legeonella in cooling tower

8/13/2019 Legeonella in Cooling Tower

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700 2006 ASHRAE.

ABSTRACT

Chemical biocide control in cooling towers is a critical

part of the overall prevention strategy against Legionellaand

Legionnaires disease. The purpose of this study was to analyzea large population of cooling towers (2,590) for the presence

and levels of Legionella and to correlate these data with

specific biocide treatments (eight different biocides in 28

different combinations). On average, Legionellawas found to

be present in 13% of the cooling towers, and 18% of these posi-

tive towers had high levels of Legionella(> 1,000 cfu/ml; 2%

of the total towers). No biocide combinations were 100% effec-

tive in preventing Legionellacolonization, but none always

failed. Oxidizing biocide (primarily bromine) combinations

with nonoxidizers were not significantly better than two nonox-

idizers and in many cases (e.g., bromine alone, bromine plus

DBNPA) were significantly worse. The nonoxidizing biocide

THPS was particularly effective in all combinations, as werethe combinations of carbamate/isothiazoline and quat/

bromine. The quat/carbamate towers, on the other hand, had

a significantly higher prevalence of Legionella. Monitoring

Legionellalevels in cooling towers may assist in determining

whether a biocide regimen is effective for a particular system.

INTRODUCTION

Legionnaires disease is a bacterial pneumonia caused by

Legionella pneumophilaand related species. The disease is

transmitted to humans via aerosols from building water

supplies. Depending on the source, the exposure may be

through indoor or outdoor routes of transmission, often lead-ing to multiple-case outbreaks of the disease. Building water

sources with a significant risk for transmission include water

cooling towers and evaporative condensers.

Thus, in the years following the identification of Legion-

naires disease and its transmission from its environment, the

maintenance of cooling towers acquired an entirely new focus.

Historically, chemical biocides were added to cooling tower

water in order to control biofouling, which could lead to

performance problems, such as slime and microbial corrosion

problems in the tower structure, heat exchanger, and plumb-

ing. With the new concern aboutLegionellaand Legionnaires

disease, the control of this specific bacterium in cooling

towers became a public health issue.

As a result of the Legionnaires disease problem, a rapid

search was begun to identify existing chemical biocides that

might be effective against this bacterium. Initially, a large

number of publications appeared from studies performed in

the laboratory using laboratory-grown Legionella, tested

against virtually every biocide approved for use in cooling

towers (Skaliy et al. 1980; Grace et al. 1981; Soracco and Pope1983; Soracco et al. 1983; Dominque et al. 1988; McCoy et al.

1986; McCoy and Wireman 1989). While many of the

biocides showed promising effectiveness against Legionella

in the laboratory, these results did not always correlate well

with field trial data when the biocides were tested in actual

cooling towers (England et al. 1982; Kurtz et al. 1982; Flier-

mans and Harvey1984; Broadbent et al. 1991; Yamamoto et

al. 1991; Pope and Dziewulski 1992; Elsmore 1993; Watanabe

1994; Bentham and Broadbent 1995; Broadbent 1999; Ganzer

2003). An excellent review of disinfectants andLegionellacan

be found in Kim et al. (2002).

Part of the explanation for these results came with the

increased understanding of the intracellular nature ofLegionella in protozoa, the overall biofilm polymicrobial

milieu that exists in nature, and the difficulty in treating

Legionellain this biofilm environment with selected biocides

LegionellaPrevalence in Cooling Towers:Association with Specific Biocide

Treatments

Richard D. Miller, PhD D. Anne Koebel

Richard D. Milleris associate professor at the Department of Microbiology and Immunology, School of Medicine, University of Louisville,

Ken. D. Anne Koebelis laboratory manager at Environmental Safety Technologies, Inc., Louisville, Ken.

CH-06-12-2

2006, American Society of Heating, Refrigerating and Air-Conditioning Engineers, Inc.

(www.ashrae.org). Reprinted by permission fromASHRAE Transactions, Volume 112, Part 1.

