EPA/600/R-17/126 I June 2017
www.epa.gov/homeland-security-research
United States
Environmental Protection
Agency
&EPA
Operational Testing of Floor Cleaning
Cloths for Household Remediation
Following a Large-Scale Biological
Contamination Incident
Office of Research and Development
Homeland Security Research Program

-------
This page left intentionally blank

-------
Operational Testing of Floor Cleaning
Cloths for Household Remediation
Following a Large-Scale Biological
Contamination Incident
Assessment and Evaluation Report
Lukas Oudejans, Ph.D.
National Homeland Security Research Center
Office of Research and Development
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
Abderrahmane Touati, Ph.D. and Eric Morris
Jacobs Technology, Inc.
Research Triangle Park, NC 27709

-------
This page left intentionally blank

-------
Disclaimer
The U.S. Environmental Protection Agency (EPA), through its Office of Research and Development's
(ORD's) National Homeland Security Research Center (NHSRC), funded and managed this investigation
through Contract No. EP-C-15-008, work assignments (WAs) 0-071 and 1-071 with Jacobs Technology,
Inc. (Jacobs). This report has been peer and administratively reviewed and has been approved for
publication as an Environmental Protection Agency document. It does not necessarily reflect the views of
the Environmental Protection Agency. No official endorsement should be inferred. This report includes
photographs of commercially available products. The photographs are included for purposes of illustration
only and are not intended to imply that EPA approves or endorses the product or its manufacturer. EPA
does not endorse the purchase or sale of any commercial products or services.
Questions concerning this document or its application should be addressed to the principal investigator:
Lukas Oudejans, Ph.D.
Decontamination and Consequence Management Division
National Homeland Security Research Center
Office of Research and Development
U.S. Environmental Protection Agency
Mail Code E343-06
109 T.W. Alexander Drive
Research Triangle Park, NC 27711
Telephone No.: (919) 541-2973
E-mail Address: Oudeians.Lukas@epa.gov
i

-------
Acknowledgments
This research effort is part of the U.S. Environmental Protection Agency's (EPA's) Homeland Security
Research Plan (HSRP) to evaluate low tech/self-help techniques for Bacillus (B.) anthracis remediation
operations. Self-help actions may be undertaken at the edge of a contamination zone of a wide-area B.
anthracis spore release to reduce indoor exposure risk outside the contamination zone. Here, a floor
cleaning/dust removal system using disposable dust cloths was evaluated to determine their propensity to
remove spores from indoor flooring materials. The results of this work would inform responders,
governments, and health departments in their guidance development for self-self-help recommendations
to the general public.
This effort was directed by the principal investigator from the Office of Research and Development's
(ORD's) NHSRC, with support of a project team consisting of staff from across EPA. The contributions of
the following individuals have been a valued asset throughout this effort:
U.S. Environmental Protection Agency Project Team
Lukas Oudejans, Principal Investigator; National Homeland Security Research Center
Sang Don Lee and M. Worth Calfee, National Homeland Security Research Center
Leroy Mickelsen and Shannon Serre, Office of Emergency Management
U.S. EPA Technical Reviewers of Report
Joseph Wood, National Homeland Security Research Center
Francisco Cruz, Office of Land and Emergency Management
U.S. EPA Quality Assurance
Eletha Brady Roberts, National Homeland Security Research Center
U.S. EPA Editorial Review
Joan Bursey
Jacobs Technology, Inc.
Abderrahmane Touati, Ph.D.
Eric Morris

-------
Table of Contents
Disclaimer	i
Acknowledgments	ii
Tables	v
Figures	v
Acronyms and Abbreviations	vi
Executive Summary	vii
1	Introduction	1
1.1	Background	1
1.2	Project Description and Objectives	1
2	Experimental Approach	2
2.1	Test Matrix	2
2.2	Experimental Methods and Materials	3
2.2.1	Swifter® Sweeper® Floor Mop System	3
2.2.2	Test Chambers	4
2.2.3	Test Coupon Preparation	5
2.2.4	Coupon and Equipment Sterilization	6
2.2.5	Spore Preparation	6
2.2.6	Coupon Inoculation	7
2.3	Operational Testing of Swiffer® Sweeper® System	7
2.3.1	Operational Testing Modes	7
2.3.1.1	Dry Sweeping Mode	7
2.3.1.2	Wet Mopping Mode	8
2.3.1.3	Dry SweepingAA/et Mopping Mode	8
2.3.2	Sample Coupon Sweeping Pattern	8
3	Sampling Approach	9
3.1	Sampling Site Environmental Conditions	9
3.2	Sampling Strategy	10
3.2.1 Sample Types	10
iii

-------
3.2.2 Sampling Frequency	10
3.3 Sampling Methods	11
3.3.1	Wipe Sampling	11
3.3.1.1	Wipe Sampling Preparation	11
3.3.1.2	Wipe Sampling Procedure	12
3.3.2	Air Sampling with Via-Cell®	14
4	Testing and Measurements	15
4.1	Analytical Procedure	15
4.2	Data Reduction	16
5	Results and Discussion	18
5.1	Post-Decontamination Recoveries	18
5.2	Swiffer® Sweeper® Decontamination Efficacy	19
5.3	Post-Decontamination Swiffer® Sweeper® Cloth Recovery	21
5.4	Spore Aerosolization	23
6	Quality Assurance and Quality Control	24
6.1	Project Documentation	24
6.2	Integrity of Samples and Supplies	24
6.3	Instrument Calibrations	24
6.4	Critical Measurements	25
6.5	NHSRC Biolab Quality Checks	26
6.6	QA Assessments and Response Actions	27
7	Summary	29
8	References	31
iv

