Variation and Characterization of Bacterial Communities Contaminating Two Saunas Operated at 64℃ and 76

Abstract

This study was performed to analyze 6 day-term variations in bacterial communities contaminating the floor of two dry saunas that were operated at 64℃ (low temp) and 76℃ (high temp). Bacteria were sampled daily from the saunas for 6 days from Monday to Saturday. Genomic DNA was isolated directly from bacteria-collected cotton swabs. The diversity of the bacterial communities collected from the saunas was analyzed using thermal gradient gel electrophoresis (TGGE). The total numbers of DNA bands separated by TGGE for bacteria collected from the low temp and high temp sauna were 20 and 18, respectively, during the 6 days. Seven of 20 bacteria in the low temp sauna and eight of 18 bacteria in the high temp sauna were detected more than three times over the 6 experimental days. Twelve of the 26 bacterial genera contaminating the saunas were cross detected. Bacteria belonging to the genera Moraxella and Acinetobacter were selectively detected in the low temp sauna, whereas those belonging to Aquaspirillum, Chromobacterium, Aquabacterium, Gulbenkiania, Pelomonas, and Aquitalea were selectively detected in the high temp sauna. Three species of bacteria contaminating both the low and high temp saunas were thermophile or thermoduric. The results indicate that the sauna-contaminating bacteria may have been transferred from outside the saunas by user traffic but did not inhabit the saunas.

Full text

Journal of Bacteriology and Virology 2013. Vol. 43, No. 3 p.195
–
203
http://dx.doi.org/10.4167/jbv.2013.43.3.195
Variation and Characterization of Bacterial Communities
Contaminating Two Saunas Operated at 64℃ and 76℃
Bong Su Kim, Jae Ran Seo and Doo Hyun Park
*
Department of Chemical and Biological Engineering, Seokyeong University, Seoul, Korea
This study was performed to analyze 6 day-term variations in bacterial communities contaminating the floor of two
dry saunas that were operated at 64℃ (low temp) and 76℃ (high temp). Bacteria were sampled daily from the saunas
for 6 days from Monday to Saturday. Genomic DNA was isolated directly from bacteria-collected cotton swabs. The
diversity of the bacterial communities collected from the saunas was analyzed using thermal gradient gel electrophoresis
(TGGE). The total numbers of DNA bands separated by TGGE for bacteria collected from the low temp and high temp
sauna were 20 and 18, respectively, during the 6 days. Seven of 20 bacteria in the low temp sauna and eight of 18 bacteria
in the high temp sauna were detected more than three times over the 6 experimental days. Twelve of the 26 bacterial
genera contaminating the saunas were cross detected. Bacteria belonging to the genera Moraxella and Acinetobacter were
selectively detected in the low temp sauna, whereas those belonging to Aquaspirillum, Chromobacterium, Aquabacterium,
Gulbenkiania, Pelomonas, and Aquitalea were selectively detected in the high temp sauna. Three species of bacteria
contaminating both the low and high temp saunas were thermophile or thermoduric. The results indicate that the sauna-
contaminating bacteria may have been transferred from outside the saunas by user traffic but did not inhabit the saunas.
Key Words: Sauna-contaminant, Thermophile, Thermoduric, TGGE, Spore-forming bacteria
INTRODUCTION
We have previously characterized a specific bacterial
community obtained by single-time and single point
sampling from a sauna operated at 75~80℃ using thermal
gradient gel electrophoresis (TGGE) (1). In that study,
selectively isolated thermophilic bacteria grew maximally
at 40℃ in both defined and complex medium but limited
growth occurred at 50℃ in a defined medium and at 60℃
in a complex medium. Bacteria isolated from a 75~80℃
sauna were thermoduric but not thermophilic. However,
the diversity and characteristics of the bacterial community
were analyzed by single-time and single-point sampling
may vary opportunistically according to the user number,
residence time, and physiological condition.