For personal use only. Additional distribution in either paper or digital form is not permitted

without ASHRAEs permission.


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ASHRAE Transactions: Symposia 701

(Coughlin and Caplan 1987; Wright et al. 1991; Green and

Pirrie 1993; Walker et al. 1994; McCall et al. 1999; Goa et al.

2001). In order to help clarify this, a research study was funded

by ASHRAE to systematically test various biocides in an

accurate, scaled-down model cooling tower system. The

results of this study (Thomas et al. 1999), while not

completely comprehensive, did conclude that oxidizing

biocides (chlorine, bromine, and ozone) better controlled

Legionellathan did the tested nonoxidizers, a result that had

been observed in another model system (McCall et al. 1999)

at about the same time.

In the real world, much cooling tower treatment is carried

out using two different biocides in order to prevent buildup of

resistant bacteria (includingLegionella). Based on the existing

information, there is a developing consensus, as noted in

OSHAs Technical Manual (OSHA 1999), that control of

Legionella in cooling towers should include an oxidizing

biocide, preferably alternated with a nonoxidizer. Specifically

mentioned in the OSHA document as effective is the oxidizer

Towerbrom 60M, especially when alternated with bromo-

chloro-dimethyl-hydantoin (BCD). On the negative side,

quaternary ammonium compounds were specifically

mentioned as not effective. The legionellosis control guide-

lines from the Cooling Technology Institute (CTI 2000) also

recommend using an oxidizing biocide as part ofLegionella

control in cooling towers.

Despite these rather specific recommendations, a wide

variety of biocides (oxidizers and nonoxidizers) continue to be

used in cooling towers, with apparent success based on the

anecdotal reports of Legionella culture results. Thus, the

purpose of the current study was to examine a large population

of cooling towers (2,590 samples) throughout the US and to

correlate the presence and numbers of Legionella pneumo-phila in each tower with the 28 different biocide treatment

combinations of eight different biocides. These results are

intended to help clarify previous biocide research and to shed

new light on effective treatment strategies againstLegionella.

METHODS

Cooling Tower Samples

All samples used in this study were obtained on a volun-

teer basis from 1998 to 2004 from approximately 1,000 differ-

ent cooling towers located throughout the United States. A

total of 2,590 water samples were collected from these towers

as part of this study by a large number of different chemicaltreatment company representatives. Typically, repeat towers

were not sampled more than twice during any year. Water

samples of 500 ml were collected in clean containers by the

chemical treatment company representatives on site and

shipped unrefrigerated to our laboratory for analysis via over-

night express delivery. Information on the specific biocides

used in the tower was included with each sample. Sampling

recommendations for the study stated that samples were to be

taken immediately prior to the addition of any slug-fed

biocides (i.e., when the biocide control would be at its lowest).

Biocides

Chemical treatment company personnel added all

biocides to the cooling towers as part of the normal mainte-

nance of these towers. The choice of biocides was made

entirely by these personnel, while the dosages (i.e., the

concentrations) of biocides to be used were administered

according to manufacturer/supplier recommendations. Infor-mation sent with each sample included the biocides used in the

tower as well as the manner and frequency of addition. For

purposes of data analysis, the biocides were grouped into eight

categories: (1) bromine, (2) quaternary ammonium

compounds (quats) including polyquats, (3) carbamate, (4)

isothiazoline, (5) glutaraldehyde, (6) hydroperoxide, (7) 2-2-

dibromo-3-nitrilopropionamide (DBNPA), and (8)

tetrakis(hydroxymethyl)phosphonium sulfate (THPS). When

used, bromine and DBNPA were added in a manner to achieve

continuous levels (via tablets), while all other biocides were

added weekly (usually automatically via timer) as a slug-fed

liquid. When two slug-fed biocides were used, they were

always alternated so that each was added weekly but three to

four days apart. Biocides from the eight categories were used

alone or in combination with another biocide, resulting in a

total of 28 different combinations.

