Bacterial contamination of new dental unit waterlines and efficacy of shock disinfection

Abstract

Background

Bacterial contamination of outlet water from dental unit waterlines (DUWLs) may lead to healthcare-associated infection. The aim of this study was to evaluate the contamination of DUWLs from new dental chairs before and after their initial shock disinfection with ICX Renew® and to determine the efficacy of this method of shock disinfection.

Methods

The microbiological quality of water samples obtained from the DUWLs of 17 new dental chairs installed at the University Hospital of Nancy, France, was assessed. Water samples were collected before and after an initial shock disinfection with one ICX Renew® treatment. Water analysis was used to assess total culturable aerobic bacteria (TCAB) at 22°C and 36°C, Legionella sp., Pseudomonas aeruginosa and total coliforms.

Results

All the DUWLs (17/17) were contaminated by viable aerobic bacteria (TCAB at 22°C and 36°C > 300 CFU/mL), including P. aeruginosa, before shock disinfection. After shock disinfection with ICX Renew®, 24% of DUWLs (4/17) remained contaminated by aerobic bacteria (TCAB at 36°C > 300 CFU/mL for 3/17) and P. aeruginosa (> 100 CFU/100 mL for 1/17).

Conclusions

To reduce the contamination of new dental chairs, initial shock disinfection of DUWLs should be performed to remove biofilms and bacteria from the DUWLs. To ensure the security of care, microbiological analysis of outlet water from DUWLs should be systematically performed before the first clinical use of a new dental chair to determine the efficacy of shock disinfection and the agreement of the results with the dental recommendations for water quality.

Clinical trial number

Not applicable.

Background

Water is crucial for dental care because it cools and rinses both the site of intervention and dental instruments, such as high-speed rotating instruments and ultrasonic scalers, to avoid overheating which can damage dental and periodontal tissues. The water of dental units may be supplied by water from the municipal water network or from a tank or a bottle connected to the dental unit; then, the water flows through the dental unit waterlines (DUWLs) and finally emerges from the dental unit instruments such as water syringes, contra-angles, turbine handpieces, and ultrasonic scalers. When water is contaminated with microorganisms, exposed individuals, especially immunocompromised individuals, are at risk of infection. Patients and dental staff are exposed to dental unit water through contact (splatters, inoculation into oral wounds), ingestion and inhalation (aerosols) [1].

Several studies published in the last decade have revealed that the water emerging from DUWLs is regularly contaminated with high densities of bacteria [2–5] and with potential human pathogenic bacteria such as Pseudomonas aeruginosa, Legionella pneumophila and Mycobacterium sp. [6–12].

The risk of healthcare-associated infections from the water of dental units is a public health concern, especially because dental care is extremely common in the population [1]. For example, 66% of adults and 87% of children in the USA had a dental visit in 2019 [13]. Several healthcare-associated infections due to contamination of dental unit water have been reported in the literature. Two case series revealed outbreaks of Mycobacterium abscessus infections, including 24 and 71 children in two pediatric dental clinics in the USA, due to the contaminated water of the dental units used for pulpotomies [8, 10]. Two deaths of elderly patients have been reported, one in Italy and the other in Sweden, following L. pneumophila infection due to exposure to water from contaminated DUWLs [9, 14]. Exposure to bacteria from DUWLs affects not only patients but also dental staff. A high prevalence of seropositivity for Legionella antibodies has been reported among dental staff [15–17], and one case of fatal pneumonia caused by Legionella pneumophila in a Californian dentist, which was probably related to contamination of the water in his dental office, has been reported [18].

To prevent these risks, the Centers for Disease Control and Prevention (CDC) recommends that the number of bacteria in water used for nonsurgical dental care should be as low as reasonably achievable and no more than 500 CFU/mL [19]. In our hospital, more restrictive criteria (namely, total culturable aerobic bacteria [TCAB] at 22 °C ≤ 300 CFU/mL; TCAB at 36 °C ≤ 100 CFU/mL; and the absence of L. pneumophila, P. aeruginosa and coliforms) are used to ensure microbiological safety [20]. To prevent and control microbial contamination of DUWLs, different disinfectants have been used and tested in dental practices over the last decade [21–29]. It has also been shown that shock disinfection of DUWLs may be used as a precautionary step to remove biofilms and reduce the quantity of bacteria before continuous treatment of DUWLs [26].

Contamination of water from DUWLs has been largely studied among dental chairs used for several years [1]. However, only one study we authored ourselves has investigated the contamination of water from DUWLs among dental chairs before their first clinical use. We performed this study on newly installed dental chairs in 2022 to evaluate the contamination of water from DUWLs after their initial shock disinfection. We observed contamination by P. aeruginosa in 3/24 dental chairs after shock disinfection, but we did not sample the water from the DUWLs before shock disinfection [6]. A second phase of renovation of our dental center allowed us to complete this first study.