-------
Tables
Table ES-1: Summary of Swiffer® Sweeper® Cleaning Tests	vii
Table ES-2: Recoveries from Swiffer® Cloths following Cleaning Treatment	viii
Table 2-1: Test Matrix for Surface Decontamination Studies	3
Table 2-2: Building Materials	5
Table 3-1: Sampling Measurements/Frequency for Swiffer® Evaluation Testing	11
Table 3-2: Sampling Materials and Equipment	12
Table 5-1: Test 1 - Dry Sweeping/Laminate	20
Table 5-2: Test 2 - Dry Sweeping/Vinyl	20
Table 5-3: Test 3 - Wet Mopping/Laminate	20
Table 5-4: Test 4 - Wet Mopping Mode/Vinyl	21
Table 5-5: Test 5 - Dry Sweeping/Wet Mopping/Laminate	21
Table 5-6: Test 6 - Dry Sweeping/Wet Mopping/Vinyl	21
Table 5-7: Post Decontamination Swiffer® Sweeper® Cloth Spore Recoveries	22
Table 5-8: Spore Recoveries in the Aerosol Samples	23
Table 6-1: Instrument Calibration Frequency	24
Table 6-2: DQIs and Acceptance Criteria for Critical Measurements	25
Table 6-3: Additional Quality Checks for Biological Measurements	27
Table 6-4: QA/QC Assessment	28
Figures
Figure 2-1: Swiffer® Sweeper® floor mop system	4
Figure 2-2: Laminate coupon located in acrylic testing chamber with Swiffer® and Via-Cell® ports	5
Figure 2-3: Coupon dimensions	6
Figure 2-4: Aerosol deposition apparatus	7
Figure 2-5: MDI and actuator	7
Figure 2-6: Swiffer® Sweeper® sampling pattern	9
Figure 3-1: Wpe sampling grid, locations, and order	13
Figure 4-1: Bacterial colonies on a spiral-plated agar plate	15
Figure 5-1: Average spore recovery (CFU) after Swiffer® treatment	18
v

-------
Acronyms and Abbreviations
ADA
aerosol deposition apparatus
ATCC
American Type Culture Collection
B.
Bacillus
Biolab
NHSRC Biocontaminant Laboratory
°C
Degree(s) Celsius
CFU
colony-forming unit(s)
cm
centimeter(s)
Dl
deionized
DQI
data quality indicator
EPA
U.S. Environmental Protection Agency
ft
foot/feet
FIFRA
Federal Insecticide, Fungicide, and Rodenticide Act
H2O2
hydrogen peroxide
HSRP
Homeland Security Research Program
in.
inch(es)
Lpm
liter(s) per minute
LR
log reduction
jjm
micrometer(s)
MDI
metered-dose inhaler
min
minute(s)
mL
milliliter(s)
mm
millimeter(s)
NHSRC
National Homeland Security Research Center
NIST
National Institute of Standards and Technology
ORD
Office of Research and Development
PBS
phosphate-buffered saline
PBST
phosphate-buffered saline with 0.05% Tween®20
PRB
polyester-rayon blend
PVC
polyvinyl chloride
QA
quality assurance
QAPP
quality assurance project plan
QC
quality control
RH
relative humidity
rpm
revolution(s) per minute
RSD
relative standard deviation
SSFMS
Swiffer® Sweeper® floor mop system
SD
standard deviation
STS
sodium thiosulfate
TSA
tryptic soy agar
VHP
vaporous hydrogen peroxide
WA
work assignment
vi

-------
Executive Summary
The U.S. Environmental Protection Agency (EPA), Office of Research and Development (ORD),
Homeland Security Research Program (HSRP) continues to strive to protect human health and the
environment from the adverse impacts resulting from acts of terror by investigating the effectiveness and
applicability of technologies for homeland security-related remediation applications.
EPA's HSRP evaluates remediation operations for Bacillus (B.) anthracis contamination, including low
tech/self-help actions that may be undertaken at the edge of a contamination zone of a wide-area B.
anthracis spore release as to reduce indoor exposure risks. Here, a floor cleaning/dust removal system
using disposable dust cloths was evaluated to determine their propensity to remove spores from indoor
flooring materials. The Swiffer® Sweeper® system was selected as a representative method of commonly
encountered dusting approaches that are readily available. The objective of this study was to evaluate the
Swiffer® Sweeper® floor mop system (SSFMS) as a low-tech method to clean indoor residential floors
contaminated with B. anthracis spores (the causative agent of anthrax). The SSFMS can be used with dry
sweeper heads or wet mopping heads.
Two types of flooring surfaces, vinyl and laminate, were contaminated with B. atrophaeus spores (used
as a surrogate for B. anthracis spores). The surfaces were inoculated using a metered-dose inhaler that
dispensed an aerosolized spore preparation (approximately 3*107 spores) onto a 12-inch (in.) x 12-in.
center surface area. Three replicate tests were performed on a total of six test combinations consisting of
the dry sweeping, wet mopping, and dry sweeping followed by wet mopping of vinyl and laminate flooring.
Samples were collected from the following:
•	Three contaminated 35-in. x 35-in. flooring coupons, after cleaning (wipe sampled in four different
areas to assess cleaning efficacy and to measure potential redistribution of spores).
•	Supplied chamber air (filter sampled to assess aerosol formation cleaning of the flooring coupon).
•	Swiffer® Sweeper® cloths used to clean the coupon surfaces.
PHYSICAL REMOVAL RESULTS
Decontamination efficacy for a test combination was expressed as the average log reduction (LR) in the
number of viable spore colony-forming units (CFU) before cleaning as compared with CFU after cleaning.
Results for each SSFMS operation mode/flooring combination are shown in Table ES-1. These data
represent the LR forthe inoculated 12-in. x 12-in. center of the coupon ("hot spot").
Table ES-1: Summary of Swiffer® Sweeper® Cleaning Tests
Swiffer® Cleaning Method
Flooring Material
CFU LR
Average Standard
(n=3) Deviation
Dry sweeping
Laminate
2.1
0.4
Vinyl
2.1
0.6
Wet mopping
Laminate
3.3
0.3
Vinyl
3.0
0.2
Dry sweeping / wet mopping
Laminate
3.4
0.2
Vinyl
3.4
0.2
vii