Thermophilic bacteria are different from thermoduric
bacteria by their ability to regenerate and grow under
thermal conditions of 45~122℃ (2, 3). The survivability
of thermoduric bacteria in a hot and dry sauna and under
heat of pasteurization depends upon their ability to sporulate
(4, 5). Spore-forming bacteria may be thermoduric or
thermophilic. But, thermophilic bacteria can't grow at
mammalian temperatures (30~40℃); however, thermoduric
bacteria can grow at mammalian temperatures and generate
spores at higher than mammalian temperatures (6~9). Dry
195
Original Article
Received: June 26, 2013/ Revised: August 21, 2013/ Accepted: August 29, 2013
*
Corresponding author: Doo Hyun Park. Department of Chemical and Biological Engineering, Seokyeong University, 16-1 Jungneung-dong, Sungbuk-gu,
Seoul 136-704, Korea.
Phone: +82-2-940-7190, Fax: +82-2-919-0345, e-mail: baakdoo@skuniv.ac.kr
○
CC
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/license/by-nc/3.0/).

196 BS Kim, et al.
saunas are generally operated at temperatures > 60℃, which
may not be a proper temperature for thermophilic bacteria,
but an environmental signal for thermoduric bacteria to
generate spores. Dry saunas may not be nutritionally proper
for bacterial growth but may be opportunistically contami-
nated with various bacteria and organic compounds by user
traffic (10). User traffic may be the major cause of sauna
contamination by bacteria, as human traffic supplies nutrients
needed by specific thermoduric and thermophilic bacteria
to opportunistically survive.
Theoretically, psychrophiles and mesophiles can't grow
and inhabit the environmental conditions inside a sauna;
however, spore-forming bacteria may survive for a particular
period, considering that endospores are often highly resistant
to chemical- and heat-treatment (11). Species belonging to
the endospore-forming bacteria are found in both natural
and artificial environments (12, 13). Endospore-forming
bacteria are widely distributed in the natural environment
such as soil and water. Most bacteria dwelling around
humans may be transferred into a sauna through direct
contact with the human body (14, 15).
In this study, the bacteria collected from two saunas on
multiple occasions was analyzed using TGGE to compare
the daily variations in the bacterial community during 6 days
from Monday to Saturday and a conditional variation in the
bacterial community based on the temperature difference
between the two saunas.
MATERIALS AND MOTHODS
Bacterial sampling
A digital thermometer on the outside wall of the two
saunas indicated that the internal temperatures were 64℃
and 76℃ in the low temp and high temp saunas, respec-
tively. Bacteria were sampled from the floor of the dry
sauna using sterilized cotton swabs. Samples were collected
around 9 o'clock in the morning on December 24~29,
2012. An aluminum foil-wrapped cotton swab was opened
immediately before sampling, and the floor was then deeply
and broadly wiped with three cotton swabs per sample area.
The sampling points were located at 30 cm from each wall
in the central part because user traffic was mainly busy
between the central part and each wall but not around the
corners. The diameter of the sampling area was about 30
cm (1). In total, 15 swabs were taken and used for DNA
extraction. The part of the cotton swab used for bacterial
sampling was placed in a sterilized conical tube immediately
after sampling, and the other end was removed and used in
a previous study (1).
16S-rDNA amplification
Total DNA was directly extracted from the bacterial cells
collected from the cotton swabs using a Genomic DNA
extraction kit (Accuprep; Bioneer, Daejeon, Korea) according
to the manufacturer's protocol. 16S ribosomal DNA was
amplified via direct polymerase chain reaction (PCR) using
the chromosomal DNA template and 16S-rDNA specific
universal primers as follows: forward 5'-GAGTTGGATC-
CTGGCTCAG-3' and reverse 5'-AAGGAGGGGATCC-
AGCC-3'. The PCR reaction mixture (50 μl) consisted of
2.5 U Tag polymerase, 250 μM of each dNTP, 10 mM
Tris-HCl (pH 9.0), 40 mM KCl, 100 ng template, 50 pM
primer, and 1.5 mM MgCl
2
. Amplification was conducted
for 30 cycles of 1 min at 95℃, 1 min of annealing at 55℃,
and 2 min of extension at 72℃ using a PCR machine (T
Gradient model, Biometra, Göttingen, Germany).
TGGE
The 16S-rDNA amplified from chromosomal DNA was
employed as the template for TGGE sample preparation. A
variable region of 16S-rDNA was amplified using the
forward primer 341f 5'-CCTACGGGAGGCAGCAG-3'
and reverse primer 518r 5'-ATTACCGCGGCTGCTGG-3'.
