This study examined the effects of a common student laundry method at Bates College on the growth of three normal skin bacteria: Staphylococcus aureus, Pseudomonas spp., and Escherichia coli. Twenty students wore clean cotton socks for a day, then one sock from each pair was washed using the common method of a cold/warm cycle with bleach-free detergent. Washing most inhibited growth consistent with S. aureus, but was less effective against E. coli and Pseudomonas spp. More growth of Pseudomonas spp. occurred in tightly-knit female socks than male socks. The study aimed to understand implications for communal laundering and bacterial transmission between students.

1. Student Laundry Methods at Bates College:
Bacterial Growth Inhibition of
Three Normal Microbiota Species
Presented to the Faculty of the
Program in Environmental Studies | Health
Bates College
In partial fulfillment of the
requirements for the degree of the
Bachelor of Science
By
James Jhun
Thesis Advisor: Karen Palin
Lewiston, Maine
December 18th
, 2015

2. 1 |
Acknowledgements
I could not have completed this thesis without the expertise and guidance of Karen Palin from
start to finish, and I can attribute much of my knowledge and interest in the fields of
microbiology and public health to her. I would also like to thank Lee Abrahamsen for the use of
her lab and the equipment within, Beth Whalon for providing materials and guidance regarding
research methods, and the twenty participants – and their feet – for their willingness and
commitment to be research samples in this study.

3. 2 |
Table of Contents
Abstract...........................................................................................................................................3
Introduction....................................................................................................................................3
Laundry at Bates College.........................................................................................................3
Cold/Warm Cycle and Bleach-Free Detergent ........................................................................4
Selected Bacterial Species........................................................................................................4
Materials and Methods..................................................................................................................6
Sampling...................................................................................................................................6
Bacterial Growth on TriPlates.................................................................................................7
Bacterial Identification.............................................................................................................8
Results.............................................................................................................................................9
Survey .......................................................................................................................................9
Bacterial Growth and Identification ......................................................................................11
Effect of Cold/Warm Cycle Washing on Selected Bacterial Species......................................12
Discussion......................................................................................................................................13
Methods | Sampling ................................................................................................................13
Methods | Machine Washing and Detergent ..........................................................................13
Results | Correlations.............................................................................................................14
Results | Physical Construction of Cotton Fabrics ................................................................15
Results | Selected Bacterial Species – Size and Sensitivity to Detergent ...............................15
Implications ..................................................................................................................................16
Communal Laundering...........................................................................................................16
Cross-Contamination .............................................................................................................16
Routes of Bacterial Transmission...........................................................................................17
Conclusion ....................................................................................................................................19
Works Cited..................................................................................................................................20
Appendix A – Consent Form........................................................................................................22
Appendix B – Survey ....................................................................................................................23

4. 3 |
Abstract
80% of student participants surveyed regularly used the preset cold/warm cycle to wash all or
part of their laundry in their regular laundry methods using machines in Bates College on-
campus residences. All of the student participants surveyed regularly used bleach-free
detergents. This common laundry method was examined for effects on growth inhibition of the
following normal microbiota species: Staphylococcus aureus, Pseudomonas spp., and
Escherichia coli. Twenty student participants (ten male and ten female) wore clean, new cotton
socks for a day; one sock from each pair was used as a control while the other was washed using
the laundry method above. Washing most inhibited growth consistent with S. aureus, but it was
not as efficient in inhibiting growths consistent with E. coli and Ps. spp. More bacterial growth
consistent with Ps. spp. occurred in the more tightly knit cotton socks distributed to female
participants than in socks distributed to male participants.
Introduction
Laundry at Bates College
The hygienic effectiveness of laundering has been well studied in domestic homes and
hospitals; however, communal laundering in college/university on-campus housing may produce
variables that may impact public health through bacterial survival and transmission. At Bates
College, on-campus houses and dormitories accommodate between 8-165 students, the vast
majority of whom launder using the Maytag brand "Extra Large Commercial Neptune Washer"
washing machines and dryers in their residences. These washing machines have three preset
options for washing: "whites" (hot cycle), "colors" (cold/warm cycle), and "bright colors" (cold
cycle). All wash cycle options have preset water temperatures and wash time (38 minutes). A
survey of 20 Bates students who participated in this study found that 80% of students regularly
used the cold/warm cycle for either all or part of their laundry. In addition, all of the students
surveyed regularly use bleach-free detergents. Therefore, this study examined the cold/warm
cycle washing with detergent not containing bleach to examine growth inhibition of three normal
microbiota species by a normal student laundry method at Bates College.