-------
One Swiffer® cleaning treatment reduced the concentration of spores on the coupon by at least two
orders of magnitude, independent of flooring type or Swiffer® cloth type (wet versus dry). However, results
within each test (three replicates) were highly variable, and none of the three cleaning methods produced
results that were significantly different from each other. Further, the wet methods (wet mopping and dry
sweeping/wet mopping) caused greater spore redistribution across the coupon, i.e., a greater spread of
contamination. Decontamination efficacy was not significantly different for vinyl and laminate flooring.
Sampling of contaminated stainless steel (reference coupons) and flooring coupons yielded spore counts
that were not significantly different from each other.
SPORES ON CLOTHS
The Swiffer® Sweeper® cloths were analyzed to determine the amount of spores collected following a
cleaning treatment. The collection efficiencies of the Swiffer® Sweeper® cloths are shown in Table ES-2.
Recoveries are with respect to the CFU recovered from the same material without the Swiffer cleaning
action (i.e., positive controls; n=3).
Table ES-2: Recoveries from Swiffer® Cloths following Cleaning Treatment
Swiffer® Cloth Condition
Flooring Material
% Recovery
Dry
Laminate
89 ±62
Vinyl
46 ± 7
Wet
Laminate
82 ± 37
Vinyl
47 ± 30
Dry and Wet sweep
Laminate
67 ± 9 [dry]
2.0 ± 0.9 [wet]
Vinyl
40 ± 4 [dry]
3.7 ± 1.2 [wet]
Although the results suggest no statistical difference between recoveries from vinyl versus laminate,
numerically more spores were recovered from the laminate surface than from the vinyl surface,
independent of treatment type. However, significant differences were observed in the average number of
spores recovered on the Swiffer® cloths (wet or dry) used during the first treatment compared to the wet
Swiffer® cloths used in a subsequent treatment. Spore recovery from the wet Swiffer® cloth used during a
dry sweeping/wet mopping operation was less than 4% of the initial spore count inoculated on the coupon
prior to treatment, independent of material type.
The % recoveries for both the wet and dry Swiffer® Sweeper® cloth at the 107 spores level (40-89%
across both materials) are comparable to currently used sponge wipe surface sampling methods 12 with
the significant benefit of the ability to sample a larger (here, 35-in. x 35-in.) surface area than the 12-in. x
12-in.) sponge wipe reference method. As such, Swiffer® Sweeper® cloths may facilitate composite
sampling from wide areas. Such sampling assessment was not part of this study which focused on the
intended physical removal / decontamination of spores from surfaces.
SPORES REAEROSOLIZATION
Aerosol samples were collected to estimate the occurrence and magnitude of aerosolization of viable
spores during each SSFMS treatment process. Less than 0.002% of the surface load was found to be
aerosolized during any of the treatment processes applied, independent of the type of material or
treatment. This study did not address the possible additional spores collected due to reaerosolization by
movements of a person who is cleaning the floor surface.
viii

-------
IMPACT OF STUDY
Floor sweeper and mop combinations such as the Swiffer® Sweeper® are most likely to be used in areas
that are not heavily contaminated. The observed 2.1-3.4 LR in viable spores across all tested cleaning
approaches indicates a reduced indoor exposure risk. A large number of spores, however, remained on
the surface after these cleaning approaches. The presence of a significant number of spores (amounts
similar to the initial spore counts) indicates that these cloths are heavily contaminated following this
cleaning approach. Reuse requires thorough inactivation of spores and may not be suitable for these
intentionally disposable cloths. A homeowner would need to dispose of not only the cloths but presumably
also the mop to avoid cross-contamination of less contaminated areas. Recommended disposal steps
should include inactivation of spores by e.g., soaking of the cloths in diluted bleach priorto disposal.
LIMITATIONS OF STUDY
The reported sweeping and mopping effectiveness should not be compared directly to the Federal
Insecticide, Fungicide, and Rodenticide Act (FIFRA) requirement for registration of a technology as a
sporicide, which requires demonstration of a greater than or equal to 6 LR in viable spores. Instead, the
sweeping and mopping effectiveness should be associated with other low-tech methods that could be
used to reduce indoor exposure potential in less contaminated areas. For example, the use of a robotic
cleaner3 on a laminate surface was reported with a similar number of spores recovered from a hot spot
location as in this study. The data in this report can assist responders, governments, and health
departments in deciding whether to recommend these common cleaning approaches.
ix

-------
1 Introduction
This project evaluated the effectiveness of floor cleaning/dust removal system using disposable dust
cloths such as the Swiffer® Sweeper® floor mop system (SSFMS) as a potential low tech/self-help method
for the removal of spores from hard-surface floors. This research supports the mission of the U.S.
Environmental Protection Agency's (EPA's) Homeland Security Research Program (HSRP) to supply
data, performance measures, and considerations that may be useful during wide-area remediation
following a biological contamination incident. The project addresses HSRP strategic goals as described in
detail in the Homeland Security Strategic Research Action Plan.4
1.1	Background
EPA's HSRP recognizes that the environmental remediation following a chemical, biological, or
radiological event involves an interconnected system of activities that requires coordinated efforts to
optimize cleanup effectiveness, minimize cost and recovery time, and reduce unintended consequences.
Currently, EPA's National Homeland Security Research Center (NHSRC) is evaluating decontamination
technologies for Bacillus (B.) anthracis as part of remediation operations. Following a wide-area
contamination incident, it is likely that homeowners at the edge of an exclusion zone, as advised by their
local governments, may consider taking self-help actions using low tech approaches that would reduce
their indoor exposure risk to biological spores migrated into their residential property. Self-help actions
could include efforts to transfer biological spores from indoor surfaces to waste collection bins by
vacuuming or dust sweeping utilizing disposable cloths or pads. Here, the Swiffer® Sweeper® was
evaluated for use in the physical removal of spores from indoor flooring materials.
1.2	Project Description and Objectives
The purpose of this project was to assess the impact of dusting/sweeping or mopping a hard, nonporous
surface as a potential self-help practice to reduce surface-bound contamination levels for the ultimate
purpose of reducing indoor exposures of home or building owners following a wide-area B. anthracis
spore release. The objectives of this study were to determine the efficacy of a common off-the-shelf dust
cleaner, the Swiffer® Sweeper® system, in removing surrogate spores for B. anthracis from hard floor
surfaces and to determine the extent of redistribution of spores from a contaminated area during a
cleaning event. Redistributed spores were considered to be either spores moved from a highly
concentrated surface area to adjacent sterile surfaces or spores present in air samples due to
reaerosolization.
1

