The ugly grasp of biofilms

by jason_cramp | April 1, 2015 12:35 pm

When the jets are turned on, water containing harmful bacteria that is living in the plumbing lines is blown into the tub where bathers are soaking.

By Ted Williams

Biofilm is a common word heard in the pool and hot tub industry lately. In the last few years, there has been an increasing amount of information via studies and tests, which can be used to educate industry members and consumers on the subject. In previous articles, there is mention of biofilm existing in wet areas on pool toys and filters etc.; however, there are conclusive tests that show biofilm continues to exist in dry areas within a pool and spa/hot tub’s plumbing and equipment as well. In fact, it will lay dormant and come back to life when it gets wet again.

That said, drain-and-fill whirlpool bathtubs are one of the worst culprits, while spas/hot tubs can also be affected when a ‘wet test’ is performed after it has been taken out of storage and filled with water. While they are in storage, biofilm and bacteria grows in the interim.

This poses the question: when most consumers get a new hot tub is it really clean? It may look clean and have clean filters, but there are many areas within the hot tub that are hidden. This is just the beginning. Not only can dormant biofilm reactivate, but it will continue to harbour and colonize bacteria once it is filled with water.

What is biofilm?

According to Wikipedia, biofilm[2] is defined as: Any group of micro-organisms in which cells stick to each other on a surface is considered a biofilm. The cells are frequently embedded within a self-produced matrix of extracellular polymeric substance (EPS). Biofilm EPS, which is also referred to as slime (although not everything described as slime is a biofilm), is a polymeric conglomeration generally composed of extracellular DNA, proteins, and polysaccharides. Biofilms may form on living or non-living surfaces and can be prevalent in natural, industrial, and hospital settings.

The microbial cells growing in a biofilm are physiologically distinct from planktonic cells of the same organism, which, by contrast, are single-cells that may float
or swim in a liquid medium.

Microbes form a biofilm in response to many factors, which may include cellular recognition of specific or non-specific attachment sites on a surface, nutritional cues, or in some cases, by exposure 
of planktonic cells to sub-inhibitory concentrations of antibiotics. When a cell switches to the biofilm mode of growth, it undergoes a phenotypic shift in behaviour in which large suites of genes are differentially regulated.

Ground zero

AhhSome brochure biofilm pics[3]
When a bather fills the tub and activates the system, normal flora, dirt, sloughed skin, body fluids, bath oils and additives, fecal matter and soap scum circulate through the system and buildup inside the plumbing lines as biofilm.

AhhSome brochure biofilm pics[4]One study by a Texas A&M University microbiologist explains how whirlpool bathtubs can become a breeding ground for bacteria, many of them potential pathogens, and as such the water can be the epicentre for infectious diseases.

As part of this study, conducted by Rita B. Moyes, PhD, 43 water samples from whirlpool bathtubs—both private and residential—were subjected to bacterial analysis within 24 hours of collection. Aseptic technique was used to collect both tap and whirlpool bathtub water samples into sterile 100-ml (3.3-oz) collection containers with sodium thiosulfate (Na2S2O3) tablets for chlorine (Cl) inactivation. One container of
tap water was collected after running the water for one to two minutes and four containers of whirlpool bathtub water was collected after a clean tub was filled and the jets were engaged for two to three minutes.

Bacteriological examinations first involved nutrient agar pour plates of water dilutions to assess relative bacterial numbers. This involved passing 100 ml (3.3 oz) of water through nitrocellulose membrane filters with a 0.45-μm pore size. Then, the filters were aseptically placed on Eosin Methylene Blue (EMB) agar, Mannitol Salts Agar (MSA), Pseudomonas Fagar, Buffered Cysteine Yeast Extract (BCYE) agar, and Sabaroud Dextrose Agar. All plates were incubated for 24 to 48 hours (72 hours for BCYE plates) at 37 C (98.6 F) except for EMB agar, which was incubated at 44 C (111.2 F). By elevating the incubation temperature for this group, many of the non-
enteric coliforms are eliminated.