The present study aimed to evaluate the initial contamination of DUWLs among new and never used dental chairs before and after initial shock disinfection and to evaluate the efficacy of initial shock disinfection with ICX Renew® (A-dec, Inc., Oregon, USA), as recommended by the manufacturer.

Methods

Dental chairs

Seventeen new dental chairs installed in 2023 at the University Hospital of Nancy, France, were included in this study. The dental chairs had an independent water bottle to supply water to their DUWLs and did not have a water cup filler (Fig. 1).

Fig. 1.

Dental chair and its independent water bottle

The dental chairs were manufactured in November 2022 and then stored by A-Dec® (A-dec, Inc., Oregon, USA) in the USA. The dental chairs were transported by boat to France over a period of seven days in February 2023. The chairs were then transported by truck over a period of two days and then stored at the distributor’s warehouse (Eurotec Dental, France). The chairs were preassembled by the distributor one month before their installation. Finally, the chairs were delivered by truck and assembled at the University Hospital of Nancy from November 6 to 15, 2023. On November 15, the DUWLs were supplied with sterile water via the independent bottles of the dental chairs to test for leaks. On November 16, the water emerging from the DUWLs was first sampled, and the DUWLs were then disinfected with ICX Renew®. On November 20, sterile water was again run through the DUWLs and then resampled (Fig. 2).

Fig. 2.

Installation, shock disinfection of DUWLs and water sampling for the 17 new dental chairs

Dental unit waterline shock disinfection

From November 16 to 20, the first disinfection of DUWLs was performed by the fitter with ICX Renew®. ICX Renew® is a DUWL shock disinfectant that consists of a simple two-part liquid solution colored blue and containing surfactants and hydrogen peroxide to reduce and remove the build-up of contaminating bacteria [30–32]. In accordance with the manufacturer’s instructions [30], solution #1 (120 mL) and solution #2 (120 mL) of ICX Renew® were mixed together in the independent empty bottle of each dental chair. Then, the disinfectant was run through the DUWLs until it flowed out of the air/water syringe and of all the handpiece tubing to coat all inside surfaces of the DUWLs. The ICX Renew® solutions remained in the independent bottle and the DUWLs for 90 ± 2 h (the manufacturer recommends extending disinfection for up to 12–96 h). Finally, all the disinfectant was purged, and then the independent bottle and DUWLs were flushed with sterile water until no trace of blue color from ICX Renew® remained.

Water sampling

Sampling was performed before (November 16) and after (November 20) DUWL shock disinfection. Water samples (500 mL) were taken simultaneously from each of the five DUWL outlets of the dental chairs (100 mL/outlet): one water syringe, two micromotor supply tubes, one turbine handpiece supply tube, and one ultrasonic handpiece supply tube. Water samples were collected aseptically in sterile containers without inhibitors and then transferred to the laboratory within 2 h in insulated boxes for microbiological analysis.

Microbiological analysis

Each water sample was analyzed in the environmental microbiology laboratory of the university hospital for the following:

• total culturable aerobic bacteria (TCAB) at 22 and 36°C according to NF EN ISO 6222 [33] using 2 × 1 mL;

• Pseudomonas aeruginosa according to NF EN ISO 16266 [34] using 100 mL;

• coliform bacteria and Escherichia coli according to NF EN ISO 9308–1 [35] using 100 mL;

• Legionella sp. according to NF EN ISO 11731 [36] using 0.2, 10, and 100 mL.

The details of the microbiological analysis were described in a previous study [20].

Statistical analysis

Microbiological data were collected in Microsoft® Excel (Microsoft Corporation, WA, USA). The data are presented as numbers and percentages. The reduction rate, R (Eq. 1), of the number of TCAB was calculated as the difference between the mean number of TCAB before (M_B) and after (M_A) shock disinfection, normalized by the value before (M_B). The number of TCAB before and after shock disinfection was compared with the Wilcoxon test for matched samples using RStudio. To perform the calculations and analysis, when the limit of quantification was exceeded (> 300 CFU/mL for TCAB), the number of bacteria was replaced by the limit value (i.e., 300). The statistical significance threshold was set at p < 0.05.

