Case Study 2 Legionella In Windscreen Wash

Case Study 2-Legionella
in windscreen wash
Group Report Summer School on Quantitative
Microbial Risk Assessment
Frantisek Kozisek, Heleen de Man, Philipp Amoah, Asli Kisikkaya, Signe, Tanja
Andersen, Nadine Czekalski
Case Study 2 Legionella in windscreen wash
1. Introduction
Legionella is a gram negative bacterium recognized as a pathogen more than 30 years ago. Since that
time a lot of information on its behavior, human exposure routes, health risks, monitoring and
management appeared, but some aspects of human health risk are still unclear. While analysis of the
outbreaks of Legionnaires' disease helped to reveal most important exposure routes (inhalation of
aerosol contaminated by Legionella produced in showers, water parks or air-conditioning systems
etc.) source of infection for sporadic cases of Legionnaires' disease is rarely identified.
It was found that professional drivers were at the highest risk of Legionnaires' disease among various
occupations (relative risk = 5.90; 95%CI: 5.04-9.60; P < 0.0001 for UK drivers in 2001-2006), but as
these were mostly sporadic cases, the source of infection and exposure route remained unknown.
New paper trying to elucidate this “mysterious” exposure pathway was recently published
(Wallensten, 2010) which attracted attention of British press. The paper based on study done in the
United Kingdom identified as possible source of infection windscreen wiper fluid without added
screenwash. Drivers or passengers of vehicles that use tap water instead of commercial screenwash
in the windscreen wiper fluid had an almost 50 times higher risk of Legionnaires' disease.
This study tries to describe in detail possible exposure routes and assess the risk related to this
source of infection as well as to suggest some risk reduction measures in a worst case scenario,
describing the risk for professional drivers.
2. Hazard identification
2.1 System under evaluation
Commercially produced screenwash contain various substances (usually propan-2-ol 10-30% and
anionic surfactant 1-5%) which improve cleaning qualities of wiper fluid and at the same time inhibit
growth of bacteria. If just plain water without addition of any disinfectant or inhibitor is used, such
water (= wiper fluid) may meet all conditions which have been recognized to promote Legionella
growth1: temperature between 20 – 50 °C (valid for most of the year if the car is parked in garage or
at least part of the year for other cars and every time when car is being used); water stagnation (days
or weeks for wiper fluid); materials supporting microbial growth (plastic reservoir); insufficient
According to the EWGLI (European Working Group for Legionella Infections).
Case Study 2 Legionella in windscreen wash
sanitation of the system (presence of biofilms, no disinfection residual);´nutritional potential of water
(tap water has usually sufficient AOC/BDOC levels not to prevent Legionella growth); pH in range of
2,0 – 8,5. It is therefore reasonable to expect Legionella growth in the wiper fluid reservoir, but no
data on concentration are available so far. The study of Wallensten only informs about unpublished
pilot study carried out by UK HPA which isolated Legionella bacteria from windscreen wiper fluid of
one car out of five which did not use screenwash, while none were identified from 16 cars that did
use screenwash. Concentration of Legionella in wiper fluid (of positive cars) may range from 1 to 106
CFU/l, based on paper by Exner et al. (1993), who found that more than 50 % of large German
buildings investigated had heavily contaminated hot water systems with Legionella (i.e. ≥ 10.000
CFU/ l), and other papers who report maximum concentration up to 106 or even 109 CFU/l in spa
water (Armstrong, 2007).
When wiper fluid is sprayed on windscreen, aerosol is created and part of it is sucked into the
passenger compartment through ventilation intake or through open windows. The study of
Wallensten et al. is based on case control study including all surviving community acquired sporadic
cases of Legionnaires' disease in England and Wales with onset between 12.7.2008 and 9.3.2009.