-------
2 Experimental Approach
2.1 Test Matrix
The SSFMS was evaluated for removing spores from surfaces contaminated with B. atrophaeus spores,
used as surrogate spores for B. anthracis. Air samples were also collected during the SSFMS operation
to determine if the SSFMS process created a potential inhalation hazard due to the reaerosolization of
spores.
Two surface types were investigated: laminate wood flooring and vinyl flooring. Known quantities of
spores were deposited on coupons made from these surfaces, and spore removal was performed using
dry sweeping, wet mopping, and dry sweeping followed by wet mopping. The laminate and vinyl surfaces
were sampled for spores, and Swiffer® Sweeper® cloths were also evaluated for viable spores as a
measure of spore removal capability.
The experimental approach used to meet the project objectives is described below:
1.	Preparation of representative coupons of test materials:
Large coupons were made from laminate or vinyl materials with dimensions of 35 inches (in.) x 35
in. for test samples and 14 in. x 14 in. for positive controls.
2.	Sterilization of the coupon materials:
Prior to use, laminate and vinyl coupons were sterilized using a hydrogen peroxide (H2O2) vapor
cycle.
3.	Contamination of coupons:
Coupons were contaminated using an aerosol deposition method, as described in Section 2.2.6.
Briefly, a known quantity of the surrogate organism (1 * 107 colony-forming units [CFU] of B.
atrophaeus spores) was deposited onto the center of a coupon using a metered-dose inhaler
(MDI) fitted into a prefabricated template. The strategy was to inoculate the center of each
coupon with a known concentration of surrogate spores.
4.	Decontamination effectiveness: The spore decontamination approach consisted of cleaning the
entire 35-in. x 35-in. surface of a contaminated coupon with a SSFMS (not sterilized). The overall
contamination reduction achieved with the Swiffer® Sweeper® was then characterized by
sampling for viable spores on the Swiffer cloth (by extraction of the cloth), the coupon surface (by
surface wipe sampling), and the chamber headspace (by air sampling). Decontamination
effectiveness was measured as log reduction (LR) in viable spores for the contaminated center
12-in. x 12-in. area. A summary of the test matrix is presented in Table 2-1.
2

-------
Table 2-1: Test Matrix for Surface Decontamination Studies
Test ID
Mop System
Material
Replicates
Blanks

Samples
1
Dry sweeping
Laminate
3
1
Surface,
air, and Swiffer® cloth
2
Vinyl
3
1
Surface,
air, and Swiffer® cloth
3
Wet mopping
Laminate
3
1
Surface,
air, and Swiffer® cloth
4
Vinyl
3
1
Surface,
air, and Swiffer® cloth
5
Dry sweeping followed
Laminate
3
1
Surface,
air, and Swiffer® cloth
6
by wet mopping
Vinyl
3
1
Surface,
air, and Swiffer® cloth
2.2 Experimental Methods and Materials
This section describes the experimental methods, including the preparation of coupons and application of
the test organism.
2.2.1 Swiffer® Sweeper® Floor Mop System
A hard surface sweeping and mopping tool, the Swiffer® Sweeper®floor mop system http://swiffer.com/en-
us/shop-products/sweepinq/swiffer-sweeper-floor-mop-starter-kit), shown in Figure 2-1, was evaluated for
its ability to remove B. atrophaeus spores from two hard, nonporous floor materials: laminate and vinyl.
Swiffer® Sweeper® is a sweeping and mopping system made by Procter and Gamble (Cincinnati, OH) that
consists of a handle and sweeping/mopping head with a disposable cloth attached. The disposable cloths
are used to remove dust and dirt from a flooring surface. According to the manufacturer, the dry cloth has
deep ridges and grooves that conform to the surface of a floor to trap and lock dirt, dust and hair. The wet
cloth dissolves dirt and grime and traps it away. A combination of both, dry sweeping and then wet
mopping, is recommended for extra dirty floor areas. Here, the SSFMS was evaluated for its ability to
remove spores from the flooring surfaces.
Both dry sweeping cloths (Swiffer® Sweeper® dry cloth refills - unscented, 26.5 * 20.3 cm [10.4 * 8.0 in.])
and wet mopping cloths (Swiffer® wet mopping cloth refills - Gain original scent, 25.4 * 20.3 cm [10.0 *
8.0 in.]) were used for spore removal.
3

-------
http://swiffer.com/en-us/shop-products/sweepinq
Figure 2-1: Swiffer® Sweeper® floor mop system
2.2.2 Test Chambers
The 35-in. x 35-in. vinyl or laminate flooring test coupons were retained in test chambers (91 centimeters
[cm] [length] x 91 cm [width] x 46 cm [height]) constructed of clear acrylic material (5-millimeters [mm]
thickness), with the inside surface coated with chemical-resistant polyvinyl chloride (PVC) Type I antistatic
film (Model # 7524T15, McMaster-Carr, Elmhurst, IL). The top of the test chamber was outfitted with a
grommet port designed to hold the Swiffer® handle and a circular sampling port for a Via-Cell® bioaerosol
sampling cassette (product no. VIA010, Zefon International, Ocala, FL). Another port, located on the front
of the chamber, was outfitted with a 0.2-rnicrometer (pm) sterilizing-grade filter (Aervent™, Millipore,
Molsheim, France) so that clean make-up air could be supplied during air sampling. The test chamber
with Swiffer® unit, sampling ports, and coupon is shown in Figure 2-2. Four test chambers (three for test
coupons and one for a procedural blank) were used for each material type/Swiffer® mode combination.
4