$$R\left(\%\right)=\frac{M_B-M_A}{M_B}$$ 1

Results

Microbiological analysis of the water samples from the 17 dental chairs revealed no coliform bacteria and no Legionella sp. (Table 1). Before shock disinfection, all the DUWLs (100%) were contaminated, with TCAB at 22°C and 36°C exceeding the limit of quantification (i.e., > 300 CFU/mL) and not quantifiable P. aeruginosa because of confluent growth (the colony-forming units covered all the Petri dish). After shock disinfection, the water quality of 13 DUWLs (76%) significantly improved (i.e., no P. aeruginosa detected, TCAB at 22°C ≤ 300 CFU/mL, and TCAB at 36°C < 100 CFU/mL) including 11 DUWLs (65%) in which no bacteria were detected. Four DUWLs (24%) still had unsatisfactory water quality levels (i.e., P. aeruginosa detected or TCAB at 22°C > 300 CFU/mL or TCAB at 36°C > 100 CFU/mL).

Table 1.

Microbiological analysis of water samples obtained from DUWLs before and after shock disinfection with ICX Renew®

	TCAB at 22°C (CFU/mL)		TCAB at 36°C (CFU/mL)		P. aeruginosa (CFU/100 mL)		Coliforms (CFU/100 mL)		Legionella sp. (CFU/100 mL)
No	Before	After	Before	After	Before	After	Before	After	Before	After
1	> 300	0	> 300	0	NC	0	0	0	ND	ND
2	> 300	0	> 300	0	NC	0	0	0	ND	ND
3	> 300	0	> 300	0	NC	0	0	0	ND	ND
4	> 300	17	> 300	93	NC	0	0	0	ND	ND
5	> 300	0	> 300	0	NC	0	0	0	ND	ND
6	> 300	0	> 300	0	NC	0	0	0	ND	ND
7	> 300	> 300	> 300	> 300	NC	0	0	0	ND	ND
8	> 300	0	> 300	0	NC	0	0	0	ND	ND
9	> 300	2	> 300	> 300	NC	0	0	0	ND	ND
10	> 300	0	> 300	0	NC	0	0	0	ND	ND
11	> 300	0	> 300	0	NC	0	0	0	ND	ND
12	> 300	0	> 300	0	NC	0	0	0	ND	ND
13	> 300	0	> 300	1	NC	0	0	0	ND	ND
14	> 300	0	> 300	0	NC	0	0	0	ND	ND
15	> 300	> 300	> 300	> 300	NC	0	0	0	ND	ND
16	> 300	0	> 300	0	NC	0	0	0	ND	ND
17	> 300	16	> 300	15	NC	> 100	0	0	ND	ND

NC colonies not countable because of confluent growth, ND Legionella sp. not detected (< 10 CFU/100 mL)

Shock disinfection resulted in > 88% and > 80% reduction rates for TCAB at 22°C and 36°C, respectively. The number of TCAB at 22°C and 36°C significantly decreased after shock disinfection (p = 0.0003 and p = 0.0005, respectively).

Discussion

In our study, the DUWLs of new dental chairs were heavily contaminated by bacteria and notably by P. aeruginosa when they were installed. As the DUWLs were supplied exclusively with sterile water before sampling, these microorganisms originated only from the DUWLs. The inner surfaces of DUWLs were likely coated with biofilms, which released bacteria into the water flowing through the DUWLs. The contamination of the DUWLs may have occurred during the manufacturing of the dental chairs and, more specifically, during quality tests performed with water from the main water supply of the factory. The contaminating bacteria were then able to grow and form biofilms during storage and delivery by boat and truck [6]. The contamination of the DUWLs may also have occurred during the dental chair preassembly step, which was carried out by the distributor one month before their installation and included quality tests performed with water from the main water supply of the distributor workshop.

DUWLs provide a favorable environment for biofilm growth considering i) the high complexity of DUWLs, consisting of approximately 6 m of narrow-bore plastic tubing with a 2 mm inner diameter, along with a number of brass connections and other inflexible plastic connections; ii) the low water flow in the DUWLs, with negligible flow at the periphery of the canal lumen; iii) the plastic materials constituting the DUWLs; iv) the water stagnation during inactive periods; and v) the average temperature of DUWLs, which may promote the proliferation of microorganisms [1, 37, 38].

