Waterline Disinfectants Reduce Dental Bioaerosols: A Multitracer Validation

Abstract

Oral microbes are dispersed during dental treatment and reduction methods have been proposed, but dental unit waterline (DUWL) disinfectants have received little attention; specifically, the effect on viruses has not been studied. This study aims to 1) investigate the effect of DUWL disinfectants on viral dispersion in dental bioaerosols and 2) establish a dual-tracer system using live bacteriophage and fluorescein supported by optical particle measurement. Bacteriophage MS2 was used as a viral tracer and fluorescein as a fluorescent tracer. Validation experiments were conducted to exclude interference of one tracer with the other or of DUWL disinfectants on detection methods. Simulated "saliva" containing the tracers was infused into the mouth of a dental mannequin during 10-min dental procedures with an air turbine handpiece (n = 3 replicates). Aerosols and droplets were sampled in an enclosed dental operatory using air samplers and settlement onto sterile filter papers. Bacteriophage was quantified using plaque assays and reverse transcription quantitative polymerase chain reaction (RT-qPCR). Fluorescein was quantified fluorometrically. The effect of DUWL disinfectants on total aerosol concentration was assessed in separate experiments using an optical particle counter. DUWL disinfectants reduced bacteriophage viability, and interference between tracers was not observed. In simulated clinical procedures, the disinfectant ICX reduced bacteriophage detection substantially (P < 0.001; 2-way analysis of variance). MS2 RNA was detected in all experimental samples but not negative controls. Samples positive on RT-qPCR but not plaque assays may indicate that virions at distant sites are nonviable. Fluorescein tracer showed good agreement with the bacteriophage tracer. DUWL disinfectants designed for continuous presence in irrigants reduce the dispersion of viable virus in dental bioaerosols during simulated procedures. Their use may therefore be important for routine infection control and as a mitigation factor during infectious disease outbreaks. Future studies should explore this using a range of viruses and other microbes.

Keywords: aerosol-generating procedure; dental disinfectants; dental facilities; dental high-speed equipment; infection control; viral plaque assay.

Figure 1.

Schematic view of sampling locations in clinical simulation experiments; not to scale. Created using BioRender.com.

Figure 2.

Results of laboratory validation experiments. (A) Effect of Alpron and ICX on viable MS2 bacteriophage detected by plaque assay (n = 3 per data point). (B) Effect of ICX at the manufacturer’s concentration on fluorescence from fluorescein (n = 3 per data point). R² = .999 for both fluorescein and fluorescein + ICX. (C) Effect of fluorescein on viable MS2 bacteriophage detected by plaque assays after incubation with 1 g/L fluorescein for 1 h or 24 h (n = 9 per condition). Error bars show standard deviation. PFU, plaque-forming units; RFU, relative fluorescence units.

Figure 3.

Results of clinical simulation experiments. Effect of ICX in dental handpiece irrigant on (A) viable MS2 bacteriophage tracer recovered from filter paper samples and detected by plaque assay and (B) fluorescein tracer recovered from filter papers and detected fluorometrically. n = 3 replicates were conducted for each experimental condition, and multiple samples were present at each distance from the procedure, thereby giving the following number of samples per condition at each distance: 0.5 m, n = 12; 1.5 m, n = 6; 3.0 m, n = 9; center and negative control, n = 3. Error bars show standard deviation. Dotted line shows lower limit of detection for fluorescein (negative control mean + 2 SD). Normalized for surface area of filter papers used for collection (7.07 cm²). PFU, plaque forming units; RFU, relative fluorescence units.

Figure 4.

Time-series data of particle number concentration (D_p 0.3–10.0 µm) for each experimental condition in optical particle counter experiments. 1× ICX = ICX at manufacturer’s concentration; 10× ICX = ICX at 10 times manufacturer’s concentration; D_p, particle diameter. The air-turbine handpiece was operated continuously beginning at 0 min. Sampling interval = 5 s shown by feint line; bold line shows 5-point moving average.