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Background <p>Contemporary molecular analytical methodologies have yielded insufficient characterization of the microbial etiology underlying chronic irreversible pulpitis; a pathological condition characterized by irreversible inflammatory alterations of the dental pulp complex necessitating endodontic intervention. This investigation employed shotgun metagenomic sequencing to comprehensively elucidate the microbiome present in affected pulpal tissues, thereby augmenting our understanding of pulpal pathogenesis.</p> Methods <p>The investigation incorporated six subjects (age range 18–35 years) presenting with clinically diagnosed chronic irreversible pulpitis according to the American Association of Endodontists diagnostic criteria. Pulpal tissue specimens were procured under rubber dam isolation utilizing stringent aseptic protocols following coronal access preparation. Genomic DNA extraction was performed via QIAamp DNA Mini Kit methodology followed by high-throughput sequencing on the Illumina Hiseq platform. Subsequent bioinformatic analysis implemented the WGSA2 pipeline for taxonomic classification, generating approximately 79.906&#xa0;million paired-end reads per specimen.</p> Results <p>Metagenomic analysis of the pulpal microbiome revealed taxonomic predominance of <i>Bacteroidetes</i> (45.095%), <i>Firmicutes</i> (17.424%), <i>Proteobacteria</i> (12.731%), and <i>Actinobacteria</i> (9.071%) at the phylum level. Notably, the investigation identified previously undocumented phyla in pulpal infections, including <i>Euryarchaeota</i>, <i>Thermoproteobacteria</i>, <i>Uroviricota</i>,and <i>Apicomplexa</i>. <i>Propionibacterium acidifaciens</i> emerged as the most consistently detected and ecologically significant species, whereas the conventionally recognized odontopathogen <i>Streptococcus mutans</i> exhibited negligible presence. Shannon diversity indices and taxonomic richness parameters demonstrated substantial inter-subject variability, with species abundance ranging from 574 to 5,468 distinct taxonomic units per pulp sample.</p> Conclusion <p>This investigation elucidated unprecedented microbial diversity within chronic irreversible pulpitis, fundamentally challenging established understanding of endodontic pathogenesis and clinical therapeutic approaches. The substantial inter-subject taxonomic heterogeneity observed herein suggests that contemporary standardized therapeutic regimens may be insufficiently targeted to address the complex polymicrobial ecosystem characteristic of pulpal pathosis. The identification of archaeal and viral constituents provides mechanistic insight into persistent endodontic infections despite technically adequate treatment modalities. These findings establish a comprehensive basis for evidence-based precision endodontics, facilitating the development of patient-specific antimicrobial strategies and novel therapeutic interventions targeting previously unrecognized microbial components. The comprehensive characterization of pulpal microbiome diversity represents a significant advancement toward molecularly informed clinical decision-making, with profound implications for treatment outcome optimization and the mitigation of therapeutic failures in contemporary endodontic practice.</p> Graphical Abstract <p></p>

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Profiling of microbes associated with chronic irreversible pulpitis using metagenomic next-generation sequencing

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2025年12月13日

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  • Preethesh Shetty,
  • Raksha Bhat,
  • Shruthi Padavu ,
  • Praveen Rai,
  • Krishna Kumar B,
  • Shishir Shetty

摘要

Background

Contemporary molecular analytical methodologies have yielded insufficient characterization of the microbial etiology underlying chronic irreversible pulpitis; a pathological condition characterized by irreversible inflammatory alterations of the dental pulp complex necessitating endodontic intervention. This investigation employed shotgun metagenomic sequencing to comprehensively elucidate the microbiome present in affected pulpal tissues, thereby augmenting our understanding of pulpal pathogenesis.

Methods

The investigation incorporated six subjects (age range 18–35 years) presenting with clinically diagnosed chronic irreversible pulpitis according to the American Association of Endodontists diagnostic criteria. Pulpal tissue specimens were procured under rubber dam isolation utilizing stringent aseptic protocols following coronal access preparation. Genomic DNA extraction was performed via QIAamp DNA Mini Kit methodology followed by high-throughput sequencing on the Illumina Hiseq platform. Subsequent bioinformatic analysis implemented the WGSA2 pipeline for taxonomic classification, generating approximately 79.906 million paired-end reads per specimen.

Results

Metagenomic analysis of the pulpal microbiome revealed taxonomic predominance of Bacteroidetes (45.095%), Firmicutes (17.424%), Proteobacteria (12.731%), and Actinobacteria (9.071%) at the phylum level. Notably, the investigation identified previously undocumented phyla in pulpal infections, including Euryarchaeota, Thermoproteobacteria, Uroviricota,and Apicomplexa. Propionibacterium acidifaciens emerged as the most consistently detected and ecologically significant species, whereas the conventionally recognized odontopathogen Streptococcus mutans exhibited negligible presence. Shannon diversity indices and taxonomic richness parameters demonstrated substantial inter-subject variability, with species abundance ranging from 574 to 5,468 distinct taxonomic units per pulp sample.

Conclusion

This investigation elucidated unprecedented microbial diversity within chronic irreversible pulpitis, fundamentally challenging established understanding of endodontic pathogenesis and clinical therapeutic approaches. The substantial inter-subject taxonomic heterogeneity observed herein suggests that contemporary standardized therapeutic regimens may be insufficiently targeted to address the complex polymicrobial ecosystem characteristic of pulpal pathosis. The identification of archaeal and viral constituents provides mechanistic insight into persistent endodontic infections despite technically adequate treatment modalities. These findings establish a comprehensive basis for evidence-based precision endodontics, facilitating the development of patient-specific antimicrobial strategies and novel therapeutic interventions targeting previously unrecognized microbial components. The comprehensive characterization of pulpal microbiome diversity represents a significant advancement toward molecularly informed clinical decision-making, with profound implications for treatment outcome optimization and the mitigation of therapeutic failures in contemporary endodontic practice.

Graphical Abstract