&EPA
United States
Environmental Protection
Agency
EPA-600/R-06-011
February 2006
Research and
Development of Risk
Management
Alternatives for
Controlling Mold
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EPA-600/R-06/011
February 2006
Research and Development of Risk
Management Alternatives for
Controlling Mold
by
Marc Y. Menetrez, Timothy R. Dean, and Doris A. Betancourt
Office of Research and Development
National Risk Management Research Laboratory
Air Pollution Prevention and Control Division
Research Triangle Park, NC 27711
EPA Project Officer: Marc Y. Menetrez
U. S. Environmental Protection Agency
Office of Research and Development
Washington, DC 20460
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Abstract
The U.S. Environmental Protection Agency, Air Pollution Prevention and Control Division (APPCD),
Indoor Environment Management Branch (IEMB) has, since 1995, conducted research into controlling
biological contamination in the indoor environment. Six areas of research have been addressed: (1)
research and development studies to quantify the effects of moisture, relative humidity (RH), and dust
and develop risk management alternatives for prevention and control of mold growth; (2) duct cleaning
effectiveness for prevention and control of microbial growth on duct materials: (3) evaluation of
antimicrobial treatments as control technologies; (4) field testing of sealants and encapsulents used in
air duct systems; (5) characterization of emission rates and modeling of exposure through heating,
ventilating, and air conditioning operation; and (6) improved methods of sampling and analysis of mold.
The conclusions resulting from this body of research are listed to summarize the accomplishments and
put into perspective the interrelationships of these areas of investigation in reducing human exposure
to biological contamination in the indoor environment. Through a cooperative research agreement, the
Research Triangle Institute (RTI) has played a major role in the development of the program described
in this report. The RTI Research Triangle Park facility houses the Microbiological Laboratory (ML) in
which the static and dynamic microbial test chamber are kept and staffed. As part of the opening of the
new EPA Environmental Resource Center Building in 2002, APPCD/IEMB has established the
Biocontaminant Laboratory (BL) for conducting applied risk management research. The BL is a multi-
functional state-of-the-art biological/molecular research laboratory engaged in intra/inter laboratory
cooperative research. A description of the RTI ML and APPCD/IEMB BL facility is included in this
report.
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Foreword
The U.S. Environmental Protection Agency (EPA) is charged by Congress with protecting
the Nation's land, air, and water resources. Under a mandate of national environmental laws,
the Agency strives to formulate and implement actions leading to a compatible balance
between human activities and the ability of natural systems to support and nurture life. To meet
this mandate, EPA's research program is providing data and technical support for solving
environmental problems today and building a science knowledge base necessary to manage
our ecological resources wisely, understand how pollutants affect our health, and prevent or
reduce environmental risks in the future.
The National Risk Management Research Laboratory (NRMRL) is the Agency's center for
investigation of technological and management approaches for preventing and reducing risks
from pollution that threaten human health and the environment. The focus of the Laboratory's
research program is on methods and their cost-effectiveness for prevention and control of
pollution to air, land, water, and subsurface resources; protection of water quality in public
water systems; remediation of contaminated sites, sediments and ground water; prevention
and control of indoor air pollution; and restoration of ecosystems. NRMRL collaborates with
both public and private sector partners to foster technologies that reduce the cost of
compliance and to anticipate emerging problems. NRMRL's research provides solutions to
environmental problems by: developing and promoting technologies that protect and improve
the environment; advancing scientific and engineering information to support regulatory and
policy decisions; and providing the technical support and information transfer to ensure
implementation of environmental regulations and strategies at the national, state, and
community levels.
This publication has been produced as part of the Laboratory's strategic long-term research
plan. It is published and made available by EPA's Office of Research and Development to
assist the user community and to link researchers with their clients.
Sally Gutierrez, Director
National Risk Management Research Laboratory
in
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EPA Review Notice
This report has been peer and administratively reviewed by the U.S. Environmental Protection Agency
and approved for publication. Mention of trade names or commercial products does not constitute
endorsement or recommendation for use.
This document is available to the public through the National Technical Information Service,
Springfield, Virginia 22161.
IV
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Table of Contents
Section Page
Abstract ii
List of Figures vi
Introduction 1
Microbiology Research Facilities 4
Research Triangle Institute Microbiology Laboratory 4
APPCD/IEMB Biocontaminant Laboratory 5
Research Program 6
Static Microbial Test Chamber 6
Dynamic Microbial Test Chamber 6
Research and Development of Risk Management Alternatives to Prevent and Control the
Growth of Mold by Studies to Quantify the Effects of Moisture, Relative Humidity,
and Dust 7
Duct Cleaning Effectiveness for Prevention and Control of Microbial Growth on
Duct Materials 9
Evaluation of Antimicrobial Treatments as Control Technologies 10
Field Testing of Sealants and Encapsulents Used in Air Duct Systems 10
Characterization of Emission Rates and Modeling of Exposure through HVAC Operation .... 12
Improved Methods of Sampling and Analysis of Mold 15
Evaluation of Microbial Volatile Organic Compounds 15
Development of Rapid Multiplex PCR 16
Discussion 19
Technical Findings 20
Conclusions 23
Future Research 24
References 27
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List of Figures
Page
Cladosporium, andAspergillus 1
Stachybotrys on drywall 1
Mixture of Cladosporium, Aspergillus, andPenicillium on painted drywall 2
Research Triangle Institute Microbiology Laboratory 4
Static Microbial Test Chamber 6
Cutaway Drawing of the Dynamic Microbial Test Chamber 7
Two Static Microbial Test Chambers 7
8 Dynamic Microbial Test Chamber 8
9 HVAC Duct Encapsulant Lining 10
10 Encapsulant Application 10
11 Room Wall Simulators 13
VI
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Introduction
The indoor environment has become an important area of
research in recent years. The past twenty years have
brought the recognition that an important factor in the
health of people in the indoor environment is the
Figure 1. Cladosporium, and Aspergillus.
Figure 2. Stachybotrys on drywall.
dampness of the buildings in which they live and work
[1-3], and the potential for mold colonization as depicted
in Figures 1 and 2. Furthermore, it is now recognized that
the principal biological organisms responsible for the
health problems in these environments are the fungi rather
than bacteria and viruses [4]. It has been estimated that
upwards of 40% of all homes in North America contain
fungal growth, while numbers in Northern Europe are in
the range of 20-40% fungal contamination [5-6].
Although traditionally, fungi in this context have been
viewed as a source of allergens (and in unusual
circumstances, pathogens), data have accumulated to show
that the adverse health effects resulting from inhalation of
fungal spores is due to a variety of factors [7]. One
characteristic associated with certain fungi, is the low
molecular weight toxins (mycotoxins) they produce.
Mycotoxins are important in human and animal health
because of their production by toxigenic fungi associated
with food and animal feed. In the indoor environment,
mycotoxins tend to concentrate in fungal spores and thus
present a potential hazard to those exposed who inhale
them.
Although only a small percentage of fungal species has
been associated with adverse health effects, increased
awareness and continuing research will likely result in the
identification of many more pathogenic and toxigenic
fungi. Organisms such as Stachybotrys chartarum,
Penicillium purpurogenum, Aspergillus versicolor, and
Cladosporium spp. are organisms that are frequently
found in buildings that are heavily contaminated with
mold and are potentially associated with adverse health
effects in humans [8-9]. These health effects may include
itchy eyes; stuffy nose; headache; fatigue; and, in severe
cases, idiopathic pulmonary hemosiderosis (IHP) in
infants [ 10-14]. In numerous cases the mold S. chartarum
has been found to be associated with idiopathic pulmonary
hemorrhage in infants [1-2]. It is also studied for toxin
production and its occurrence in water damaged buildings.
Growth of S. chartarum on building materials such as
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drywall has been frequently documented. Indoor exposure
to mold has also been linked to pulmonary disease,
including allergies and asthma. Given this significant risk
of exposure and frequency of occurrence, environmental
factors regarding the growth of mold have been studied.
The U.S. EPA, Air Pollution Prevention and Control
Division (APPCD), Indoor Environment Management
Branch (IEMB) has conducted research since 1995, into
controlling biological contamination in the indoor envi-
ronment. The IEMB Biocontaminant Laboratory (BL)
conducts research into biological contamination in the
indoor environment. The goal of this research is the devel-
opment of engineering guidelines for the prevention,
mitigation, and control of biocontaminants which can be
problematic in the indoor environment as shown in Figure
3. Mold contamination can also cause significant damage
to buildings. The last ten years of research findings have
contributed greatly to understanding the threat of mold
and the development of effective solutions.
Biological contamination in the indoor environment is
recognized as a major health concern [15]. Exposure to
airborne biocontaminants or their metabolites can induce
irritational, allergic, and infectious responses, including
acute reactions such as vomiting; diarrhea; hemorrhage;
convulsions; and, in some cases, death [1, 3, 15-17].
Reducing occupant exposure to indoor air pollutants is the
primary goal of the majority of indoor air quality (IAQ)
research. For many indoor biocontaminants (e.g.,
microorganisms), the main growth locations are the
structural and finishing materials and furnishings of the
building [18,19]. The application of effective engineering
controls within the building is essential to prevent
biological pollution in the indoor environment.
It is well recognized that fungi can colonize and amplify
on a variety of building materials if sufficient nutrients
and moisture are present. Mold contamination has been
associated with a variety of building and furnishing
materials including carpet, ceiling tile, gypsum wallboard,
flooring, insulation, and heating and air-conditioning
components [18, 19].
The goal of this research publication was to provide
industry, academia, and regulators with an organized
presentation of the many developments accomplished in
risk management research in the last decade within US
EPA/APPCD/IEMB. It can ultimately provide engineering
guidelines for the prevention, mitigation, and control of
biological contaminants in the indoor environment.
The objectives were to: (1) provide a scientific basis for
studying building material colonization by microorgan-
isms, (2) conduct research on source management and
climate control, (3) evaluate engineering solutions and
Figure 3. Mixture of Cladosporium, Aspergillus, and Penicillium on painted drywall.
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control techniques, and (4) develop molecular charac-
terization techniques for identifying biological contam-
inants.
