State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China; Department of Operative Dentistry and Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu, China; Department of Orthodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
Arch Oral Biol. 2017 Nov;83:174-180. doi: 10.1016/j.archoralbio.2017.07.023. Epub 2017 Jul 29.
The aim of this study was to analyze the characteristics of the clustered regularly interspaced short palindromic repeats (CRISPR) sites in 45 clinical Streptococcus mutans strains and their relationship to the clinical manifestations of early childhood caries (ECC).
Forty-five S. mutans strains were isolated from the plaque samples taken from sixty-three children. CRISPR sites were sequenced and BLAST was used to compare these sites to those in the CRISPRTarget database. The association between the distribution of CRISPR sites and the manifestation of caries was analyzed by Chi-Square test. Further, biofilm formation (by crystal violet staining) and the synthesis of polysaccharide (by anthrone-sulfuric method) of all clinical isolated S. mutans strains with both CRISPR sites and no CRISPR site were comapared. Finally, acidogenicity and acidurity of two typical strains were determined using pH drop and acid tolerance assays. Biofilm formation and EPS synthesis by two typical strains were compared by 3D CLSM (Confocal Laser Scanning Microscope) assays and the expression of gtf genes were evaluated using qPCR.
We found that most of the spacers in the clinical S. mutans strains were derived from Streptococcus phages APCM01 and M102. The number of CRISPR sites in these strains was associated with the clinical manifestations of ECC. Moreover, we found that the biofilm formation and EPS synthesis ability of the S. mutans strains with both CRISPR sites was significant improved.
An association was found between the distribution of CRISPR sites and the clinical manifestations of caries. The CRISPR sites might contribute to the cariogenic potential of S. mutans.
本研究旨在分析 45 株临床变形链球菌（Streptococcus mutans）中簇状规则间隔短回文重复序列（CRISPR）位点的特征及其与婴幼儿早期龋（ECC）临床表现的关系。
从 63 名儿童的牙菌斑样本中分离出 45 株 S. mutans。对 CRISPR 位点进行测序，并使用 BLAST 将这些位点与 CRISPRTarget 数据库中的位点进行比较。采用卡方检验分析 CRISPR 位点的分布与龋病表现的关系。进一步比较所有具有 CRISPR 位点和无 CRISPR 位点的临床分离 S. mutans 菌株的生物膜形成（结晶紫染色）和多糖合成（蒽酮-硫酸法）。最后，采用 pH 值下降和耐酸试验测定两株典型菌株的产酸能力和耐酸性。通过 3D CLSM（共聚焦激光扫描显微镜）试验比较两株典型菌株的生物膜形成和 EPS 合成，采用 qPCR 评价 gtf 基因的表达。
我们发现临床 S. mutans 菌株中的大多数间隔子来源于噬菌体 APCM01 和 M102。这些菌株中的 CRISPR 位点数量与 ECC 的临床表现有关。此外，我们发现具有 CRISPR 位点的 S. mutans 菌株的生物膜形成和 EPS 合成能力显著提高。
CRISPR 位点的分布与龋病的临床表现之间存在关联。CRISPR 位点可能有助于 S. mutans 的致龋潜能。