5. 4 |
Cold/Warm Cycle and Detergent without Bleach
In the past few decades, the average temperature of washing has steadily decreased due to
the cost-efficiencies of low-temperature washing and the need to wash modern fabrics and dyes
at lower at high temperatures (Zoller, 2004). Cold-wash laundering is great for ease of
laundering and energy conservation; however, many studies, including those of Larson & Gomez
(2001) and Elias et al. (2010), have shown that, in comparison to hotter temperatures of washing,
cold-wash laundering with bleach-free detergent significantly reduces the bactericidal effect.
Wicksell et al. (1973) conducted a study using bleach-free detergents with cotton swatches
inoculated with S. aureus and found that washing at 68°C produced a 4 log reduction while
washing at 24°C produced a 1.5 log reduction. Blaser et al. (1984), indicate that the bactericidal
effect is dependent on temperature in the absence of bleach, although the inclusion of bleach
makes both hot and cold cycles equally bactericidal.
Liquid laundry detergents contain surfactants that work to rid fabrics of dirt, oils, and
microbes (Bhat et al., 2011). The inclusion of bleach drastically increases bactericidal and
bacteriostatic properties (Blaser et al., 1984); however, the widespread use of detergent without
bleach by students - survey responses indicated that 100% of study participants regularly use
detergent without bleach - shifts the focus of our study to the effects of cold/warm cycle washing
with detergent not containing bleach.
Selected Bacterial Species
This study examined bacterial growth of the normal human microbiota, Staphylococcus
aureus, Pseudomonas aeruginosa, and Escherichia coli before and after washing. Scott et al.
(1982) investigated microbial contamination in domestic environments and found that the most
frequently isolated bacterial species was S. aureus; Ps. aeruginosa and E. coli were also
commonly found. Furthermore, Blaser et al. (1984) found that the most common Gram-negative

6. 5 |
rod found in used sheets and clothing included Pseudomonas spp., while the most common
Gram-positive coccus was S. aureus. The growth of these three selected bacterial species are
examined in many studies on laundering (Gerba & Kennedy, 2007; O’Toole et al., 2009; Blaser
et al., 1984), which is why this study examined growth of these organisms as well.
S. aureus is a Gram-positive, non-motile coccus found in about 30% of healthy humans
in the nose, throat, and on the skin (Wertheim et al., 2005). It is important to note that S. aureus
may also be present as MRSA (methicillin resistant S. aureus), a dangerous pathogen (Pottinger,
2013). Infections are caused by S. aureus penetrating a wound or broken skin, and this can cause
minor skin infections (like pimples or impetigo), boils, scalded skin syndrome, meningitis,
osteomyelitis, endocarditis, and even lead to pneumonia (Wertheim et al., 2005). For
immunocompromised individuals, an infection can be life-threatening.
P. aeruginosa is a Gram-negative aerobic rod. It may be part of normal human skin flora
and can infect the urinary tract, airway, burns, and wounds (Ryan, 2004). It is a versatile
bacterium, as it grows at a variety of temperatures and can utilize a diverse range of nutrients,
making it a potentially dangerous pathogen (Ryan, 2004). Furthermore, Legnani & Leoni (1997)
found that Ps. aeruginosa survives well inside washing machines. It is the most common cause
of infections of the outer ear and burn wounds, as well as a common cause of dermatitis, skin
rash, and swimmer's ear as a result of contact with contaminated water (Balcht & Smith, 1994).
In addition, infections of the kidneys and lungs can be fatal, especially in immunocompromised
individuals (Balcht & Smith, 1994). It is used for laundering studies because it can grow in water
and can cause serious infections (Balcht & Smith, 1994).
E. coli is a Gram-negative, anaerobic facultative rod. It is found in the mammalian gut,
along with many other beneficial bacteria (Eckburg et al., 2005). Most strains of E. coli are not