Cases were contacted by phone and controls were consecutively recruited by sequential digital
dialing matched by area code, sex and age group. Those who consented were sent a questionnaire
asking questions on driving habits, potential sources in vehicles and known risk factors, this resulted
in 75 cases and 67 controls to be included in the study. The risk factors identified were (single
variable analysis):
Not use of screenwash in wiper fluid (odds ratio /OR/ 22.06)
More than two hours driving with open window per day (OR 19,82)
Number of hours in vehicle for job a day 2-4(OR 5,5), 4-6(OR 10,5), more than 8(OR 15,0)
Smoking (OR 11.86)
None showers taken at home (OR 6.37)
Always driving with open window (OR 6.04)
Use of van (OR 4.42)
Year of manufacturing is before 2000 (OR 3.27)
Driving through industrial areas (OR 2.25)
After multiple variable analyses (adjustment for age, sex, season and smoking) the main risk factor
proved to be significant was not using screenwash in wiper fluid (OR 47.24).
Case Study 2 Legionella in windscreen wash
2.2 Characteristics of Legionella pneumophila
Legionella are gram negative, aerobic, motile and non-spore forming rods (width: 0.3-0.9 μm, length:
2-20 μm, Dennis, 1990) commonly found in surface waters (Muraca et al. 1988) and have also been
detected in cooling towers (Garbe et al., 1985), air conditions (Levy et al., 1980), shower heads
(Cordes et al., 1981) and drinking water (Hsu et al., 1984, Tison and Seidler, 1983). In these
environments Legionella mostly parasitize in different protozoa, especially amoeba. When growing
extracellular they are often associated with microbial biofilms but can also occur freely in the water
column. Legionella are temperature sensitive organisms, not growing below 20°C. They prefer hot
stagnant waters (between 25-46°C) or elastomeric surfaces (Fields, 2007) and reach their highest
growth rates and also highest virulence at 35-37°C (Mauchline et al., 1994, Fliermans et al. 1981).
Further, Legionella need neutral pH, soluble iron and the amino acid l-cysteine for growth(Ellis 1993).
Under optimal conditions extracellular Legionella can double in number in eight hours (Cooling
Tower Institute 1990). Intracellular growth may differ somewhat, e.g. Holden et al. (1984) reported
an increase of Legionella in Acanthamoeba castellanii by 3 to 4 orders of magnitude in 48 to 72
hours. Factors that inhibit growth of Legionella are Na-ions (no growth in seawater), surfaces of
copper or stainless steel as well as low temperatures (Fields, 2007). Chlorine compounds (Chlor
dioxide, Monochloramine), H2O2 and copper-silver ionization can be used for chemical disinfection
as well as UV and ultra sonic as physical measures.
2.3 Health outcome
Legionella contains at least 40 species (Samrakandi et al. 2002) of which 21 is possibly associated
with human disease (Fang et al. 1989). Legionella were firstly linked to human disease after the
bacterial species L. pneumophila was identified as the cause of an atypical pneumonia among
persons at an American Legion convention in Philadelphia in 1976 (Fraser et al. 1977). L.
pneumophila is the most prevalent species associated with Legionnaires disease, causing
approximately 90% of the diagnosed cases of Legionellosis (Yu et al. 2002). The species L.
pneumophilia has multiple serotypes, and different strains demonstrate differing virulence
(Fitzgeorge et al. 1983). L. pneumophila are classified as intracellular pathogens, due to their ability
to survive and multiply in human monocytes/macrophages (Swanson and Hammer 2000).
Legionnaires’ disease is an atypical pneumonia caused by Legionella, and is most commonly caused
by the species L. pneumophila. The term Legionellosis includes Legionnaires’ disease and a milder
illness called Pontiac Fever.
Case Study 2 Legionella in windscreen wash
Legionnaires’ disease was first identified in 1976 and continues to be a significant cause of human
morbidity and mortality. It occurs mainly in the elderly or infirm (Millar 1997) but can also develop in
young and healthy individuals (European Working Group on Legionella Infections, 2005). The most
susceptible groups are: males (75% of all cases), smokers, immuno suppressed, people with chronic
diseases (typically lung disease) and professional drivers. The reported incidence rates of
Legionnaires’ disease in the general population from various countries are typically in the range of 3
to 16 per 1,000,000 per year (Cameron et al. 1991; Lim et al. 2003). Legionnaires' disease has an
incubation period commonly cited as of 2 to 10, , days from exposure to the appearance of clinical
symptoms, but an incubation period up to 19 days has been reported (Boer et al. 2002). Symptoms of
Legionnaires disease include: a high fever, chills, headache, muscular pains, dry cough, difficulty
breathing and diarrhea (Ellis 1993), but not all symptoms occur in all cases. Most Legionellosis cases
arise sporadically, meaning they are not linked to a common source of the infection and fewer, some
are clustered and linked to a common (but not always identified) source of infection (Joly, 1993). The
fatality rate for Legionella Disease ranges from 5-30% during various outbreaks and is less than 5% if
therapy is started quickly.
