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APPLIED AND ENVIRONMENTAL MICROBIOLOGY, Apr. 1980, p. 840-844 Vol. 39, No. 4
0099-2240/80/04-0840/05$02.00/0
Microbial Prevalence in Domestic Humidifiers
HARRIET A. BURGE,* WILLIAM R. SOLOMON, AND JEAN R. BOISE
Allergy Division, Department of Internal Medicine, University of Michigan, Ann Arbor, Michigan 48109
The prevalence of viable thermophilic bacteria and actinomycetes and meso-
philic fungi was examined in 145 samples from 110 domestic humidifiers. A total
of 72 and 43% of furnace and console humidifier samples, respectively, contained
viable thermophilic bacteria, whereas 60 and 72% of these samples produced
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mesophilic fungal growth. Thermophilic actinomycetes were recovered from
seven humidifier samples. Efforts to detect thermophilic actinomycete antigens
in 15 humidifier fluid samples were not successful. Antifoulants added to humi-
difier fluid reservoirs had no apparent effect on microbial frequency. Airbome
microbial recoveries did not reflect patterns of humidifier contamination with
respect to either kinds or numbers of microorganisms in 20 homes in which
volumetric air samples were obtained during humidifier operation.
Like many other microorganisms, thermo- (Becton, Dickinson & Co., Rutherford, N.J.), a Zip-
philic actinomycetes are known to colonize home lock plastic bag, and a form on which they provided
humidification systems. Thermoactinomyces the following information: name, address, telephone
species were isolated from 17 of 44 humidifier number, date sample taken, age of home, years occu-
pied by subject, heating type and fuel, dwelling type
samples (39%) as well as from a number of other and construction; indoor conditions (i.e., very humid,
household substrates by Kurup et al. (11) and humid, comfortable, rather dry, parched); and make
from 12 of 20 humidifier water samples by Fink and model number of humidifier, type, capacity, how
et al. (9). Banazak et al. (1) also reported acti- long in use in current season, date(s) and type(s) of
nomycete contamination in an unspecified num- most recent cleaning, antifoulants used if any, and
ber of humidifiers as well as in 50 to 75% of home location (consoles). Patients were asked to fill the
air samples. Additional reports have dealt with syringe with humidifier fluid, replace the plastic screw
specific humidifiers implicated in cases of hyper- cap on the syringe hub, place the syringe in the plastic
sensitivity pneumonitis from which these micro- bag, and refrigerate the sample, bringing it to their
next (weekly or biweekly) clinic visit.
organisms were isolated (4, 8, 12, 19). These This procedure produced 63 patient-collected sam-
documented recoveries have stimulated a justi- ples in 1975-1976 and 46 in 1976-1977. Samples col-
fiable concern over the possible risks of home lected by laboratory personnel during home visits (10
humidifying systems especially for people with in 1977, 28 in 1977-1978) included a 50-ml sample of
respiratory disease. undisturbed humidifier fluid, a 20-ml sample of hum-
The studies to be presented here were de- idifier fluid taken after vigorous scraping of internal
signed to provide a quantitative survey of micro- reservoir surfaces, and a 50-ml sample of tap water
bial prevalence in domestic humidifying units, from the home. In addition, fluid temperature and pH
to assess determinants of such contamination, measurements (pHydrion Papers, Microessential Lab-
and to examine the potential for aerosolization oratories, Brooklyn, N.Y.) were made in situ, and the
same information was elicited as for the patient-col-
of viable microbial particles from these potential lected samples. Home visits also permitted air sam-
sources. pling to be done during humidifier operation. Single
MATERLALS AND METHODS plates of malt extract agar (32 g of malt extract, 16 g
of agar, 1 g of yeast extract [all from Difco Laborato-
Humidifiers for study were identified by approach- ries, Detroit, Mich.], and 1,000 ml of distilled water)
ing patients who received immunotherapy injections were simultaneously exposed beneath the sixth stage
for respiratory allergy at the University of Michigan of each of two upright Andersen samplers (Andersen
allergy clinics. These subjects, all of whom lived within Samplers, Inc., Atlanta, Ga.) as previously described
50 miles of Ann Arbor, were familiarized with the (16). Air was drawn into the samplers at a rate of 0.03
objectives of the study and asked for their cooperation. m3/min for 1 min. Subsequently, single plates of Tryp-
Volunteers initially collected a sample of humidifier ticase soy agar (BBL, Cockeysville, Md.) were simi-
reservoir fluid which was brought to the clinic (1975- larly exposed. Air temperature and relative humidity
1976, 1976-1977). In the second and third seasons of measurements were made with a sling psychrometer
the study (1977 and 1977-1978), a similar patient group during these sampling procedures.