-------
Figure 2-2: Laminate coupon located in acrylic testing chamber with Swiffer® and Via-Cell® ports
2.2.3 Test Coupon Preparation
Two types of coupons, vinyl and laminate flooring, with dimensions of 35 in. x 35 in, for test samples and
14 in. x 14 in. for positive controls, were used in this study (Table 2-2). Thin painter's tape was used to
section the large coupons into nine areas. A 14-in. x 14-in. center sampling area (Figure 2-3) was
designated as the placement area for the aerosol deposition apparatus (ADA), as described in
Section 2.2.6. Both the test coupons and the positive control coupons had an effective inoculation area of
12 in. x 12 in. Only the center area was inoculated to allow for measurement of cross contamination due
to the cleaning from a local hot spot to the surrounding area.
Table 2-2: Building Materials
Material
Description
Manufacturer/
Supplier Name
(location)
Coupon
Surface Size,
Length x
Width (in.)
Inoculated
Surface Size,
Length x
Width (in.)
Material Preparation and
Sterilization
Vinyl
flooring
Vinyl flooring 8 feet
(ft) x 12 ft Casa
Grande beige, precut
sheet vinyl, residential
grade, low gloss, stain
resistant, scratch
resistant, 0,157 in. (4
mm) thick
Casa Grande,
item #L91118X12,
Lowe's Home
Improvement,
Morrisville, NC
35 x 35 or
14X14
12x12
Remove incidental dust and
grime with alcohol wipes (P/N
21910-110, VWR International,
LLC, Radnor, PA).
Sterilize (vaporous hydrogen
peroxide, VHP).
Laminate
flooring
Laminate wood
locking flooring 7-5/8
in, x 50-3/4 in.
Item #103553,
Lowe's Home
Improvement,
Morrisville, NC
35 x 35 or
14x14
12x12
Fabricate coupons using
established procedures.
Remove incidental dust and
grime with alcohol wipes.
Sterilize (VHP).
5

-------
35"



14"









14"

10.5"





10.5"


Figure 2-3: Coupon dimensions
2.2.4	Coupon and Equipment Sterilization
Prior to use, laminate and vinyl coupons were sterilized with 400 parts per million hydrogen peroxide
(H2O2) vapor for four hours using a STERIS VHP® ED1000 generator (STERIS, Mentor, OH) and then
held at room temperature for a minimum of 2-3 days to allow off-gassing of H2O2. Stainless steel
coupons were sterilized with a 250 degrees Celsius (°C) gravity cycle using an autoclave (Amsco Century
SV 120 Scientific prevacuum sterilizer, STERIS, Mentor, OH). Sterility was evaluated by swab sampling
one coupon from each sterilization batch. Prior to each test, the test chambers were sterilized using the
following procedure:
1.	Don sterile gloves, wipe the inner surfaces using a bleach towel (Dispatch® hospital cleaner
disinfectant towels with bleach, concentration of sodium hypochlorite: 0.65%; model number
69260; Caltech Industries, Inc., Midland, Ml).
2.	Wait for at least 5 minutes (min) and then wipe the surface again using a new bleach towel.
3.	Discard the bleach towels and wipe the surface using 3% sodium thiosulfate (STS) wipes. These
wipes are prepared in-house by adding 90 milliliters (mL) of sterile 1N STS and 310 mL of sterile
deionized (Dl) water to a canister of clean wipes (Fisherbrand™ dry Clean-Wipes™, cat. no. 06-
664-14, Fisher Scientific Pittsburgh, PA).
4.	Immediately follow by wiping the surface using alcohol wipes (premoistened alcohol/DI water
Clean-Wipes™, cat. no. 06-665-24, Fisher Scientific, Pittsburgh, PA).
2.2.5	Spore Preparation
The test organism for this work was a powdered spore preparation of B. atrophaeus American Type
Culture Collection (ATCC) 9372 mixed with silicon dioxide particles. This preparation was obtained from
the U.S. Army Dugway Proving Ground Life Sciences Division. The preparation for this procedure is
described in Brown et al.5 In short, after 80-90% sporulation, the suspension was centrifuged to generate
a preparation of approximately 20% solids. A preparation resulting in a powdered matrix containing
6

-------
approximately 1 * 1011 viable spores per gram was prepared by dry blending and jet milling the dried
spores with fumed silica particles (Degussa, Frankfurt am Main, Germany).
2.2.6 Coupon Inoculation
Coupons (test and positive controls) were inoculated with B. atrophaeus spores using an MDI.6 Briefly,
each coupon was contaminated independently with a separate ADA designed to fit one 14 in. x 14 in.
coupon. The MDI and actuator were attached to an ADA through an opening in the center top of the
ADA'S stainless steel hood, which is sized to cover the square inoculation area exactly. The MDI was
discharged one single time into the ADA. The spores were allowed to settle on the coupon surfaces for a
minimum of 18 hours. At the end of the minimum 18-hour period, the ADA was removed immediately
before sampling. Photographs of an ADA, an MDI, and an MDI actuator used in this project are shown in
Figures 2-4 and 2-5.
Figure 2-4: Aerosol deposition apparatus	Figure 2-5: MDI and actuator
2.3 Operational Testing of Swifter® Sweeper® System
A sampling team of three people, designated as one sampler and two support persons, was
used. The sampler was responsible for taking the mop and wipe samples, while a support person was
responsible for assembling the sampling equipment and receiving and securing samples taken by the
sampler. Another support person handled testing materials such as contaminated coupons and ADAs and
operated the air sampling equipment.
2.3.1 Operational Testing Modes
The Swiffer® Sweeper® was evaluated for spore removal using three operation modes: sweeping,
mopping, and a combination of sweeping followed by mopping.
2.3.1.1 Dry Sweeping Mode
The following procedure was used for evaluation of the Swiffer® in dry sweeping mode:
7