A GC clamp (5'-CGCCCGCCGGCGGGCGGGCGGGG-
CGGGGGCACGGGGGG-3') was attached to the 5'-end
of the 341f primer (16). The PCR and DNA sequencing
procedures were identical to the 16S-rDNA amplification
conditions, with the exception of annealing temperature.
The TGGE system (Bio-Rad, Dcode
TM
, Hercules, CA,
USA) was operated in accordance with the manufacturer's
specifications. Aliquots (45 ml) of the PCR products were
electrophoresed on gels containing 8% acrylamide, 8 M urea,

Variation and Characterization of Bacteria Contaminating Dry Saunas
197
and 20% formamide in 1.5 ×TAE (Tris, acetate, and
EDTA) buffer system at a constant voltage of 100 V for
12.5 h, followed by 40 V for 0.5 h, with a temperature
gradient of 39~52℃. Prior to electrophoresis, the gel was
equilibrated to the temperature gradient for 30~45 min.
Amplification and identification of the TGGE band
DNA was extracted from the TGGE band and purified
with a DNA gel purification kit (Accuprep, Bioneer). The
purified DNA was then amplified with the same primers
and procedures used for TGGE sample preparation, except
that the GC clamp was not attached to the forward primer.
The amplified DNA was sequenced to identify the bacteria
based on 16S-rDNA sequence homology using GenBank
database.
RESULTS
TGGE patterns
The daily variations in the bacterial community collected
from the floors of the low and high temp saunas were
analyzed by the TGGE technique. The TGGE pattern for
the bacteria contaminating the low temp sauna was not
significantly different from that contaminating the high temp
sauna (Fig. 1). A range of 5~10 species was observed daily
in the low temp sauna, whereas 8~12 species contaminating
the high temp sauna based on the DNA band number.
Variations in DNA band number and DNA band position
(migration distance on electrophoretic gel) were not patterned
by specific days of the week. This result shows that the
bacteria contaminating the saunas may be opportunistic
and related to both user traffic and operating temperature of
a sauna.
Identification of DNA separated by TGGE
Twenty and 18 bacterial species were separated by
TGGE and identified by sequence homology in the low
and the high temp saunas, respectively (Tables 1 and 2).
Bacteria in Moraxella and Acinetobacter were selectively
detected in the low temp sauna, whereas bacteria in
Aquaspirilum, Chromobacterium, Aquabacterium, Gulben-
kiania, Pelomonas, and Aquitalea were selectively detected
in the high temp sauna during the 6-day sampling from
Monday-Saturday. The bacterial diversity contaminating the
saunas may be directly influenced by individual bacteria-
contaminated users. An uncultured bacterium (JN883765)
was detected daily in the low temp sauna, whereas Neisseria
flava and uncultured bacterium (JN883765) were detected
daily in the high temp sauna. The two bacteria that were
detected daily in the low or high temp saunas commonly
belonged to the normal microflora of specific organs of the
human body (Table 3), suggesting that the bacteria detected
in the saunas may be contaminants from user bodies but not
residents in the saunas. The user's body may be the main
source of bacteria which contaminate a sauna.
Figure 1. Thermal gradient gel electrophoresis (TGGE) profiles
of 16S-rDNA isolated from bacteria that were collected from floors
of two different saunas operated at 64℃ and 76℃. Each numbe
r
on electrophoresis gel indicates DNA band originated from
a
specific bacterial species. Abbreviations: M, Monday; T, Tuesday;
W, Wednesday; Th, Thursday; F, Friday; S, Saturday.

198 BS Kim, et al.
General characteristics of the bacteria detected by
TGGE
The general characteristics of the bacteria collected from
the low and high temp saunas was tabulated based on genus
or species information reported by other researchers or
from a database. As shown in Table 3, most of the bacteria
identified based on 16S-rDNA extracted from the TGGE
gel and collected from both the low and the high temp
saunas were not thermophiles except bacterium ODP-
193-27 (17), Bacillus megaterium (18), and Deinococcus
geothermalis (19). Bacillus megaterium is a spore-forming
thermoduric bacterium but not a thermophile. Thermophilic
and thermoduric bacteria were detected one to four times
over the 6 experimental days, whereas mesophilic bacteria
were detected less than three times during the entire sampling
period. In contrast, Neisseria flava (20) was detected in the
high temp sauna everyday over 6 days and an uncultured
bacterium (JN883765) was detected in both saunas every
day over the 6 days (21). Both bacterial species detected
each day are known to contaminate specific organs of
human body (20, 21); whereas most of the bacteria detected
in both the high and the low temp sauna are known to
inhabit natural habitats such as spring water, wastewater,
soil, seawater, and lake water (22~31). From this result, it
can be presumed that the human body may be the most
suitable carrier for bacteria because the human body is a
proper habitat for bacteria to temporarily survive or grow.
