General Protocol for Testing Systems that Treat Dental Unit Water

(Goal = < 200 cfu/ml)

17 March, 2000

Return to Dental Unit Water Menu

Research Question: Does the unit, disinfectant, or unit-disinfectant combination maintain bacterial levels in the dental unit water within the ADA goal of less than or equal to 200 colony forming units per milliliter (cfu/ml) at the exit port of handpieces and air/water syringes?

Methods

  1. Installation of dental unit waterline disinfection units
    1. Install disinfectant unit onto dental unit according to manufacturer directions.
    1. The unit installation is to be performed by either by CRA personnel, maintenance personnel of the office’s choice, or by the manufacturer.
    2. As per manufacturer’s directions, install only part of the disinfecting unit (such an injection port). Installation by CRA personnel, maintenance personnel of the office’s choice, or by the manufacturer.
  1. In office use of dental unit waterline disinfection unit, disinfectant, or unit-disinfectant combination
    1. Dental Unit Waterline Disinfection Unit Only.
    1. Operate dental unit waterline disinfection unit according to manufacturer’s directions. Operation to be conducted either by CRA staff members or by office staff members with CRA supervision.
    2. Maintenance or servicing of the dental unit waterline disinfection unit to be performed according to manufacturer’s directions, either by CRA personnel, the office staff, maintenance personnel of the office’s choice, or by the manufacturer.
    1. Disinfectant Only.
    1. The following options, per manufacturer’s directions, can be employed to introduce the disinfectant into the dental unit waterlines:
    1. Disinfectant is added through an injection port.
    2. Disinfectant is added to the reservoir of an independent water system that is either already in use by the office or has been previously evaluated by CRA.
    1. After the recommended contact time, flush the disinfectant from the lines, as per the manufacturer’s directions, by one of or by a combination of the following methods:
    1. For colored disinfectants, flush until disinfectant is no longer visible.
    2. For colorless disinfectants, flush until disinfectant cannot be detected by recommended monitoring device (pH strips, etc.).
    3. Flush until the smell and/or feel of the disinfectant is no longer detected.
    4. Flush for recommended time (usually 30 seconds to 1 min).
    1. Unit-Disinfectant Combination.
    1. Operate the dental unit waterline disinfection unit according to manufacturer’s directions. Operation by either CRA staff members or by office staff members with CRA supervision.
    2. Add the disinfectant recommended by the manufacturer. Addition of the disinfectant performed by either CRA staff members or by office staff members with CRA supervision.

    3. If disinfectant is automatically added to the dental unit waterlines by the unit, monitor disinfectant concentration and made sure the reservoir(s) are kept filled with disinfectant.
    4. If there is a recommended contact time, flush disinfectant from the lines, as per the manufacturer’s directions, by one of or a combination of the following methods:
    1. For colored disinfectants, flush until disinfectant is no longer visible.
    2. For colorless disinfectants, flush until disinfectant cannot be detected by recommended monitoring device (pH strips, etc.).
    3. Flush until the smell and/or feel of the disinfectant is no longer detected.
    4. Flush for recommended time (usually 30 seconds to 1 min).
  1. Treat patients, per normal procedures
  2. Collection of Water Samples
    1. Attach sterile handpiece or air/water syringe tip to end of line, then spray 5 - 50 ml of water into properly labeled sterile bottles. Avoid touching the tips of handpieces on the bottle lip. Use of sterile gloves is recommended.
    2. Send water samples to the lab either:
    1. Overnight in refrigerated pack or on ice if travel time will be more than 30 minutes.
    2. At room temperature, if travel time will be less than 30 minutes.
    1. Once samples are received at the lab, refrigerate until microbial evaluation. Evaluate for microbial load as soon as possible after receipt.
  1. Microbial Evaluation of Water Samples
    1. Controls.
    1. Negative control: sterile distilled water.
    2. Positive control: tap water sample.
    3. Neutralization controls: place duplicate 1-ml aliquots of test water or sterile PSS ([Physiological Sterile Saline] for a neutralization system control) into 9 ml of appropriate sterile neutralizer. Add a known titer of Staphylococcus aureus. Allow 30 minute contact time and then filter entire neutralization control (see step V.C.).
    1. Serially dilute water samples and positive control 1:10, first in appropriate sterile neutralizer (see step V.G.c.) and then further in PSS.
    2. Aseptically filter duplicate 1-ml aliquots through a 0.45-m m filter (unless sample is a neutralization control, see V.A.c.).

    3. Place filter onto R2A (water samples) or TSA (S. aureus controls) agar plates. Use plates that have been poured within the week and refrigerated.
    4. Incubate TSA agar plates upside down for 18 – 24 hours at 37° C.
    5. Place the R2A plates upside down inside a plastic bag and close with a twist tie (to prevent loss of moisture from the plates). Incubate R2A agar plates for 7 days at room temperature (~ 20 - 25oC).
    6. Count control colonies and calculate total cfu/ml of the sample.
    1. Negative control is acceptable only if no colonies are found.

    2. Positive control is acceptable only if colonies are growing on the R2A agar. The positive controls are compared over the testing period, to verify that the titers are within the expected range and that the colonies appear to be typical of what is normally found in city tap water samples.
    3. Neutralization controls are acceptable only if the S. aureus titer is close to the titer added. If there is a significant drop in the cfu/ml, then the neutralizer did not stop the action of the disinfectant in the water sample, and another, more effective neutralizer should be found and used.
    1. Count water sample colonies and calculate total cfu/ml of the sample.

Return to Top 

Return to Dental Unit Water Menu