In comparison to the tap water samples, the findings indicated the bacterial numbers were much higher in the whirlpool bathtub samples. Further, the results were not significantly different based on the tub’s location (i.e. private versus hotel), while normal (non-jetted) bathtub water samples were not significantly different from tap water samples.

The study’s results also showed all whirlpool bathtub samples yielded microbial growth, whereas 68 per cent of the tap water samples showed no growth under the experimental conditions used in this trial. No data correlating the number of viable organisms in water with the risk of acquiring infection is currently available. However, the analysis of 100-ml (3.3-oz) filtered samples yielded too numerous to count or greater than 300 colony-forming units (CFUs) in 61 per cent of the whirlpool bathtub samples tested, indicating the bacterial load for a 100-ml (3.3-oz) sample was fairly high in a majority of cases.

AhhSome brochure biofilm pics[5]
Biofilm is a complex structure of bacteria which adheres to surfaces that are in regular contact with water. Biofilm consist of colonies of bacteria and other micro-organisms (such as yeasts, fungi, and protozoa) that secrete a mucilaginous protective coating (EPS matrix) in which they are encased.

The plated filters yielded the following results

Growth on EMB was followed by the use of Enterotube II, a commercial testing system for identification of gram negative, oxidase negative rods belonging to the enterobacteriaceae family.

Escherichia coli (E. coli), Proteus mirabilis, Yersinia pseudotuberculosis, Shigella sp, Serratia sp., and Klebsiella sp., were among the organisms identified in this group.

Gram positive, catalase positive cocci that formed yellow colonies on MSA followed by a positive rabbit plasma coagulase test confirmed the presence of Staphylococcus aureus, Pseudomonas F agar, and OxiFerm tubes, a commercial test system for gram negative, oxidase positive rods were used to identify the presence of various Pseudomonas species. Growth on Sabaroud Dextrose agar, colony morphology, and gram stain indicated the presence of fungi.

BCYE agar with polymyxin-anisomycin-vancomycin (PAV) was used for enhanced growth of presumptive Legionella species. PAV inhibits gram positive organisms, while polymyxin B inhibits many gram negative bacilli, and anisomycin suppresses yeast. On BCYE agar, Legionella pneumophila produces green colonies and Legionella micdadei produces blue colonies and suspected Legionella sp., was further tested serologically using a latex bead agglutination test.

Association of infections with whirlpool bathtubs has been recognized for a number of years, but due to the increased popularity of hydrotherapy the matter should be brought to public attention. There are several explanations for this lack of association between whirlpool bathtub use and subsequent clinical disease in the public eye. The implication of whirlpool bathtub aerosol exposure may represent 
an unrecognized source of infection in 
the medical history of a high proportion 
of clinical cases.

Whirlpool bathtubs remain popular with homeowners

A survey by the National Association of Home Builders (NAHB) shows 58 per cent of owners want a hot tub or whirlpool bathtub, with the latter being increasingly popular in hotels, hospitals, and health resorts.

Due to this popularity, not only does the public need to be educated on the possible risks associated with them, but the industry also needs to explore more effective ways to prevent the growth of biofilm in these environments through advanced cleaning methods and new design technologies.

Ideally, every time a dormant hot tub 
is filled with water it should be purged, drained, cleaned, and refilled until there is no more evidence of biofilm. Hot tubs are constructed in such a way where they
do not gravitationally drain so water can remain in lower lying pipe areas making them even more susceptible to biofilm growth. Once hot water is circulating within the hot tub, there is a new breeding ground for continued bacteria and biofilm growth.

What is lurking in the plumbing?

According to Moyes, whirlpool bathtubs are almost always a prime area for potentially harmful microbes, with the main reason being the lining of the pipes.