Table 1. Bacterial species identified based on sequence homology of 16S-rDNA extracted from electrophoresis gel (figure 1, 64℃). Therma
l
gradient gel electrophoresis was performed with the DNA extracted from the bacteria daily collected from low temp sauna from Monda
y
to Saturday.
Week: band number in TGGE gel
Bacteria (Genus or species)
Accession
code
Homology
(%)
Mon Tue Wed Thu Fri Sat
Bacterium ODP-193 AB084523 98 1
Uncultured bacterium HE649228 98 1 1 1
Enhydrobacter aerosaccus JX845725 98 2 2 2 2
Neisseria flava HF558370 98 3
Moraxella cuniculi AJ247221 99 1
Moraxella sp. KC119125 98 2
Uncultured bacterium JN883765 98 3 1 3 4 3 3
Moraxella pluranimalium NR042666 98 4 4
Acinetobacter radioresistens JF919868 97 5 4 4
Acinetobacter sp. GU430989 98 6 5 5
Bacillus megaterium JX311358 99 2 6
Acinetobacter seohaensis EU420936 98 5 3 5 7
Uncultured bacterium JF237408 99 4
Acinetobacter beijerinckii JN644620 98 5
Uncultured Acinetobacter JN866218 99 6 8
Leptothrix sp. JQ946028 97 6
Uncultured bacterium AB732642 98 7 7 9 7
Uncultured bacterium JF661828 97 8
Deinococcus geothermalis EU600161 97 9 6
Uncultured bacterium JN882031 99 10 7
*Bold letters: Bacteria selectively detected in 64℃ sauna

Variation and Characterization of Bacteria Contaminating Dry Saunas
199
DISCUSSION
Bacteria are ubiquitous organisms based on the diversity
of their habitats and natural ecosystems, such as animals
(intestine, skin, and genital organs), plants (leaves and roots)
and man-made structures (32~36). The most popular
man-made structures are buildings utilized as dwellings,
businesses, and manufacturing plants, which may be habitats
for heterotrophic bacteria due to the plentiful organic
compounds and the proper environmental conditions (37).
A sauna is a hot, dry room located inside a building, and the
environmental conditions are not suitable for any organisms
except thermophilic bacteria (1). Temperature is one of the
major environmental factors influencing bacterial growth
and has been artificially controlled to cultivate fermentation
bacteria, suppress harmful bacteria, or destroy pathogenic
bacteria (38~40). Generally, the temperature of a sauna is
60~80℃, which is sufficient to inhibit or stop growth of
mesophilic bacteria except spore-formers (41).
The mesophilic bacteria that contaminated both the low
and the high temp saunas may neither survive nor grow but
the thermophilic bacteria (bacterium ODP-193-27, B.
megaterium, D. geothermalis) can survive and grow in both
saunas based on their characteristics (17~19). However,
the growth of thermophilic bacteria contaminating both
saunas must be experimentally verified because a sauna is
an artificial location that satisfies only the temperature
conditions for thermophilic bacteria (42). The ecological
niche of bacteria is mineralization of organic compounds
by parasitism, symbiosis, and saprophytism (43~45). The
thermophilic bacteria that contaminated the saunas may
Table 2. Bacterial species identified based on sequence homology of 16S-rDNA extracted from electrophoresis gel (figure 1, 76℃). Therma
l
gradient gel electrophoresis was performed with the DNA extracted from the bacteria collected from high temp sauna from Monday to
Saturday.