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Microbiology Research Facilities
In response to increasing examples of mold contamination
and the potential threat to human health, the U.S. EPA,
APPCD/IEMB has conducted research into controlling
biological contamination in the indoor environment. This
capstone report of "Research and Development of Risk
Management Alternatives for Controlling Mold" covers
the "where, why, and how" this research has been
performed; the findings and conclusions that have been
accomplished; as well as a preview of upcoming research.
Research Triangle Institute's Micro-
biology Laboratory
The Research Triangle Institute (RTI) microbiology lab-
oratory (ML) (as shown in Figure 4) designs and conducts
applied and basic research in environmental microbiology
and aerobiology. Specializing in biological aerosols, they
research biological contaminants isolated from the envi-
ronment to identify environmental causes of illness and to
recommend methods for preventing such biological con-
tamination and its associated adverse health effects. RTFs
microbiology research program is fully equipped to
collect, characterize, study, inactivate, and control micro-
bial populations. Particular emphasis is placed on cooper-
ative research among microbiologists, aerosol scientists,
engineers, and chemists.
Figure 4. Research Triangle Institute Microbiology
Laboratory.
Laboratory and field research includes environmental
microbiological assessment, environmental biological
pollution studies, and antimicrobial/biocide efficacy eval-
uations. Projects include Homeland Security defense pro-
jects, evaluating equipment for detecting bioterrorism
agents, and decontamination protocols.
RTFs microbiologists work in a uniquely designed micro-
biology laboratory applying expertise in: (1) methods de-
velopment, (2) system or process evaluation, (3) environ-
mental monitoring and exposure assessment, (4) tech-
nology transfer assistance, (5) bioaerosol penetration and
containment, (6) aerosol sampler evaluations, (7) product
evaluations and assessments, (8) materials biodeteri-
oration, (9) bioburden assessments, (10) static and dy-
namic chamber studies, (11) antimicrobials/biocides as-
sessments (gas-phase, aqueous, bound, and UVC irradi-
ation), (12) antimicrobial "As-Used" evaluations, (13)
cleanroom fabric microbial penetration evaluations, (14)
biofiltration assessments, and (15) Cleaning effectiveness.
RTFs microbiology research program is home to a unique
microbiology research laboratory. Functioning as a Class
10,000 or better cleanroom, the 1,500 ft2 biological safety
level 2 (BSL2) laboratory contains standard microbio-
logical equipment such as steam autoclave; incubators;
refrigerators; centrifuges; light, fluorescent, and phase
contrast microscopes; colony counters; spectrophoto-
meters; fluorometers; analytical balances; and biological
safety cabinets. A low-temperature freezer houses an
extensive collection of fungi and bacteria.
In addition, RTI has a full range of bioaerosol samplers
including 1-, 2-, and 6- stage Andersen impactors,
Mattson-Garvin samplers, glass impingers, and a high
volume surface sampler (FFVS3). They have the required
facilities and expertise for the collection of membrane
filter sam-ples. Located within the laboratory is a large
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microbio-logical aerosol test facility, which contains a
microbial dynamic test chamber with controlled air flow,
temper-ature, and relative humidity conditions. Additional
chambers include small, bioaerosol test chambers that are
used to evaluate bioaerosol sampler collection efficiencies
for recovery of airborne viruses. A walk-in environ-
mentally controlled room houses 21 static chambers that
are used primarily for ASTM 6329 assessments.
The RTI Microbiology Laboratory, including the static
and dynamic microbial growth chambers, were developed
through a cooperative agreement with EPA/APPCD/
IEMB.
APPCD/IEMB Biocontaminant Laboratory
In 2002, the APPCD/IEMB Biocontaminant Research and
Development Program created an in-house biocontaminant
laboratory utilizing state-of-the-art analytical evaluation of
biological contaminants by means of both cultural and
molecular biology techniques. The analytical capabilities
encompass both viable and non-viable mold and bacteria,
fragments of organisms or spores, and biological
particulate matter in indoor and ambient air. Samples of
indoor and outdoor air can be analyzed quantitatively for
species specific identification of mold, [and mycotoxins,
and |3-(1,3) glucan assays], bacteria, (and endotoxins),
allergens, particulate matter (PM), and microbial volatile
organic compounds (MVOC). Characterization of
microbial populations can be processed from soils, waters,
air, dusts, and materials. Microscopy, electron microscopy
and photomicrography are performed. Allergen
identification by antigen assay or other enzymelinked
immunosorbent assays (ELISAs) are also performed.
Laboratory capabilities include two standard BSL 2 hoods
and two custom BSL 2 hoods for inoculation, deposition,
and exposure studies; gas chromatography/mass spectro-
metry (GC/MS) (Agilent 6890 Gas Chromatograph,
equipped with a 5973N Mass Selective Detector and 7683
Autosampler, DB-WAX polyethelene glycol bonded phase
column) for performing MVOC studies; biological
contaminant analysis performed by polymerase chain
reaction (PCR) (Bio-Rad and ABI Laboratories, Thermal
Cycler, and PCR detection system); and genetic sequen-
cing analysis by ABI PRISM 3100 Genetic Analyzer,
multi-color florescence based DNA capillary electro-
phoresis analysis system. Laboratory analysis includes
vegetative and spore forms of bacteria and mold; allergen
identification including dust mites, cockroach, dog and cat
dander, and ragweed and tree pollen; human, animal, and
plant virus and prion identification. Sample collection of
viable and non-viable bioaerosols by air-filtration and
Andersen viable and non-viable impactors, and filtration.
BL personnel developed a method for measuring bio-
logical particulate material (BioPM), has studied bio-
aerosol penetration and containment, and conducted
evaluations of wall simulation and distribution of con-
taminants. The laboratory is capable of analyzing bio-
logical samples for unknown pathogenic, toxic, immuno-
suppressant and carcinogenic contaminants.
With the crucial human health and safety issues of micro-
bial contamination that affect the public, it is critical to
understand our surroundings and how we interact. Bio-
logical contaminants—such as bacteria, fungi, viruses,
protozoa, and algae as well as spores or other components
of such organisms and products of organism growth such
as mycotoxins—have been linked to a variety of adverse
health effects. These effects include asthma and allergic
reactions, infectious diseases, and a range of symptoms
from sneezing to dizziness and digestive problems.
The research involving the collaborative work in both the
RTI ML and the IEMB BL has resulted in a body of re-
search publications which are summarized. The specific
research accomplishments have also been grouped into
areas of investigation. A discussion of the interrela-
tionships of these areas of investigation is included. Re-
ducing human exposure to biological contamination has
been the overall objective through a comprehensive
programmatic approach.
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Research Program
Six areas of research were identified by EPA/APPCD/ IEMB
for allocation of program resources: (1) research and
development of risk management alternatives to prevent
and control the growth of mold by studies to quantify the
effects of moisture, relative humidity (RH), and dust, (2)
duct cleaning effectiveness for prevention and control of
microbial growth on duct materials, (3) evaluation of
antimicrobial treatments as control tech-nologies, (4) field
testing of sealants and encapsulents used in air duct
systems, (5) characterization of emission rates and
modeling of exposure through heating, ventilating, and air
conditioning (HVAC) operation, and (6) improved
methods of sampling and analysis of mold. Each of the
six research areas is described below. How-ever, prior to
the discussion of these research areas, the development of
the static and dynamic microbial growth chambers are
described. The development of these chambers has
impacted all six research areas by providing unique
research tools which facilitated many of these biological
investigations.
Static Microbial Test Chamber
The static microbial test chamber (SMTC), depicted in
Figure 5, is constructed from acrylic sheeting (measures
32 x 39 x 51 cm) with shelves for samples and a saturated
salt solution on the chamber bottom to control the
equilibrium RH (ERH) [20]. The SMTC was tested using
ASTM 6329-98 [21]. This method was developed as part
of on-going indoor air biocontaminant research. Multiple
SMTCs were used to evaluate fungal growth on 2.5 to 3.8
cm square sections of fiberglass duct liner (FGDL) at
various environmental conditions. The SMTC was de-
veloped to assess potential microbial growth on a variety
of common building materials. Temperature and RH are
controlled to simulate the desired environmental con-
ditions. Prior to chamber testing, materials can be treated
by soaking to simulate a wetting event or treated with an
antimicrobial to simulate mitigation practices [20 - 28].
Figure 5. Static Microbial Test Chamber.
Dynamic Microbial Test Chamber
To allow for experiments which involve air movement
over contaminated surfaces and the release of
biocontaminant particles into the air, research was
conducted in the dynamic microbial test chamber (DMTC)
depicted in Figure 6. The stainless steel and glass chamber
is a room-sized 2.44 m3 cube, designed and constructed
under a cooperative agreement between EPA and RTI.
Chamber air is conditioned by an air-handler unit (AHU)
which keeps the chamber at a temperature of 18-32 °C, a
controlled RH ranging from 55% to 95%, and an air
circulation rate of 1.4 to 4.8 m3/min. Air temperature and
RH can be either raised or lowered depending on the
requirement of the experiment.
The DMTC has been constructed to study the growth,
emissions, and transport of biological contaminants. The
DMTC allows for a variety of microbiological research to
be performed involving biological growth on building
materials, evaluation of emission and deposition of bio-
aerosols, the impact of HVAC mechanical system com-
ponents on biological contaminants, and in-duct tests of
air cleaners. The chamber permits a contained and highly
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Figure 6. Cutaway Drawing of the Dynamic Microbial
Test Chamber.
controlled approach to the study of bioaerosol
characterization [29].
Research and Development of Risk
Management Alternatives to Prevent and
Control the Growth of Mold by Studies to
Quantify the Effects of Moisture, Relative
Humidity, and Dust
This research used a static chamber test method (SCTM)
with the SMTC (see Figures 5 and 7) described above, and
laboratory equipment, materials, and reagents to provide
controlled environments, which allow scientific investi-
gations of physical conditions and environmental factors
favorable to the growth of biological contamination in
indoor spaces[20].