Isolation and Quantitation of LegionellaSpecies

Legionellawere isolated using a modification of the low-

pH treatment and selective media protocol as described by the

Centers for Disease Control (Barbaree et al. 1992). Briefly, a

1 ml portion of the sample was acidified to pH 2.2 by addition

of 1 ml of 0.2 M HCl-KCl buffer. After a 5 min period at room

temperature, the sample was neutralized by addition of 1 ml of

0.1 N KOH. Portions (10:1 and 100:l) from the acid-treatedsample were spread-plated onto both buffered charcoal-yeast

extract (BCYE) agar and glycine-vancomycin-polymyxin

(GVP) selective agar medium.1In addition, 10-1, 10-2, 10-3,

and 10-4dilutions of the original sample were also plated on

the same two media. All incubations were at 35C. After three

to five days of incubation, typical Legionellacolonies were

counted and their identification confirmed by lack of growth

on BCYE agar media without L-cysteine as well as immunof-

luorescence microscopy using a monoclonal antibody reagent

specific forL. pneumophila.2No additional serogrouping of

the L. pneumophila isolates was performed. The levels of

culturableL. pneumophilawere expressed as cfu/ml of origi-

nal sample, with a level of sensitivity of 10 cfu/ml.

Statistical Evaluation

A control group in this study without biocide treatment

was not possible because of ethical limitations. Thus, for each

biocide and biocide combination, estimates of the prevalence

1. Both BBL Prepared Media, Becton Dickinson and Company,Cockyesville, MD.

2. Genetic Systems Corp., Redmond, WA.


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702 ASHRAE Transactions: Symposia

ofLegionellacolonization and highLegionella (> 1,000/ml)

colonization were calculated. Exact binomial 95% confidence

intervals were generated for each of the 28 treatments and

compared to the overall prevalence estimates. Treatments with

a 95% confidence interval that did not overlap with the overall

Legionellaprevalence 95% confidence interval (131) or the

overall high Legionellaprevalence 95% confidence interval

(21) were considered significantly different.

RESULTS

Prevalence of Legionella

in Cooling Towers Using Bromine

Bromine was the most common oxidizing biocide used in

this study (572 towers). The effectiveness of bromine in

controlling the prevalence of Legionella in towers varied

considerably according to the specific biocide combination, as

illustrated in Table 1. For example, the combination of

bromine/quat had a very lowLegionellaprevalence, with only

4% of the 135 towers colonized with Legionella, which wasstatistically well below the 13% average ofLegionella in the

entire population of towers (341/2590; 95% confidence inter-

val of 1%). Bromine/carbamate (11%) and bromine/isothia-

zoline (16%) were close to average, while bromine alone and

bromine/DBNPA were significantly less effectivethat is,

well above average (23% and 42%, respectively)in terms of

Legionellacolonization. Bromine/glutaraldehyde also had a

high Legionellaprevalence (20%) compared to the average,

but it did not quite achieve a statistical difference. Bromine/

THPS was actually the best combination (0% colonization)

although the number of towers was too small to achieve signif-

icance. Overall, the effectiveness of bromine as a biocide cate-

gory tended toward being worse than the average for all

towers, with a prevalence rate of 89/572 (163%), although

there was a slight overlap in the 95% confidence intervals.

When the data were reexamined to detect the prevalence

of high levels (> 1,000/ml) ofLegionellain the cooling towers,

the results for biocide combination effectiveness were similarto the overall prevalence discussed above, despite the small

number of positive towers precluding significance with any of

the biocide treatments. An exception was that no towers with

highLegionellawere found in the bromine/DBNPA category

(0%), despite the overall high Legionella prevalence rate

(42%) mentioned above. Overall, bromine as a biocide cate-

gory had an average prevalence of highLegionella.


in Cooling Towers Using Quats

The effectiveness of quats in controlling the prevalence of

Legionellain the towers also varied considerably, according to

the specific biocide combination (Table 2). The significanteffectiveness of the combination of quats/bromine (4%) was

discussed above. The combinations of quats/isothiazoline

(4%) and quats/hydroperoxide (2%) tended toward effective-

ness but were used in too few towers for the results to achieve

significance. In contrast, the combination of quats/carbamate

(23%) was significantly less effective and well above average

in terms ofLegionellacolonization. Towers with quats/glut-

araldehyde (16%) and quats alone (20%) tended toward

above-average prevalence but did not achieve significance.