Week: band number in TGGE gel
Bacteria (Genus or species)
Accession
code
Homology
(%)
Mon Tue Wed Thu Fri Sat
Bacterium ODP-193 AB084523 98 1 1 1 1
Enhydrobacter aerosaccus JX845725 98 2 2 2 2
Neisseria flava HF558370 98 3 3 3 3 2 1
Uncultured bacterium JN883765 98 4 4 4 4 3 2
Aquaspirillum serpens AB680863 100 5 5 4 3
Acinetobacter radioresistens JF919868 97 6 6 5 5 4
Acinetobacter sp. GU430989 98 6 6 5
Bacillus megaterium JX311358 99 7 7 6
Chromobacterium sp. HQ234407 98 5
Uncultured Aquabacterium sp. JQ288705 98 6
Gulbenkiania mobilis NR042548 97 8
Pelomonas sp. AB730488 97 7 7
Leptothrix sp. JQ946028 97 8 8
Acinetobacter beijerinckii JN644620 98 7 9 8 9 9
Uncultured bacterium AB732642 98 8 10 9 10 10
Aquitalea sp. JN208179 97 11
Deinococcus geothermalis EU600161 97 12 7
Uncultured bacterium JN882031 99 8
*Bold letters: Bacteria selectively detected in 76℃ sauna

200 BS Kim, et al.
grow by saprophytism because most bacteria were randomly,
occasionally, and discontinuously detected from both saunas
over the 6 experimental days.
Individuals travel domestically or in foreign countries and
Table 3. General characters of bacterial genus or species identified based on sequence homology of DNA extracted from electrophoresis
gel (Fig. 1). DNA used for thermal gradient gel electrophoresis was directly extracted from bacteria collected from 64℃ and 76℃ of dr
y
saunas.
Bacteria (Accession code)
Detection days
(LTS/HTS)
General characters or released information
Bacterium ODP-193-27 1/4
A thermophile capable of growing at 60~90℃, which were found in subsurface
of hydrothermal vent (17)
Enhydrobacter aerosaccus 3/4
A heterotrophic, mesophilic, non-pathogenic, gas-vacuolated, and facultative
anaerobic bacterium (22)
Neisseria flava 1/6
An non-pathogenic, mesophilic, and anaerobic bacterium that is often found in
the upper respiratory tract surface in humans (20)
Moraxella sp. 1/0
A mesophilic and opportunistically infective bacterium, and some species
belonging to this genus are commensal of mucosal surface (43)
Moraxella pluranimalium 2/0
Gram-negative, mesophilic, and heterotrophic bacterium that was isolated from
sheep and pig (44)
Acinetobacter radioresistens 3/3 A mesophilic, non-spore-forming, aerobic, and soil-dwelled bacterium (23)
Chromobacterium sp. 0/1
A mesophile that inhabits in soil and natural water is sometime is found in foods
(24)
Uncultured Aquabacterium sp. 0/1 It has been isolated and found from drinking water and freshwater spring (25)
Bacillus megaterium 2/3
This bacterium is able to survive in some extreme conditions such as desert
environments due to the spores it forms. (18)
Acinetobacter seohaensis 4/0
A Gram-negative, non-motile, and mesophilic bacterium that was isolated from
sea water of the Yellow Sea in Korea (26)
Deinococcus geothermalis 2/2 This is an extremely radiation resistant, moderately thermophilic bacterium (19)
Gulbenkiania mobilis 0/1 A mesophilic bacterium that was isolated from treated municipal wastewater (27)
Pelomonas sp. 0/1
A mesophilic, Gram-negative, non-spore-forming bacterium that was isolated
from industrial wastewater and haemodialysis water (28)
Leptothrix sp. 1/2
A filamentous and mesophilic bacterium that resides in organic matter-plentiful
aquatic environments (29)
Aquitalea sp. 0/1
A mesophilic, Gram-negative, and non-spore-forming bacterium that was isolated
from humic-lake samples (30)
Uc. Bacterium (HE649228) 3/0
A bacterium that was detected in a uranium mine tailing sediment-water interface
at Key Lake, Northern Saskatchewan
Uc. Bacterium (JN883765) 6/6 A bacterium belonging to normal gut microflora in human (21)
Uc. Bacterium (JF237408) 1/0
A bacterium belonging to human skin microflora related with atopic dermatitis
(45)
Uncultured Acinetobacter 2/0
A Gram-negative, non-motile, and mesophilic bacterium that was isolated from
Tibetan Plateau.