With the use of multiple SMTCs and the development of
the SCTM, three varieties of FGDL and ceiling tile mate-
rials were evaluated for their ability to support the growth
of the fungus Penicillium chrysogenum.[22]. Fungal
growth was evaluated on 2.5 to 3.8 cm square sections of
FGDL. Wetting clean samples of FGDL materials was
found to not increase amplification of the P. chrysogenum
over levels seen without wetting [23]. Soiling FGDL
samples with dust accumulated and previously harvested
from FfVAC systems exhibited a significant association
with the growth of P. chrysogenum [23]. At moderate
soiling levels (0.4-0.7 mg/cm2), growth occurred on
FGDL ductboard and flexible ductboard but not galva-
nized steel [24]. At heavy soiling levels (9-18 mg/cm2),
growth was seen on all three types of duct liner [24]. This
suggests that dust accumulation should be properly con-
trolled in any FfVAC duct to prevent the growth of P.
chrysogenum [22, 24].
The same SMTC environmental chambers (Figures 5 and
7) were used to study the impact of different levels of
moisture and RH on the ability of ceiling tiles to support
the growth of Penicillium glabrum. Amplification oc-
curred at RH levels above 85%. Lower RH was demon-
strated as effective in controlling fungal contamination on
ceiling tiles [25].
Most of the existing standard test protocols for evaluating
antimicrobial efficacy focus on applying the active
chemical compound (antimicrobial or biocide) to the
surface of a building material [26, 27]. VanOsdell, et al.
[29], provides a practical hands-on evaluation protocol
that is important to SCTM testing of materials under
realistic environmental conditions (i.e., temperature and
humidity) in which soiling with sterilized dust is a factor.
The dust was obtained from the National Air Duct
Cleaners Association (NADCA) and was gathered from
actual HVAC systems which were cleaned by member
companies. The use of this method enables the generation
Figure 7. Two Static Microbial Test Chambers.
7
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of a quantitative endpoint for growth in a we 11-controlled
environment with improved repeatability and com-
parability between tests and materials. This method was
developed for evaluating fungal growth (as measured by
sporulation) on indoor materials and has been used
successfully to evaluate the ability of different types of
materials to sustain the growth of Penicillium glabrum,
Aspergillusniger,A. versicolor,andP. chrysogenum [20].
Resistance to fungal growth was demonstrated to vary for
three types of newly purchased FGDL (FGDL, FGDL
with biocide, and FDGL ductboard) inoculated with
Penicillium chrysogenum. Of these types of FGDL tested,
the FGDL ductboard demonstrated growth after inocu-
lation and 6 weeks of static chamber isolation at 97% RH;
in analogous testing, wetting FGDL produced growth on
FDGL ductboard and FGDL, and soiling FGDL with dust
collected from residential heating and air-conditioning
systems caused growth on all three types of FGDL,
including one of which contained a manufacturers applied
fungal biocide [26, 27]. When considering these findings
and the expected soiling which is produced by normal
FfVAC operation, the importance of maintaining low
indoor RH is demonstrated to be imperative.
In another project utilizing the DMTC, the impact of RH,
air velocity, and surface growth on the emission rates of
fungal spores were measured. The DMTC (see Figures 6
and 8) was operated at 23.5 °C and 95% RH and utilized
a separate AHU that forced conditioned air through a high
efficiency particulate air (HEPA) filter and eight mini-
ducts to simulate HVAC duct use with single pass air
velocity rates of 250 cm/s. Each 28.0 x 84.0 cm miniduct
contained a single sheet of FGDL material to be tested.
Eight miniducts were used to simultaneously test three
encapsulant coatings applied to: one set of three FGDL
samples soiled with duct dust obtained from NADCA,
three unsoiled FGDL samples, and two control samples,
for a total of eight miniduct samples. HEPA filtered con-
ditioned air passed over the surface of the FGDL samples,
traveling horizontally within the miniduct apparatus, and
exited the duct and returned to the external air-handler for
filtration and conditioning. The research indicates that
emission rates are inversely proportional to RH but
directly related to air flow and surface loading [28].
Figure 8. Dynamic Microbial Test Chamber.
In another set of experiments, SMTC and DMTC test
results were generated under conditions of constant
temperature, varying degrees of RH and conditions of
wetting. Microorganisms (Penicillium glabrum,
Aspergillus niger, A. versicolor, and P. chrysogenum)
were used to evaluate the extent of biological growth upon
building materials of differing moisture content. Dry and
wet, used and new FGDL and ceiling tile materials were
evaluated. Emphasis was on correlating the moisture
content of building materials with microbial growth.
Growth was determined to be a function of organism, RH,
and the degree of soiling. The extent of soiling or dust
deposited on FGDL and ceiling tile materials was also
shown to be a significant determinant of growth [22-29].
The research showed that emission rates for these
materials for A. versicolor and P. chrysogenum are
inversely proportional to RH but directly related to air
flow rate and surface loading [24-28, 30-32].
In another experiment, the environmental factors leading
to the growth of Stachybotrys chartarum on building
materials was investigated given the significant risk of
exposure and frequency of reported occurrence.
Commonly used building materials were sterilized,
inoculated with S. chartarum, and exposed to controlled
levels of relative humidity and wetting. A quantitative
analysis of viable S. chartarum was performed on the
building materials during a seven month period. The
research indicates that, for environments with a relative
humidity below total saturation, wetting was necessary for
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visible growth to occur. Conversely, high levels of relative
humidity without wetting did not initiate growth. Porous
materials after becoming sufficiently wet and measuring
saturation on a moisture meter, exhibited mold growth in
every experiment conducted [33].
Duct Cleaning Effectiveness for Preven-
tion and Control of Microbial Growth on
Duct Materials
HVAC systems have been shown to act as a collection
source for dust, and the accumulated dust can consist of
such contaminants as mold, fungi, and bacteria. The po-
tential health risks associated with exposure to these
contaminants make removal of the dust a consideration,
especially if improving indoor air quality is required. Be-
cause of their potential to rapidly spread contamination
throughout a building, ventilation systems materials are of
particular significance as potential microbial contami-
nation sources. Portions of ventilation systems near
cooling coils and drain pans are known to be exposed to
high moisture levels for extended periods, and fibrous duct
insulation materials are known to have become sources of
microbial contamination in some buildings. The evaluation
of duct cleaning as a means of control or prevention of
microbial growth on insulated and galvanized duct
surfaces has been conducted. Although duct cleaning is
effective in removing accumulated dust and contaminants
from the inner duct surface, the effects on air quality have
not been substantiated. In addition, the abrasive action of
rotary brushing used in duct cleaning could weaken the
integrity of interior duct insulation. Possible erosion of
duct insulation can however be avoided if a surface
encapsulant is applied. Field evaluation of duct cleaning
was performed as part of a larger proj ect which focused on
the use of antimicrobial encapsulants [18, 22, 21]. Most
commercial applications of duct cleaning (rotary brushing
or vacuuming) include the use of an antimicrobial encap-
sulant on FGDL. This improves the integrity or structure
of the surface, helps to reunite any loose fibers which may
have become partly dislodged by the actions of the
cleaning process, and deposits a surface which should be
hostile to potential microbial growth. The testing of three
commercially available antimicrobial encapsulents/
sealants were monitored after being applied to a FGDL
surface that was contaminated with mold and cleaned. The
field experiment was conducted in the EPA test house,
Gary, NC [23]. The results of the study of antimicrobial
encapsulants efficacy is discussed in the next section,
Evaluation of Antimicrobial Treatments as Control
Technologies, and in the section titled Field Testing of
Sealants and Encapsulants Used in Air Duct Systems.
As covered in the previous section, concurrent laboratory
testing was performed which revealed that if dust is
allowed to build on FGDL within the air distribution duct
network, mold growth can become established at elevated
RH levels even when an antimicrobial encapsulant is
applied to duct surfaces [22, 23].
Understanding the cause of microbial contamination, the
means of controlling or preventing microbial growth, and
the consequential effects of the uncontrolled spread of
microbial growth in typical operating conditions has been
addressed [22-29]. To facilitate biological research on
duct materials, the static and dynamic chambers were
designed and constructed, and the methods of testing
microbial growth under constant temperature and RH and
conditions of static or dynamic air movement was
developed [20, 22, 29]. The evaluation of fungal growth
on FGDL and ceiling tiles were discussed above under
Research and Development of Risk Management
Alternatives to Prevent and Control the Growth of Mold
by Studies to Quantify the Effects of Moisture, Relative
Humidity (RH) and Dust. The impact of RH, air velocity,
and surface growth on the emission rates of fungal spores
from the surface of contaminated material have been
studied and are addressed under the section titled
Characterization of Emission Rates and Modeling of
Exposure through HVAC Operation.
Findings confirm that fungal growth on FGDL is intrusive
throughout the materials and that guidelines which
recommend discarding microbially contaminated porous
duct material should be followed [30-32]. Mechanical
cleaning by HEPA air-vacuuming was able, at best, to
reduce imbedded fiber soiling and temporarily decrease
fungal levels. These fungal populations were able to
reestablish growth within six weeks [31, 32].
-------
Evaluation of Antimicrobial Treatments as
Control Technologies
The efficacy of antimicrobial treatments (see Figure 9) to
eliminate or control biological growth in the indoor envi-
ronment can easily be tested on nonporous surfaces. How-
ever, the testing of antimicrobial efficacy on porous
surfaces, such as those found in the indoor environment
(i.e., gypsum board, HVAC duct-liner insulation, and
wood products) can be more complicated and prone to
incorrect conclusions regarding residual organisms and
nonviable allergens [30-34]. Research to control biolog-
ical growth using three separate antimicrobial encap-
sulants on contaminated duct-liner insulation has been
performed in both field and laboratory testing. The results
indicate differences in antimicrobial efficacy for the period
of testing [34].
Figure 9. HVAC Duct Encapsulant Lining.
