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Testimony before the Chairman, Subcommittee on National Security, 
Emerging Threats, and International Relations, House Committee on 
Government Reform, House of Representatives:

United States Government Accountability Office:


For Release on Delivery Expected at 2:00 p.m.

Tuesday, April 5, 2005:

Anthrax Detection:

Agencies Need to Validate Sampling Activities in Order to Increase 
Confidence in Negative Results:

Statement of Keith A. Rhodes, Chief Technologist, Center for Technology 
and Engineering, Applied Research and Methods:


GAO Highlights:

Highlights of GAO-05-493T, a testimony before Chairman, Subcommittee on 
National Security, Emerging Threats, and International Relations, House 
Committee on Government Reform, House of Representatives:

Why GAO Did This Study:

In September and October 2001, letters laced with Bacillus anthracis 
(anthrax) spores were sent through the mail to two U.S. senators and to 
members of the media. These letters led to the first U.S. cases of 
anthrax disease related to bioterrorism. In all, 22 individuals, in 
four states and Washington, D.C., contracted anthrax disease; 5 died. 
These cases prompted the Subcommittee to ask GAO to describe and assess 
federal agencies' activities to detect anthrax in postal facilities, 
assess the results of agencies' testing, and assess whether agencies' 
detection activities were validated.

What GAO Found:

GAO recommends that the Department of Homeland Security (DHS) develop a 
coordinated approach to working with federal agencies, so that 
appropriate validation studies of various activities involved in 
detecting anthrax are conducted. The DHS Secretary should also ensure 
that an agreed-on definition of validation is developed; appropriate 
investments are made to explore improved sampling strategies; and 
agencies' policies, procedures, and guidelines reflect the results of 
all these efforts. DHS stated that while it has the overall 
responsibility for coordination, EPA and HHS have the lead roles in 
responding to biological attacks. DHS said that it would coordinate 
with EPA to ensure that appropriate investments are made to explore 
improved sampling.

What GAO Recommends:

The U.S. Postal Service, Centers for Disease Control and Prevention 
(CDC), and Environmental Protection Agency (EPA) conducted several 
interdependent activities, including sample collection and analytic 
methods, to detect anthrax in postal facilities in 2001. They developed 
a sampling strategy and collected, transported, extracted, and analyzed 
samples. They primarily collected samples from specific areas, such as 
mail processing areas, using their judgment about where anthrax would 
most likely be found--that is, targeted sampling. The agencies did not 
use probability sampling in their initial sampling strategy. 
Probability sampling would have allowed agencies to determine, with 
some defined level of confidence, when all results are negative, 
whether a building is contaminated. This is important, considering that 
low levels of anthrax could cause disease and death in susceptible 

[See PDF for image]

[End of figure]

The results of the agencies' testing in 286 postal facilities were 
largely negative--no anthrax was detected. But negative results do not 
necessarily mean that a facility is free from anthrax. In addition, 
agencies' detection activities (for example, sample collection and 
analytical methods) were not validated. Validation is a formal, 
empirical process in which an authority determines and certifies the 
performance characteristics of a given method. Consequently, the lack 
of validation of agencies' activities, coupled with limitations 
associated with their targeted sampling strategy, means that negative 
results may not be reliable.

In preparing for future incidents, the agencies have (1) made some 
changes based on what has been learned about some of the limitations of 
their sampling strategies, (2) made some revisions to their guidelines, 
and (3) funded some new research. In addition, the Department of 
Homeland Security (DHS) has taken on the role of coordinating agencies' 
activities and has undertaken several new initiatives related to 
anthrax and other biothreat agents. However, while the actions DHS and 
other agencies have taken are important, they do not address the issue 
of validating all activities related to sampling. Finally, the agencies 
have not made appropriate and prioritized investments to develop and 
validate all activities related to anthrax and other biothreat agents.

To view the full product, including the scope and methodology, click on 
the link above. For more information, contact Keith Rhodes, 202-512-

[End of section] 

April 5, 2005:

Mr. Chairman and Members of the Subcommittee:

We are pleased to participate in this hearing by presenting our 
assessment of the federal agencies'--U.S. Postal Service (USPS), 
Centers for Disease Control and Prevention, and Environmental 
Protection Agency (EPA)--activities conducted to detect anthrax in 
postal facilities in 2001. My statement is based on our report, 
entitled Anthrax Detection: Agencies Need to Validate Sampling 
Activities in Order to Increase Confidence in Negative Results, which 
was issued on March 31, 2005.[Footnote 1]

As you know, in September and October 2001, contaminated letters laced 
with Bacillus anthracis, or anthrax spores,[Footnote 2] were sent 
through the mail to two senators, Thomas Daschle and Patrick Leahy, and 
members of the media. The postal facilities in New Jersey and 
Washington, D.C., that processed the senators' letters became heavily 
contaminated.[Footnote 3] Other mail routed through these facilities, 
as well as additional ones in the postal network, also became 
contaminated. In addition, numerous federal facilities in the 
Washington, D.C., area were later found to be contaminated. The letters 
led to the first cases of anthrax disease related to bioterrorism in 
the United States. In all, 22 individuals contracted anthrax disease in 
four states (Connecticut, Florida, New Jersey, and New York) as well as 
in Washington, D.C. Five of these 22 individuals died.