The less severe disease Pontiac Fever is a non-pneumonic influenza-like disease with a short
incubation period (2 to 72 hours), and illness duration 2 to 5 days, with symptoms including
headache, nausea, vomiting, aching muscles, malaise, and cough (Ellis 1993). In contrast to
Legionnaires' disease, no fatalities have been reported for Pontiac Fever (Ellis 1993). It has been
conjectured to be due to avirulent strains of Legionella (Fields et al. 1990), or to endotoxins from
non-viable organisms (Fields et al. 2001).
2.4 Exposure route
Inhalation of airborne droplets or droplet nuclei containing Legionella is generally thought to be the
commonest mode of transmission. The aerosols may be generated by mechanical devices (e.g.
cooling towers of air-conditioning systems) or by the use of potable water, especially from domestic
hot-water installations (e.g. showers) (Breiman et al., 1990). Aerosol formation is deemed necessary
to cause pneumonic disease, but in some cases aspiration following ingestion of contaminated water,
ice, and food has also been implicated as the route of infection (Marrie et al., 1991; Blatt et al., 1993;
Venezia et al., 1994). Even when it is possible to demonstrate that the disease strain and the strain
colonizing a plumbing system are identical, the exact route of transmission sometimes remains a
Case Study 2 Legionella in windscreen wash
matter of speculation. For L. pneumophila there is no evidence of person-to-person transmission
(Fraser, 1977; Yu, 1983).
3. Analysis
3.1 Exposure assessment
All assumptions in this case are based on the driving habitats of professional drivers (driving app. 10
hours per day) which are assumed to be the worst case scenarios. Additionally the model is only for
vehicles with no open window and no air-condition turned on. In Table 1 it is seen that 4% of the
professional drivers has a chance of having Legionella in their windscreen fluid, based on the
knowledge that 20% uses tap water as wiper fluid and 20% of this tap water contains Legionella (
Wallensten et al, 2010). In the tap water with Legionella it is estimated that a concentration of 1-106
cfu/l of Legionella is present as described in the hazard identification (Exner et al., 1993).
To examine the frequency of screenwash use on yearly basis, a survey was done, of which
nationality, driving hours per day, age and wiping frequency was taken into account. Based on the
results for the “ordinary” driver, the wiping frequency of a professional driver was multiplied with
five, since an ordinary driver drives 1-2 hours per day, and it was assumed that a professional driver
drives 8 hours on work plus 1-2 hours off work. Based on this the screenwash use was estimated to
be 6 times per day.
No data was found on the volume of screenwash used per wiping, therefore a small experiment was
conducted in the summer school in delft 2010, with one car of which is was found to be
approximately 30 ml, however this is highly dependent on the vehicle type, hence the assumption
was a volume in the range of 20-50 ml of fluid per wipe. Of this water it was estimated that; 10% is
aerosolized with a range between 5-15% and of these aerosols it is assumed that 66% is reaching the
vehicle with a range of 8-104% (Luke et al).
In the article of Armstrong and Haas (2007) the dilution coefficient from water to air is given to be on
average: 2,3x10-5 cfu/m3 per cfu/l which was included in the model.
Allan et al. (year?) describes the inhalation rate for people to be 0.0167 m3 per hour. Since no
quantitative experimental data was available for the duration of wiper fluid in the car with active
Legionella is available the assumption that the duration of inhalation is approximately 60 seconds is
Case Study 2 Legionella in windscreen wash
Further variability is the volume of the vehicle, which varies depending on the type of vehicle that is
driven (bus, car, limousine etc.). Therefore it is estimated that the volume of the car on average is 25
m3 and minimum 3 m3 and maximum 50 m3 (Luke et al., year?).