Three common HVAC antimicrobial encapsulants were
evaluated for their use on FGDL in both laboratory and
field application experiments. The antimicrobial encap-
sulants tested are manufactured for use in HVAC system
components and on duct surfaces (see Figure 10). Coating
I was a polyacrylate copolymer containing 9% barium
metaborate and 0.16% iodo-2-propynyl butylcarbamate;
Coating II was an acrylic coating containing decabromo-
diphenyl oxide and antimony trioxide; and Coating III was
an acrylic primer containing phosphoric acid compounds
with a phosphated quaternary amine complex [diethanol-
amine n-coco alkyl derivatives; 2,2'-(coco ankylimino)bis-
ethanol]. Although the field test was inconclusive and
truncated, the findings are discussed in the next section.
Laboratory SMTC experiments showed differences in
Figure 10. Encapsulant Application.
degrees of efficacy for the three antimicrobial coatings.
Two antimicrobial coatings limited fungal contamination
for the duration of testing. The effective coatings
(Coatings I and III) contained zinc oxide/borates and a
phosphated quaternary amine, respectively. The ineffec-
tive coating (Coating II) contained decabromodiphenyl
oxide and antimony trioxide. Although all three anti-
microbials are registered with the EPA, they were not
equally effective, nor should they be expected to perform
equally in field use [33, 34].
Methods of testing antimicrobial efficacy are needed to
evaluate differences in products having or seeking EPA
registration. The series of experiments described in
Menetrez et al. (2002), are adequate tests for viable mold
[34]. However, in addition to viable mold antimicrobial
efficacy, additional testing may be needed for viable
bacteria (bacteria cells and spores), nonviable mold
(mycotoxins) and bacteria (endotoxins), as well as viruses
and some forms of allergens (e.g., dust mites) to determine
whether the manufacturers' claims apply [34-36].
Field Testing of Sealants and Encap-
sulants Used in Air Duct Systems
Under favorable conditions, biocontaminants are able to
grow and multiply on a variety of building materials and
indoor surfaces. All antimimicrobial manufacturers'
claims should be verified through the testing of efficacy
performance. Potential biocontaminants and their me-
tabolites that are claimed to be controlled should be tested
on the appropriate materials and under the appropriate
conditions of use. Methods of testing for antimicrobial
10
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efficacy are incomplete and may be misleading if (1)
testing is performed only on nonporous materials, (2)
testing involves a limited number (one or two) of micro-
bial species, and (3) testing of microbial fragments is not
included. Analytical methods are available to accomplish
a comprehensive determination of antimicrobial efficacy
and should be made part of the evaluation process.
The overall goal of antimicrobial efficacy research should
be to contribute to developing better methods of under-
standing indoor air bioaerosol contamination and to en-
hance the ability to prevent and control indoor exposure to
all forms of biocontaminants. Specific objectives include
(1) establishing standardized test methods and procedures
for evaluation, (2) developing techniques and equipment
to conduct test evaluations, and (3) identifying materials,
microbial populations, and constituent biological entities
(viable and nonviable metabolites and microbial volatile
organic compounds) to be tested with antimicrobial
treatments [34].
Three commercially available biocidal encapsulants/
sealants (as previously described in the section titled
Evaluation of Antimicrobial Treatments as Control
Technologies) were also field tested and monitored after
being applied to fiberglass duct liner surfaces that were
contaminated with mold and cleaned. The field experiment
was conducted in the EPA test house, Gary, NC. Par-
ticipating members of the NADCA rotary-cleaned and
spray-coated according to the manufacturers specifications
the fiberglass duct liner in the trunk-lines of the EPA test
house with three popular brands of encapsulants/sealants.
The encapsulant/sealant efficacy was field-tested under
normal residential conditions for cooling and heating. The
test environment was representative of the area for a
HVAC system located in a residential crawl space. During
the cooling season, the HVAC system was cool and had a
high humidity when running (especially in the area of the
cooling coils and drain pan where condensate flow was
constant and the air remained near saturation) and had
some intermediate condition when not running. The results
suggest that dust and high humidity should be properly
controlled in any HVAC system to prevent the growth of
P. chrysogenum [23, 33, 34].
Field study measurements of FGDL surfaces prior to
coating averaged approximately 1,000 CPUs/10 cm2 of
fungal contamination. This concentration of mold con-
tamination did not substantially change during the experi-
ment. Part of the reason for this was attributed to the fact
that after 4 months of monitoring, the cooling coils were
found to be leaking condensate into the supply duct and
needed to be replaced. The conveyance of moisture into
the adjacent duct may have been the reason for the initial
growth of mold. After replacing the defective cooling
coils, only background concentrations of fungal contami-
nation were found on FGDL surfaces. The reduced
moisture level created by the new cooling coils lessened
the potential for the growth of mold and effectively
terminated the test house field experiment before
differences in antimicrobial efficacy could be demon-
strated. After monitoring the test house for 7 months (a
complete cooling season) without evidence of fungal
concentration exceeding background levels, the experi-
ment was stopped [34, 35].
Dynamic chamber laboratory experiments of untreated
FGDL removed from the test house as bulk samples
indicated that the population of A. versicolor increased by
3 logs (1,000 fold) by the end of the first month and
remained approximately level through the 3.5-month study
[34]. A slight variation increase followed by a decrease in
the A. versicolor population was observed for Coatings I
and III on FGDL, for the period of 3.5 months. This was
in contrast to the increased populations observed in
samples with Coating II. The increase in fungi population
for Coating II was comparable to that observed in the
untreated samples, as compared with successful limiting of
growth accomplished by Coatings I and III [34]. The
laboratory testing of these three antimicrobial encapsulants
are described in the previous section titled Evaluation of
Antimicrobial Treatments as Control Technologies for
laboratory testing.
Soiled FGDL experiments (described in the previous
section) resulted in similar populations of A. versicolor for
untreated and Coating II samples. The results were again
similar for Coatings I and III, in which fungi populations
were observed to increase in the first month, and then
11
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decrease. The results indicates that antimicrobials can
remain effective with moderate dust loading [34].
A comparison of mold growth on FGDL demonstrated in
the SMTC results (conducted at 70%, 85%, 90%, and 94%
RH) with DMTC (conducted at 94 % RH) results for the
three types of antimicrobials agreed. Increased fungi
growth for Coating II was comparable to that observed in
the untreated samples, whereas limited growth was ob-
served on Coatings I and III [34]. These studies demon-
strated that SMTC fungi growth decreases with de-
creasing RH. Both SMTC and DMTC methods indicate
greater effectiveness in controlling growth with Coatings
I and III [34].
Characterization of Emission Rates and
Modeling of Exposure through HVAC
Operation
Biological contaminants are known to be indoor air
pollutants which carry a substantial health risk with
exposure [1, 3, 15-18]. Biological allergens (mold, bac-
teria, pollen, animal dander, dust mites, roaches) and their
fragments in the size range of 0.3 to 10.0 |im make up a
component of airborne particulate matter (BioPM),
addressed by Menetrez, et al. [36, 37], and Foarde, et al.
[38]. BioPM is composed of a large variety of viable and
non-viable organisms, some of which can be infectious
bacteria and fungi, as well as fragmented pieces of biolog-
ical organisms which can be allergenic, toxic, immuno-
suppressant or can produce inflammatory responses [1,3,
15-18]. To limit exposure to BioPM in the ambient and
indoor environment, the development of control technol-
ogies are required. Antimicrobial agents and biocides have
long been used to control, prevent, and remediate micro-
bial growth for many different applications in the environ-
ment. This research deals with BioPM as the main phy-
sical mechanism for pulmonary exposure to biocontam-
inants. The objective of this research was to evaluate the
sample collection and analysis of BioPM in the size range
of 0.3 to 10.0 |im [32, 36-38]. An additional part of this
work quantified the emission rates of mold (a substantial
constituent of BioPM) and used a computer model to
project its distribution throughout a school with a HVAC
dominated indoor environment.
The importance of minimizing exposure to BioPM of
indoor origin in the indoor environment is well
established. However, the importance of minimizing
exposure to BioPM in the ambient fraction of particulate
matter with aerodynamic diameters less than 2.5 |im
(PM25) has not been well studied. In 1998, North
American Research Strategy for Tropospheric Ozone and
Aerosols (NARSTO) listed a number of ambient PM con-
stituents that need further study. One of these is BioPM,
now thought to be a previously unrecognized causative
agent for adverse health effects.
Biological agents potentially play two important roles in
influencing the adverse health effects that have been
associated with PM25 exposures. The first role is as a
constituent of PM. The second is as an agent to exacerbate
adverse health effects in sensitive individuals. In addition
to mold, yeast, and pollen, bacteria are present in both
ambient and indoor air from a large variety of human
activities. Agricultural activities such as plowing or hog
production, manufacturing operations such as cotton mills
or grain storage, and waste treatment activities such as
wastewater treatment release airborne bacteria. The size
range of airborne bacteria is from 0.5 to 2.0 |im (Bacillus
spp., Pseudomonas spp., Xanthomonas spp., and
Arthrobacter spp.), and submicrometer fragments of these
gram-negative organisms can contain toxins that are
combined with their cell wall. These fragments are known
as endotoxins and, when inhaled, have been shown to
increase non-specific bronchial reactivity in asthmatics
[33,36-38,40].
The evaluation and control strategy of BioPM had been
complicated by the lack of methods that will allow us to
quantitatively assess the BioPM fraction of PM2 5. At issue
is how much of occupant exposure originates indoors, how
much is derived from outdoor sources, and what is the
interaction. Because PM exposure indoors can originate
from both indoor and outdoor sources, it was determined
that information was needed to scientifically quantify the
relationship between indoor and outdoor levels of PM
aerosols [33, 36-40].
An investigation has been conducted into the feasibility of
12
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developing sampling methods and analysis techniques for
quantifying BioPM and its relative distribution indoors
and outdoors. Andersen non-viable impactor samples of
the indoor and outdoor air were collected over a 5-week
period from three sites. Then the samples were analyzed
for total protein, ragweed, and fungal antigens (3-1,3
glucan and endotoxins. A preliminary assessment of a
variety of sampling methods for the measurement of the
BioPM fraction of indoor and outdoor PM was performed
for the purpose of method optimization [33, 36-40]. The
results indicated that sizable fractions of the fine PM
sampled in both indoor and outdoor samples were of bio-
logical origin. These results are preliminary, and although
they establish the presence of abiological component with
indoor and outdoor fine PM, additional research is needed
to (1) further develop the measurement method, (2) quan-
tify the relationship between indoor and outdoor levels of
the BioPM, and (3) determine the fraction of ambient and
indoor PM that is biological [33, 36-40].