The threat of bioterrorism has been recognized for a considerable time. 
Long before the anthrax incidents, several hoax letters indicating the 
presence of anthrax had been mailed to federal and state agencies, as 
well as to private sector organizations. In calendar year 2000, the 
Federal Bureau of Investigation (FBI) responded to about 250 cases 
potentially involving weapons of mass destruction. Of these, 200 were 
related to anthrax, although all turned out to be hoaxes. Nevertheless, 
these events raised the possibility that facilities could become 
contaminated and would therefore have to be evaluated for environmental 
contamination. However, federal agencies have not been fully prepared 
to deal with environmental contamination, that is, anthrax released 
through the mail, including the potential for multiple dispersals in 
indoor environments.[Footnote 4]

Before I discuss our assessment, let me first present some background. 
(See appendix I for a discussion of our scope and methodology.)


Although anthrax can infect humans, it is most commonly found in plant- 
eating animals. Human anthrax infections are rare in the United States, 
and when infection does occur, it usually results from occupational 
exposure to infected animals or contaminated animal products, such as 
wool, hides, or hair. Anthrax infection can occur (1) cutaneously, 
usually from a cut or abrasion on the skin; (2) gastrointestinally, by 
ingesting undercooked, contaminated meat; and (3) through inhalation, 
by breathing aerosolized, or airborne, spores into the lungs.

The response to the incident in the American Media Incorporated 
building in Florida in September 2001 led to the identification of mail 
as the potential source of contamination; eventually, it led to the 
sampling of the postal facilities. The agencies began sampling on 
October 12, 2001, in Florida and stopped on April 21, 2002, when the 
Wallingford, Connecticut, facility was sampled for the last time. Four 
contractors conducted USPS sampling.

The mission of USPS is to provide affordable, universal mail service. 
As of May 28, 2004, more than 800,000 workers processed more than 200 
billion pieces of mail a year. The USPS headquarters office is in 
Washington, D.C. USPS has nine area offices; approximately 350 P&DCs; 
and about 38,000 post offices, stations, and branches; the P&DCs vary 
widely in size and capacity. The USPS mail system is involved in 
collecting, distributing, and delivering letters, flats (that is, 
catalogs and magazines), and parcels, as well as other items that vary 
in size and capacity.

The federal agencies involved in the response in the postal facilities 
had differing responsibilities. The Centers for Disease Control and 
Prevention (CDC) and state and local health departments primarily 
provided public health advice and assistance to USPS. CDC has had 
primary responsibility for national surveillance of specific diseases, 
including anthrax; it has also conducted epidemiologic investigations 
to determine, among other things, the source of the disease. The FBI 
has been responsible for criminal investigations involving interstate 
commerce and the mail and crimes committed on federal property. The 
Environmental Protection Agency (EPA) has been the nation's lead agency 
for responding to a release of hazardous substances into the 

On October 8, 2001, the President created the Office of Homeland 
Security to develop and coordinate a comprehensive national strategy 
for dealing with domestic terrorist threats or attacks. The office, 
which had limited involvement in the 2001 response, was superseded by 
the Homeland Security Act of 2002, which transferred many of its 
functions to the Department of Homeland Security (DHS); it became 
operational in 2003. DHS was created by combining many previously 
separate agencies and is assigned a lead role in coordinating the 
efforts of federal agencies that respond to acts of terrorism in the 
United States.

In addition, the Laboratory Response Network (LRN) was developed in 
1999 to coordinate clinical diagnostic testing for bioterrorism. The 
primary purpose on the biological side was to detect the presence of 
biothreat agents in a number of specimen and sample types. These 
laboratories function as first responders that can perform standard 
initial tests to rule out, but not definitively confirm, anthrax.

Now I will discuss our assessment of the following federal agencies' 
activities: (1) federal agencies' activities to detect anthrax 
contamination in the postal facilities; (2) the results of the federal 
agencies' testing in the postal facilities; and (3) whether agencies' 
activities were validated and, if not, discuss any issues that arose 
from the lack of validation and any actions they took to address these 

Federal Agencies' Activities to Detect Anthrax Contamination in the 
Postal Facilities:

CDC, EPA, and USPS, the federal agencies involved in sampling the 
postal facilities in 2001 to detect anthrax, undertook several 
activities: (1) sampling strategy development, followed by (2) sample 
collection, (3) transportation, (4) extraction, and (5) analysis of the 
samples (see fig. 1).