7. 6 |
dangerous, but a pathogenic strain like E. coli O157:H7, which produces Shiga toxin, can cause
serious, often life-threatening, infections (Karch et al., 2005). Other virulent strains can cause
meningitis, gastroenteritis, and urinary tract infection (Karch et al., 2005). Most cases of
transmission occur through the consumption of contaminated foods; however, transmission can
also occur through laundering (Fig. 3). The results of a study using low-temperature washing
with bleach-free detergents showed that there was an estimated 0.1 grams of fecal matter per pair
of laundered underwear (Gerba & Kennedy, 2007). Fecal matter is of concern because it may not
only contain E. coli, but also hepatitis A virus, Salmonella spp., and norovirus (Gerba &
Kennedy, 2007).
Materials & Methods
Sampling
Pairs of socks were distributed to 10 male and 10 female students to be worn for an entire
day, from when they woke up to when they slept. Target brand scks distributed to male students
weighed about 23 g and Hanes brand socks distributed to female students weighed about 16 g.
All socks were made of about 70% cotton and other fibers, including spandex and polyester. A
survey was distributed to participants in order to gather information about each participant’s
activities while wearing the socks and about their regular laundering processes (see Appendix A
for consent form and Appendix B for survey).
Socks were collected the next day and one sock from each pair was washed in a
cold/warm cycle. Each wash contained a dirty sock and clean clothes - four cotton t-shirts and 2
large cotton shower towels (Target) - to simulate a normal laundering process. Approximately 25
mL of bleach-free detergent (Era 2X Ultra Active Stainfighter liquid detergent) was used in
every wash. One of three different washing machines (Maytag “Extra Large Commercial
Neptune Washer” Model: MAH21PRAWW) in an on-campus residence was used for each wash.

8. 7 |
Upon completion of the wash cycles, washed socks were removed from the machine using sterile
tongs, placed in a clean bag, and transported to the laboratory.
Bacterial Growth on TriPlates
The washed socks and control socks were immersed in 250 mL of nutrient broth
(Acumedia - Neogen, Lansing, MI) for 24 hours at room temperature. After incubation, each
broth was inoculated onto a TriPlate (Hardy Diagnostics, Santa Maria, CA), containing Vogel-
Johnson, cetrimide, and MacConkey agars (Fig. 1). Each agar was inoculated using the streak-
plate method for isolation. Control cultures on TSA obtained from the Bates College Department
of Biology (courtesy of Beth Whalon) were streaked onto TriPlates as follows: S. aureus on
Vogel-Johnson, Ps. aeruginosa on cetrimide, and E. coli on MacConkey (Fig. 2). All inoculated
TriPlates were incubated at 37°C for 24 hours.
Figure 1. New TriPlate before inoculation. Media types Figure 2. Control cultures of (clockwise from top) S. aureus,
clockwise from top) are Vogel-Johnson, Cetrimide, and Ps. aeruginosa, and E. coli on a TriPlate after incubation at
MacConkey. 37°C for 24 hours.