agreed to an in-home study conducted by allergy re- A 0.1-ml portion of each humidifier sample was
search staff. Patients volunteering to collect their own cultured on Trypticase soy agar, Trypticase soy agar
samples were given a sterile 50-ml disposable syringe with 30 Lg of novobiocin per ml (Sigma Chemical Co.,
840VOL. 39, 1980 MICROBIAL PREVALENCE IN DOMESTIC HUMIDIFIERS 841
St. Louis, Mo.) and malt extract agar with 0.8 ug of never effected automatically. A total of 36 of the
chlortetracycline (Sigma). Trypticase soy agar cul- furnace humidifiers (FH) were of the rotating-
tures were incubated at 50°C in a humidified incuba- drum type, having a circular belt of plastic mesh
tor; malt extract agar was incubated at room temper- which is passed continually through a fluid col-
ature. Samples that produced confluent growth on
either medium later were diluted 1:1, 1:2, and 1:4 and lection. The belt is applied to a drum which
recultured. Humidifier solids recovered after scraping revolves in the air stream to be moistened. Com-
were scattered on the surface of Trypticase soy agar monly, a sump was provided to prevent overfill-
with novobiocin and incubated at 50°C. ing of the unit. Thirteen FH were of the tric-
Colony counts were expressed as colony-forming kling-column type in which fluid is pumped to
units (CFU) per milliliter of fluid or CFU per cubic the top of a rectangular screen of coarse metal
meter of air. Bacteria were Gram stained and classified fibers, through which it then percolates. Air
only by shape and Gram stain reaction. Thermophilic passing through the mesh is moistened in an
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actinomycetes were identified by the method of Kurup
and Fink (10). Mesophilic fungi were identified with operation that effectively obviates a pool or res-
the aid of keys and descriptions in Barnett and Hunter ervoir of fluid. The types for the remaining eight
(2), Barron (3), and Ellis (7). FH were not specified.
For double immunodiffusion studies of humidifier Temperature and pH data and water quality
fluid (13) glass microslides coated in Noble agar measurements for the humidifier fluids (Table
(Difco) were employed. Center wells were filled with 2) were obtained only on the 38 occasions where
0.01-ml aliquots of humidifier fluid; antisera were home air samples also were obtained. Microbial
placed in outer wells in dilutions from 1:1 to 1:128 recoveries from all humidifier samples are sum-
(Greer Laboratories, Lenoir, N.C.). A 1:1 mixture of marized in Table 3. FH were more frequently
humidifier fluid and an appropriate antigen (Greer) free of microorganisms detectable in our system
was used in central wells on control slides. Antisera
used were Thermoactinomyces vulgaris, Thermoac- than CH. Mesophilic fungi were recovered from
tinomyces candidus, Saccharomonospora viridis, and CH with slightly greater frequency than from
Micropolyspora faeni. FH, whereas the reverse was true for thermo-
Percent dry weight was measured by oven drying in philic bacteria. Thermophilic actinomycetes
preweighed containers of 2 ml of humidifier fluid. were recovered from only four FH and three
Hardness, calcium, nitrate, phosphate, and silica were CH. Of the fluid samples from the 36 rotating-
measured with water quality testing kits (Ecologic, drum FH, 17% were sterile compared with 31%
Bohemia, N.Y.). of 13 trickling-column FH units. A total of 70
and 83% of the rotating-drum units produced
RESULTS mesophilic and thermophilic cultures, respec-
Categories of humidifier fluid and concurrent tively, whereas 46% of the trickling-column FH
home air samples taken are summarized in Table were similarly contaminated.
1 by humidifier type. All the console humidifiers Antifoulants apparently had no effect on
(CH) were of a similar design in which a fan either the frequency or level of microbial con-
blows air past a felted fiber belt moistened by tamination in either humidifier type (Table 3).
repeated passage through a shallow reservoir of The most frequently used antifoulant was house-
fluid. Water is added manually from above and, hold bleach (four CH, one FH). Sixteen other
in general, drainage of the unit is difficult and treatments were used, no one in more than three
humidifiers.