Figure 1: Agency Sampling Activities:

[See PDF for image]

[End of figure]

Neither these activities nor the overall process has been validated for 
anthrax testing. Consequently, the agencies had only limited 
information available for reliably choosing one method over another and 
no information on the limits of detection to use when evaluating 
negative results. In addition, the sampling strategy used by the 
agencies could not provide any statistical confidence with regard to 
the basic question: Is this building contaminated? Therefore, in the 
future, in the absence of a positive result, a different strategy is 
needed that will provide statistical confidence, at a defined level, to 
answer this question.

Activity 1: Sampling Strategy Development:

The first activity involved agencies' developing a sampling strategy, 
which included deciding how many samples to collect, where to collect 
them from, and what collection methods to use. The agencies primarily 
used a targeted strategy: They collected samples from specific areas 
considered more likely to be contaminated, based on judgments. Such 
judgments can be effective in some situations, for example, in 
determining (1) the source of contamination in a disease outbreak 
investigation or (2) whether a facility is contaminated when 
information on the source of potential contamination is definitive. 
However, in the case of a negative finding, when the source of 
potential contamination is not definitive, the basic question--Is this 
building contaminated?--will remain unanswered.

Agencies Primarily Used a Targeted Strategy:

The targeted strategy the agencies used was reflected in their site- 
specific sampling activities. Sample sizes varied by facility and 
circumstances, increased over time, and excluded probability sampling. 
In the beginning, in each USPS facility, 23 samples were to be 
collected from specific areas relating to mail processing and up to 20 
additional "discretionary" samples were to be collected, depending on 
the type and size of the facility. Later, USPS increased the number of 
samples required to a minimum of 55, with up to 10 additional 
discretionary samples for larger facilities. Consequently, the number 
of samples collected varied by facility, from a low of 4 to a high of 
148. CDC's and EPA's site-specific strategies were primarily 
discretionary. The number of samples CDC collected varied by facility, 
ranging from a low of 4 to a high of 202. The number of samples EPA 
collected ranged from a low of 4 to a high of 71.

According to CDC, a targeted sampling strategy may be effective in 
detecting contamination in a facility when sufficient site-specific 
information exists to narrow down the locations in which the release 
and contamination are most likely to have occurred. CDC's assumptions 
for this strategy are that at the outset, (1) a scenario where all 
locations have an equal chance of being contaminated is generally the 
exception rather than the rule; (2) information collected about the 
event, combined with technical judgment about exposure pathways, can be 
used to identify locations where contamination is most likely to be 
found; (3) contamination levels of the highest public health concern 
can usually be detected using a variety of available methods, despite 
their limitations; and (4) there is important public health value in 
quickly identifying contaminated locations. However, these assumptions 
may not always apply. For example, there may be limitations in the 
available information that restrict the ability to reliably identify 
target locations. The method of contamination spread could conceivably 
be via a mechanism where there is an equal chance of any area being 
contaminated. Lastly, all results may be negative, which will lead to a 
requirement for additional testing, as was the case in Wallingford. 
This, in turn, will result in the loss of the critical time needed for 
public health intervention.

CDC and USPS officials said that they used a targeted strategy for 
several reasons, including limitations on how many samples could be 
collected and analyzed. They also said that in 2001 they lacked the 
data necessary to develop an initial sampling strategy that 
incorporated probability sampling. We disagree with this 
interpretation. Probability sampling is statistically based and does 
not depend solely on empirical criteria regarding the details of 
possible contamination.

Incorporating Probability Sampling Would Allow Greater Confidence in 
Negative Results:

We consider probability sampling to be a viable approach that would 
address not only the immediate public health needs but also the wider 
public health protection, infrastructure cleanup, and general 
environmental contamination issues. We recognize that in a major 
incident, the number of samples that may need to be collected and 
analyzed may challenge available laboratory resources. Accordingly, 
there is a need to develop innovative approaches to use sampling 
methods that can achieve wide-area coverage with a minimal number of 
individual samples to be analyzed. For example, high-efficiency 
particulate air (HEPA) vacuum techniques, in combination with other 
methods, appear to be one such approach that could achieve this. In 
addition, because of limited laboratory capacity, samples may need to 
be stored after collection for subsequent analysis, on a prioritized 

The situation in 2001 was unique, and the agencies were not fully 
prepared to deal with environmental contamination. In the future, if 
the agencies decide to use a targeted rather than a probability 
sampling strategy, they must recognize that they could lose a number of 
days if their targeted sampling produces negative test results. In this 
case, additional samples would need to be collected and analyzed, 
resulting in critical time, for public health interventions, being 
lost. This was so at the Wallingford postal facility in the fall of 
2001, when about 3 weeks elapsed between the time the first sampling 
took place and the results of the fourth testing, which revealed 
positive results. Furthermore, about 5 months elapsed between the time 
of the first sampling event and the time anthrax was found in the 
Wallingford facility's high-bay area.