-------
1.	The support person sets the inoculated coupon, including the ADA, inside the test chamber prior
to sampling.
2.	The support person opens the bag containing a dry Swiffer® cloth (Swiffer® Sweeper® dry cloth).
3.	Using aseptic technique, the sampler removes the cloth from the bag and assembles it on the
Swiffer® handle, which is already inserted into the sampling port of the test chamber.
4.	The support person then removes the ADA and simultaneously starts the air sampling period of
25 min, corresponding to one full air exchange, and the sweeping of the coupon surface.
5.	When the air sampling is completed, the support person holds open the lid of the chamber, and
the sampler retrieves the mop cloth.
2.3.1.2	Wet Mopping Mode
Wet mopping was conducted as outlined above with the following exception: the dry sweeping cloth
(Swiffer® Sweeper® dry cloth) was replaced with a wet mopping cloth (Swiffer® Sweeper® wet mopping
cloth). Sampling and collection of the mop cloth were conducted as before.
2.3.1.3	Dry Sweeping/Wet Mopping Mode
After conducting testing using either dry or wet mop Swiffer® cloths, surface cleaning was conducted
using a combination of the two methods. In this approach, the coupon surface is first cleaned using the
dry cloth and then cleaned using the wet cloth as follows:
1.	The support person sets the inoculated coupon, including the ADA, inside the test chamber prior
to sampling.
2.	The support person opens the bag containing a dry Swiffer® cloth (Swiffer® Sweeper® dry cloth).
3.	Using aseptic technique, the sampler removes the dry cloth from the bag and assembles it on the
Swiffer® handle, which is already inserted into the sampling port of the chamber.
4.	The support person then removes the ADA and simultaneously starts the air sampling period of
25 min, corresponding to one full air exchange, and the sampler begins to sweep the surface of
the coupon.
5.	When the air sampling period is completed, the support person holds open the lid of the chamber
and the sampler retrieves the dry Swiffer® Sweeper® cloth.
6.	At the end of the sweeping phase sampling period, both the support person and the sampler
change their gloves and repeat steps 2 through 5 using a wet mopping cloth (Swiffer® Sweeper®
wet mopping cloth).
The order of dry sweeping followed by wet mopping is the recommended order according to the
manufacturer (http://swiffer.com/en-us/tips-and-articles/how-to-use-swiffer-sweeper).
2.3.2 Sample Coupon Sweeping Pattern
All sample coupons, 35 in. * 35 in., were swept using an up and down and left to right movement (see
Figure 2-6) in the following sweeping pattern:
1. Place the loaded Swiffer® Sweeper® in the lower left-hand corner of the sample coupon.
8

-------
2.	Start sweeping with a stroke straight toward the top of the coupon. The Swiffer® head will be
parallel with the top of the coupon. After reaching the top of the coupon, pull the sweeper straight
down, retracing the path swept in the first stroke. The Swiffer® head will be parallel with the
bottom of the coupon. This set of strokes sweeps the non-inoculated area.
3.	In the next stroke, overlap the previous stroke approximately 5 in. and go diagonally from left to
right. When the top of the coupon is reached, sweep the return stroke straight down with the
sweeper head parallel to the top edge of the coupon until the bottom of the coupon is reached.
4.	Repeat the coupon sweeping strokes as in step 3 until the end of the test coupon is reached.
5.	When the right side of the coupon is reached, perform a straight sweeping stroke toward the top.
The Swiffer® head on this stroke is parallel to the top edge. When the top is reached, pull the
sweeper straight down toward the bottom of the coupon, retracing the path of the up stroke. The
Swiffer® head on this stroke is parallel to the bottom edge. This completes the coupon sweeping.
Figure 2-6: Swiffer® Sweeper® sampling pattern.
The brown rectangle in the lower left corner represents the Swiffer® mop. The blue area is the 12-in. x 12-
in. area inoculated with spores. Brown lines represent movements of the Swiffer® mop head.
3 Sampling Approach
This section discusses the sampling approach, including sampling site environmental conditions,
sampling media, sampling frequency, wipe sampling procedures, and prevention of cross-contamination
during sampling.
For each sampling event (or test), a sampling event log sheet was maintained that included each
sampling team member's name, the date, the run number, and all sample codes with corresponding
coupon codes. The coupon codes were preprinted on the sampling event log sheet before sampling
began. The following sections discuss the sampling strategy and methods used for sampling coupon
surfaces and the test chamber air.
3.1 Sampling Site Environmental Conditions
Ambient environmental conditions such as temperature and relative humidity (RH) can affect the
evaporation rate of liquids from surfaces. All tests were conducted at room temperature and ambient RH.
Three strategically placed HOBO® temperature and RH sensors (HOBO® data logger U12 series, Onset
9

-------
Computers, Bourne, MA) recorded RH and temperature. The devices are calibrated by the on-site
Metrology Laboratory (Metlab) at EPA -RTP. All coupons were conditioned at ambient conditions for one
week before use.
3.2 Sampling Strategy
Each test consisted of four large material coupons (three test coupons and one procedural blank) and
three 14-in. x 14-in. material coupons (positive controls). Additionally, three 14-in. x 14-in. stainless steel
coupons were inoculated with the test set and served as inoculation control coupons. Swabs were taken
of each surface type prior to each test to monitor for contamination. The following sections discuss the
sample types and frequency of sampling and monitoring events.
3.2.1	Sample Types
Each testing sequence resulted in the following samples:
•	Wipe samples: Polyester-rayon blend (PRB) wipe samples were collected from each material in
sets of four, positive control surface samples were collected in sets of three for both stainless
steel and material type, and one was collected for a procedural blank surface sample.
•	Swiffer® Sweeper® cloth (dry and wet): cloth samples were collected for each material
type/Swiffer® Sweeper® operational mode, including a procedural blank.
•	Aerosol samples: Aerosol samples were collected using Via-Cell® bioaerosol cassettes during
each decontamination and procedural blank test. Results for these samples were used to
estimate the occurrence and magnitude of fugitive emissions of viable B. atrophaeus spores
during the sweeping and/or mopping process.
•	Swab Samples: BactiSwab™ samples were collected to check the sterility of the material and
equipment prior to sampling.
3.2.2	Sampling Frequency
The Swiffer® dry cloth tests and the Swiffer® wet cloth tests each generated 20 samples. The test using
dry and then wet cloths generated 34 samples. One air sample was generated during the Swiffer® testing
sequence for a sampling period of 25 min at an air sampling flow rate of 15.3 L/min, corresponding to one
full air exchange. Samples were recovered, extracted, and plated. Plates were incubated to grow the B.
atrophaeus spores, and the resulting colonies were counted.
Table 3-1 shows the sampling frequencies for each material type/Swiffer® mode combination test (as
shown in Table 2-1) required for this project. Tests 5 and 6 samples were doubled since these tests
involved a combination of sweeping and mopping operations.
10