TABLE 1. Summary of humidifier and associated Fifteen FH and nine CH were sampled more
home air samples than once (Table 1). Most of these repeat sam-
ples followed a period of nonuse during which
No. of the humidifiers were cleaned. Thermophiles
samples were recovered two or more times in nine FH
Furnace and in three CH and were consistently absent in
Various types sampled 58 four FH and four CH. Mesophiles were consist-
Sampled twice ......... ......... 8 ently present in seven FH. Of these, the taxa
Sampled three times .. 6 recovered were consistent in three. Mesophiles
Sampled four times ...... .............. 1
were consistently present in three CH, two of
Total samples ............ 80
Associated home air samples .30 which were consistent with respect to taxa re-
covered.
Console Thernophilic microorganisms recovered from
Various types sampled ... 53 humidifier fluids were primarily Bacillus spe-
Sampled twice ......... 6 cies. In addition, Saccharomonospora viridis
Sampled three times . 3 and T. vulgaris were recovered from two and
Total samples .65 four humidifiers, respectively. To test the pos-
Associated home air samples. 8 sibility of inhibitors in the humidifier fluids or842 BURGE, SOLOMON, AND BOISE APPL. ENVIRON. MICROBIOL.
TABLE 2. Water quality parameters measured on 30 furnace and 8 console humidifier samples
Parameter
SampleNirt
from: Temp Hardness (mg/ Calcium Nitrate Phosphorus Silica Dry wt (mg/li-
(0C) liter) (mg/li ter) (mg/liter) (mg/liter) ter)
Console
humidifer
13-17a 6-10 75-1,035 50-475 0-440 0.1-1.0 10-25 700-19,000
14.7b 7.5 537 244 114 0.4 15 8,900
15C 6.5 440 175 53 0.15 12.5 4,800
Fumace
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humidifier
14-29 4-10 150-12,350 0-6,000 0-440 0.1-10.0 5-60 400-206,900
21 7 4,316 1,206 118 1.1 29 34,000
21 7.2 2,200 570 88 0.3 20 12,000
Tap water
100-160 50-95 0-2.6 0.1-1.0 1.5-10
131 63 1.4 0.17 2.6
145 55 1.8 0.1 5.5
a Range.
b Mean.
'Median.
TABLE 3. Frequency of microbial contamination in TABLE 4. Mesophilic taxa recovered from
humidifier fluid samples humidifier fluids
Console Furnace
Humidifier fluid na "Ster- % Me- Ther- % Acti-
sample ile" b sophilic mo- cetes Mesophilic taxa % Posi- Range % Posi- Range
~~philic
~~ tive (colonies/ tive (colonies/
Furnace total 57 23 60 72 7
With antifoulant 10 20 60 70 Alternaria 3.5 10
Without 47 23 60 72 Aspergillus 3.8 10-30 3.5 10
antifoulant Aureobasidium 1.9 10
Console total 53 11 72 43 6 Candida 1.9 10
With antifoulant 21 9 76 52 Cladosporium 5.7 10-80 10.5 10-970a
Without 32 13 69 38 Cylindrocarpon 1.7 10
antifoulant Epicoccum 1.9 10
Fusarium 5.7 10-120 15.8a 10-590a
'Number of humidifier fluid samples. Gliocladiuma 3.8 10-60 3.5 20-260a
b With no microorganisms recovered in our culture system. 10-80 7.0 10-20
Penicillium 9.4
Phialophoraa 15.1a 10-2,180a 15.8a 30-1,260a
Rhodotorula 11.3a 10-100a 14.00 10-1,230a
in (local) Ann Arbor tap waters or both, T. Scolecobasidium' 3.5 20-3,000a
vulgaris and T. candidus were inoculated into Sepedonium' 1.7 980a
Sordaria 1.9 20
five apparently sterile humidifier samples and Sphaeropsidales 1.9 40
two samples of tap water. Both organisms were Sporobolomyces 1.7 20
readily recovered from all inoculated samples. Verticillium 3.5 10-20
Fifteen humidifier fluid samples were tested by Yeast 28.30 10-2,5000 7.0 10-870a
double immunodiffusion with antisera to T. can- Nonsporulating 1.9 10 5.3 10-1,500a
didus, T. vulgaris, S. viridis, and M. faeni. No a Refers to unusual taxa, frequency, or level.