Uc. Bacterium (AB732642) 4/5 A bacterium detected in arsenic sediment
Uc. Bacterium (JF661828) 1/0
A bacterium belonging to microbial community in anaerobic digestion of carrot
waste (37)
Uc. Bacterium (JN882031) 2/1 A bacterium that was found in crude oil (31)
*UC, Uncultured bacterium; LTS, Low temp sauna; HTS, High temp sauna

Variation and Characterization of Bacteria Contaminating Dry Saunas
201
carry various bacteria between locations without realizing
that they have contaminated their bodies, clothes, and
baggage except in an emergency. Travelers are contaminated
with various microorganisms during travel by contacting
others, touching, visiting, and buying merchandise. The
surface of the human body is a temporary habitat for
bacterial growth due to the organic compounds excreted
with sweat and sebum. Mesophilic bacteria detected from
the saunas may have originated from natural or artificial
environments; however, the thermophilic bacteria could
have originated from a foreign country based on their
general characteristics (Table 1).
In a previous study (1), bacterial samples were collected
from a sauna operated at 75~80℃ on July 25, 2012 and
most were thermoduric and spore-forming bacteria belonging
to the Bacillus genus. The bacterial community collected
from saunas during the summer is significantly different
from that collected during the winter, which may be caused
by differences in environmental conditions between summer
and winter (46). Plentiful organic compounds, high tem-
perature, and high humidity of a natural environment during
the summer may be a more proper condition for Bacillus
sp. than other bacteria. The seasonal differences in the
bacterial communities that contaminated the saunas may
not be a general phenomenon because sampling period,
number, and area in each study were not equivalent.
In this study, the daily variation in bacteria contaminating
the saunas may not be proportional to the number, frequency
of use, or cleanliness condition of users but may be
randomly and opportunistically influenced by the bacterial
species contaminating the user's' bodies. The bacteria
commonly and frequently detected from both saunas can
survive on human body for a relatively longer time than
those rarely detected in one of the saunas. General house
conditions may be a more proper than natural environment
for mesophilic bacteria to grow and for thermophilic bacteria
to survive, considering the plentiful amount of organic
compounds wasted from food.
Our results indicate that both the mesophilic and
thermophilic bacteria detected from the saunas could have
originated from the user's house and not their body because
most people wash their bodies one or two times per day. No
human pathogenic or harmful bacteria were in either sauna
in this study. The bacteria detected from both saunas over
the 6 days provided useful information about the origin of
bacterial communities contaminating saunas because the
saunas are not a proper and stable habitat for mesophilic
bacteria to grow. Saunas may be a specific place to collect
various bacteria from the human body but not permit
growth of mesophilic bacteria. Most pathogens can grow at
mammalian temperature but do not grow at the temperature
of a sauna. Bacteria can convert from saprophytism to
virulence only while growing continuously (47).
REFERENCES
1)
Lee JY, Park DH. Characterization of bacterial com-
munity contaminating floor of a hot and dry sauna. J
Bacteriol Virol 2012;42:313-20.
2) Bott TL, Brock TD. Bacterial growth rates above 90℃
in Yellowstone hot springs. Science 1969;164:1411-2.
3) Brock TD, Freeze H. Thermus aquaticus gen. n. and sp.
n., a non-sporulating extreme thermophile. J Bacteriol
1969;98:289-97.
4)
Walsh C, Meade J, McGill K, Fanning S. The bio-
diversity of thermoduric bacteria isolated from whey. J
Food Safe 2012;32:255-61.
5)
Banykó J, Vyletelová M. Determining the source of
Bacillus cereus and Bacillus licheniformis isolated from
raw milk, pasteurised milk and yoghurt. Lett Appl
Microbiol 2009;48:318-23.
6)
Brock TD, Boylen LK. Presence of thermophilic
bacteria in laundry and domestic hot-water heaters. Appl
Microbiol 1973;25:72-6.
7) Pask-Hughes R, Williama RA. Extremely thermophilic
gram-negative bacteria from hot tap water. J Gen
Microbil 1975;88:321-8.
8) Oshima T. Imahori K. Description of Thermus thermo-
philus (Yoshida and Oshima) comb. Nov. A non-
sporulating thermophilic bacterium from a Japanese
thermal spa. Int J Syst Bacteriol 1974;24:102-12.
