RNA-based Assay Demonstrated Enterococcus faecalis Metabolic Activity after Chemomechanical Procedures

Area of Science:

• Microbiology
• Molecular Biology
• Endodontics

Background:

• Ribosomal RNA (rRNA) reflects metabolic cell activity.
• Enterococcus faecalis is a common cause of persistent endodontic infections.
• Distinguishing viable from non-viable bacteria is crucial in endodontic treatment assessment.

Purpose of the Study:

• To compare the sensitivity of rRNA-based quantitative polymerase chain reaction (RT-qPCR) with ribosomal DNA (rDNA)-based assays for identifying active E. faecalis.
• To evaluate the persistence of active E. faecalis in root canals after chemomechanical preparation.

Main Methods:

• Samples were collected from root canals of 18 patients before (S1) and after (S2) chemomechanical preparation.
• RNA and DNA were extracted, with RNA converted to complementary DNA (cDNA) via RT-PCR.
• Both cDNA and genomic DNA were analyzed using qPCR with primers specific to E. faecalis 16S rRNA.

Main Results:

• E. faecalis detection rates were higher with the rRNA assay (77.8% in S1, 61.1% in S2) compared to the rDNA assay (72.2% in S1, 33.3% in S2).
• The rRNA-based method demonstrated significantly higher sensitivity in both S1 and S2 samples.
• A high rRNA to rDNA ratio in S2 samples indicated the persistence of metabolically active E. faecalis post-treatment.

Conclusions:

• RT-qPCR targeting rRNA is a sensitive method for detecting active E. faecalis in endodontic samples.
• The rRNA-based assay identified persistent viable E. faecalis in treated root canals.
• This assay may be valuable for monitoring microbial load during endodontic therapy.