This study demonstrated that initial shock disinfection of new dental chairs before their first clinical use is crucial to decrease the bacterial load inside DUWLs but it may be insufficient to ensure patient safety. The manufacturer of the dental chairs installed in our hospital advises a shock disinfection of the DUWLs with one ICX Renew® treatment before the first use of the new dental chairs to clear organic deposits and bacterial contamination [30]. In the present study, we observed that one shock disinfection with ICX Renew® treatment was insufficient for 24% of the new dental chairs, in line with our previous study, which reported 25% noncompliant water samples for new dental chairs after only one ICX Renew® treatment [6]. Taken together, these data indicate that the manufacturer’s recommendations should be changed. From this perspective, the systematic use of repeated shock treatments could be considered. It is noteworthy that three consecutive ICX Renew® treatments are already recommended by the manufacturer for existing dental chairs when the water quality test results exceed the water quality action level [30]. Moreover, in a laboratory test performed by the manufacturer, the use of two ICX Renew® treatments resulted in a > 99.999% reduction in planktonic bacteria, and a third ICX Renew® treatment resulted in little to no observable bacteria in the DUWL tubing [31]. However, further research under real conditions is necessary to determine the number of repeated ICX Renew® treatments needed to ensure microbiological safety. Due to the necessity of putting the dental chairs into operation by a set date for imperative functional reasons related to patient care, we were not able to test the required number of ICX Renew® cycles to remove the biofilm. Therefore, the other option that we considered is the use of another shock disinfectant. In the present study, the four DUWLs with unsatisfactory water quality levels were secondarily treated with BRS® (Alpro Medical GmbH, Germany), another shock disinfection, which led to decontamination of the DUWLs. The efficacy of BRS® was previously demonstrated in two studies performed in our hospital [6, 20].

Whatever the shock disinfection method used, microbiological analysis of a water sample appears to be required before the first clinical use of the dental chairs to confirm the efficacy of disinfection and to prevent healthcare-associated infections.

Shock disinfection removes biofilms from DUWLs, rapidly reduces the quantity of bacteria in outlet water and allows satisfactory results for the continuous disinfection of in-use DUWLs [26]. The three main factors influencing the contamination of water from DUWLs under in-use conditions were found to be the DUWL disinfection frequency, the quality of the water supplied, and the efficacy of the antiretraction valve [3]. Concerning DUWL disinfection, it has been reported that daily or weekly disinfection is more efficient than no disinfection [3], that continuous disinfection is more efficient than intermittent disinfection [24], and that not all continuous disinfection systems have the same efficacy [25, 39]. Concerning the quality of the water supplied, the use of purified water significantly reduced the concentration of bacteria in the outlet water from DUWLs [3]. This effect was comparable to that observed when sterile or 0.22 μm-filtered water was used to supply the independent water bottle of dental chairs [20, 40]. Finally, the absence or dysfunction of the antiretraction valve may also significantly increase the risk of retrograde contamination of DUWLs by oral fluids [3, 4]. Oral pathogens were isolated from DUWL samples, confirming a possible retrograde contamination route from a patient’s mouth to the DUWL [41]. In addition to the use of suitable valves, one way to reduce this risk and transiently reduce bacterial contamination is to flush DUWLs for 20–30 s between each patient [37]. Unfortunately, all these measures are poorly followed by dentists, especially in France, where 65% of DUWLs are supplied by the main water of the municipal network, 30% of DUWLs are not disinfected, and only 20% of dentists report flushing their DUWLs between patients [42].

A limitation of this study was the limit of quantification of the total culturable aerobic bacteria (TCAB) at 22 and 36°C, which was limited to 300 CFU/mL because 1 mL of an undiluted water sample was cultured on 90 mm Petri dishes [33]. To enable a more accurate determination of DUWL outlet water contamination by TCAB, another study including the analysis of diluted samples is needed.

Conclusions

The DUWLs of new dental chairs are contaminated by numerous bacteria, including P. aeruginosa, which is an opportunistic human pathogen. Initial shock disinfection of DUWLs before their first clinical use is crucial for removing biofilms from the inner surfaces of DUWLs, reducing the contamination of outlet water, and reducing the risk of healthcare-associated infection.

The efficacy of initial shock disinfection is not guaranteed, and microbiological analysis of the outlet water from DUWLs seems crucial to ensure its microbiological quality before the use of a new dental chair.

Abbreviations

CFU

Colony-forming unit

DUWLs

Dental unit waterlines

EDTA

Ethylene diamine tetra acetic acid

TCAB

Total culturable aerobic bacteria

Authors’ contributions

Conceptualization, A.B., A.F., J.L. and E.M.; Methodology, A.B., A.F., A.L. and E.M.; Formal Analysis, A.B. and E.M.; Investigation, A.B., J.L. and E.M.; Data Curation, A.L. and E.M.; Writing—Original Draft Preparation, A.B.; Writing—Review & Editing, A.B., A.F., A.L., E.M. and J.L.; Visualization, A.B.; Supervision, E.M.; Project Administration, E.M.

Funding

The authors received no funding for this work.

Data availability

Data fully available in the article.

Declarations

Ethics approval and consent to participate

Not applicable.

Competing interests

The authors declare no competing interests.