Overall, the effectiveness of quats as a biocide category was

statistically within the range of average, with a prevalence

rate of 68/484 (14%).

Table 1. Prevalence of Legionella in Cooling TowersUsing Bromine Alone or in Combination

with Another Biocide (N=572)

Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

Bromine alone 30/130 (237) 6/130 (53)

+ quat 5/135 (43) 1/135 (14)

+ carbamate 4/36 (118) 1/36 (312)

+ isothiazoline 21/128 (166) 3/128 (25)+ glutaraldehyde 21/103 (207) 7/103 (74)

+ DBNPA 8/19 (4222) 0/19 (018)

+ THPS 0/21 (016) 0/21 (016)

Total bromine 89/572 (163) 18/572 (31)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for high Legionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower than theaverage (21%).395% confidence intervals.

Table 2. Prevalence of Legionella in Cooling Towers

Using Quaternary Ammonium (Quat) Biocides Alone

or in Combination with Another Biocide (N=484)

Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

Quats alone 4/20 (2014) 0/20 (017)

+ bromine 5/135 (43) 1/135 (14)

+ carbamate 54/239 (236) 7/239 (32)+ isothiazoline 1/26 (416) 0/28 (012)

+ glutaraldehyde 3/19 (1613) 0/19 (018)

+ hydroperoxide 1/45 (210) 0/45 (08)

Total quats 68/484 (143) 8/484 (21)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for high Legionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower than theaverage (21%).395% confidence intervals.


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the results for biocide effectiveness were strikingly different,

with very few towers containing any high levels ofLegionella,

although the small number of positive towers again precluded

any significance differences to any of the biocide treatments.

Overall, quats as a biocide category had an average prevalenceof highLegionella.


in Cooling Towers Using Carbamates

Carbamates were the second most common biocide used

in this study (929 total tower samples). As illustrated in

Table 3, the effectiveness of carbamates in controlling the

prevalence ofLegionellain towers also varied according to the

specific biocide combination. Biocide combinations with

carbamate/isothiazoline and carbamate/THPS were signifi-

cantly more effective than average, with Legionella preva-

lence rates of 7% and 4%, respectively. In contrast, the

carbamate/quat combination (23%), as mentioned above, was

significantly worse than average. Carbamate/glutaraldehyde,

carbamates/hydroperoxide, and carbamates/DBNPA all

tended toward above-average (17%, 15%, and 33%, respec-

tively)Legionellaprevalence, but the number of towers in each

case was too small to achieve statistical significance. Simi-

larly, carbamates/bromine (11%) and carbonate alone (5%)

tended toward effectiveness but were used in too few towers to

achieve significance. Overall, the effectiveness of carbamate

as a biocide category was statistically within the range of aver-

age, with a prevalence rate of 115/929 (12%).



the results for biocide combination effectiveness had similar

trends to the overall prevalence as discussed above. The only

exception was that the carbamates/quat combination waswithin range of average in terms of high Legionella (2%)

despite the high overall prevalence rate for this combination

(23%). Overall, carbamates as a biocide category had an aver-

age prevalence of highLegionella.


in Cooling Towers Using Isothiazoline

Isothiazoline was the most commonly used biocide in the

study (1224 total tower samples) and was variably effective in

controlling the prevalence ofLegionellain towers (Table 4).