Table 1. Uncertainties and variability’s of used to create the model in Crystal Ball.
Chance to get Legionella in screenwash wiperfluid in the car
Concentration Legionella in wiper fluid (if question above is yes)
Wiping frequency
volume needed to clean window
% of aerolisation
% of outside to inside
dilution coefficient water to air
inhalation rate
duration of inhalation wiper fluid aerosols
Volume of the car
Value Average St. Minimum Maximum Distribution
1.E+06 Triangular
6 1
0.05 Uniform
15% Triangular
104% Triangular
1.60E-05 3.10E-05 Triangular
0.0556 Triangular
200 Triangular
50 Triangular
3.2 Dose response
The dose-response relation is found in literature of Armstrong and Haas (2007) where they evaluate
a quantitative microbial risk model for Legionnaires’ disease for human exposures for selected spa
outbreaks. For these outbreaks they have estimated the air concentrations near the whirlpool spa to
be five to 18 colony forming units per cubic meter (CFU/m3) and 50 to 180 CFU/m3 for each of the
alternate assumptions. The estimated 95th percentile ranges of Legionella dose for workers within
15 m of the whirlpool spa were 0.13-3.4 CFU and 1.3-34.5 CFU, respectively. The modeling for hot
springs Spas 1 and 2 resulted in estimated arithmetic mean air concentrations of 360 and 17 CFU/m3,
respectively, and 95 percentile ranges for Legionella dose of 28 to 67 CFU and 1.1 to 3.7 CFU,
respectively. There are limited reports on air concentrations of Legionella therefore it is assumed
that the model parameter values of Muller et al. (1983) can be used as a dose-response relation,
that is: the probability of risk of infection= 1-e-rd, r= 0,06, d= dose (cfu/day). The exponential model
was chosen, since Muller et al. (1983) found no difference between the distributions examined.
3.3 Sensitivity analysis
The most sensitive parameter is the chance of getting Legionella in the wiper fluid when plain water
is used as wiper fluid and the concentration of Legionella in wiper fluid, the other assumptions
contributed with less sensitivity.
Based on the sensitivity analysis the data need identified is: the concentration of Legionella in tap
water, the percentage of aerosolization, the actual percentage of cars in which growth actually do
occur, more about the type of ventilation systems in cars.
Case Study 2 Legionella in windscreen wash
4. Risk characterization
Based on the simulation with 10.000 trials, the mean probability of infection relating to the exposure
assumptions was found to be 2.04*10^-7, with a total number of infections per year to be;
Mean probability of infections*population density*drivers in the population (assumed)*year giving
the result: 2.04*10^-7*61000000*0.02*365 = 90.8412
(is the risk acceptable or not? Should have been compared to the literature)
We supposed triangular distribution of data defining concentration of Legionella in wiper fluid, but
this assumption is not supported by any data – just because such data are missing. The same is valid
for our estimates of most likelihood and maximum values, which we deduced from published results
from hot water systems (in buildings) and hot spas (maximum) and information that densities above
104 to 105 CFU/liter represent a potential increased threat to human health (Joly, 2006; etc.). On the
other hand it was surprising for us that under such ideal growth supporting conditions the Legionella
was found only in one out of five cars not using screenwash. Specially this information needs further
Information on proportion of aerosol (forming outside a car) which can get in the cabin, was deduced
from Australian study investigating effect of cabin ventilation rate on ultrafine particle (UFP)
exposure inside automobiles based on simultaneous monitoring UFPs outside and inside the cars
(Knibbs, 2010). The group of five different cars going different speed and with different ventilation
regime in the busy tunnel was included in the study. Significant variability in average median incabin/on-road (I/O) UFP ratios was observed (0.08 to about 1.0), strongly related to their choice of
ventilation setting, vehicle type and speed of the vehicle. Beside substantial variability of the factor
itself, there is uncertainty about our extrapolation, because concentration and distribution of UFPs in
the tunnel was rather homogenous over time, but concentration of Legionellas in aerosol “cloud”,
once created (in few seconds), is rapidly decreasing due to immediate start of aerosol dilution. We
also did not take into account the worst case scenario of driving with window open which seems to
increase the risk according to the study of Wallensten et al. (2010).