Biocontaminants such as mold spores are capable of being
released into the indoor air from the site of growth and
being transported in a viable ornon-viable form. Exposure
to toxic mold and the mycotoxins contained in the spore
and vegetative body have been shown to produce adverse
health effects resulting from inhalation, ingestion, and
dermal contact. A study of the release of Stachybotrys
chartarum spores from contaminated gypsum wallboard
showed the effects of environmental conditions on the
transport of viable and non-viable spores and fragments.
The findings of S. chartarum spore emissions with low air
velocity flow conditions were found to be directly
proportional to airflow and indirectly proportional to
relative humidity. These emission findings corroborate
previous observations involving Penicillium and
Aspergillus [33, 36, 37]. Viability of S. chartarum spore
emissions was also discussed with respect to culturable
and commonly used field measurement techniques [37].
Spore emission rates from FGDL for Penicillium
chrysogenum and Aspergillus versicolor at four RH levels
(95%, 85%, 75% and 65%) of conditioned input air and
two velocities (35 and less than 10 cm/s) were used to test
emission rates. Emissions increased as the air velocity
increased across the contaminated FGDL surface. The
published experiments also found that as the RH was
lowered, the emission rate increased for both organisms.
These relationships were investigated to determine
whether the spore emission rate for S. chartarum would
follow a similar pattern. The emission rates did confirm
that S. chartarum spore emissions increase with increased
airflow and as the humidity is lowered [37, 38].
A second set of experiments was designed to quantify
emissions rates of four room-wall simulators (RWS) con-
taining gypsum wallboard (see Figure 11), at 65% RH
over an extended period of time. The RWSs (17.1 x 17.1
x 161.7 cm duct section) were constructed of 16 gauge
stainless steel, and the front cover is made from glass to
permit visual monitoring of growth on the test material.
The top inlet and bottom discharge ends of the RWS tran-
sition to 2.5 cm diameter tubes. Each RWS was con-
nected to a pressurized plenum fed by a blower drawing
HEPA filtered and conditioned air from the DMTC. A
single piece of gypsum board 107 x 42 cm (4494 cm2) was
scored length-wise to permit it to be folded into a three-
sided trough that fit into and formed the interior walls of
the RWS. Air from the plenum entered each RWS through
the inlet transition, flowed vertically down along the glass
and three-sided gypsum board wall, and discharged
through the outlet transition. Bioaerosol samples were
drawn from the RWS discharge air through the bottom end
into the tubing which penetrated the DMTC wall and into
the sampling equipment. Surface concentrations of spores
Figure 11. Room Wall Simulators.
13
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on the gypsum wallboard in the four RWS were essen-
tially the same. However, the age of the growth on the
RWS 1, 2, and 3 was 2 months old when the emission ex-
periments were initiated. RWS 4 was 3 months old and
exhibited a higher sustained rate of emissions. RWS 1
through 3 started at levels between 500 and 1,000
CFU/m2/hr and tapered off by day 2 or 3. RWS 4 started
at the same initial level as RWS 1, 2, and 3 but continued
to emit that same level of spores for the entire first week
before the level started to decrease. Emission of spores at
an RH below 65% and an air velocity of 35 cm/s exhibited
rates of both culturable colony forming units (CPUs) and
total spores per square meter that were sustained at a high
level of emission for an 80 day period. For 30 days CPUs
appeared to be about 10% of the number of total spores.
After 30 days, the levels of culturable CPUs and total
spores approach being equal. For the final 25 days, the
percentage of culturable CPUs was lower than the second
30 days, but higher than the first 30 days [37].
To better understand the implications of the emission rates
measured in these experiments, an IAQ model was used to
estimate concentration and potential exposure, as has been
done with other indoor air contaminants. The goal was to
estimate the potential indoor concentration levels using a
typical emission rate from these experiments and reason-
able ventilation parameters. The model used was RISK
IAQ Model for Windows [38], which is a completely
mixed room model incorporating source/sink behavior that
can generate concentration and exposure estimates as
functions of time. The ventilation flows and pollutant
emission rates were set at desired levels for each modeled
room. For this study, a school was modeled having 10
rooms distributed over a floor area of 371.6 m2 with 2.44-
m ceilings and served by one HVAC system. The total
HVAC airflowtothe area was 56.6 nrVmin (2000 ftVmin).
The school was assigned three separate HVAC scenarios:
(1) source with the HVAC always on and no outdoor air,
(2) source with HVAC on from 0800 to 1700 hours, 9-
hour duty cycle, and no outdoor air, and (3) the same 9-
hour duty cycle as scenario 2 with the addition of an
outdoor air intake level of 5% of the recirculating airflow
to the HVAC system as is the case in atypical school. The
source of emission was calculated from an area of mold
contamination of 9.29 m2 (100 ft2), selected as repre-
sentative of a modest area of contamination. The actual
areas of mold contamination in a problem building would
be expected to emit S. chartarum spores at a similar con-
centration per unit of contaminated surface area.
Concentration profiles were run, and the exposure levels
were calculated for an average person (breathing at 0.83
m3/h and, for simplicity, staying in the room 24 h/day) for
all scenarios. The model specified a building dominated
by the HVAC system and having a building air exchange
rate of 3.8 air changes per hour (ach). Various surfaces
within a building, as well as components of a HVAC
system may support microbial growth. The area of con-
tamination was located within the school building, which
was served by the HVAC system as defined by each of the
three scenarios modeled. From the evaluation, we can
conclude that running the HVAC system constantly will
result in the maximum transport of spores and the greatest
concentration of both total spores and CPUs and is the
worst scenario of the three. A nine hour HVAC duty cycle,
which is 37.5% of the time, was calculated by the model
to have a proportional decrease in total spores and CPUs,
and an air filter that removes 40% of the particulate matter
from the air stream can also achieve a significant decrease
in total spores and CPUs. This indicates that air filtration
is able to dramatically reduce spore concentrations from
unacceptably high levels to background levels. For this
simulation using the RISK IAQ Model for Windows [38],
mold spore concentrations were reduced from 680 to 60
CFU/m3 by use of a 40% particle filter, effecting a one
order of magnitude decrease. It should be stressed that the
model treats the entire area of the school as a well mixed
reactor, having equal concentrations regardless of the
proximity to the source of contamination, and only con-
siders entire spores in the 4 to 6 |im size range [37].
Understanding the factors that govern spore release,
aerosolization, and transport allows prediction of potential
exposure. S. chartarum spore emissions from gypsum
board at low flow are directly proportional to airflow and
inversely proportional to RH, which supports previous
research with Penicillium and Aspergillus. The relation-
ship between the culturable CPUs and total spores needs
14
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further investigation. However it demonstrates the dif-
ficulty in providing a correlation between airborne field
measurements (using culturable CPUs) and human ex-
posure. Air filtration can be a helpful tool for minimizing
exposure and containing mold contamination while
remediation is performed [37].
Improved Methods of Mold Sampling and
Analysis
Mold contamination exists in either solid (intact spores
and fine non-viable particles) or gaseous state (microbial
volatile organic compounds). Fine particulate matter is a
significant pollutant both indoor and outdoors, and
measurements of indoor exposures are needed for eval-
uating total long-term personal exposures to PM25 origi-
nating from both ambient outdoor and indoor sources.
Bacteria, fungi, viruses, and allergens are important com-
ponents of outdoor and indoor aerosols. Desiccated, non-
viable fragments of these organisms are also ubiquitous.
These fragments can remain pathogenic, toxic, or aller-
genic (depending upon the specific organism or organism
component) [36, 37, 39, 40]. The collection and charac-
terization of these fragments were addressed as BioPM in
the previous section, Characterization of Emission Rates
and Modeling of Exposure Through HVAC Operation.
The identification of gases produced by the metabolic
activity of mold and the development of molecular iden-
tification techniques of mixed samples of mold spores are
addressed in the following sections.
Evaluation of Microbial Volatile Organic Com-
pounds
Analytical methods are available to comprehensively
identify microbial populations and metabolites (particles
and gases). A study by Menetrez et. al. [39] examined
MVOC emissions from six fungi (Aspergillus glaucus,
Aspergillus versicolor, Cladosporium sphaerospermum,
Penicillium chrysogenum, Penicillium italicum, Rhodo-
torula glutinis) and one bacterial species (Streptomyces
spp.) commonly found in indoor environments. Data were
presented on peak emission rates from inoculated agar
plates loaded with surface growth, ranging from 33.5
|lg/m2/24 hr for Cladosporium sphaerospermum (pre-
dominantly 8-Humulene, Tetramethyltetrahydrofuran, and
a-Humulene) to 515 |ig/m2/24 hr for Rhodotorula glutinis
(predominantly 3-Methyl-2-butanol, Phenyl ethyl alco-
hol, and 2-Methyl-l-propanol). Furthermore, changes in
MVOC emission levels over the growth cycle of two of
the microorganisms are examined. In addition, a calcu-
lation of the impact of MVOC emissions on indoor air
quality in a typical house is made, as well as an appli-
cation of an exposure model used in a typical school
environment [42].
A unique method was developed and utilized for the
purpose of this research in which microbial growth was
allowed on agar plates (glass) enclosed within glass
blocks. Consecutive growth periods of 24 hr were
followed by gas collection of analytes adsorbed on the
Tenax-TA tubes which were thermally desorbed at 250 °C
into a GC equipped with a flow splitter [42].
The measured emission rates reflect single-day emissions
from the tested microorganisms. In a growth environment,
fungi experience an exponential increase in biomass
(normally in hours) followed by a period of time where
biomass either remains constant, or drops because of
autolysis. Thus, the MVOC peak emission rate variations
observed may be a direct result of normal microbial meta-
bolism. It was anticipated that observed emission varia-
tions between replicates would be significant. However,
the actual MVOC peak emission rates were relatively
consistent between replicates, possibly due to the 24-hour
sampling time chosen for this investigation. The long
sampling period may have minimized the influence of any
short-time variations in MVOC emissions inside the
chambers. The variation from the mean average for the test
microorganisms as a group was ±28%; the largest single
variation was 76%, and the lowest was 5.4% [44].