Therefore, in the future, strategies that include probability sampling 
need to be developed in order to provide statistical confidence in 
negative results. Further, even if information on all the performance 
characteristics of methods is not yet available, a probability sampling 
strategy could be developed from assumptions about the efficiency of 
some of the methods. And even if precise data are not available, a 
conservative, approximate number could be used for developing a 
sampling strategy. This would enable agencies and the public to have 
greater confidence in negative test results than was associated with 
the sampling strategy used in 2001.

Activity 2: Collecting Samples:

The agencies used a variety of sample collection methods. USPS 
primarily used the dry swab method. CDC and EPA used premoistened and 
dry sterile, synthetic (noncotton) swabs, wet synthetic wipes, and HEPA 
vacuums for sampling. To determine whether anthrax was airborne, CDC 
performed air sampling in the Brentwood facility 12 days after the 
contaminated letters were processed. Airborne anthrax spores pose a 
health risk because they can cause inhalational anthrax, the most 
serious form of the disease. Agency officials stated that laboratory 
requirements had influenced the choice of sample collection methods. 
For example, in the New York area, CDC used only dry swabs, following a 
requirement by New York public health laboratories.

The majority of the samples were collected by the dry swab method, 
which experts and others we interviewed considered the least effective. 
Single methods were involved in 304 sampling events--that is, CDC and 
USPS collecting dry swab samples (185) and CDC and others collecting 
premoistened swabs (119).[Footnote 5] However, for some sampling 
events, CDC used wet wipes, HEPA vacuum, and air samples at Brentwood 
and swabs, wet wipes, and HEPA vacuum samples at Wallingford.

USPS officials said that the choice of dry swabs was based on advice 
from CDC and an APHL working group, which had coordinated with the head 
of LRN. CDC stated that the reason for the use of swabs was an 
accommodation USPS had reached with APHL. According to APHL officials, 
the working group consulted with CDC's NCID in November 2001. APHL said 
that an NCID official, who was a member of the group, agreed that the 
dry synthetic swab method could be used but that premoistened swabs 
would pick up more spores.

During our fieldwork, we tried to determine what specific advice CDC 
gave the Association of Public Health Laboratories (APHL) on using dry 
swabs. In responding to our inquiry, CDC did not specifically deny 
APHL's statement that an official from CDC's National Center for 
Infectious Diseases (NCID) told APHL that dry swabs could be used. 
However, an official from CDC's National Institute for Occupational 
Safety and Health (NIOSH), which was not a member of the working group, 
said that CDC has always recommended using premoistened swabs. 
Nevertheless, according to APHL, "the NIOSH recommendation was not 
known by the NCID working group members, nor did they advocate on its 

The decision to use dry rather than premoistened swabs stemmed partly 
from the concern of some public health officials, including APHL 
officials we interviewed, that moistened swabs would allow anthrax 
spores to germinate, growing into vegetative cells instead of remaining 
as spores. Other public health officials we interviewed said it was 
highly unlikely that anthrax spores would germinate into vegetative 
cells in a premoistened swab. APHL officials said that it was feared 
that such vegetative cells would be destroyed during certain analytic 
procedures. However, none of the agencies' collection methods were 
evaluated for anthrax detection in environmental samples. In the 
absence of empirical research, agencies had no information available 
for reliably choosing one method over another and no information on the 
limits of detection to use when evaluating negative results.[Footnote 6]

Activity 3: Transporting Samples:

Agencies transported samples by land or air to laboratories for 
extraction and analysis (activities 4 and 5). The USPS sample 
collection plan included shipping instructions that were based on 
regulations for shipping infectious substances and designed to prevent 
their inadvertent release. EPA's sample collection plan did not refer 
to transportation requirements. According to CDC's guidelines, anthrax 
samples were to be considered infectious substances and packaged 
according to applicable federal regulations enforced by the Department 
of Transportation. These regulations were aimed at "ensuring that the 
public and the workers in the transportation chain are protected from 
exposure to any agent that might be in the package."[Footnote 7] Among 
other potential requirements, infectious material must be contained in 
a securely sealed, pressure resistant, watertight, primary receptacle 
surrounded by an absorbent and cushioning material. This material must, 
in turn, be enclosed in a securely sealed, watertight, and durable 
secondary packaging, which has to be enclosed in an outer packaging 
constructed of fiberboard or equivalent material, as well as shock 
absorbent material if more than 50 milliliters are shipped in one 