The combinations of isothiazoline/carbamate (7%) were

discussed above as being significantly more effective thanaverage, and the combination of isothiazoline/THPS (7%)

also had a significantly lower than averageLegionellapreva-

lence. The isothiazoline/bromine and isothiazoline/hydroper-

oxide tended to have higher than average Legionella

prevalence (16% and 18%, respectively), although the number

of towers in which they were used was too small to be signif-

icant. All of the remaining biocide treatment regimens were

also statistically similar to average. Overall, the effectiveness

of isothiazoline as a biocide category was statistically within

the range of average, with a prevalence rate of 142/1224

(12%).Table 3. Prevalence of Legionella in Cooling Towers

Using Carbamates Alone or in Combination


Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

Carbamates alone 1/22 (518) 0/22 (015)

+ bromine 4/36 (118) 1/36 (312)

+ quat 54/239 (236) 7/239 (32)

+ isothiazoline 24/338 (72) 5/338 (12)

+ glutaraldehyde 10/58 (178) 1/58 (27)+ THPS 6/147 (42) 1/147 (13)

+ hydroperoxide 11/74 (157) 4/74 (54)

+ DBNPA 5/15 (3321) 1/15 (725)

Total carbamates 115/929 (122) 20/929 (21)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for highLegionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower than theaverage (21%).395% confidence intervals.

Table 4. Prevalence of Legionella in Cooling TowersUsing Isothiazoline Alone or in Combination


Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

Isothiazoline alone 7/59 (127) 1/59 (27)

+ bromine 21/128 (166) 3/128 (25)

+ quat 1/28 (416) 0/28 (012)

+ carbamate 24/338 (72) 5/338 (12)+ glutaraldehyde 66/486 (143) 5/486 (11)

+ THPS 7/97 (74) 2/97 (25)

+ hydroperoxide 16/88 (187) 4/78 (54)

Total isothiazoline 142/1224 (122) 20/1224 (21)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for highLegionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower than theaverage (21%).395% confidence intervals.


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the results for biocide combination effectiveness had similar

trends to the overall prevalence as discussed above. Overall,

isothiazoline as a biocide category had an average prevalence

of highLegionella.


in Cooling Towers Using Glutaraldehyde

Glutaraldehyde was a commonly used biocide in the

study (843 towers), and almost all of the combinations trended

toward higher prevalence ofLegionellain the towers (Table 5).

The combination of glutaraldehyde plus bromine had the high-

est Legionella prevalence of 20%, but the 95% confidence

interval had a slight overlap with the overall average. With the

exception of glutaraldehyde/THPS (13% prevalence), all of

the remaining combinations, plus glutaraldehyde alone, had

higher than averageLegionellaprevalence values, but each of

the populations were too small to achieve significance. Simi-

larly, the overall effectiveness of glutaraldehyde as a biocide

category tended toward being worse than the average of all

towers, with a prevalence rate of 126/843 (152%), although

there was a slight overlap in the 95% confidence intervals.

When the glutaraldehyde data were reexamined to detect

the prevalence of high levels (> 1,000/ml) ofLegionellain the

cooling towers, many glutaraldehyde combinations appeared

to trend toward better control. However, three of the combi-

nations (glutaraldehyde alone, glutaraldehyde plus bromine,

and glutaraldehyde plus hydroperoxide) still had elevated

levels that were close to being statistically significant (6%,

7%, and 9%, respectively). Overall, glutaraldehyde as a

biocide category had an average prevalence of high

Legionella.


in Cooling Towers Using THPS

While used less frequently (397 towers) than somebiocides, THPS was the most effective biocide category in this

study in terms ofLegionellaprevalence (Table 6). All of the

biocide combinations hadLegionellaprevalences of average

or better, although only two, THPS/carbamate (4%) and

THPS/isothiazoline (7%), had populations large enough to

achieve statistical significance at the 95% confidence level.

Overall, the effectiveness of THPS as a biocide category was

statistically better than average, with a prevalence rate of

28/397 (7%).