-------
Table 3-1: Sampling Measurements/Frequency for Swiffer® Evaluation Testing
Testing Sequence
Measurement
Sample Type
Frequency

Procedural blank coupon CFU
Swiffer® cloth
1 (2 for Tests 5 and 6)

Wipe
4 (8 for Tests 5 and 6)
For each material type/
Swiffer® dry, wet and
combination test
Laboratory blank solution CFU
Swiffer® cloth
1 (2 for Tests 5 and 6)
Test coupons post-sweep/mop
Swiffer® cloth
1 (2 for Tests 5 and 6)
CFU
Wipe
4 (8 for Tests 5 and 6)

Positive control CFU
Wipe
6 (3 stainless steel and 3 test
material)

Air sampling
Via-Cell®
3 (6 for Tests 5 and 6)
3.3 Sampling Methods
The following sections discuss the methods used for wipe sampling and air sampling.
3.3.1 Wipe Sampling
The general approach for the wipe sampling was to use a moistened, sterile, non-cotton PRB gauze wipe
to wipe an area to recover spores. A three-person team (consisting of a sampler, a coupon handler, and a
support person) was used to collect wipe samples. Aseptic techniques were used throughout.
3.3.1.1 Wipe Sampling Preparation
All materials needed for sampling were prepared before the sampling event began. Table 3-2 lists the
materials used for sampling. Team members wore non-powdered surgical gloves during sampling.
Individually wrapped premoistened bleach wipes, used for sample bag decontamination, were placed in
sterile sampling bags. A sampling material bin was stocked for each sampling event based on the sample
quantity needed for that test. The bin contained enough wipe-sampling kits to accommodate all required
samples for a specific test event.
11

-------
Table 3-2: Sampling Materials and Equipment
Material or Equipment
Description
Non-powdered, sterile surgical
gloves
KIMTECH PURE* G3 Sterile STERLING™ Nitrile Gloves, Kimberly-Clark
Professional® (VWR P/N HC61110 for extra-large, VWR P/N HC61190 for
large, and VWR P/N HC61180 for medium)
Non-powdered, non-sterile
surgical gloves
Examination gloves (Fisherbrand™ Powder-Free Nitrile Exam Gloves (Fisher
Scientific P/N 19-130-1597D for large and 19-130-1597C for medium sizes)
Disposable laboratory coats
Kimberly-Clark Kleenguard A10 light-duty apparel (P/N 40105)
Phosphate-buffered saline (PBS)
PBS with PBST (Sigma Aldrich USA, P/N: P3563-10PAK)
50-mL conical tubes
BD Falcon™ BlueMax graduated tubes, 15-mL (Fisher Scientific P/N 14-959-
70C)
Sterile sampling bags
Fisherbrand™ Sterile Sampling Bags (TWIRL'EM) Overpack size 10-in. by 14-
in.
Inner bag size: 5.5-in. by 9-in. (for wipe)
Sample bag size: 5.5-in. by 9-in.
Bleach wipes
Dispatch® bleach wipes (Clorox® Co., Oakland, CA) or
Hype-Wipes (Current Technologies, Indianapolis, IN)
Wipes for sampling
Kendall Curity Versalon absorbent gauze sponge, 2-in. by 2-in., sterile packed
(ravon-polvester blend). http://www.mfasco.com/ (last accessed December 5.
2016)
Swabs
BactiSwab®. httD://www. remelinc.com/lndustrial/CollectionTransDort/
BactiSwab.asDX (last accessed December 5. 2016)
Analytical filter units
150-mL Nalgene analytical filter units, 0.2-|jm cellulose acetate (Fisher
Scientific cat. no. 130-4020)
Aerosol sampling cassettes
Via-Cell® VIA010 bioaerosol samplinq cassette. http://www.zefon.com/store/
via-cell-bioaerosol-samDlina-cassette.html (last accessed December 5. 2016)
Sterilizing-grade filter
Aervent™0.2-micron sterilizing-grade cartridge filter (Millipore SAS, Molsheim,
France)
Sampling pump
Isokinetic Method 5 Source Sampling Console,
httD://www. aDexinst.com/Droduct/xc-50-method-5-source-samDlina-console

(last accessed April 25, 2017)
3.3.1.2 Wipe Sampling Procedure
After the coupon surface was swept with the SSFMS, wipe samples were collected at four locations to
determine the level of spores remaining on the center of the coupon and spore redistribution across the
coupon by the Swiffer® cloth. The four locations sampled are indicated in Figure 3-1 and consist of the
following surface areas (corresponding to the numbers shown in ascending order): 147, 110, 147, and
196 in.2. The wipe sampling started at the lowest anticipated spore concentration area (Location 1) and
proceeded to the highest anticipated spore concentration (Location 4). The wipe sampling for both the dry
and the wet sweeping mode started when the aerosol sampling was completed (25 min). This sampling
procedure was used also for sampling the procedural negative coupons following a Swiffer® Sweeper®
treatment. Wipe samples were extracted in 20 mL of phosphate-buffered saline with 0.05% TWEEN®20
(PBST), sonicated, vortexed, and subjected to serial 10-fold dilution and plating.
12