Rhodotorula glutinis yielded the highest average peak
emission rate (515 |ig/m2/24 h); whereas, Cladosporium
sphaerospermum produced the lowest (33.5 |ig/m2/24 h).
The average peak emission rate for all the microorganisms
studied was 206 ±57.8 |ig/m2/24 h, based on a total
colonized surface area of 1 m2 [42].
Experiments were performed to evaluate peak emission
rate variations over the life of the biocontaminant
15
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colonies. Agar plates inoculated withPenicillium chryso-
genum and Streptomyces spp. were introduced into the
glass chambers, followed by headspace collection. Head-
space samples were collected when the colonies were new
(2-4 days), mature (6-14 days), and middle-aged (14-18
days). MVOC emissions increased to a maximum at the
later sample periods [42].
Except for Penicillium italicum, the primary MVOC
emissions from each microorganism were alcohols and
ketones. Percent emissions were calculated by dividing
each MVOC's mass concentration by the total mass
quantified from each biocontaminant after background
subtraction. Alcohols and ketones, as well as reduced
sulfur compounds (e.g., dimethyl disulfide, dimethyl
trisulfide), have been identified previously as odiferous
compounds associated with biocontaminated indoor
environments. These compounds account for 32-93% of
the total MVOC emissions found in individual samples.
The reduced sulfur compounds are classified as hetero-
compounds in this article, which also include the nitrogen-
containing compounds identified throughout the experi-
ment (e.g., 2-acetylthiazole, methylpyrazine). Predomi-
nant emissions from Penicillium italicum were identified
as sesquiterpenes, consistent with the many sesquiterpene
compounds. The identification of a-humulene in the head-
space of Cladosporium sphaerospermum suggests its pos-
sible role as a microbe-specific growth indicator, yet the
emission of this sesquiterpene by C. sphaerospermum
growing on building materials needs to be ascertained
before it is identified as such [42].
Volatile metabolic by-products from seven common in-
door biocontaminants have been studied in an attempt to
establish MVOC peak emission rates from each individual
microorganism and to evaluate their impact on overall
IAQ. Many of the MVOCs identified in this study have
been reported; however, identification of 8-humulene and
a-humulene in the headspace of Cladosporium sphaero-
spermum suggests their possible role as microbe-specific
growth indicators [42] which could be used as an indicator
of indoor mold growth.
To better understand the implications of the emission rates
measured in these experiments, an IAQ model was used to
estimate concentration and potential exposure as has been
done with other indoor air contaminants. The goal was to
estimate the potential indoor concentration levels for using
a typical emission rate from these experiments and
reasonable ventilation parameters [38]. As in other ex-
periments, the RISK IAQ Model for Windows was used,
which assumes a completely mixed room model con-
ditions and incorporating source/sink behavior that can
generate MVOC concentration and exposure estimates as
function of time [38].
For this study, a school was modeled having 10 rooms
distributed over a floor area of 371.6 m2 with 2.44-m
ceilings and served by one HVAC system. The source of
MVOC emissions was calculated from an area of mold
contamination of 9.29 m2 (100 ft2), selected as repre-
sentative of comparable emission rates.
Emission modeling for the different scenarios of HVAC
operation demonstrated concentrations of MVOCs similar
to those reported in the literature for known problem
buildings [38]. The modeling results suggest that, as more
MVOC emissions are identified, future modeling might
serve as an aid in assessing the impact of MVOCs on
indoor environments by providing field investigators a
tool that can be used to assess the potentially deleterious
effect of microbiological metabolic by-product emissions
to overall IAQ [42].
Development of Rapid Multiplex PCR
Characterization of mold has previously been limited to
visual identification or morphology [43, 44]. However,
questions of accuracy and reproducibility have revealed a
need to go beyond visual identification and growth
morphology methods. The use of molecular biology has
brought about significant change in microbiology which
we have focused on the characterization of mold.
The growth of filamentous fungi (mold) in the indoor
environment occurs in a dynamic setting. Environmental
conditions are constantly changing allowing a vast diver-
sity of different organisms to establish a stronghold and
flourish in the built environment. These different organ-
16
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isms produce a myriad of MVOCs, toxins, and allergenic
components. Although only a small percentage of fungal
species has been directly associated with adverse health
affects, increased awareness and continuing research will
likely result in the identification of many more pathogenic
and toxigenic species. In order to properly assess the fun-
gal exposure to building occupants and the implement an
effective remediation strategy, it is imperative that fungal
screening, isolation, and characterization be performed.
Research has been conducted utilizing molecular methods
of fungal identification and characterization that are rapid,
easy to perform, and cost sensitive. Molecular character-
ization research uses the PCR as the backbone of these
applied strategies. The PCR allows for the amplification
of virtually unlimited quantities of specific target DNA
from the organism of interest. This amplified DNA can
then be used to identify the organism or coupled to other
technologies such as restriction fragment length poly-
morphism (RFLP) analysis, or genetic sequencing to un-
ambiguously identify the organism of interest.
Reducing the time necessary for accurate fungal identi-
fication is an important aspect of rapid exposure esti-
mation. To reduce the time necessary to extract and purify
fungal DNA suitable for the PCR, research was performed
to simplify existing technologies while maintaining high
quality results [45]. It was determined that a simple
mechanical disruption of the fungal spores followed by a
phenol: chloroform-ethanol concentration produced highly
purified DNA suitable for molecular analysis. The pro-
cedure, taking only one hour to complete, greatly enhances
the rapidity of the subsequent molecular identification
regime [45].
When attempting to identify numerous organisms, mul-
tiple PCR reactions are needed, increasing both the time
and cost of identification. Additional PCR research has
focused on developing and optimizing a multiplex PCR
that is capable of identifying four organisms in a single
reaction. Research efforts have developed a simple method
to identify multiple fungal species, Stachybotrys char-
tarum, Aspergillus versicolor, Penicillium purpurogenum,
and Cladosporium spp. by performing multiplex PCR and
distinguishing the different reaction products by their
mobility during agarose gel electrophoresis [46]. Positive
identification made by multiplex PCR can be made within
24 hours following receipt of the samples [46].
Additional research has been carried out by coupling the
PCR to RFLP patterns. Following the amplification of the
target sequence, the amplified DNA is enzymatically
broken down, or digested, into smaller components. The
specific smaller components are a direct result of the DNA
sequence of each of the organisms. Identification can then
be made by analysis of the different reaction products
following their mobility during agarose gel electro-
phoresis. The results indicate that organisms belonging to
the genuses Stachybotrys, Cladosporium, Aspergillus, and
Penicillium can accurately be identified using their mo-
bility patterns following treatment with only four digestion
enzymes [47]. These identifications are unambiguous,
allowing for rapid and positive identification of fungal
contaminants [45-47].
PCR coupled to genetic sequencing has also been a major
research focus. The overarching goal of this research is to
positively identify as many organisms at a time as rapidly
as possible. An environmental sample may contain hun-
dreds of fungal spores and fragments: therefore, following
PCR with multiple organisms, the amplified DNA is in-
serted into a mobile genetic element and inserted into bac-
terial cells. Each bacterial cell will contain only a single
piece of PCR amplified DNA or the DNA from only a
single organism of the environmental sample. The bac-
terial DNA can then be recovered and genetic sequencing
carried out specifically for the PCR amplified fragment.
The individual genetic sequences are then used to posi-
tively identify each organism present in the sample. Initial
research results have shown the methodology capable of
identifying Stachybotrys, Cladosporium, Aspergillus, and
Penicillium species from amixed culture environment and
the generation of a patent application for this technology
[45-47].
Fungal sampling and collection technology from various
building materials has also produced results that save time
and are inherently more accurate and cost effective. The
17
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objective of this research was to develop a sample test population [48]. This research showed that homogenizing
method for porous building materials (ceiling tile) to samples of bulk ceiling tiles with the masticator blender is
detect and quantify mold growth utilizing a masticator an effective method to recover mold spores from porous
blender. By comparing results to previously used tech- surfaces and can be a helpful tool when applied to
nologies, statistical analysis showed that the masticator antimicrobial efficacy testing.
method yielded significantly higher estimates of the mold
18
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Discussion
The significant technical findings of the research discussed
in this report are summarized and listed below. They often
overlap into more than one of the six research areas. An
example of this is the development of the static and
dynamic chambers which were utilized in all six research
areas addressed in this report. The discussions of each of
the six research areas and the technical findings are only
meant to summarize the referenced publications and not to
restate technical results from those publications. The
findings listed below fall into one or more of the following
areas: (1) research and development of risk management
alternatives to prevent and control the growth of mold by
studies to quantify the effects of moisture, RH, and dust,
(2) duct cleaning effectiveness for prevention and control
of microbial growth on duct materials, (3) evaluation of
antimicrobial treatments as control technologies, (4) field
testing of sealants and encapsulents used in air duct
systems, (5) characterization of emission rates and
modeling of exposure through HVAC operation, and (6)
improved methods of sampling and analysis of mold.
19
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Technical Findings
Static Microbial Growth Chamber
• Establishing that SCTM was adapted into ASTM
Standard 6329-98 [21]. The SCTM defines how to
conduct microbial testing in a well-controlled envi-
ronment. The SCTM is essential to understanding
mechanisms and improving the repeatability and
comparability of data [20, 21].
Dynamic Microbial Growth Chamber
• Developing the DMTC, a room-sized dynamic
chamber, designed and constructed under a co-
operative agreement between EPA and RTI, that can
test HVAC ducts (mini-ducts) scaled to simulate
horizontal duct velocities and duct materials in the
mini-duct apparatus [29], or vertical low velocity air
flow against gypsum wallboard with room wall
simulators [37].