However, these regulations did not address one of the most important 
issues--maintaining the biological integrity of samples while being 
transported. Failure to do so could result in false negative test 
results. For example, analysis by culture requires that spores can 
germinate, divide and multiply, so that tests can determine whether a 
sample contains anthrax. Temperature and exposure to certain kinds of 
light, such as ultraviolet light, can be deleterious to some 
microorganisms. Therefore, it is important that every sample collected 
retain its original physical form before and during transportation.

We did not attempt to ascertain (1) the specific transit times for 
delivering all the samples to laboratories, (2) whether sample 
transportation was delayed, and (3) if it was, how long it was delayed. 
We also did not attempt to ascertain the environmental conditions the 
samples were shipped under or when they were received at the 
laboratories. Finally, we did not attempt to ascertain the degree to 
which spores could have been exposed to varying environmental 
conditions from the time of release to the time of sample collection, 
which could have affected sample integrity. Anthrax spores are robust, 
compared with other pathogenic microorganisms, but whether 
transportation affected their viability cannot be known because the 
conditions of their transportation were not validated. Transport 
conditions, once validated, would have to be standardized to ensure 

Activity 4: Extracting Samples:

LRN protocols required that sample material be extracted with specific 
extraction procedures and fluids (such as sterile saline or water) and 
that the extracted fluid be subjected to specific analytic methods. For 
the samples USPS collected under the APHL agreement, the extraction 
methods included adding a sample processing solution to the conical 
tubes containing the dry swabs before "plating." This process was 
adapted from LRN protocols for extracting swabs. However, the private 
laboratory (not part of LRN) that originally analyzed the samples for 
USPS did not use an extraction fluid; it inoculated the noncotton, 
rayon-tipped dry swab directly onto a culture plate.

Several factors could have affected extraction efficiency. For example, 
according to public health officials and other experts, the degree to 
which swabs or wipes can retain spores depends on the material they are 
made of. Cotton is more retentive than some artificial fibers like 
rayon and may be more difficult for extraction of spores for analysis. 
Other factors affecting spore extraction are the physical nature of the 
collection device and surface properties. For example, swabs are easier 
to manipulate and immerse in extract fluid than more bulky wipes are. 
CDC has acknowledged that "the recovery efficiency of the analytical 
methods has not been adequately evaluated."

The reproducibility of the results when an extraction fluid is used can 
also be an issue. For example, a U. S. Army Medical Research Institute 
for Infectious Diseases (USAMRIID) official we interviewed told us of 
an unpublished USAMRIID study conducted to determine the efficiency of 
extracting anthrax from swabs; the study showed that even if the same 
procedure was followed, the results were not always the same.[Footnote 
8] Although the importance of reproducibility has been recognized, 
definitive scientific information regarding extraction efficiency is 
lacking. In its absence, it is not clear whether sampling results were 
affected, particularly with respect to samples that may have contained 
few spores. Without knowing the extraction efficiency, a false negative 
result may potentially be seen as a true negative.

Activity 5: Analyzing Samples:

Analyzing the samples involved a variety of methods and required two 
steps--preliminary and confirmatory--to generate a final result. The 
laboratory analytic methods that were used for detecting anthrax in 
clinical samples already existed, but they had not been used for 
environmental samples. As a result, different analytic approaches were 
taken at the preliminary step, involving adaptations of such protocols. 
Samples deemed positive at the preliminary step were not always 
confirmed as positive, as was to be expected. However, this could cause 
problems for the agencies. In addition, some agencies considered 
preliminary analyses by field-based instruments unreliable, while 
others maintained that they were reliable but had been used 
inappropriately. However, once sample extracts were subjected to the 
required confirmatory tests, a positive result was indeed a positive.

In analyzing the postal samples, laboratories used a variety of methods 
for preliminary and confirmatory testing. Preliminary tests included 
colony morphology, Gram's stain, hemolysis, and motility 
tests.[Footnote 9] Any culture isolates that could not be ruled out in 
the preliminary step of testing were considered presumptively positive 
and referred for confirmatory testing. Confirmatory tests included 
culture analyses (traditional microbiological and biochemical 
analyses), gamma phage lysis (a test that identifies the susceptibility 
of the organism to anthrax-specific viruses that create a kill zone in 
anthrax cultures), and direct fluorescent antibody assay, or antibody 
analyses employing a two-component test that detects the cell wall and 
capsule, or outer covering, produced by vegetative cells of anthrax.