9) Ward J, Cockson A. Studies on a thermophilic bacillus:
its isolation, properties, and temperature coefficient of

202 BS Kim, et al.
growth. J Bacteriol 1972;112:1040-2.
10) Metzger WJ, Patterson R, Fink J, Semerdjian R, Roberts
M. Sauna-takers disease. Hypersensitivity pneumonitis
due to contaminated water in a home sauna. JAMA
1976;236:2209-11.
11) Scheldeman P, Pil A, Herman L, De Vos P, Heyndrickx
M. Incidence and diversity of potentially highly heat-
resistant spores isolated at dairy farms. Appl Environ
Microbiol 2005;71:1480-94.
12) Murai R, Yoshida N. Geobacillus thermoglucosidasius
endospores function as nuclei for the formation of single
calcite crystals. Appl Environ Microbiol 2013;79:3085
-90.
13)
Yokoya F, York GK. Effect of several environmental
conditions on the "thermal death rate" of endospores of
aerobic, thermophilic bacteria. Appl Microbiol 1965;
13:993-9.
14)
Martin PA, Travers RS. Worldwide abundance and
distribution of Bacillus thuringiensis isolates. Appl
Environ Microbiol 1989;55:2437-42.
15)
Stefanic P, Mandic-Mulec I. Social interactions and
distribution of Bacillus subtilis pherotypes at microscale.
J Bacteriol 2009;191:1756-64.
16) Cheung PY, Kinkle BK. Mycobacterium diversity and
pyrene mineralization in petroleum-contaminated soils.
Appl Environ Microbiol 2001;67:2222-9.
17) Kimura H, Asada R, Masta A, Naganuma T. Distribution
of microorganisms in the subsurface of the manus
basin hydrothermal vent field in Papua New Guinea.
Appl Environ Microbiol 2003;69:644-8.
18) Vary PS, Biedendieck R, Fuerch T, Meinhardt F, Rohde
M, Deckwer WD, et al. Bacillus megaterium--from
simple soil bacterium to industrial protein production
host. Appl Microbiol Biotechnol 2007;76:957-67.
19) Ferreira AC, Nobre MF, Rainey FA, Silva MT, Wait R,
Burghardt J, et al. Deinococcus geothermalis sp. nov.
and Deinococcus murrayi sp. nov., two extremely
radiation-resistant and slightly thermophilic species from
hot springs. Int J Syst Bacteriol 1997;47:939-47.
20) Scott RM. Bacterial endocarditis due to Neisseria flava.
J Pediatr 1971;78:673-5.
21) Schmidt B, Mulder IE, Musk CC, Aminov RI, Lewis
M, Stokes CR, et al. Establishment of normal gut
microbiota is compromised under excessive hygiene
conditions. PLoS One 2011;6:E28284.
22)
Staley JT, Irgens RL, Brenner DJ. Enhydrobacter
aerosaccus gen. nov., sp. nov., a gas-vacuolated,
facultatively anaerobic, heterotrophic rod. Int J Syst
Evol Microbiol 1987;37:289-91.
23)
Nishimura Y, Ino T, Lizuka H. Acinetobacter radio-
resistens sp. nov. isolated from cotteon and soil. Int J
Syst Evol Microbiol 1988;38:209-11.
24) Koburger JA, May SO. Isolation of Chromobacterium
spp. from foods, soil, and water. Appl Environ Microbiol
1982;44:1463-5.
25) Chen WM, Cho NT, Yang SH, Arun AB, Young CC,
Sheu SY. Aquabacterium limnoticum sp. nov., isolated
from a freshwater spring. Int J Syst Evol Microbiol
2012;62:698-704.
26) Yoon JH, Kim IG, Oh TK. Acinetobacter marinus sp.
nov. and Acinetobacter seohaensis sp. nov., isolated
from sea water of the Yellow Sea in Korea. J Microbiol
Biotechnol 2007;17:1743-50.
27)
Vaz-Moreira I, Nobre MF, Nunes OC, Manaia CM.
Gulbenkiania mobilis gen. nov., sp. nov., isolated from
treated municipal wastewater. Int J Syst Evol Microbiol
2007;57:1108-12.
