Microbiological testing, specifically for total bacteria and Legionella bacteria in cooling tower water systems, has been using a methodology that basically has not changed for many years. The recent Legionnaires’ disease outbreaks due to cooling tower water systems has prompted a closer look at the methodology to determine if there is a need to consider a change that would be more representative of the true microbiological levels in these systems. Testing and measuring of the biofilm instead of water samples suggests that this is a much better method. Implementing this method will provide better indication of total bacteria and Legionella bacteria levels in the cooling tower water system, which would reduce outbreaks.

The solution to improved cooling tower operations is a different control method. This article discusses a field-friendly testing procedure that can be used in cooling water systems.

Bacteria and Cooling Tower Water

Cooling tower water systems are an ideal environment for the growth of many types of bacteria, including Legionella bacteria. These systems have an environment of warm water (60 to 100°F [15 to 38°C]), aeration and nutrients. These nutrient sources can be from the cooling tower makeup water, from the air scrubbed out by the cooling tower water, and from the water treatment chemicals used for scale and corrosion control. Bacteria can come from the makeup water or from the air scrubbed by the cooling water. Therefore, for successful cooling system operation, it is important to not only treat to prevent corrosion and scaling but to use biocontrol treatments to prevent pathogens such as Legionella bacteria.

Control of these bacteria is done to prevent slime deposits and corrosion that can reduce the operating efficiency of the cooling water-contacted surfaces (including the heat transfer equipment), the cooling tower and the piping. Legionella bacteria control is needed to prevent any mist or drift from the cooling tower that may spread Legionnaires’ disease.

Recently, there have been several Legionnaires’ disease outbreaks because of cooling tower water systems.[1] This has resulted in a surprising number of deaths and illnesses.[2] This resulted in a need to examine if common testing procedures provide inadequate information about the presence of these bacteria.

Microbiological Testing

The determination of both total bacteria[3] and Legionella bacteria[4] has always involved taking or dipping into the cooling tower water to determine the level of bacteria present. The Legionella bacteria levels considered as good control is essentially zero, or as limited by the test to, or less than 10 colony forming units per milliliter (cfu/mL).[5]

The total bacteria levels considered to have good control in the cooling tower water is 104 cfu/mL or less, which is 10,000 cfu/mL as established by industry and organizations such as the Cooling Technology Institute.[6]

biofilm-fouled heat exchanger

FIGURE 1. A biofilm-fouled heat exchanger is shown. Total bacteria and Legionella bacteria are found in these slime deposits.

This means that there still will be some total bacteria present in the cooling water. Incidentally, the total bacteria test does not detect Legionella bacteria or other pathogens, or even those that can cause corrosion.

biofilm-fouled cooling tower

FIGURE 2. The biofilm-fouled cooling tower at the HVAC installation profiled in the first case history is shown.

We have learned that testing of cooling-water samples for corrosive bacteria such as sulfate-reducing bacteria (SRB) does not provide accurate information of their presence. This group of bacteria can cause serious metal corrosion known as microbiologically induced corrosion (MIC).[7] It was found that water samples taken for detection of the sulfate-reducing bacteria can be zero or extremely low; while at the same time, taking a swab sample of the biofilm in the same equipment would show very high levels of this corrosive bacterium.

This resulted in changing the method of detecting these corrosive bacteria. Instead of taking water samples, we found that taking swab samples of the biofilm was much more meaningful and accurate as to the amount of this bacteria present and contributing to corrosion. We also are aware that total bacteria and Legionella bacteria are found in the slime deposits known as biofilm. So, why do we not test the biofilm to obtain a much better and more accurate information about the amount of both the total bacteria and the Legionella bacteria present in the cooling system?

Figures 1 and 2 show examples of biofilm in cooling tower water equipment.

Perhaps changing the current method from using water for testing would require certified laboratories to modify their testing for Legionella bacteria and develop a new certification or testing procedure to use for sampling biofilm. Even the test for total bacteria levels of cooling water has used the industry standard of 104 or less cfu/mL for years as good total bacteria control in cooling water systems, which should be changed. New standards should be developed for biofilm testing. Thus, we have developed a field-friendly and inexpensive method to capture the biofilm to test for total bacteria and for Legionella bacteria independently. The industry should consider and use it to develop new standards with acceptable levels.

Field-Friendly Biofilm Coupon-Testing Procedure

A field friendly biofilm-monitoring procedure was developed by the authors.[8] This procedure is a modification and expansion of that originally suggested by Lui, et al.[9] This procedure involves the use of a stainless-steel wire-mesh biofilm coupon as a substrate to capture some biofilm that would represent what is found on the surfaces of the cooling water-contacted equipment. This biofilm coupon is similar in size to typical corrosion coupons. It is placed in a corrosion-coupon rack and exposed to cooling water flowing at a rate of 3 to 5 ft/sec for 30 days.


FIGURE 3. The closeup shows how biofilm typically grows on stainless steel wire-mesh coupons.

Figure 3 provides a view of how biofilm could appear on the wire-mesh coupon. After exposure, the biofilm coupon is carefully removed and immersed in 10 mL of sterile water or sterile Butterfield’s Buffer. The biofilm is dispersed into the sterile water by a 15-sec sonication. The water containing the dispersed biofilm then is analyzed using the standard total bacteria test or sent to a certified laboratory for Legionella bacteria analysis. The results are expressed as cfu per square centimeter (cfu/cm2), which is the surface area of the wire mesh coupon. They also are expressed as cfu/mL from the sterile water containing the dislodged biofilm from the mesh coupon. Either result can be used to determine the presence of the biofilm bacteria levels.