“They are full of inaccessible air, and water tends to get trapped in these pipes, often for long periods of time,” she explains. “When the jets are then switched on, this water, containing harmful bacteria, gets blown into the tub where bathers are soaking.”

A typical whirlpool bathtub incorporates a system of inaccessible airlines and plumbing lines. When a bather fills the tub and activates the system, normal flora, dirt, sloughed skin, body fluids, bath oils and additives, fecal matter and soap scum circulate through the system and buildup inside the plumbing lines as biofilm. Biofilm is abundant in nutrient-containing aquatic environments and, due to physiological co-operation, is inherently more resistant to various antimicrobial treatments and cleaning methods. Manufacturers recommend flushing the system with automatic dishwasher detergent, bleach, vinegar, or baking soda, but the effectiveness of these products are highly doubtful. In fact, most systems permit dirty bathwater to backfill the airlines when the pump is turned off.

Unlike plumbing lines, airlines will not admit fluid while the pump is operating. Even if industry-recommended cleaning agents were effective, they cannot reach the airlines, making it impossible to completely clean the entire system, although whirlpool bathtubs are typically drained after each use, it appears the circulation system does not fully drain. The American National Standards Institute (ANSI) has adopted a standard that permits the typical circulation system to retain more than 295 ml (10 oz) of bathwater when the whirlpool tub is fully drained. Stagnant, organic-containing bathwater trapped inside a system already rich in biofilm provides an ideal environment for bacteria to flourish. These factors combine to expose the bather to potentially pathogenic organisms.

To get an idea of how much bacteria can be in the pipelines of a whirlpool bathtub, the study shows a normal teaspoon of tap water contains an average of 138 types of bacterium, with many samples not having any bacteria at all; however, the same teaspoon of water from a whirlpool bathtub contains an average of more than 2.17 million bacterium.

As well as leading to numerous diseases (e.g. urinary tract infections, septicemia, pneumonia, and several types of skin infections), because of the aerosol mist created by the whirlpool action, microbes are forced into bather’s lungs or even open wounds. One bacterium, Legionella pneumophila, can cause Legionnaires’ disease, of which 90 per cent of all cases can be traced back to bacteria developed from a warm environment.

Moyes says studies such as this were performed in the ’70s, but the evidence collected did not show any sufficient reasons for concern as whirlpool bathtubs, or even spas/hot tubs for that matter, cannot be clearly identified as the source of infection. For instance, should someone develop 
a respiratory problem, a doctor can diagnose a respiratory infection, but he/she cannot determine how they got it. Therefore, the best way to prevent biofilm from forming is to scrape the pipes clean.

Establishing health guidelines

The five stages of biofilm development
  1. Attachment: Bacteria attaches to the surface. It wants a place to call home and grow. Bacteria want to be in relationships, so they find a nice surface to settle down and join up with a few of their closest friends.
  2. Colonization: Takes place as bacteria multiply and divide, growing in number. It is at this crucial point this attachment is ‘irreversible.’ The bacteria colony is there to stay unless purposefully removed. This stage is typically accomplished in a matter of minutes or hours at most.
  3. Protection: The bacteria colony or biofilm begins protecting itself against invasion from ‘lethal’ chemicals (e.g. chlorine or bromine), predators, or anything that wants to destroy it. In doing this, bacteria begin to excrete a protective coating called an ‘exopolysaccharide’ film. This film is sticky or slimy, and very hearty. Now the biofilm is ready to experience explosive growth.
  4. Growth: The biofilm gets bigger and tougher. Super colonies of biofilm are actually absorbing certain chemicals that were meant to destroy them.
  5. Distribution: Broken parts begin to attach to other surfaces or different parts of the same surface and the cycle begins anew.Figure_2_redraw[6]

The results of the Texas A&M University study has prompted the Public Health Laboratory Service Water Committee in the U.K. to investigate the link between the use of whirlpool bathtubs and infections further so health guidelines can be established. Likewise, the Dutch government has launched a plan to combat Legionnaires’ disease 
by implementing water safety measures after 242 cases of the disease occurred in 1999 due to the exposure to aerosolized bacteria from a whirlpool spa at the Westfriese Flower Exhibition in the Netherlands.