Other specialized tests, such as molecular subtyping, were also 
conducted to determine what strain of anthrax was involved. The test 
results were reported as positive--anthrax was found--or negative-- 
anthrax was not found. Traditional microbiological analyses require 18 
to 24 hours before a result can be generated, depending on the 
laboratory protocols and procedures. In a few instances, results were 
also reported as number of colony forming units (CFU) per gram of 
sample material.

According to CDC guidelines, LRN laboratories were to analyze samples 
by appropriate LRN protocols. According to CDC, all LRN laboratories 
were qualified to perform the preliminary tests, and most could perform 
confirmatory and other specialized tests. While a lower level of LRN 
laboratory could analyze swab samples for preliminary testing, all 
other samples--such as bulk, wipes, air samples, or vacuum samples-- 
were to be analyzed at a higher level of LRN laboratory. Samples could 
also be analyzed at CDC laboratories. Presumptive positives found at a 
lower level LRN laboratory had to be referred to an appropriately 
qualified laboratory for confirmatory testing.

The problems agencies encountered in preliminary testing included 
issues related to training and quality control, as well as problems 
with using field-based analytic methods with limitations that were not 
well understood. In preliminary testing, a suspect organism must first 
be selected; at this point, human error or quality control issues can 
affect the results. For example, we identified a problem involving 
culture in the preliminary tests--that is, a reliance on the naked 
human eye to identify and select the growth of anthrax on the petri 
dish. Many different types of organisms could be growing that looked 
like, but were not, anthrax. This is significant because when negative 
results were obtained during preliminary testing, no further testing 
was to be done.

The agencies were also faced with problems when deciding how to respond 
to preliminary positive results that might eventually turn out to be 
confirmed otherwise. For example, agencies did not have clear criteria 
for when to close facilities. In addition, although hand-held assays 
(HHA) were considered preliminary tests, concerns were raised that the 
negative results might lead to a false sense of security.[Footnote 10] 
During the 2001 incidents, USPS kept the Brentwood facility open, 
following CDC's advice that closing it was not warranted. According to 
USPS officials, the correctness of this advice appeared to be confirmed 
by the HHA results obtained on October 18, 2001. When CDC confirmed a 
case of inhalation anthrax in a Brentwood employee on October 21, 2001, 
the facility was closed that day. According to USPS, it was not until 
October 22, 2001, that the laboratory's culture tests of the other 
samples, collected on October 18, revealed positive results. In a more 
recent instance, on November 6, 2003, USPS shut down 11 postal 
facilities in and around Washington, D.C., after a preliminary test-- 
not a confirmed result--from a routine air sample taken on November 5 
indicated that a naval mail processing facility might be contaminated 
with anthrax. USPS tracked the flow of mail through its own facilities 
and closed 11 postal facilities that delivered mail to the naval 
facility. The subsequent confirmatory tests were negative, and the 
facilities were reopened about 3 days later.

All the activities discussed above are interdependent, and many 
variables for each one can affect the results. Further, problems 
associated with any one of these activities could affect the validity 
of the results generated by the overall process. Given that there are 
so many variables, the use of different sample collection strategies, 
reflected in site-specific plans, could yield different results. For 
example, three potential sample collection plans could be used in one 
facility--plan A, using one collection method (for example, a swab); 
plan B, using two methods (for example, a swab and wipe); and plan C, 
using three methods (for example, swab, wipe, and HEPA vacuum). How 
these collection methods are to be applied--that is, how they are 
physically used and how much area each sample covers--is a variable. 
Within each plan, sample transportation protocols could differ, 
involving variables such as temperature--plans A and B might require 
transporting at ambient temperature, while plan C might require 
freezing temperature--the sample collection method's moistness during 
transport, and the size and construction of the packaging.

In addition, within each plan, laboratory extraction and analysis 
protocols could differ, involving variables such as (1) different 
manufacturers' different formulations of extraction fluids, (2) 
different ways to physically release spores from a particular 
collection method (such as a swab) into the liquid extract (such as by 
shaking or vortexing), and (3) a combination of analytic methods, such 
as culture or polymerase chain reaction (PCR) for deoxyribonucleic acid 
(DNA) amplification to identify anthrax. Any problems experienced with 
any of these variables across any of these plans could affect the final 

The Sampling Results Were Largely Negative:

The results of the CDC, EPA, and USPS testing in 286 postal facilities 
were largely negative. Of 286 facilities, 23 tested positive. For 2 of 
these 23 facilities, test results were negative at first but positive 
on a subsequent testing. However, in 1 of these facilities--the 
Wallingford, Connecticut, facility--it was not until the fourth testing 
that positive results were obtained.

