Rapid Bacterial Detection during Endodontic Treatment

Abstract

Bacteria present in the root canal (RC) space following an RC treatment (RCT) can lead to persistent infections, resulting in treatment failure and the need for reintervention or extraction. Currently, there are no standardized methods in use to clinically detect bacterial presence within RC spaces. The use of paper point sampling and fluorescence staining was shown to be a rapid method, able to detect residual bacteria following treatment. The study demonstrated that Calcein acetoxymethyl (AM) proved to be a suitable dye for detecting vital bacteria within mature endodontic biofilms, with an improved sensitivity over colony-forming unit counting in a stressed biofilm model. Furthermore, in a clinical trial with primary RCTs, 53 infected teeth were sampled in vivo, and increased detection of vital cells was found when compared with colony-forming unit counting, highlighting the sensitivity of the technique in detecting low cell numbers. By combining fluorescent staining and microspectroscopy with software-based spectral analysis, successful detection of vital cells from RCs was possible after 5 min of Calcein AM incubation. Application of this technology during RCT has the potential to reduce persistent infections through vital cell detection and additional treatment. Furthermore, this technique could be applied to antimicrobial research and disinfection control in clinical settings ( ClinicalTrials.gov NCT03055975).

Fluorescence fingerprints of oral bacteria

“…Fluorescent labeling is a very common methodology, and it can be applied to compounds that fluoresce from ultraviolet to near‐infrared wavelengths . Fluorescent probes are widely used such as viability staining using SYTO 9 and propidium iodide , and in situ hybridization to distinguish specific bacteria . Although the detection of specific bacteria based on fluorescence is sensitive and valuable, it is also costly and time‐consuming.…”

Rapid Chairside Microbial Detection Predicts Endodontic Treatment Outcome

“…Each canal was then sampled with two paper points to remove excess moisture and collect the maximum amount of residual bioburden. Samples were then assessed using a fluorescent spectral analysis as described by Herzog et al [ 5 ]. Calcein AM is a vital fluorescent stain, which only becomes fluorescent in the presence of vital cells.…”

“…As the detection was strongest at the paper points tip, the first three measurements from the tip were averaged and the value gained from the proportional Calcein fluorescence was used as an indicator of the percentage score [ 5 ].…”

“…Conventional techniques for bacterial detection and quantification such as colony-forming unit (CFU) counting [ 3 ] and DNA extraction, followed by polymerase chain reaction using universal bacterial primers [ 4 ] are time-consuming and not practicable for use in routine RCTs. Our group has developed and validated a methodology based on the use of a fluorescent vital dye for the chairside detection of vital cells present in the canal space, sampled with sterile endodontic paper points during RCTs [ 5 ]; this technique was found to be more sensitive than bacterial culture in detecting bacteria in vitro and in vivo [ 5 ].…”

Novel Approaches to Detect and Treat Biofilms within the Root Canals of Teeth: A Review

“…The dye calcein-AM (Calcein acetoxymethyl ester) has been used for detecting vital bacteria recovered from root canals on adsorbent paper points [18]. The dye is incubated with the sample for 5 min and the sample then evaluated using fluorescence and micro-spectroscopy with software-based spectral analysis.…”