Due to the presence of pathogenic and potentially pathogenic organisms, public education on the hazards of piped whirlpool bathtub use should become a priority. Immunocompromised and post-operative individuals should discontinue use, and all individuals should avoid submersion of the head and possible ingestion of the water. Another concern, particularly in the hospital setting, is a whirlpool bathtub could act as a reservoir of antibiotic resistant micro-organisms. Exploration of potential preventative measures against whirlpool bathtub acquired infections should be a main focus regardless of the apparent lack of clinical evidence. New technology in design and the use of professional cleaning systems would be beneficial in reducing the risks associated with whirlpool bathtub microbial exposure.

What does this mean for hot tubs?

In this author’s opinion, due to the warm water within the plumbing lines, spa/hot tubs may in fact contain more bacteria and biofilm than a whirlpool bathtub. In this author’s 35 years in the hot tub industry, biofilm was not known as a main contributor of bacteria in water systems. Biofilm has been around forever, but it is not visible as it is hidden in pipelines and equipment. Sure, bacteria are introduced by bathers, but biofilm continues to reproduce as it clings to the pipelines like glue. Biofilm acts as a sheet of armour shielding bacteria from chemicals, allowing it to continue to colonize, breakout, and release into the body of water. This is why many hot tub owners find they have to constantly add more chemicals to sanitize the water even though they have not been using it very often. Even shocking the water with high doses of chlorine (which is bad for bather health and the environment) does not penetrate the biofilm wall and kill 
the bacteria.

By Health Canada standards there needs to be a sanitizer present to kill harmful bacteria, but what about the harmful chemicals left behind? It becomes an ‘if this, than this’ scenario. For example, if there is one part per million (ppm) of chlorine, than the water has no bacteria, which seems to be the lesser of two evils. It is great for killing bacteria, but it makes more sense to be able to obtain the same sanitation level while using less chlorine.

Test kits will measure the quality of water only from where the sample was taken in the hot tub, but bacteria is continually being fed into the hot tub through the jets from the plumbing. If one were to insert a blade into the pipes, scrape off some biofilm and add it to the water sample, the result would be different. Continuing to treat just the visible body of water is no longer the answer; one must remedy the root of the problem. This means removing biofilm so chemicals can attack the freed bacteria in a much more efficient manner.

Historically, there have been products on the market to clean hot tubs, but they only clean part way. Research has found most of them are ineffective in removing biofilm and may only remove some scale buildup. Other products have come along that focus on biofilm and water maintenance, but they are packaged with other products and chemicals making them too expensive for consumers.

Removing biofilm and reducing chemicals is an environmentally responsible move, which will also reduce health risks. If a product is too expensive, the consumer may shy away from it not ever knowing the real value it presents. Another problem may be converting customers to newer, better products as it can be time consuming and, therefore, easier to stick with the ‘if it is not broke, do not fix it’ approach. However, savvy retailers will have the foresight to understand that keeping customers happy is the number one goal. This is especially important for retailers looking to beat their competition by being the first or only retailer with a proven product in addition to having knowledge on water care. Customers need to be presented with the facts and solutions and they appreciate a supplier who is able to offer both.


Moyes, Rita B., “Microbial Loads in Whirlpool Bathtubs An Emerging Health Risk,” Department of Biology, Texas A&M University, College Station, Texas

Williams_Headshot[7]Ted Williams is president of 
Elm Sales & Equipment Inc., 
in Mississauga, Ont. Since 1980, he has been involved in all facets of the Canadian hot tub industry, including wooden and acrylic hot tub manufacturing and design, electro-mechanical and electronic controls design, manufacturing, and distribution.
 He can be reached via e-mail at[8].

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