When the THPS data were reexamined to detect the prev-

alence of high levels (> 1,000/ml) ofLegionellain the cooling

towers, the results for biocide effectiveness had similar trends

to the overall prevalence. No high levels of Legionellaweredetected in towers with THPS/bromine or THPS/glutaralde-

hyde and in only one tower out of 147 with THPS/carbamates

and 2 towers out of 97 with THPS/isothiazoline. Overall,

THPS as a biocide category had a better than average preva-

lence of highLegionella, although because of the overall aver-

age low number of positives, there was a slight overlap in the

95% confidence intervals.


in Cooling Towers Using Hydroperoxide

Hydroperoxide was not a commonly used biocide in the

study (294 towers), and with one exception, most combina-tions trended toward a slightly higher prevalence ofLegionellaTable 5. Prevalence of Legionella in Cooling TowersUsing Glutaraldehyde Alone or in Combination


Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

Glutaraldehyde alone 11/66 (178) 4/66 (644)

+ bromine 21/103 (207) 7/103 (74)

+ quat 3/19 (1613) 0/19 (018)


+ THPS 9/68 (137) 0/68 (05)

+ hydroperoxide 6/43 (149) 4/43 (96)

Total glutaraldehyde 126/843 (152) 20/843 (21)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for high Legionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower than theaverage (21%).395% confidence intervals.


Using THPS Alone or in Combination


Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

THPS alone 6/64 (95) 0/64 (06)

+ bromine 0/21 (016) 0/21 (016)+ carbamate 6/147 (42) 1/147 (13)

+ glutaraldehyde 9/68 (137) 0/68 (05)

+ isothiazoline 7/97 (74) 2/97 (25)

Total THPS 28/397 (72) 3/397 (11)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for high Legionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower than theaverage (21%).395% confidence intervals.


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in the towers (Table 7). The combination of hydroperoxide/

quat had a lowLegionellaprevalence of only 2%, although the

95% confidence interval overlapped slightly with the overall

Legionellaaverage in all towers. All the remaining combina-

tions of hydroperoxide had slightly higher than average

Legionella prevalence values (range 14%18%), although

each of the populations was too small to achieve statistical

significance. Overall, the effectiveness of hydroperoxide as a

biocide category was statistically within the range of average,

with a prevalence rate of 41/294 (14%).

When the hydroperoxide data were reexamined to detect

the prevalence of high levels (> 1,000/ml) ofLegionellain the

cooling towers, the results for biocide combination effective-

ness were similar to the overall prevalence, with higher than

average prevalence of elevatedLegionellain all towers except

for hydroperoxide/quat and hydroperoxide alone, despite the

small number of positive towers precluding statistical signif-

icance with any of the biocide treatments. Overall, hydroper-

oxide as a biocide category also had a higher than average

prevalence of high Legionella (5%), although there was aslight overlap in the 95% confidence intervals.


in Cooling Towers Using DBNPA

DBNPA was the least commonly used biocide in the study

(61 towers), so the sample size was relatively small for all

combinations (Table 8). Nevertheless, as mentioned above, the

combination of DBNPA plus bromine was much less effective,

with a significantly higher rate of Legionella prevalence

(42%). DBNPA/carbamate and DBNPA/hydroperoxide also

had higher prevalence rates (33% and 15%, respectively), but

the populations were not large enough to achieve statistical

significance. As one might predict from these data, the overall

effectiveness of DBNPA as a biocide category was also

significantly worse than average, with a prevalence rate

of 17/61 (28%).

When the DBNPA data were reexamined to detect the

prevalence of high levels (> 1,000/ml) of Legionella in the

cooling towers, the results for biocide combination effective-ness were similar to the overall prevalence, with higher than

average prevalence of elevatedLegionellain all towers except

for DBNPA/bromine, despite the small number of positive

towers precluding statistical significance with any of the

biocide treatments. Overall, DBNPA as a biocide category

also had a higher than average prevalence of highLegionella

(5%), although the total population of DBNPA towers was too

small to determine statistical significance.