9. 8 |
Bacterial Identification
Vogel-Johnson agar was used to select for growth of S. aureus, identified on the medium
as small black colonies producing an agar color change to yellow. Cetrimide agar was used to
select for growth of Ps. aeruginosa, identified on the medium as white or transparent colonies
producing an agar color change to blue, blue-green, or yellow-green. Bacterial species other than
Ps. aeruginosa grew on cetrimide agar; Biolog GEN III Microplates (Biolog Inc., Hayward,
CA), were used to further characterize bacterial species that grew on cetrimide agar.
MacConkey agar was used for the selective growth of Gram-negative, lactose-fermenting
enteric bacilli. They appear as pink/purple colonies, which were further differentiated on Eosin
methylene blue (EMB) agar (Acumedia - Neogen, Lansing, MI) to identify E. coli, which
appears as a dark purple colonies with a metallic green sheen. Control cultures of E. coli
(courtesy of Beth Whalon, Bates College Department of Biology) were grown on EMB agar for
comparative identification. Colonies that appeared dark purple with a metallic green sheen were
consistent with an identification of E. coli. Colonies that did not present phenotypes consistent
with the control culture of E. coli on EMB were tested via indole, methyl red, and Voges-
Proskauer tests (TSB medium from Sigma-Aldrich, St. Louis, MO; MR-VP medium from
Remel, Lenexa, KS; reagents from BD, Sparks, MD). An IMViC result of + + - - was
presumptively identified as E. coli. Colonies identified as E. coli, Ps. spp. and others, and S.
aureus were quantified relative to a control in Table 1.

10. 9 |
Table 1. Bacterial growth was quantified using a scale from 1+ to 4+ relative to growth on the control
culture TriPlate. 1+ indicates up to about 25% of growth on control. 2+ indicates about 50% of growth on
control. 3+ indicates about 75% of growth on control. 4+ indicates the same amount of growth as control.
1+ 2+ 3+ 4+
Vogel-Johnson
Cetrimide
MacConkey
Results
Survey
The survey collected information about participants’ daily activities possibly affecting
bacteria present on their socks. Responses show that the average length of time that socks were
worn was generally consistent between men and women (Table 2). There were two outliers;
participants M1 and W7 wore their socks for only about half of the total average time (Table 2).
Men tended to wear either casual or athletic shoes, whereas women wore mostly boots.
Sixteen participants used regular soap/body wash on their feet, three participants did not indicate
that they regularly used any products on their feet, and one participant (W8) used antibacterial
soap/body wash on her feet. Bacterial growth from socks worn by W8 was similar to that of
other participants.

11. 10 |
Table 2. Survey responses about participants’ daily activities possibly affecting bacteria present on their socks. Listed here are
participants’ average amount of time wearing the socks and the number of participants for each type of shoe worn and each
regularly used product on feet. n=20 (10 men and 10 women)
Time Socks Worn Types of Shoes Worn Regularly Used Products on Feet
Average (hours) Boots Casual Athletic
Regular
Soap/Body Wash
Antibacterial
Soap/Body Wash None
Men 11.88 0 8* 6* 8 0 2
Women 12.82 7* 3* 3* 8 1 1
Total 7* 11* 9* 16 1 3
* = Different shoe types sometimes worn while wearing socks. These numbers indicate the net total of types of shoes worn.
Students were also surveyed about laundry methods they regularly used. 80% of
participants regularly use Tide® or Tide® Pods, while the remainder used a variety of other
detergents (Table 3). 55% of participants regularly used only “colors” cycles, while 25%
regularly used “whites” and “colors” cycles, and 20% regularly used “bright colors” (Table 3).
Table 3. Laundry methods used by student participants in the study. Numbers of male and female participants who
regularly use the listed laundry detergent and wash cycles. n=20 (10 men and 10 women)
Laundry Detergent Brand Wash Cycle
Tide® Tide® Pods Other Colors Whites/Colors Bright Colors
Men 5 4 1 8 1 1
Women 5 2 3 3 4 3
Total 10 6 4 11 5 4
Bacterial Growth and Identification
Bacterial growth was assessed qualitatively on TriPlates (Table 4). Bacterial species other
than Ps. aeruginosa grew on cetrimide agar. Some of these species were presumptively
identified using Biolog GenIII as Pantoea spp., Enterobacter spp., and Serratia spp.