28) Gomila M, Bowien B, Falsen E, Moore ER, Lalucat J.
Description of Pelomonas aquatica sp. nov. and
Pelomonas puraquae sp. nov., isolated from industrial
and haemodialysis water. Int J Syst Evol Microbiol
2007;57:2629-35.
29)
Nelson YM, Lion LW, Ghiorse WC, Shuler ML.
Production of biogenic Mn oxides by Leptothrix
discophora SS-1 in a chemically defined growth
medium and evaluation of their Pb adsorption char-
acteristics. Appl Environ Microbiol 1999;65:175-80.
30) Lau HT, Faryna J, Triplett EW. Aquitalea magnusonii
gen. Nov., sp. nov., a novel Gram-negative bacterium
isolated from a humic lake. Int J Syst Evol Microbiol
2006;56:867-71.
31) Gong XC, Liu ZS, Guo P, Chi CQ, Chen J, Wang XB,
et al. Bacteria in crude oil survived autoclaving and
stimulated differentially by exogenous bacteria. PLoS
One 2012;7:e40842.
32)
Loper JE, Lindow SE. A biological sensor for iron
available to bacteria in their habitats on plant surfaces.
Appl Environ Microbiol 1994;60:1934-41.

Variation and Characterization of Bacteria Contaminating Dry Saunas
203
33) Pearson HA. Rumen microbial ecology in mule deer.
Appl Microbiol 1969;17:819-24.
34) Pernthaler J, Amann R. Fate of heterotrophic microbes
in pelagic habitats: focus on populations. Microbiol
Mol Biol Rev 2005;69:440-61.
35) Romero-Steiner S, Witek T, Balish E. Adherence of skin
bacteria to human epithelial cells. J Clin Microbiol
1990;28:27-31.
36) Bowers RM, Sullivan AP, Costello EK, Collett JL Jr,
Knight R, Fierer N. Source of bacteria in outdoor air
across cities in the midwestern United State. Appl
Environ Microbiol 2011;77:6350-6.
37) Garcia SL, Jangid K, Whitman WB, Das KC. Transition
of microbial communities during the adaption to
anaerobic digestion of carrot waste. Bioresour Technol
2011;102:7249-56.
38)
Taylor WI, Schelhart D. Effect of temperature on
transport and plating media for enteric pathogens. J
Clin Microbiol 1975;2:281-6.
39)
Ratkowsky DA, Olley J, McMeekin TA, Ball A.
Relationship between temperature and growth of
bacterial cultures. J Bacteriol 1982;149:1-5.
40)
Ratkowsky DA, Lowry RK, McMeekin TA, Stokes
AN, Chandler RE. Model for bacterial culture growth
rate throughout the entire biokinetic temperature range.
J Bacteriol 1983;154:1222-6.
41)
Nakasaki K, Sasaki M, Shoda M, Kubota H. Char-
acteristics of mesophilic bacteria isolated during thermo-
philic composting of sewage sludge. Appl Environ
Microbiol 1985;49:42-5.
42) Brock TD, Freeze H. Thermus aquaticus gen. n. and sp.
n., a nonsporulating extreme thermophile. J Bacteriol
1969;98:289-97.
43) Verduin CM, Hol C, Fleer A, Van Dijk H, Van Belkum
A. Moraxella catarrhalis: from emerging to established
pathogen. Clin Microbiol Rev 2002;15:125-44.
44) Vela AI, Arroyo E, Araqón V, Sánchez-Porro C, Latre
MV, Cerdà-Cuéllar M, et al. Moraxella pluranimalium
sp. nov., isolated from animal specimens. Int J Syst
Evol Microbiol 2009;59:671-4.
45)
Kong HH, Oh J, Deming C, Conlan S, Grice EA,
Beatson MA, et al. Temporal shifts in the skin
microbiome associated with disease flares and treatment
in children with atopic dermatitis.Genome Res 2012;
22:850-9.
46) Morita RY. Psychrophilic bacteria. Bacteriol Rev 1975;
39:144-67.
47)
Toledo-Arana A, Dussurget O, Nikitas G, Sesto N,
Guet-Revillet H, Balestrino D, et al. The Listeria
transcriptional landscape from saprophytism to virulence.
Nature 2009;459:950-6.