Testing results differed between the primary facilities and 
Wallingford. First, in the three primary facilities, results were 
positive each time a facility was tested, with the important exception 
of the two quick tests in Brentwood. In Wallingford, considered less 
likely to be contaminated, results were positive only on the fourth 
sampling. Second, in the primary facilities, sampling with a single 
method produced some positive results, regardless of the sample 
collection method. In Wallingford, neither dry nor premoistened swabs 
produced any positive results. Third, in the primary facilities, both 
single and multiple methods produced positive results; in Wallingford, 
only multiple methods produced positive results.

When comparing the positive results, obtained with dry swabs, across 
the primary facilities, the proportions differed. For example, in one 
sampling event in Brentwood, out of 29 samples collected using dry 
swabs, 14 were positive (48 percent), whereas in Morgan, out of 56, 
only 7 were positive (13 percent). In addition, for the West Palm 
Beach, Florida, facility, sampled several times during one sampling 
event, out of 38 dry swab samples collected, only 1 was positive (about 
3 percent). While we did not define this facility as primary, it was 
suspected of processing a contaminated letter, although none was found. 
However, the use of both wet and dry swabs produced positive results in 
this facility.

USPS and CDC sampled facilities that processed mail from the primary 
facilities to determine whether any other facilities had become 
contaminated. The majority of test results from these facilities were 
negative: Of 286 facilities sampled, 23 tested positive, including the 
3 primary facilities, and 263 tested negative.

For some of the positive facilities, excluding the primary ones:

* Generally, only 1 or 2 of the total samples collected for each 
facility were positive, such as several post offices that received mail 
from Brentwood, including Dulles (11 samples collected, 1 positive), 
Friendship Station (32, 1 positive), Pentagon Station (17, 2 positive), 
and Raleigh, North Carolina (42, 1 positive). These facilities were 
considered cross-contaminated.

* West Palm Beach and Wallingford tested positive only on retesting, 
whereas initially they had tested negative. The West Palm Beach 
facility tested positive on the second testing. According to CDC, the 
sampling strategy used in this facility was found to have limitations 
and was not used again. However, Wallingford did not test positive 
until the fourth testing. These results underscore the importance of 
retesting and cast doubt on the efficiency of the testing.

Of the 263 facilities that tested negative, only 9 were sampled more 
than once. A facility in West Trenton tested negative, even though an 
employee had contracted cutaneous anthrax. The facility in West Trenton 
was tested twice by the FBI and once by CDC, during which a total of 57 
samples were collected, with negative results.

Final, or confirmed, results will be negative if contamination is not 
present in a facility. However, a result can be negative for several 
other reasons, such as (1) the sampling method was not efficient 
enough, (2) samples were not collected from places where contamination 
was present, (3) not enough samples were collected, (4) not enough 
spores were recovered from the sample material, or (5) analysis of the 
sample extract was not sensitive enough to detect anthrax spores that 
were present (that is, the result was a false negative).

Agencies' Activities Were Not Validated:

None of the agencies' activities to detect anthrax contamination in the 
postal facilities were validated. Without validation, the sampling 
activities could have been based on false assumptions. Using an 
ineffective method or procedure could result in a finding of no 
contamination when in fact there is contamination--a false negative. 
Because the sampling methods are not validated, it is not known to what 
extent they will underestimate contamination. Thus, in the case of a 
negative result, agencies would have no sound basis for taking public 
health measures for the occupants of the contaminated facility.

Validation, as it is generally understood, is a formal, empirical 
process in which the overall performance characteristics of a given 
method are determined and certified by a validating authority as (1) 
meeting the requirements for the intended application and (2) 
conforming with applicable standards. Because the agencies did not use 
an empirical process to validate their testing methods, the agencies 
had limited information available for reliably choosing one method over 
another and no information on the detection limit to use when 
evaluating negative results.

Validating the overall process is important because operational and 
health-related decisions are made on the basis of testing results 
generated by that process. In addition, validation would offer 
assurance that the results of using a particular method, which is part 
of that process, are robust enough to be reproduced, regardless of 
which agency, contractor, or laboratory is involved. Thus, agencies and 
the public could be reasonably confident that any test results 
generated by a process that includes that method would be reliable and, 
in particular, that any negative results would mean that a sample was 
free from contamination (within the method's limits of detection).

In preparing for future incidents, the agencies have (1) made some 
changes based on what has been learned about some of the limitations of 
their sampling strategies, (2) made some revisions to their guidelines 
to reflect some of this knowledge and experience or developed new ones, 
(3) funded some new research, and (4) planned or conducted conferences 
addressing some of the issues we have identified. In addition, DHS has 
taken on the role of coordinating agencies' activities and has 
undertaken several new initiatives related to dealing with anthrax and 
other biothreat agents.