12. 11 |
Table 4. Growth of S. aureus, Ps. spp., and E. coli on TriPlates for unwashed (control) and washed socks. Socks were
cultured in nutrient broth for 24 hours at room temperature, streaked onto TriPlates, and incubated for 24 hours at 37°C.
Growth was assessed relative to 4+ control cultures on TriPlates (Table 1). n=20 (10 men and 10 women)
Table 4A. Male participants
S. aureus
Unwashed Washed
Ps. spp. and others
Unwashed Washed
E. coli
Unwashed Washed
M1 2 0 2 0 0 0
M2 4 0 4 2 0 2
M3 3 0 2 0 0 0
M4 4 0 2 0 3 0
M5 3 0 3 0 0 1
M6 4 0 2 0 3 0
M7 4 0 1 0 0 0
M8 2 0 2 0 2 1
M9 3 0 3 0 1 0
M10 4 1 2 0 0 1
Table 4B. Female participants
S. aureus
Unwashed Washed
Ps. spp. and others
Unwashed Washed
E. coli
Unwashed Washed
W1 0 0 2 1 0 0
W2 2 0 3 1 2 1
W3 3 0 3 2 0 0
W4 3 1 3 2 0 0
W5 0 0 2 1 1 1
W6 0 0 2 2 0 1
W7 0 0 2 0 2 0
W8 3 0 3 1 0 1
W9 4 0 1 1 0 0
W10 4 0 2 0 0 1

13. 12 |
Effect of Cold/Warm Cycle Washing on Selected Bacteria
For men, growth consistent with S. aureus was cultured from all unwashed (control)
socks and only on 1 of the 10 washed socks. Growth consistent with Ps. spp. was cultured from
all unwashed (control) socks and only on 1 of the 10 washed socks. Growth consistent with E
coli was cultured from 4 of the 10 unwashed (control) socks and 4 of the 10 washed socks (Fig.
4).
For women, growth consistent with S. aureus was cultured from 6 of the 10 unwashed
(control) socks and only on 1 of the 10 washed socks. Growth consistent with Ps. spp. was
cultured from all unwashed (control) socks and on 8 of the 10 washed socks. Growth consistent
with E coli was cultured from 3 of the 10 unwashed (control) socks and 5 of the 10 washed
socks.
Figure 4. The number of male and female participants’ unwashed and washed socks contaminated with Staphylococcus aureus,
Pseudomonas spp. and others, and Escherichia coli. n=20 pairs of socks from men and women.

14. 13 |
Discussion
Methods | Sampling
Research in earlier studies have inoculated cotton swatches with determined amounts of
S. aureus, Ps. aeruginosa, and E. coli to assess bacterial survival rates after cold cycle washing
with detergent not containing bleach (Gerba & Kennedy, 2007; O’Toole et al., 2009). In this
study, it was not possible to wash purposefully contaminated clothes in communal washing
machines to examine the survival of pathogens. Here, a sampling method was used to examine
bacterial survival under a commonly used laundry method; pairs of socks were worn by
participants and one sock served as a control while the other was washed. We assumed that both
socks in each pair worn by participants contained similar amounts and species of bacteria.
However, growths consistent with E. coli were present on more washed socks than unwashed
(control) socks for female participants (Fig. 4), which may indicate a difference in E. coli
presence between socks in a pair.
Methods | Machine Washing and Detergent
Socks were washed with four clean cotton t-shirts and two clean cotton shower towels to
replicate a normal laundry process. Three washing machines were used interchangeably to
account for potential differences in performance; no differences were found. The detergent used
in each cold/warm cycle, Era detergent, was chosen because it does not contain bleach and it is
available to students at the Bates school store. Survey responses from participants in the study
indicated that 80% of the twenty participants regularly use Tide® detergent or Tide® Pods
(Table 3). Using Tide® detergent or Tide® Pods would have more accurately replicated
commonly used laundry methods at Bates College; however, the most important factor in
determining which detergent to use was the lack of bleach, and 100% of participants indicated
that they regularly used bleach-free detergents (Table 3). Studies have shown that, in the absence