humidifier fluid produced precipitin lines against
any of the antisera. Mesophilic recoveries from these homes, similar taxa were encountered in
humidifier fluids are summarized in Table 4. the air and the humidifier samples. In none of
Spearman rank order correlations between these these cases could air contamination by humidi-
mesophilic recoveries and other humidifier fluid fier organisms be clearly demonstrated since all
parameters (temperature, pH, percent dry taxa recovered in both fluid samples and in air
weight, hardness) were not significant. are commonly present in both indoor and out-
In 17 homes in which air samples were made door air in our area. None of the unusual humi-
concurrently with humidifier samples, the hum- difier organisms (e.g., Phialophora) were en-
idifiers were contaminated with unusual num- countered in room air. Thermophilic actinomy-
bers or types of organisms (Table 5). In seven of cetes (Thermomonospora fusca, T. candidus,VOL. 39, 1980 MICROBIAL PREVALENCE IN DOMESTIC HUMIDIFIERS 843
TABLs 5. Mesophilic recoveries from 17 humidifier fluid and associated air samples
Humidifier fluid Air
Pink yeast, Phialophora Penicillium, pink yeast
Pink yeast Penicillium, Aspergillus, Alternaria, Cladosporium,
arthrospores
Bacteria, pink yeast Penicillium, Aspergillus, Cladosporium, Alternaria
Bacteria, Cladosporium Aspergillus, Penicillium, pink yeast
Fusarium Penicillium, Sporobolomyces, Aspergillus, Cladosporium
Scolecobasidium, Fusarium, Penicillium, Aspergillus, pink yeast
Alternaria
Fusarium, nonsporulating, Pink yeast, Penicillium, Cladosporium, Aspergillus
arthrospores
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Nonsporulating, Cladosporium, Aspergillus, Wallemia, Cladosporium, pink yeast, Penicillium,
Fusarium, Aspergillus, bacteria Myriococcum, Olpitrichum, Epicoccum
Phialophora, Fusarium, Penicillium Aspergillus, Penicillium, nonsporulating, yeast
Fusarium, yeast Aspergillus, Penicillium, pink yeast, Cladosporium
Pink yeast, Verticillium, Aspergillus, pink yeast, Penicillium, Cladosporium,
nonsporulating Tritarachium
Phialophora Penicillium, Cladosporium, pink yeast, Alternaria,
Aspergillus, Epicoccum
Cladosporium, Scolecobasidium Penicillium, Aspergillus, pink yeast, nonsporulating
Bacteria, Penicillium Aspergillus, Penicillium, pink yeast
Yeast, pink yeast Nonsporulating, yeast, Cladosporium, Penicillium, Alternaria,
arthrospores
Fusarium, Penicillium Penicillium, Cladosporium, yeast, Aureobasidium, Candida,
sphaeropsids, Alternaria
Cylindrocarpon Cladosporium, Alternaria, yeast, Aspergillus, Aureobasidium,
Epicoccum, Penicillium
Streptomyces sp., S. viridis) which were rare (or idifier samples they analyzed. The health effects
absent) in humidifier fluids were recovered from of these saprophytes remain unknown, as does
the air of all homes studied in levels from 50 to the potential for exposure to pathogenic bacteria
5,000/m3 of air sampled. from domestic humidifying systems.
Temperature conditions in CH and most FH
DISCUSSION were well within the optimum range for growth
These studies, which admittedly are based on of mesophilic fungi, and these organisms were
a circumscribed geographic area, suggest that recovered from 72 and 60% of such units. Many
humidifiers, in fact, do not commonly contribute of the taxa recovered are common in either
thermophilic actinomycetes to home air. In spite indoor or outdoor air in our area (e.g., species of
of considerable effort to detect contaminating Penicillium and Cladosporium). However, Phi-
thermophilic actinomycetes, we were able to alophora, which we have not isolated from air
recover actinomycetes from only seven humidi- (18), was recovered from 15% of all humidifying
fiers or 6% of those studied. On the other hand, units in levels ranging above 2,000 CFU/ml of
these organisms were recovered from the air of fluid. Other fungi recovered from humidifier
100% of homes studied. Disease-provoking anti- fluid which are rarely isolated from air in our
gens could, of course, be present in fluids unas- area are Sepedonium, Scolecobasidium, and
sociated with currently viable microorganisms Gliocladium. The allergenic potential of these
or where organisms are growing on inaccessible organisms is unknown, although Phialophora
surfaces, as reported by Miller (12). However, species are known to cause invasive human dis-
such antigens were not detected in the 15 fluids ease by inoculation into peripheral tissues.
we tested. The risks of air contamination during opera-
The maximum reservoir temperature meas- tion of contaminated humidifiers are not clear.
