Pinheiro E T, Anderson M J, Gomes B P F A, Drucker D B
Endodontic Area, Dental School of Piracicaba, University of Campinas - UNICAMP, Piracicaba, São Paulo, Brazil.
Oral Microbiol Immunol. 2006 Jun;21(3):137-44. doi: 10.1111/j.1399-302X.2006.00285.x.
The objectives of the present study were to identify enterococcal species isolated from the canals of root-filled teeth with periapical lesions using biochemical and molecular techniques, and to investigate the genetic diversity of the isolates. Twenty-two Enterococcus strains, isolated from the canals of root-filled teeth with persisting periapical lesions, were identified to species level using rapid ID 32 STREP galleries and partial 16S rDNA sequencing. To subtype the strains, genomic DNA from the isolates was analyzed by pulsed field gel electrophoresis (PFGE) after digestion with SmaI. Intragenic regions of two genes, ace and salA, were sequenced for further differentiation of the isolates. All strains were identified as Enterococcus faecalis by both commercial kit and partial 16S rDNA sequencing. PFGE with SmaI of 22 isolates demonstrated 18 macrorestriction profiles, whereas 13 distinct genotypes were identified after analysis of the ace and salA composite sequences. Most of the isolates from distinct patients had different PFGE profiles. Moreover, in two cases, different E. faecalis strains were found in different root-filled teeth from the same mouth. E. faecalis was the only enterococcal species isolated from the canals of root-filled teeth with periapical lesions. Genetic heterogeneity was observed among the E. faecalis isolates following PFGE and sequence-based typing method. Furthermore, the genetic diversity within root canal strains was similar to previous reports regarding E. faecalis isolates from different clinical and geographic origins.
本研究的目的是使用生化和分子技术鉴定从患有根尖周病变的根管充填牙根管中分离出的肠球菌种类，并研究分离株的遗传多样性。使用快速ID 32 STREP鉴定板和部分16S rDNA测序，将从患有持续性根尖周病变的根管充填牙根管中分离出的22株肠球菌鉴定到种水平。为了对菌株进行亚型分析，用SmaI消化后，通过脉冲场凝胶电泳（PFGE）分析分离株的基因组DNA。对两个基因ace和salA的基因内区域进行测序，以进一步区分分离株。通过商业试剂盒和部分16S rDNA测序，所有菌株均被鉴定为粪肠球菌。22株分离株经SmaI酶切后的PFGE显示出18种宏观限制性图谱，而在分析ace和salA复合序列后鉴定出13种不同的基因型。来自不同患者的大多数分离株具有不同的PFGE图谱。此外，在两例中，在同一口腔的不同根管充填牙中发现了不同的粪肠球菌菌株。粪肠球菌是从患有根尖周病变的根管充填牙根管中分离出的唯一肠球菌种类。在PFGE和基于序列的分型方法之后，在粪肠球菌分离株中观察到遗传异质性。此外，根管菌株内的遗传多样性与先前关于来自不同临床和地理来源的粪肠球菌分离株的报道相似。