Figure 4 is an example of the 316L stainless steel, size 80 wire-mesh biofilm coupon that is about 3 inches by 0.5 inch and is approximately 0.0625 inch thick. The stainless steel wire-mesh is an ideal surface to capture any biofilm if present in the cooling tower water system. Stainless steel is used to eliminate any corrosion deposits to interfere with biofilm. If sea or brackish water is being tested, titanium wire mesh should be used. In the first case history, which will be further described later in this article, this mesh coupon was exposed in for 30 days and has not been cleaned (as discussed in the case history).

bacteria bacteria

The levels of total bacteria and of Legionella bacteria found on the biofilm coupon should be zero or very low to ensure there is none present in the cooling tower water system. This may require much greater use of microbicides to obtain these low levels of bacteria when using the biofilm coupon procedure; however, it would provide much greater assurance of minimizing Legionella bacteria or fouling/corrosion due to total bacteria.

stainless steel wire-mesh biofilm

FIGURE 4. A stainless steel wire-mesh biofilm coupon is shown before cleaning and exposure for 30 days to cooling tower water.

Case Histories Illustrate Biofilm Testing Method

Several case histories involving field-operated cooling tower water systems were used to demonstrate the use of the biofilm coupon compared to standard microbiological water testing.

negative total bacteria

FIGURE 5. Dip slide results show the negative total bacteria counts at the Colorado HVAC cooling tower detailed in the second case history.

Case 1: California HVAC Cooling Tower. The first case history is an HVAC cooling tower water system in California that operates continuously, seven days per week, and uses city water as makeup. This cooling tower water system was chosen because it does not use any chemical water treatment that could influence the microbiological testing results. The water treatment is by a physical water treatment system[10] that has been in use for more than a year with good results for scale, corrosion and microbiological control to the end-users’ complete satisfaction.

The microbiological testing was for both total bacteria and Legionella bacteria, including water samples as well as biofilm coupons and their comparable results. Table 1 summarizes the results.

The dip slides and the laboratory tests of the water samples were analyzed for total bacteria. The slides and water sample testing results are quite low but do show some bacteria. This value is considered excellent since it is far below the industry accepted value of 104 (10,000) total bacteria.

The biofilm mesh coupon water results reported as 1,500 cfu/mL shows much higher total bacteria levels due to the biofilm compared to the water samples. Still, the data was much lower than the industry standard.

The biofilm mesh coupon results of 1.3 cfu/cm2 is also a very low level of total bacteria, but it still had the presence of biofilm. The levels of total bacteria reported as cfu/cm2 and as cfu/mL are extremely low, indicating essentially very little biofilm on the mesh coupon. This is apparent as seen in figure 2, which shows the coupon from this case study prior to cleaning. An acceptable cfu/cm2 and/or cfu/cm2 needs to be established by the industry.

The Legionella test of the cooling water sample showed none detectable. No Legionella bacteria test was performed on the mesh coupon water due to an error in communications. It will be done on future tests.

biofilm coupon

FIGURE 6. The biofilm coupon is shown (prior to cleaning) after a 30-day exposure in the Colorado HVAC cooling tower.

Case 2: Colorado HVAC Cooling Tower. The second case history is a chemically treated HVAC cooling tower water system in Denver, Colo. It operates continuously, seven days per week. It has been using a combination scale/corrosion phosphonate-based inhibitor formulation with liquid stabilized bromine and glutaraldehyde biocides for several years with good results from scale/corrosion. Test results from dip slides from this unit usually show zero for microbiological results.

Prior to the field test, microbiological monitoring by dip slides of the cooling tower water has shown zero total bacteria present on weekly testing results. This testing was continued during this case history evaluation. Figure 5 shows typical results.

Legionella bacteria testing had been taken on water samples twice yearly, but the testing had not shown any Legionella present. These results were based on the limitations of the test as performed by a certified laboratory.

It was decided to install a stainless steel wire-mesh biofilm coupon to see if there was biofilm present in the cooling tower water. The coupon was sterilized, placed into the corrosion-coupon rack and exposed for 30 days. Cooling water flow through the coupon rack was at 3 to 5 ft/sec.

Table 2 shows the weekly dip slide results and the biofilm mesh coupon results for both total bacteria and for Legionella bacteria testing. Figure 6 shows the appearance of the test coupon after a 30-day test.

This case history provided some interesting results. The dip slides showed no total bacteria present; at the same time, the total bacteria dislodged from the stainless steel wire-mesh biofilm coupon showed a very high level at 100,000 cfu/mL.

The biofilm mesh coupon reported 300 cfu/cm2 compared to 100,000 cfu/mL, which indicated that biofilm is present while the dip slides on water were negative. The tests for Legionella bacteria on both the water sample and the stainless steel wire-mesh coupon for presence in the biofilm also were negative. It would have been nice to see Legionella bacteria present in the biofilm; however, the client was pleased that none was present.


This study certainly indicates that the use of water samples for testing of total bacteria and most likely for Legionella bacteria is not providing accurate results of their presence. Water testing currently indicates that bacteria control is acceptable and under good control, when that may not be true.

The use of stainless steel wire-mesh biofilm coupons for biofilm testing of total bacteria and Legionella bacteria should be considered because more accurate results can be obtained by this low cost and field-friendly procedure. This procedure would ensure much better total bacteria control as well as greater assurance in the elimination of Legionella bacteria. Further testing needs to be done to help verify that the field-friendly procedure is acceptable and can help establish new industry acceptable standards for total bacteria and for Legionella bacteria control in cooling tower water systems.

If this biofilm testing method had been used, it is likely that those cooling tower water systems that had recent Legionnaires’ disease outbreaks may not have occurred. Using the field-friendly corrosion coupon method, the microbiological control program would have been more aggressive and effective in Legionella bacteria elimination. PC