micros (40%,8/20)were the prevalent species.The other 13 species were found in a few cases respec-tively,rummeliibacillus,exiguobacterium aurantiacum,pseudomonas veronii,phocaeicola abscessus were respectively de-tected in one patient.CONCLUSION:A high diversity of the microbial flora is associated with endodontic treatment failures.Enterococcus faecali is the most frequently identified species by 16S rRNA gene clone library analysis.%目的：基于16S rRNA，构建治疗失败根管内微生物的16S rRNA基因文库，并通过分子生物学方法分析其微生物的多样性。方法：选取经完善根管治疗2年后失败的患牙20例，根管内取样提取细菌DNA；然后用PCR扩增细菌16S rRNA基因，并构建其基因文库；最后通过测序比对构建系统发育树以确定微生物种类。结果：PCR检测显示，所有样本中有细菌出现；20个样本中共发现15种不同的系统发育型细菌（phlylotype），分属4个细菌门（phylum）：厚壁菌门（Firmicutes）、变异菌门（Proteobacteria）、拟杆菌门（Bacte-roidetes）、放线菌门（Actinobacteria）；每个样本中检出1～6种不同的细菌系统发育型，平均每个样本中可检测到2．7种；在检出的15种细菌中，以粪肠球菌（Enterococcus faecali）的出现频次最高（60％，12／20），微小消化链球菌（Peptostreptoccus micros）次之（40％，8／20），而另外13种细菌的出现频次较低，其中有4种系统发育型细菌（Rummeliibacillus、Exiguobacterium aurantiacum、Pseudomonas veronii、phocaeicola abscessus ）只出现了1次。结论：根管治疗失败主要是由多种细菌感染所致，其中以粪肠球菌的检出率最高。