Research and Development of Risk
Management Alternatives to Prevent and
Control the Growth of Mold by Studies to
Quantify the Effects of Moisture, Relative
Humidity, and Dust
• Determining that water incursion or standing water is
not required for growth on materials [23-26]. For
some species of mold, humidity alone can provide
sufficient moisture to permit growth on building
materials (material and organism dependent), relative
to the hygroscopicity of the material [24-26].
• Developing a method for artificially soiling materials
[22]. Allowing fungal growth characterization or
antimicrobial efficacy testing methods to simulate
realistic environmental conditions will result in lab-
oratory experiments that more closely resemble real-
world applications [22].
• Confirming that, under equilibrium conditions, RH
and moisture content correlate well with mold growth
(depending on the moisture requirements of the test
organisms) [24, 25]. However, undernonequilibrium
conditions, mold growth correlates better with in-
creasing moisture content (in duct liner, ceiling tile
and gypsum wallboard) than with RH [24-28].
• Finding that variations in the characteristics of similar
building materials can impact the fungal resistance of
that material [23-27, 34]. Both new and used ma-
terials are capable of supporting mold growth, but
generally used materials (soiled) were more sus-
ceptible [25-27].
• Establishing that reducing the moisture content of
wet materials (within 3 days) before fungal growth
became established provided effective source man-
agement [24, 28]. However, established microbial
growth may continue even after the moisture content
of a particular material is lowered below that required
to initiate growth [28].
• Finding that growth of S. chartarum was not detected
in environments with a relative humidity below total
saturation and no wetting occurred. Wetting was ne-
cessary for visible growth to occur. Porous materials,
after becoming sufficiently wet, exhibited mold
growth at all relative humidity levels tested [33].
• Acutely impacting control and remediation practices
by showing that RH is inversely related to fungal
spore emissions [22, 25, 32]. Lowering uncontrolled
humidity is almost always a recommended practice
which will lead to increased airborne contamination.
This strategy points out the need for containment of
contaminated areas to prevent the spread of con-
taminants [32].
Duct Cleaning Effectiveness for Preven-
tion and Control of Microbial Growth on
Duct Materials
• Confirming that fungal growth is intrusive throughout
porous materials and that guidelines that recommend
discarding microbially contaminated porous duct
20
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material should be followed [30-32]. Mechanical
cleaning by HEPA air vacuuming was able, at best, to
reduce imbedded fiber soiling and temporarily
decrease fungal levels. These fungal populations
experienced regrowth within six weeks [31, 32].
• Finding suggest that dust and high humidity should
be properly controlled in any HVAC system to pre-
vent the growth of mold [23, 33, 34].
Evaluation of Antimicrobial Treatments as
Control Technologies
• Finding that significant variation in antimicrobial
efficacy of encapsulents to limit or eliminate bio-
logical growth indicates a need for widespread
product testing and for the development of an effi-
cacy testing protocol [34].
• Developing a method for artificially soiling materials,
as previously discussed, allowed for fungal growth
conditions of antimicrobial efficacy testing to sim-
ulate realistic environmental real-world applications
[22].
Field Testing of Sealants and Encap-
sulents Used in Air Duct Systems
• Finding differences in degrees of efficacy for three
antimicrobial coatings. Two antimicrobial coatings
limited fungal contamination for the duration of
testing. The effective coatings contained zinc oxide/
borates and a phosphated quaternary amine. The in-
effective coating contained decabromodiphenyl oxide
and antimony trioxide. Although all three antimicro-
bials are registered with the EPA, they were not
equally effective and would not be expected to per-
form equally in field use [34, 35].
Characterization of Emission Rates and
Modeling of Exposure Through HVAC
Operation
• Finding that fine PM sampled in both indoor and
outdoor samples were of biological origin. This
establishes the presence of a biological component
with indoor and outdoor fine PM [36-39, 41].
• Determining that the emission rates of fungal spores
from the surface of contaminated material result from
a complex interaction of factors. Emission rates differ
between organisms and are inversely proportional to
RH but directly related to air flow and surface
loading. Potential indoor concentrations were
modeled using RISK IAQ Model for Windows [37,
38]. The modeled levels related well to the values
reported in the literature for known problem
buildings, suggesting that, once microbial emission
rates are well enough understood, models may be
useful in predicting exposure and, eventually, risk for
individual organisms [37].
• Understanding the factors that govern spore release,
aerosolization, and transport allow prediction of
potential exposure. S. chartarum spore emissions
from gypsum board at low flow are directly
proportional to airflow and inversely proportional to
RH, which supports previous research with Peni-
ciIlium andAspergillus. The relationship between the
culturable CPUs and total spores needs further
investigation but suggests one reason that correlation
between airborne field measurements (using cultur-
able CPUs) and possible exposure is so difficult. Air
filtration can be a helpful tool for minimizing
exposure and containing mold contamination while
remediation is performed [36, 37, 40].
Improved Methods of Sampling and
Analysis of Mold
• Identifying 8-humulene and a-humulene (among the
many MVOCs from mold growth that have been
reported) in the headspace of Cladosporium sphaero-
spermum suggests their possible role as microbe-
specific growth indicators [42].
• Establishing that simple mechanical disruption
followed by standard phenol :chloroform-ethanol
concentration produces highly pure fungal DNA
suitable for use in subsequent molecular biology
applications [45].
• Developing a method of multiplex PCR that is
capable of identifying 4 environmentally relevant
fungi (Stachybotrys chartarum, Aspergillus
versicolor, Penicillium purpurogenum, and
Cladosporium spp.). The method is preferable due to
savings in time and cost with increased identification
accuracy [46].
• Developing a method of PCR followed by RFLP to
21
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generate DNA patterns that enable the identification
of medically relevant fungal organisms [46, 47].
Developing a fungal screening technology that is
capable of identifying potentially hundreds of organ-
isms from a single PCR reaction. The methodology
has been shown to work with Stachybotrys, Clado-
sporium, Aspergillus, and Penicillium species from a
mixed culture environment and has resulted in the
initiation of patent rights [46, 47].
Developing a method of sampling and collecting
fungal spores from porous building materials. The
method, involving mastication, provides more
complete analysis and accuracy by homogenizing the
sample rather than relying on surface sampling [50].
The findings listed above cover a broad expanse of re-
search related to detecting and controlling mold con-
tamination. Additional work is needed to further reduce
human exposure to biological contaminants.
22
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Conclusions
The six areas of research identified by EPA/APPCD/
IEMB for allocation of program resources were (1)
research and development of risk management alternatives
to prevent and control the growth of mold by studies to
quantify the effects of moisture, relative humidity and
dust, (2) duct cleaning effectiveness for prevention and
control of microbial growth on duct materials, (3)
evaluation of antimicrobial treatments as control
technologies, (4) field testing of sealants and encapsulents
used in air duct systems, (5) characterization of emission
rates and modeling of exposure through HVAC operation,
and (6) improved methods of sampling and analysis of
mold. These areas of research were investigated, and the
most significant findings are summarized in the discussion
section above. Advances in research and development of
prevention and control as well as mitigation practices for
a variety of molds were achieved. Understanding the
growth requirements of mold, developing the test
methodology ASTM Standard 6329-98 [21] and the static
and dynamic microbial test chambers for determining
antimicrobial efficacy, and determining the most effective
technique to identify, handle, and mitigate contaminated
materials will ultimately improve the ability to control
biological contaminants and reducing human exposure.
23
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Future Research
Future areas of microbiological research are being
planned. Progress in these areas of research will be subject
to change and re-evaluation overtime. However, these six
research areas are a logical next step from past and present
accomplishments.
Microbial Resistant Building Materials
Product Evaluation - Gypsum Wai I board
When building materials becomes exposed to moisture by
weather events, leaks in the building envelope, or in-
adequate control of relative humidity, absorption and
transport of moisture through that material often renders
it susceptible to the growth of biological contaminants.
Microbial colonization and the rapid growth and dis-
persion of mold can expose building occupants and pro-
duce severe illnesses including pulmonary, immunologic,
neurological and oncogenic disorders. Removing sub-
strates from building materials or incorporating anti-
microbial agents in the manufacture of building products
may prevent mold growth and the spread of contaminants.
The manufacture of microbial resistant building materials
(such as wallboard, ceiling tiles and flooring) can inhibit
or prevent mold growth. Limiting or preventing mold
growth by the manufacture of microbial resistant building
materials creates a product which can sustain temporary
adverse conditions and is less likely to become a source of
biological contamination, or need replacement than the
current products.
Possible methods of gypsum wallboard improvement are
being studied, including treatment with antimicrobials,
ozone, and heat during the manufacturing process. The
ability to remove viable mold from the inner sections of
the wallboard may impair the ability of mold to germinate
and grow following an isolated water incident. This re-
duction in mold growth could prevent contaminated wall-
board from having to be removed and land filled, fitting
well with the sustainability focus supported by the EPA.
The manufacture of microbial resistant gypsum wallboard
has been initiated by a number of companies producing
building material. Each company has established their own
individual manufacturing strategy for producing this
material. The resultant building material could potentially
have a longer product life and be both environmentally
friendly and less likely to need replacing than the current
products.
The evaluation of gypsum wallboard would test (1) micro-
bial growth, (2) moisture absorption, and (3) VOC emis-
sion. Established methods would be used to form the basis
of evaluation. Each product evaluation result would then
be evaluated.
Gypsum wallboard has been the selected to be the first
building material to be evaluated by this process. Eval-
uation of joint compound and tape should be addressed in
the future. The impact to the building product industry and
the consumer public can be significant, with both gaining
advantages through the sale of better products.
Microbial Resistant Building Materials
Product Evaluation - Ceiling Tiles and
Flooring
Ceiling tiles and flooring that have a greater ability to
withstand moisture and prevent mold growth will be less
problematic and in need of replacement than the current
products. Possible methods of ceiling and flooring im-
provement should be studied, including treatment with
antimicrobials, ozone, and heat during the manufacturing
process. The ability to remove viable mold from the inner
sections of the building products may impair the ability of
mold to regrow following an isolated water incident. This
reduction in mold growth could prevent contaminated
ceiling and flooring systems from having to be removed
and landfilled, fitting well with the sustainability focus
supported by the EPA.