15. 14 |
of bleach, bacterial growth is dependent on the cold/warm cycle’s temperature of washing
(Blaser et al., 1984), thus the type of bleach-free detergent may not matter.
Results | Correlations
Survey responses were compared to bacterial analyses to investigate correlations between
participants’ activities while wearing the socks and bacterial growth on control socks. There
were no correlations between the total length of time that the socks were worn and growth of S.
aureus, Ps. spp. and others, and E. coli from unwashed socks (Figure 5). Participants with
outlier lengths of time socks worn (M1, W7, and W8) did not produce outlier amounts of growth
for any of the selected bacterial species. In addition, no differences in bacterial analyses were
found between participants who engaged in sports/physical activities and those who did not. All
bacterial growths were included in the analysis.
Figure 5. Growth of selected bacterial species vs. total time socks were worn by male and female participants in the
study. n = 20 participants (10 male and 10 female)

16. 15 |
Results | Physical Construction of Cotton Fabrics
Growth consistent with Ps. spp. was cultured from eight female washed socks and only
on male washed sock (Fig. 4). Most variables that could have caused this difference were
controlled for (i.e. cold/warm wash cycle, detergent used, and mostly cotton sock material).
However, a notable difference between male and female socks was the physical construction of
the sock, in which the size of the stitching in socks distributed to male participants was larger
than that of socks distributed to female participants. Leonas (1993) studied the antimicrobial
properties of surgical gowns and found that variations in fabric construction influence microbial
adherence. A tighter construction of the cotton sock could act as a filter for bacteria, resulting in
more bacterial retention post-wash. This may explain the findings reported in this study.
Results | Selected Bacterial Species – Size and Sensitivity to Detergent
Differences in growths consistent with the three selected bacterial species may also be
explained by differences in bacterial size. O’Toole et al. (2009) found that larger microorganisms
were retained more on cotton swatches during machine washing. S. aureus is only 0.5-1.0 µm in
diameter, while E. coli can be ~2.5 µm long and ~1.5 µm in diameter, and Ps. spp. are 1.5-5.0
µm long and 0.5-1.0 in diameter (Osiro et al., 2012; Siegrist, 2010). S. aureus is the smallest of
the three selected bacterial species (though it may cluster), and it is more sensitive to metabolic
inhibition by detergents than Gram-negative rods (Baker et al., 1941). This may explain why
washing with bleach-free detergent inhibited more growth consistent with S. aureus than growth
consistent with E. coli and Ps. spp. The size of the bacteria and the sensitivity to detergent may
have been factors in bacterial growth inhibition.

17. 16 |
Implications
Communal Laundering
All on-campus residences at Bates College have at least one washing machine and one
dryer which are used communally; communal laundering can be associated with increased risk of
infection. A study of 238 New York City households found that the use of a communal laundry
and non-bleach detergents was associated with increased risk of infection, while non-communal
laundering methods did not produce evidence of association with risk of infection (Larson et al.,
2004). An investigation into an outbreak of MRSA skin infections in a U.S. county jail reported
an immediate decline in skin infections after correcting their communal laundry methods;
machines were overloaded and used cold cycles with bleach-free laundry soap (Elias et al.,
2010). Based on these previous studies, a recommendation from this study is that students avoid
overloading their laundry and utilize the hot “whites” cycle and bleach when possible.
Cross-Contamination
Using the cold/warm cycle to launder allows a student to wash all of his/her laundry
together, regardless of color or type. This allows for cross-contamination within a load of
laundry; bacterial transmission can occur from fabrics that come into frequent contact with the
environment and body cavities, such as socks and undergarments, to fabrics that come into
frequent contact with the face, like pillow sheets and towels. Cross-contamination may also
occur person-to-person in the event that a single wash cycle is shared between friends or
roommates that combine their laundry.
Cold/warm cycle washing using bleach-free detergent has been found to be ineffective in
reducing bacterial growth, and as a result, may produce cross-contamination to other fabrics in a
load. In a study using cotton swatches inoculated with S. aureus, cold cycle washing (30°C) with
liquid non-bleach detergent produced only a 1-2 log reduction in bacterial growth, as well as