ured in any humidifier was 290C, well below the Mist generators have been shown clearly to pro-
45 to 600C range optimal for thermophilic acti- duce viable particle aerosols (6, 17). Bacterial
nomycetes. Yet, thermophilic bacteria with a emissions from evaporation-type humidifiers
similar optimum temperature range were iso- (including all units reported here) also may oc-
lated from 72% of our FH units and 43% of our cur when viable cells apparently are shaken from
CH units. Seabury et al. (15) also reported large moist surfaces of fibrous material; however, re-
numbers of thermophilic bacteria from all hum- sulting airbome levels apparently fall rapidly844 BURGE, SOLOMON, AND BOISE APPL. ENVIRON. MICROBIOL.
after turn-off (6). Schultz et al. (14) reported or both, the continued use of such systems seems
that clearly contaminated evaporative humidi- defensible.
fiers did not routinely produce bacterial aerosols,
and our failure to demonstrate manifest airborne ACKNOWLEDGMENTS
contamination argues against extensive intro- This work was supported by Public Health Service grant
duction of viable microbes into room air during AI 10181 from the National Institute of Allergy and Infectious
operation of the units studied. A potential for Diseases.
acute exposure during cleaning does exist. LITERATURE CITED
Cartwright and Aargrave (5) discussed humidi-
fier scale as a nidus for humidifier microbial 1. Banazak, E. F., J. Barboriak, J. Fink, D. Schlueter,
contamination. This material often is scraped, A. Sosman, W. Thiede, and G. Unger. 1974. Epide-
miologic studies relating thermophilic fungi and hyper-
scrubbed, and otherwise violently disturbed dur-
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sensitivity lung syndromes. Am. Rev. Respir. Dis. 110:
ing cleaning; hosing also may spread contami- 585-591.
nated droplets into the air. 2. Barnett, H. K., and B. B. Hunter. 1972. Illustrated
Antifoulants that had been added to the fluid genera of imperfect fungi, third edition. Burgess Pub-
lishing Co., Minneapolis.
reservoirs in 17% of FH and 40% of CH studied 3. Barron, G. L. 1968. The genera of hyphomycetes from
did not produce clearly evident effects on the soil. The Williams & Wilkins Co., Baltimore.
microbial flora. Covelli et al. (6) studied the 4. Burke, G. W., C. B. Carrington, R. Strauss, J. Fink,
impact of a sodium hypochlorite antifoulant and and E. Gaensler. 1977. Allergic alveolitis caused by
home humidifiers. J. Am. Med. Assoc. 238:2705-2708.
reported that changes in Pseudomonas popula- 5. Cartwright, Y., and P. R. Aargrave. 1970. Pseudomo-
tions lasted, at most, 24 h. By asking our vol- nas in ventilators. Lancet i:40.
unteers not to disturb their humidifiers within 6. Covelli, H. D., J. Kleeman, J. Martin, V. Landau, and
24 h of our visit, we virtually insured that eva- R. Hughes. 1973. Bacterial emission from both vapor
and aerosol humidifiers. Am. Rev. Respir. Dis. 108:
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ticed. However, a more sustained influence of 7. Ellis, M. B. 1971. Dematiaceous hyphomycetes. Common.
additives, in general, on microbial populations Mycol. Inst. Kew, Surrey.
also was nowhere evident. These findings, com- 8. Fink, J. N., E. Banazak, W. Thiede, and J. Barboriak.
1971. Interstitial pneumonitis due to hypersensitivity to
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addition of currently available agents to humi- 9. Fink, J., A. Resnick, and J. Salvaggio. 1971. Presence
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systems. Appl. Microbiol. 22:730-731.
Long-term effects of physical cleaning on 10. Kurup, V. P., and J. N. Fink. 1975. A scheme for the
humidifier microbial contamination are not well identification of thermophilic actinomycetes associated
documented. Covelli et al. (6) reported that thor- with hypersensitivity pneumonitis. J. Clin. Microbiol.
2:55-61.
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sterility after resumption of operation. However, Mycologia 68:662-666.
only one-third of the CH examined more than 12. Miller, M. M. 1976. Chronic hypersensitivity lung disease
once in our study were consistently contami- with recurrent episodes of hypersensitivity pneumonitis
due to a contaminated central humidifier. Clin. Allergy
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Sanford. 1967. Studies of a new humidifying device as
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sistently contaminated with thermophiles and pir. Dis. 96:517-519.
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samples and were rarely cleaned during the pe- 16. Solomon, W. R., and J. A. Gilliam. 1970. A simplified
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which were cleaned frequently and kept com- airborne fungus particles. J. Allergy 45:1-13.
pletely free of scale, were consistently sterile by 17. Solomon, W. R. 1974. Fungus aerosols arising from cold-
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