DISCUSSION

The information presented in this study of results from

2,590 cooling towers and 28 biocide combinations is the firstlarge-scale correlation comparingLegionellaprevalence with

specific biocide use. The authors recognize that with this type

of study there are inherent weaknesses in the numerous uncon-

trolled variables, including actual concentrations of biocides

in the tower water, the cycles of concentration operative in

each tower, the water temperature, pH, filtration, and other

chemicals used for scale control, and biodispersants. Never-

theless, assuming that (1) the biocides were being added

according to manufacturer recommendations and (2) that the

samples were being taken, as directed, immediately prior to

addition of any slug-fed biocides (when Legionellanumbers

should be at their highest), then a sufficiently large population

size should average out the other variables, and the biocidesthemselves would become the predominant variable in each

group. Thus, the significance of these data is that they are


Using Hydroperoxide Alone or in Combination


Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

Hydroperoxide alone 3/17 (1814) 0/17 (020)

+ quat 1/45 (210) 0/45 (08)


+ glutaraldehyde 6/43 (149) 4/43 (96)

+ DBNPA 4/27 (1511) 2/27 (76)

Total Hydroperoxide 41/294 (144) 14/294 (52)1Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for highLegionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower thanthe average (21%).395% confidence intervals.


Using DBNPA in Combination with

Another Biocide (N=61)

Biocides

Legionella

Prevalence,1

(%Cl)3

HighLegionella

Prevalence2

(%Cl)3

(Legionella

1,000 cfu/ml)

DBNPA alone Not used Not used + bromine 8/19 (4222) 0/19 (018)

+ carbamate 5/15 (3321) 1/15 (725)

+ hydroperoxide 4/27 (1511) 2/27 (76)

Total DBNPA 17/61 (2811) 3/61 (541Number positive forLegionella/total number of towers in that cate-gory. Percentages in bold are significantly higher or lower than the av-erage (131%).2Number positive for highLegionella/total number of towers in thatcategory. Percentages in bold are significantly higher or lower thanthe average (21%).395% confidence intervals.


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representative of the way that these biocides are being used in

real-life situations and theLegionellacontrol that is a result of

their use. With the large groups such as the isothiazoline/glut-

araldehyde combination (486 samples) or the carbamates/

isothiazoline combination (338 samples), this assumption is

more likely to be reliable. On the other hand, data from combi-

nations with small numbers (such as most of the DBNPA data)would need to be evaluated with much more caution.

As one might expect from this study, there were no

biocide combinations that were considered to be statistically

100% effective in preventing Legionella colonization, nor

were there any that always failed. Thus, the comparisons of

biocide combinations in terms ofLegionellaprevalence were

all relative to each other and also as compared to the average

colonization rate for all 2,590 towers (13%) and average high

Legionellarate (2%). While most of the biocide combinations

were close to this average, there were some that clearly were

statistically better or worse than the average, both in terms of

overall Legionella prevalence and the occurrence of high

levels ofLegionella.

Perhaps the most unexpected result from the study was

the data from oxidizing versus nonoxidizing biocides. Despite

the reported findings from other studies on the effectiveness of

oxidizing biocides (see Kim et al. 2002), the results from these

2,590 towers indicated that the prevalence ofLegionella(and

also highLegionella) in cooling towers treated with bromine

(alone or combined with another nonoxidizer) was no better,

and in many cases significantly worse, than towers that alter-

nated two nonoxidizers. For example, bromine alone and

bromine/DBNPA were significantly more likely to be associ-

ated with Legionella colonization (including high levels of

Legionella) than were towers treated with carbamates plusisothiazoline or any combination of THPS. The hydroperox-

ide biocide combinations were also somewhat disappointing

(except for hydroperoxide/quat) compared to many of the

nonoxidizing combinations, although the numbers of towers

in these combinations were too small to achieve statistical

significance. Finally, chlorine was not used in enough towers

to be included in the study, but the few towers that did use chlo-

rine had an unusually large number of highLegionellaoccur-

rences (data not shown). Perhaps future updates of this study

will provide a complete comparison of chlorine to the other

biocide combinations.

A second surprise from the study was the relative effec-tiveness of some combinations of quaternary ammonium

(quat) biocides despite the fact that they were disparaged in

previous studies and by OSHA. While the quat/carbamates

combination was indeed less effective than average, the

combination of quat plus bromine (135 towers) had a very low

prevalence of both Legionella and high Legionella levels.