18. 17 |
cross-contamination between contaminated and sterile swatches (International Scientific Forum
on Home Hygiene, 2010). Furthermore, another study, using cotton swatches inoculated with S.
aureus and E. coli, found that cold/warm washing at 20-30°C with detergent not containing
bleach produced a 2-3 log reduction for these selected bacterial species, as well as cross-
contamination of sterile cotton swatches and sterile cotton T-shirts and underwear (Gerba &
Kennedy, 2007). Cross-contamination is a possibility with the selected bacteria in this study, and
it is magnified by low log reductions produced by cold/warm cycle washing with bleach-free
detergent.
Figure 6. Routes of bacterial transmission from the environment to a person through laundering. Adapted from “The
infection risks associated with clothing and household linens in home and everyday life settings, and the role of
laundry,” by Bloomfield et al. (2011), International Scientific Forum on Home Hygiene.
Routes of Bacterial Transmission
The process of manually transporting washed laundry from the washer to the dryer
exposes skin to microbes that survive the washing process. Routes of bacterial transmission from
the environment to the body can occur through hand-to-laundry contact and can transmit to the
three major portals of entry (Fig. 6). Through hand-to-laundry contact, bacterial transmission can
occur between people when a student removes another student’s washed laundry from the
washing machine in order to use the machine themselves. Personal observations indicate that this

19. 18 |
is a common occurrence when the washing machines are used frequently, like on weekends.
Without washing hands after touching laundry, bacterial transmission may occur to the skin,
mouth, and eyes & nose (Fig. 3).
This study found that washing socks through cold/warm cycle with bleach-free detergent
did not inhibit growths consistent with S. aureus on all socks. Sattar et al. (2001) found that the
rate of transfer of S. aureus from moist cotton fabric to dry hands was moderate (0.4%), and
Sattar & Springthorpe (1996) found that S. aureus survives well on hands and can withstand
drying on hands and fabrics. However, we found that only few washed socks produced growth
consistent with S. aureus, which raises little concern for bacterial transmission despite the
moderate transfer rate from moist cotton fabrics to hands and survival of S. aureus (Sattar et al.,
2001; Sattar & Springthorpe, 1996).
Growth consistent with E. coli occurred on more washed socks, but this is of little
concern for bacterial transmission to the body since Mackintosh & Hoffman (1984) found that E.
coli did not survive well on hands, indicated by a <1% recovery on hands contaminated by moist
cotton fabrics after five minutes. Growth consistent with Ps. spp. may present the most concern
for bacterial transmission among the bacteria studied here. Growth consistent with Ps. spp. was
found on 8 washed socks worn by female participants; Mackintosh & Hoffman (1984) found a
moderate rate of transfer (0.36%) from moist cotton fabric to dry hands and a 49% recovery of
Ps. aeruginosa on contaminated hands after five minutes. Hand-to-laundry contact may produce
moderate transfer of Ps. aeruginosa, which was shown to survive well on hands (Mackintosh &
Hoffman, 1984); hands should be washed after handling wet laundry.

20. 19 |
It is important to note that the laundering process can involve tumble hot drying, which
reduces the numbers of bacteria that survive the washing process (Tano & Melhus, 2014).
Microbes have been shown to survive the drying process, especially if the clothes are still damp
upon completion of the drying cycle (McNeil, 1963). In on-campus residences, Maytag dryers
run at a set time of 60 minutes but personal observations have shown that clothes can remain
damp if the laundry load is too large. If clothes remain damp after the dry cycle, a student must
either run the dryer again or remove the clothes. During periods of frequent laundry machine use,
students may remove another student’s laundry in order to use the drying machine, if clothes
remain damp, bacterial transmission may occur from person to person.
Conclusion
This study has shown that when socks washed in the cold/warm cycle with bleach-free
detergent are cultured, bacterial growth consistent with an identification of E. coli and Ps. spp.
occurs in more socks than that of S. aureus. Because machine washing at warmer temperatures
reduces bacterial survival and bleach is an effective antimicrobial agent (Blaser et al., 1984), the
hot “whites” cycle and bleach may be more effective in inhibiting bacterial growth.
The results of this study are consistent with those of a study on the survival of S. aureus,
E. coli, and Ps. aeruginosa in a cold-wash cycle with non-bleach detergent, which found that E.
coli and Ps. aeruginosa survived in greater numbers than S. aureus (Munk et al., 2001).
However, Munk et al. (2001) also found that growth of Ps. aeruginosa and E. coli was inhibited
after machine drying at 53 C for 60 min (similar to Bates on-campus drying machines), though S.
aureus was able to survive this drying process. This study found that few washed socks produced
bacterial growth consistent with S. aureus. Therefore, the process of cold/warm cycle washing
with non-bleach detergent and machine drying should be effective against the three selected
bacterial species.