Assumptions on breath rate (Alan, 1998) and volume of fluid sprayed within one cleaning run
(verified by personal experiments of study group member and relative) may be considered as reliable
and also with acceptable variability. The simulation (Alan, 1998) suggested that most age groups’ 24hour inhalation rates can be represented with log-normal probability density functions. Arithmetic
mean values and standard deviations for these distributions are as follows: approximately 9.3 ± 2.4
Case Study 2 Legionella in windscreen wash
m3/day for toddlers; 14.6 ± 3.0 m3/day for children; 15.8 ± 3.7 m3/day for teenagers; 16.2 ± 3.8
m3/day for adults; and 14.2 ± 3.3 m3/day for seniors.
4.2 Risk management
Wallensten et al. (2010) estimated that around 20 % of community acquired sporadic cases could be
attributed to this way of exposure, which represents about 70 annual cases for UK. Even if number of
cases would be lower, management of this risk is justifiable as seems to be easily preventable. Since
the risk management is based on the balance between the safety and costs of the consumers the
need of risk management is low and should therefore be of low costs. Alternatively other suitable
chemicals/biocides for plain water used as windscreen fluid or cheaper alternatives to windscreen
fluid with screenwash should be produced. Obviously not all known and widely available biocides
could be used and therefore it is questionable if suitable alternative would be really cheaper then
screenwash concentrate and such research is worthwhile.
The risk should be communicated to the professional drivers educate all drivers not to use just plain
water as wiper fluid, but fluid with screenwash either already pre-diluted or prepared by driver from
screenwash concentrate, either by folders handed out by the employers from the working place or
by the learning books used when taking a driver license. Another solution would be to add
information about the proper windscreen fluid use in the car manual, add a warning sign on the
wiper fluid reservoir in the car, or create campaigns at gas stations etc.
In the meantime the communication of this risk could be quite easy to explain: If you want to prevent
health problems with Legionella, use screenwash in your windscreen wiper fluid.
5. Conclusion
In this case many assumptions were made, since there is a major lack of experimental data, however,
the conclusion is still clear; it is better to use commercially screenwash rather than plain water,
because the commercial screenwash also include a disinfectant so Legionella cannot grow in the
reservoir of wiper fluid. Nevertheless, even so plain water is used as windscreen wiper fluid, the risk
of a response according to the estimated dose is very low and therefore the chances of getting
Legionnaires disease is minimal, even in the worst case scenario where you are a professional driver.
Based on this it is concluded that the risk management should be minimal and cheap.
Case Study 2 Legionella in windscreen wash
6. References
Allan M, Richardson GM. (1998). Probability Density Functions Describing 24-Hour Inhalation Rates
For Use in Human Health Risk Assessments. Human and Ecological Risk Assessment: An International
Journal, 4: 2, 379 — 408.
Armstrong TW, Haas CN (2007). “Quantitative Microbial Risk Assessment Model for Legionnaires'
Disease: Assessment of Human Exposures for Selected Spa Outbreaks. J Occup Environ Hyg, 4: 8, 634646.
Cooling Tower Institute. (1990). "Legionnaires' Disease Bacteria." CTI Journal, 11(1), 70-71.
Dennis, P. J. (1990). "Reducing the risk of Legionnaires' disease." Ann.Occup.Hyg., 34, 189-193.
D L Tison and R J Seidler. 1983. Legionella incidence and density in potable drinking water supplies.
Appl Environ Microbiol. 1983 January; 45(1): 337-339.
Ellis, K. V. (1993). "Legionellosis: A Concise Review." J Inst Water and Envir Management, 7, 416-430.
Exner M, Tuschewitzki GJ, Langer B, Wernicke F, Pleischl S. (1993). [Incidence and evaluation of
Legionella in hospitals and other large buildings] (in German). Schriftenr Ver Wasser Boden Lufthyg.