While the number of towers in this latter combination was not

as high as some combinations, one would not have expected

such a lowLegionellaprevalence if the quats were as ineffec-

tive as reported. Perhaps the surfactant quality of the quats

serves to aid in the removal of biofilm, increasing the effec-

tiveness of certain other biocides.

One particular biocide that seemed to stand out in terms

ofLegionellacontrol in these towers was THPS. While it was

not perfect, THPS alone or in any combination had generally

very low Legionella colonization rates and very few high

levels of Legionella. In fact, the worst THPS combination(THPS/glutaraldehyde) was right at average in terms of

Legionellaprevalence. What was even more surprising was

that the THPS/bromine combination had lowLegionellaeven

though THPS is supposed to be inactivated by oxidizing

biocides and is only recommended for alternating with another

nonoxidizer. Perhaps the levels of free halogen in these towers

at the time of the THPS addition were not sufficient to inacti-

vate this biocide completely in the slug-fed dose. On the nega-

tive side, both glutaraldehyde and DBNPA trended toward

being somewhat disappointing, based on their worse than

average ability to control Legionella colonization in the

towers.

Finally, while the information in this study may be helpful

in examining biocide trends, it would be unwise to use these

data to indict any particular biocide group or combination of

biocides as unsafe for use againstLegionellaor to promote one

above all others at this time. Larger populations of each

biocide category need to be studied to achieve statistical

significance with each group. Many of the treatment catego-

ries showed interesting trends, but larger numbers of cooling

towers will be required in order to assign significant differ-

ences. The reason a chemical treatment fails in some situations

is unknown. One theory is that the bacteria in the tower

(including Legionella) build up resistance to a particular

biocide (much like antibiotic resistance in the clinical field).Thus, the popular use of two alternating biocides has been

promoted to address this possibility. The final message from

the present study is that any of the common biocide combina-

tions could work effectively (some more effective than others),

but that failures to controlLegionellacan be observed with any

biocides. When they occur, these situations need to be identi-

fied and remediated quickly. The results of this study suggest

that monitoring may assist in determining whether a regimen

is effective for a particular system.

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DISCUSSION

Robert MacLeod-Smith, Director, Baltimore Aircoil Ltd.,

New Denham, UK: This is a comment on the effectiveness of

biocides in controllingLegionella. In the UK, cooling towers

are required to be controlled withLegionellalevels typically

103times lower than accepted levels in the US. Is there a new

ASHRAE standard to try to address this inconsistency as towhat are safe and acceptable levels of Legionellain cooling

tower water? Also, the same types of biocides as included in

the study are the same as the biocides being used to control

Legionellato much lower levels in Europe.

Richard D. Miller:The speaker recognizes that in the UK the

cooling towers are required to maintainLegionellaat very low

levels, using the same biocides mentioned in this paper. Since

our limit of sensitivity was only 10/ml, it is likely that many of

the samples that were recorded as None Detected in our

study may have had detectableLegionellaif limits of sensitiv-

ity were increased to those required in the UK. It is unlikely

that a new ASHRAE standard will propose a limit on

Legionella in cooling towers, although the issue was hotly

debated during the development of ASHRAE Guideline 12-

2000 and currently in the newLegionellastandard committee,

SPC 188P. Nevertheless, the point of the paper was to demon-

strate the relative effectiveness of biocide combinations, with

a major point that all of the different biocide combinations

occasionally failed to preventLegionellacolonization (even at

10/ml), thus having an impact on decisions of cooling tower

operators whether to monitor in order to verify biocide effec-

tiveness. The results of the paper also contradict generally

accepted conclusions that quat biocides are ineffective against

Legionella, and also that an oxidizing/non-oxidizing combi-

nation is superior to two non-oxidizers forLegionellacontrol

Finally, the treatment that stood out in terms of relative

effectiveness againstLegionellawas the THPS biocide group.


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