21. 20 |
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Appendix A
Consent Form
This is a senior thesis study being conducted by
James Jhun ‘16
for the
Environmental Studies - Health major
under the advisorship of
Prof. Karen Palin
This is a study on the effectiveness of laundering in Bates College dormitories. It is common for students
living in Bates dormitories to regularly launder using cold wash cycles, which has been shown to be less
effective in killing bacteria than hot wash cycles.1 This study examines whether the “typical” laundry
process in Bates College dormitories eliminates Escherichia coli, Staphylococcus aureus, and
Pseudomonas aeruginosa, which are commonly found as normal microbiota, and evaluates their survival
in the cold wash cycle. Pairs of socks will be distributed to Bates College students to be worn for one day.
Socks will be collected and cultured to compare the presence of the above microorganisms before
washing and after cold-cycle washing with bleach-free detergent with the Maytag brand laundry machines
in Bates dormitories.
As a participant in this study, you will be asked to:
1. Wear the pair of new socks provided for you from when you wake up until you sleep
2. After you wear the socks, put them back into the plastic bag that they came in
3. Fill out the attached questionnaire
While wearing the socks:
- Go about your day as you normally would
- Do not get them wet or contaminated by food, drinks, feces, urine, or anything else
- Keep the socks on, but if you choose to shower or partake in physical activities/sports, indicate so
in the attached questionnaire
- You may walk around with socks on and without shoes i.e. around your room
- If you have any questions, email jjhun@bates.edu or call/text at 781-640-0661
Your participation is completely voluntary and can be withdrawn at any time, there is no risk to you, and
only the researcher will store/keep the questionnaires until the end of the semester when the thesis is
completed. Data will be used and presented in aggregate only and no participants will be individually
identified.
As a participant in this study, I agree to the above statements.
Signature ______________________________________ Date ________________
1
Blaser, M. J., P. F. Smith, H. J. Cody, W.-L. L. Wang, and F. M. Laforce. "Killing of Fabric-Associated Bacteria in Hospital Laundry by Low-
Temperature Washing." Journal of Infectious Diseases 149, no. 1 (1984): 48-57.

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Appendix B
Questionnaire
Date: ____________
Sock ID: __________
1. I wore the socks from ____:______ AM to ____:_______ PM on _________________, 2015
2. While wearing the socks, I wore these types of shoes (check all that apply):
__ Casual (sneakers, boat shoes)
__ Dress
__ Athletic (running, sports, cleats)
__ Boots
__ Open-toed
Other: ______________________________
3. Check all that apply:
__ I took a shower before wearing the socks
__ I took the socks off in order to shower and then put them back on
__ I took the socks off for ________ minutes
__ I engaged in sports/physical activities (specify: _______________________________)
for ___________ hour(s) while wearing the socks
__ I walked around with the socks on but without shoes. The location was _________________
4. I regularly use the following products on my feet (check all that apply):
__ Regular soap/body wash
__ Soap/body wash labelled as antibacterial
__ Regular foot powder/spray/lotion
__ Foot powder/spray/lotion for treatment/prevention of athlete’s foot or warts
5. If you got the socks wet, stained, etc… please specify: ______________________________
6. What brand of laundry detergent do you use most often: _____________________________
7. Which cycle on the Bates washing machines do you regularly use? Check all that apply.
__ Whites
__ Colors
__ Bright Colors
8. Would you like to be informed of the results of this study?
__ No
__ Yes, my email is: _______________________@bates.edu