24
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The evaluation of ceiling tiles and carpet flooring would
test (1) microbial growth, (2) moisture absorption, and (3)
VOC emission. Established methods would be used to
form the basis of evaluation.
Acoustic ceiling tiles and carpeting used as building
materials should be evaluated by this grading process. The
impact to the building product industry and the consumer
public can be significant, with both gaining advantages
through the sale of better products.
HVAC Biological Film (Biofilm) Research
The presence of moisture on HVAC system cooling coils
and drip pan from condensate flow establishes conditions
favorable for microbial growth. The established microbial
growth can then be responsible for releasing gases
(MVOCs, Dirty Socks Syndrome) or particles (BioPM)
into the conditioned airstream. The presence of a micro-
bial film on the cooling coils is also responsible for loss of
heat transfer efficiency and over-all component operation
and is the possible cause of condensate "blow-by" into the
supply air duct. The characterization of fungal organisms
and their byproducts (MVOCs and BioPM) that are re-
sponsible for this condition and the most effective form of
treatment (UV irradiation) would give building owners,
building occupants, and building remediators accurate
information for identifying and dealing with this problem.
The use of UV to destroy any biofilm that has been
established on HVAC surfaces and not allow re-growth is
the most efficacious manner of long-term treatment. The
use of UV to deactivate airborne biological contaminants
transported within existing HVAC systems adds additional
benefit to this treatment alternative. Also, the prevention
of biofilm build-up on cooling coil surfaces increases
thermal transfer efficiency and decreases HVAC system
energy use at the same time that it prevents organisms
from establishing a foothold in the indoor environment,
increasing the sustainability of current building systems.
MVOC characterization of gases and PCR identification
of particles will be used to determine the best way to
identify biofilms. An HVAC system that has an estab-
lished biological film will be treated with UV radiation.
Measurements of biological contaminants and heat trans-
fer efficiency from before and after treatment will be
conducted. Laboratory and field demonstrations will be
performed concurrently.
Asthma Triggers
Research will focus on developing optimization tech-
niques for collection and analysis of fine and ultra fine
biological particles (fungal, bacterial, pollen, animal dan-
der, and dust mite and cockroach allergens), which make
up the entire range of biological particulate matter that are
known asthma triggers. Improving methods of sampling
for these size fractions with filter or impactor collection
(such as the Andersen non-viable sampler) and subsequent
analysis via scanning electron microscopy, polymerase
chain reaction, mycotoxin presence, and gravimetric
measurements can result in a better understanding of the
potential exposure to viable and non-viable particles that
are biological asthma triggers.
Homeland Security: Tests on Thermal
Destruction Using Biocontaminated
Building Materials (Bacillus anthracis
surrogates): Bench Scale and Rotary Kiln
Incinerator
A significant amount of contaminated building material
may need to be disposed of after a bio-terrorism attack.
The efficacy of disposal of building materials con-
taminated with biological agents by incineration is compl-
icated by matrix effects associated with the contaminant
and the material. This project is examining the destruction
of surrogate biological agents inoculated on several com-
mon building materials including ceiling tiles and wall-
board. A laboratory-scale reactor and a rotary kiln are
being used in this project to examine building material,
heating temperature, and residence time affecting the
destruction of surrogate biological contaminants, including
Bacillus subtilis and Geobacillus stearothermophilus, both
surrogates for Bacillus anthracis. Work is focusing on the
thermal destruction of biological contaminants as well as
stack sampling to monitor containment of contaminant
organisms during incineration. The results from these
studies can be used to evaluate incineration technologies
for appropriateness for disposal of contaminated building
materials.
25
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Reduced Infectious Disease
The spread of infectious disease in humans can be
attributed to communication by touch and inhalation of
the infectious agents (virus and bacteria). Because air
conveyance through the HVAC can transport infectious
agents along with other particles throughout a building,
one infectious person can spread viable organisms to many
other through the conveyance of conditioned air.
Research will focus on developing optimal air treatment
techniques of fine and ultra fine biological particles (viral,
bacterial) which are responsible for the spread of infec-
tious disease. Biological surrogates of viral and bacterial
organisms will be inoculated into the air stream traveling
through an HVAC system. Improving methods of treat-
ment can result in less transmission of infectious disease
as well as better biological indoor air quality.
Research will involve UV irradiation to destroy viral and
bacterial microorganisms on a surface and in a moving
stream of air. Although the ability of UV to destroy
bacillus spores and mold has been demonstrated, the
antiviral or anti-pneumonia efficacy of UV on surfaces
and in a moving airstream is largely unknown. The use of
UV to clean air within an existing HVAC systems can
have other beneficial effects such as preventing fungal
growth and decreasing the energy use.
26
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Indoor/outdoor ratios of biological PM: A preliminary study
(Extended abstract). Proc. Air and Waste Manage Assoc
Specialty Conference, PM2000: Particulate Matter and
Health - The Scientific Basis for Regulatory Decision-
Making, Charleston, SC, VIP-94.
40. Menetrez, M.Y.; Foarde, K.K.; Ensor, D.S., 2000. Fine
biological PM: understanding size fraction transport and
exposure potential (Poster). Proc. Air and Waste Manage
Assoc Specialty Conference, PM2000: Particulate Matter
and Health - The Scientific Basis for Regulatory Decision-
Making,, Charleston, SC, VIP-94.
41. Menetrez, M.Y.; Foarde, K.K.; Ensor, D.S., 2000 Com-
parison of analytical methods for the measurement of non-
viable biological PM. Proc. of the Air and Waste Manage.
Assoc. Specialty Conference, Engineering Solutions to
Indoor Air Quality Problems.
42. Menetrez, M.Y.; Foarde, K.K., 2002, Microbial volatile
organic compound emission rates and exposure model.
Indoor and Built Environ. DOL10.1159/000066016.
11:208-213.
43. Burge, H.A., 1995 Bioaerosols in Residential
Environments. Bioaerosols Handbook, Chapter 21. CRC
Press, Boca Raton, FL, pp 579-591.
44. Willeke K., Macher J.M., 1999. Air sampling: bioaerosols,
assessment and control. American Conference of
Governmental Industrial Hygienists, Kemper Woods
28
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Center, Cincinnati, OH. pp 11/1-25. 2005. A simple polymerase chain reaction/restriction
45. Dean, T.R.; Betancourt, D.; Menetrez, M.Y., 2004. A rapid fragment length polymorphism assay capable of identifying
DNA extraction method for PCR identification of fungal medically relevant filamentous fungi. Molecular
indoor air contaminants. J. Microbiol Methods. 56:431-434. Biotechnology. 31:21-28.
46. Dean, T.R.; Roop, B.; Betancourt, D.; Menetrez, M.Y., 48. Betancourt, D.A.; Dean, T.R.; Menetrez, M.Y., 2005.
2005. A simple multiplex polymerase chain reaction assay Method for evaluating mold growth on ceiling tile. J.
for the identification of four environmentally relevant Microbiol Methods. 61:343-347.
fungal contaminants. J. Microbiol Methods. 61:9-16.
47. Dean, T.R.; Kohan, M.; Betancourt, D.; Menetrez, M.Y.;
29
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
1. REPORT NO.
EPA-600/R-06/011
2.
3. RECIPIENT'S ACCESSION NO.
4. TITLE AND SUBTITLE
Research and Development of Risk Management Alternatives for
Controlling Mold
5. REPORT DATE
February 2006
6. PERFORMING ORGANIZATION CODE
7. AUTHORS
Marc Y. Menetrez, Timothy R. Dean, and Doris A. Betancourt
8. PERFORMING ORGANIZATION REPORT NO.
9. PERFORMING ORGANIZATION NAME AND ADDRESS
See Block 12
10. PROGRAM ELEMENT NO.
11. CONTRACT/GRANT NO.
In-house
12. SPONSORING AGENCY NAME AND ADDRESS
U. S. EPA, Office of Research and Development
Air Pollution Prevention and Control Division
Research Triangle Park, North Carolina 27711
13. TYPE OF REPORT AND PERIOD COVERED
Capstone; 1995-2005
14. SPONSORING AGENCY CODE
EPA/600/13
15. SUPPLEMENTARY NOTES
EPA Project Officer is Marc Y. Menetrez, Mail Drop E305-03, Phone (919) 541-7981, e-mail
menetrez.marc@epa.gov
16. ABSTRACT
The report discusses research conducted since 1995 into controlling biological contamination in the indoor
environment. The areas that have been addressed are (1) research and development studies to quantify
the effects of moisture, relative humidity (RH), and dust and develop risk management alternatives for
prevention and control of mold growth; (2) duct cleaning effectiveness for prevention and control of
microbial growth on duct materials: (3) evaluation of antimicrobial treatments as control technologies; (4)
field testing of sealants and encapsulents used in air duct systems; (5) characterization of emission rates
and modeling of exposure through heating, ventilating, and air conditioning operation; and (6) improved
methods of sampling and analysis of mold. The conclusions resulting from this body of research are
summarized and the interrelationships of these areas of investigation in reducing human exposure to
biological contamination in the indoor environment are put into perspective. As part of the opening of the
new EPA Environmental Resource Center Building in 2002, the EPA's Air Pollution Prevention and Control
Division, Indoor Environment Management Branch has established the Biocontaminant Laboratory (BL) for
conducting applied risk management research. The BL is a multi-functional, state-of-the-art
biological/molecular research laboratory engaged in intra/inter laboratory cooperative research. A
description of the BL facility is included.
17.
KEY WORDS AND DOCUMENT ANALYSIS
a. DESCRIPTORS
Air Pollution
Biological Laboratories
Fungi
Stachybotrys chartarum
Penicillium chrysogenum
HVAC
Wallboard
18. DISTRIBUTION STATEMENT
Release to Public
b. IDENTIFIERS/OPEN ENDED TERMS
Pollution Control
Stationary Sources
19. SECURITY CLASS (This Report)
Unclassified
20. SECURITY CLASS (This Page)
Unclassified
c. COSATI Field/Group
13B
14B
06C
06F
06M
13A
13C
21. NO. OF PAGES
36
22. PRICE
30
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United States
Environmental Protection
Agency
National Risk Management
Research Laboratory
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