Hoffmann, P., Bendinelli, M., Friedmann, H., Fields, BS. (2007).“Legionella pneumophila
Pathogenesis: Lessons learned from Genomics“.Springer US.85-94
Holden, E. P., Winkler, H. H., Wood, D. O., and Leinbach, E. D. (1984). "Intracellular growth of
Legionella pneumophila within Acanthamoeba castellanii Neff." Infect.Immun., 45, 18-24.
Hsu SC, Martin R, Wentworth BB. (1984). “Isolation of Legionella species from drinking water”. Appl
Environ Microbiol. 1984 October; 48(4): 830-832
Joly P, Falconnet P-A, André J et al (2006). Quantitative Real-Time Legionella PCR for
Environmental Water Samples: Data Interpretation. App Environ Microbiol, 72(4): 2801-2808.
Knibbs LD, de Dear RJ, Morawska L. (2010). Effect of Cabin Ventilation Rate on Ultrafine Particle
Exposure Inside Automobiles. Environ. Sci. Technol., 44 (9): 3546–3551.
SHANDS, M.D.; and DAVID W. FRASER, M.D. 1981. “Isolation of Legionella pneumophila from Hospital
Shower Heads”. Ann Intern Med February 1,94:195-197;
Muller, D., Edwards, M.L., & Smith, D.W. 1983. Changes in iron and transferring levels and body
temperature in experiomental airborne legionellosis.
Case Study 2 Legionella in windscreen wash
Muraca, P., Yu, V., and Stout, J. (1988). "Environmental Aspects of Legionnaires' Disease." J
AWWA(February), 78-86.
Paul L. Garbe, DVM; Barry J. Davis, MSEHE; Jay S. Weisfeld, MD; Lauri Markowitz, MD; Patricia Miner,
MPH; Frank Garrity, MD; James M. Barbaree, PhD; Arthur L. Reingold, MD. 1985. “Epidemiologic
Demonstration of Cooling Towers as a Source”. JAMA. 254(4). p521-524.
Wallensten A, Oliver I, Ricketts K, Kafatos G, Stuart JM, Joseph C (2010). Windscreen wiper fluid without added
screenwash in motor vehicles: a newly identified risk factor for Legionnaires' disease. Eur J Epidemiol :DOI
W S Mauchline, B W James, R B Fitzgeorge, P J Dennis, and C W Keevil . 1994.“Growth temperature reversibly
modulates the virulence of Legionella pneumophila”. Infect Immun. 62(7): 2995–2997.
Case Study 2 Legionella in windscreen wash
Conceptual flow scheme of the risk assessment
Inoculum concentration
Optimal growth conditions
Tapwater is used as windscreen wiper fluid by 20 % of
the population instead of screenwash. The distribution
of Legionella pneumophila in the tapwater is:
0 CFU/L in 80 % of tapwater used
1 to 106 CFU/L in 20 % of tapwater used
All the optimal growth conditions can be met in
car wiper fluid
Transmission route
Host characteristics
From windscreen wiper fluid to air trough windscreen
wiping generating aerosols inhaled by driver. The
contaminated air is transferred into the car through:
Ventilation systems
Open windows
Other routes
Stagnant water
Temperature 35°C
Nutrients supporting growth
pH 2-8.5
Males (app. 75%)
Elderly (mean age 45 years)
Immuno suppressed
Chronic disease (typically lung disease)
Professional drivers, aged 18-80 year
Legionellosis: Mortality rate 5-30%
Frequency and volume of inhalation
Initial concentration in wiper fluid
Duration time of the pathogen
Air volume in car (car model dependant)
Dilution factor (water to air, 1/10 assumed)
Frequency of exposure event; frequency of
windscreen wiping (seasonal) and time spent in
car (ordinary or professional driver)
Legionnaire’s disease; a severe illness
causing pneumonia
Pontiac fever; a less severe respiratory
Risk characterization
Combine data from exposure and effect
assessment into characterization of risk
Risk management
Inform population about risk by media
Cheaper alternative to screenwash
Add ins of biocide to water
Variability and sensitivity analysis
Identify if outcome answers the problem
Identify data needs
Define/estimate risk for defined groups
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