根管消毒对原发性牙髓感染中细菌组的影响：下一代测序研究。

Impact of root canal disinfection on the bacteriome present in primary endodontic infection: A next generation sequencing study.

机构信息

Department of Advanced Oral Sciences and Therapeutics, School of Dentistry, University of Maryland, Baltimore, Maryland, USA.

Department of Periodontics and Oral Implantology, Dr D.Y. Patil Dental College and Hospital, Dr D Y Patil Vidyapeeth, Pune, India.

出版信息

Int Endod J. 2024 Aug;57(8):1124-1135. doi: 10.1111/iej.14074. Epub 2024 May 3.

DOI:10.1111/iej.14074

PMID:38700876

Abstract

AIM

To investigate the bacteriome present in teeth with primary endodontic infection (PEI) and apical periodontitis (AP) and to determine quantitatively and qualitatively the impact of chemomechanical preparation (CMP) using 2.5% sodium hypochlorite NAOCl on the bacteriome found in PEI with AP using the Illumina MiSeq platform.

METHODOLOGY

Thirty-six paired samples from 18 patients were successfully sequenced and analysed. Samples were collected at two sampling times: before (s1) and after (s2) CMP using 2.5% NaOCl. The DNA was extracted from s1 and s2 samples and quantified using quantitative PCR (qPCR). All 36 samples were sequenced using the Illumina MiSeq platform. Raw V3-V4 amplicon sequencing data were processed with the DADA2 pipeline to generate amplicon sequence variants (ASVs). Alpha diversity metrics representing abundance (Chao1) and diversity and evenness (Shannon, Simpson) were computed. The paired-sample Wilcoxon's test was used to compare alpha diversity metrics and qPCR counts between s1 and s2. The PERMANOVA method (with 999 permutations) was applied to compare community composition between sample types (s1 versus s2) and between patient IDs. ALDEx2 (ANOVA-like differential expression tool for high-throughput sequencing data) to investigate differentially abundant taxa between s1 and s2. A paired-sample Wilcoxon's test was used to compare alpha diversity metrics and qPCR counts between s1 and s2.

RESULTS

The qPCR counts were significantly higher in s1 compared to s2 (p = .0007). The Chao1 index indicated no difference in alpha diversity (p < .7019); whereas Shannon (p = .0056) and Simpson (p = .02685) indexes showed higher values in s2. The PERMANOVA test using Adonis2 showed a significant effect of sample time on community composition (R = .0630, p = .012). Patient ID also showed a significant effect on community composition (R = .6961, p = .001). At the genus level, Dialister, Mogibacterium, Prevotella, and Olsenella were differentially enriched at s1, while Actinomyces, Stenotrophomonas_unclassified, Enterococcus_unclassified, and Actinomyces_unclassified were differentially enriched in s2.

CONCLUSION

The bacteriome present in teeth with PEI with AP is complex and diverse. CMP using 2.5% NaOCl showed a high quantitatively and qualitatively disinfectant impact on the bacteriome present in PEI with AP.

摘要

目的

研究原发性牙髓感染（PEI）和根尖周炎（AP）牙齿中的细菌组，并使用 Illumina MiSeq 平台定量和定性地确定使用 2.5%次氯酸钠（NaOCl）的化学机械预备（CMP）对 AP 中发现的 PEI 细菌组的影响。

方法

成功对 18 名患者的 36 对样本进行了测序和分析。在两次采样时间采集样本：CMP 前（s1）和后（s2）使用 2.5%NaOCl。从 s1 和 s2 样本中提取 DNA，并使用定量 PCR（qPCR）进行定量。使用 Illumina MiSeq 平台对所有 36 个样本进行测序。使用 DADA2 管道处理原始 V3-V4 扩增子测序数据，以生成扩增子序列变异体（ASVs）。计算代表丰度（Chao1）和多样性和均匀度（Shannon、Simpson）的 alpha 多样性指标。使用配对样本 Wilcoxon 检验比较 s1 和 s2 之间的 alpha 多样性指标和 qPCR 计数。应用 PERMANOVA 方法（999 次随机排列）比较样本类型（s1 与 s2）和患者 ID 之间的群落组成。使用 ALDEx2（用于高通量测序数据的 ANOVA 样差异表达工具）研究 s1 和 s2 之间差异丰度的分类群。使用配对样本 Wilcoxon 检验比较 s1 和 s2 之间的 alpha 多样性指标和 qPCR 计数。

结果

qPCR 计数在 s1 中明显高于 s2（p = .0007）。Chao1 指数表明 alpha 多样性无差异（p < .7019）；然而，Shannon（p = .0056）和 Simpson（p = .02685）指数在 s2 中显示出更高的值。使用 Adonis2 的 PERMANOVA 检验显示样本时间对群落组成有显著影响（R = .0630，p = .012）。患者 ID 也对群落组成有显著影响（R = .6961，p = .001）。在属水平上，Dialister、Mogibacterium、Prevotella 和 Olsenella 在 s1 中差异丰富，而 Actinomyces、Stenotrophomonas_unclassified、Enterococcus_unclassified 和 Actinomyces_unclassified 在 s2 中差异丰富。