-------

3

1
4



2
Figure 3-1: Wipe sampling grid, locations, and order
The sampling team used aseptic techniques and followed a strict sampling protocol to avoid any cross-
contamination among coupons or among samples. The support person logged observations into the
laboratory notebook and made sampling kit items (premoistened all-purpose sponge, conical tube,
sampling bags, etc.) available to the other team members.
The following sequence was followed for obtaining a sample:
•	The support person opened the test chamber to allow access to the coupon and removed the
Swiffer® head, being careful not to touch the coupon.
•	The sampler removed the Swiffer® cloth and put it in the sample bag that was handed to the
sampler by the support person.
•	The support person handed a sampling sponge to the sampler.
•	The sampler wiped the surface of the coupon horizontally, using a consistent amount of pressure
and using S-strokes to cover the designated sample area of the coupon.
•	The sampler folded the sponge in half, concealing the exposed side and then wiped the same
surface vertically using the same S-stroke technique.
•	The sampler folded the sponge again and rolled up the folded sponge so that it would fit into a
conical tube.
•	The support person opened a conical tube, holding the tube inside a sterile sampling bag.
•	The sampler placed the folded sponge into the conical tube that the support person was holding,
being careful not to touch the surface of the tube or the plastic sterile sampling bag.
For each separate test (each of the two flooring types and each of the three Swiffer® methods), surface
sampling of the materials was completed for all procedural blank coupons before sampling of any test
material. Positive controls were sampled last.
13

-------
3.3.2 Air Sampling with Via-Cell®
Air sampling was conducted concurrently with Swiffer® sweeping using an EPA Method 5 dry gas meter
box to measure the volume of air added, a Method 5 pump to force airflow (at 15 liters per minute, Lpm),
and Via-Cell® bioaerosol sampling cassettes to sample for aerosolized spores. Air sampling events were
conducted for one full air exchange of the sampling chamber (13.45 ft3, added to the test chamber in
approximately 25 min). A Millipore filter was installed on the air inlet of each chamber to prevent any
contamination present in the laboratory air from contaminating the air introduced into the chamber.
14

-------
4
Testing and Measurements
4.1 Analytical Procedure
Spores were extracted from the wipe samples, swabs, aerosol filters, and Swiffer® Sweeper® cloths.
Spores in these extracts were assayed by growth on nutrient agar plates in the NHSRC Biocontaminant
Laboratory (Biolab). The wipe samples and aerosol filters were extracted in 20 mL of PBST and vortexed
for two minutes in 10-second bursts. The Swiffer® Sweeper® cloths were extracted in 80 mL of PBST
and stomached at 230 revolutions per minute (rpm) for two minutes. The samples were analyzed either
qualitatively for spore presence (quality control [QC] swab samples) or quantitatively for the number of
viable spores recovered per sample (CFU). Details of the analysis procedures are provided below.
The extracts for all sample types were plated in triplicate using a spiral plater (Autoplate® spiral plating
system, Advanced Instruments Inc., Norwood, MA), which deposits a known volume of sample in three
10-fold serial dilutions on each plate. Plates were then incubated at 35 ± 2 °C for 16 to 19 hours. During
incubation, the colonies develop along the lines where the liquid was deposited on the rotating plate in
decreasing amounts from the center to the edge of the rotating agar plate (see Figure 4-1). The number
of spores was estimated using a QCount colony counter (Advanced Instruments Inc., Norwood, MA).
Figure 4-1: Bacterial colonies on a spiral-plated agar plate
Positive control samples were diluted 100-fold (10 2) in PBST prior to spiral plating, and samples with
unknown concentrations were plated with no dilution and following a 100-fold dilution. Samples with
known low concentrations were plated with no dilution. The QCount instrument automatically calculates
the CFU/mL in a sample based on the dilution plated and the number of colonies that develop on the
plate and records the data in an MS Excel spreadsheet. Only plates containing between 30 and 300 CFU
were used for spore recovery estimates.
15

-------
If a low count (< 30 CFU per plate) occurs for a sample that was spiral plated with a neat (undiluted)
aliquot, then two options are available:
1.	Spread plate an undiluted aliquot with a larger volume (100, 200, and 400 microliters, each in
triplicate).
2.	Filter plate an undiluted aliquot with a larger volume (e.g., 1, 2, or 10 mL).
Filter plating is done using the Pall MicroFunnel unit with 0.45 jjm GN-6 Metricel white membrane (P/N
4804, Pall Corporation, Port Washington, NY). The sample aliquot is added to 10 mL Dl water, which is
then poured over the filter. The vacuum system is opened and the liquid is tunneled through the filter,
trapping the spores on the filter/membrane. The filter is then washed with another 10-mL aliquot of sterile
Dl water. The filter is removed from the plastic housing and placed onto a tryptic soy agar (TSA) plate.
Plates are incubated at 35 ± 2 °C for 18-24 hours prior to manual enumeration.
The swabs are removed from sample packaging and held by the wooden end while the inoculated end is
rolled across a fresh TSA plate in an S-motion. The swab is rolled over the TSA plate so that the entire
circumference of the inoculated tip touches the plate. Plates are incubated overnight at the appropriate
temperature and then evaluated for growth.
4.2 Data Reduction
Each test series was composed of a flooring type (vinyl or laminate) and a Swiffer® Sweeper® mode (dry,
wet, or dry/wet), positive controls and procedural blanks (negative controls). The raw data for this study
are the colony counts from each of the three test coupons, each of the six positive control coupons (three
flooring coupons and three stainless steel coupons), and each procedural blank (one negative control
coupon) from each of the six test series (flooring type/Swiffer® Sweeper® mode combination).
Average and standard deviation (SD) were calculated for counts from the replicate test coupons and
positive control coupons. The LR of spores was calculated for the inoculated center area only; the
redistribution of the spores to other areas outside the center of the coupon was not included in this
calculation. LR is defined in this project as the difference in the average of the logarithm of the number of
viable spores (determined by CFU) recovered on the material control coupons minus the average of the
logarithm of the number of viable spores (determined by CFU) recovered on the center, 12 by 12 in. area
of the test coupons.
Efficacy, defined as LR in CFU count after a Swiffer® Sweeper® treatment, was calculated using Equation
4-1 for each material within each combination of decontamination procedure (/) and test material (J) as
follows:
C
Z(log,„C,JO)
c=1	
'"HVJ
f \
(4-1)
where
16

-------
Cijc is the number of viable organisms recovered from C control coupons for the P
decontamination procedure and 7th test material,
Nijc is the number of control coupons for the P decontamination procedure and 7th test material,
Nijk is number of viable organisms recovered on the /