However, while the actions DHS and other agencies have taken are 
important, they do not address the issue of validating all activities 
related to sampling. Since the fall of 2001, studies have been 
performed, or are under way, that may contribute to the validation of 
the individual activities. Nonetheless, these studies address only some 
aspects of an individual activity rather than the overall process. 
Finally, the agencies have not made appropriate and prioritized 
investments to develop and validate all activities related to anthrax 
and other biothreat agents.


The lack of validated methods for assessing contamination in postal 
facilities impeded the agencies in responding to the incidents. The 
need that all methods, from sampling to final analysis, be validated, 
so that their performance characteristics can be clearly understood, is 
not in doubt. But any combination of methods that makes up the overall 
process should also be validated because the effect of different 
permutations of methods may not be predictable. It must be recognized, 
however, that an inability to validate the entire process reduces, to 
some degree, the level of confidence in the results. To assess the 
impact of relying on the validation of individual activities, 
experiments could be performed with a limited number of processes, 
combining different methods.

The issues we have raised in this report apply to any anthrax incident, 
including the March 2005 incident involving DOD facilities in the 
Washington, D.C. area. In addition, while the 2001 events involved 
anthrax, many other biothreat agents exist. Differences in their 
characteristics mean different solutions. Accordingly, efforts to 
develop sampling strategies and to validate methods should address 
requirements specific to those biothreat agents as well. However, since 
addressing other agents would consume resources and time, these efforts 
should be prioritized in a long-term strategy.

The several agencies that dealt with the anthrax attacks generally 
worked well together, but we have identified areas that would have 
benefited from one agency's taking the lead in coordinating the 
response. Given the mission of DHS and its responsibilities, it appears 
that DHS is now well positioned to take a lead role in promoting and 
coordinating the activities of the various agencies that have technical 
expertise related to environmental testing. In addition, it is 
important that all participating agencies recognize and support DHS in 
that role and that they have an effective structure for participating 
in identifying and addressing the appropriate issues.

Recommendations for Executive Action:

Accordingly, in our report, we recommended that to improve the overall 
process for detecting anthrax and to increase confidence in negative 
test results generated by that process, the Secretary of Homeland 
Security develop a coordinated approach. This approach would include 
working with agencies to ensure that appropriate validation studies of 
the overall process of sampling activities, including the methods, are 
conducted. Specifically, the Secretary should (1) take a lead role in 
promoting and coordinating the activities of the various agencies with 
technical expertise related to environmental testing; (2) ensure that a 
definition of validation is developed and agreed on; (3) guarantee that 
the overall process of sampling activities, including methods, is 
validated so that performance characteristics, including limitations, 
are clearly understood and results can be correctly interpreted; (4) 
see that appropriate investments are made in empirical studies to 
develop probability-based sampling strategies that take into account 
the complexities of indoor environments; (5) ensure that appropriate, 
prioritized investments are made for all biothreat agents; and (6) 
ensure that agency policies, procedures, and guidelines reflect the 
results of such efforts.

We obtained written comments on a draft of this report from CDC, DHS, 
and USPS. We also obtained written comments from APHL on excerpts from 
the draft that pertained to its role in anthrax testing. Although we 
requested comments from DOD and EPA, DOD said it had no comments and 
EPA provided only technical comments.

CDC, DHS, and USPS, as well as APHL, agreed with our conclusion-- 
methods for detecting anthrax contamination in facilities were not 
validated--and with the thrust of our recommendations--calling for a 
coordinated, systematic effort to validate the methods to be used for 
such testing.

In response, DHS stated that while it has the overall responsibility 
for coordination for future biological attacks, EPA has "the primary 
responsibility of establishing the strategies, guidelines, and plans 
for the recovery from a biological attack while HHS has the lead role 
for any related public health response and guidelines." DHS further 
stated that EPA "is developing specific standards, protocols, and 
capabilities to address the risks of contamination following a 
biological weapons attack and developing strategies, guidelines, and 
plans for decontamination of persons, equipment, and facilities." DHS 
pointed out that in the Conference Report on H.R. 4818, the conferees 
expressed their expectation that EPA will enter into a comprehensive 
MOU [memorandum of understanding] with DHS no later than August 1, 2005 
that will define the relationship and responsibilities of these 
entities with regard to the protection and security of our Nation. The 
Conferees expect the MOU to specifically identify areas of 
responsibilities and the potential costs (including which entity pays, 
in whole or part) for fully meeting such responsibilities. EPA shall 
[is to] submit to the House and Senate Committees on Appropriations a 
plan no later than September 15, 2005 that details how the agency will 
meet its responsibilities under the MOU, including a staffing plan and 

Finally, DHS stated, "Even though DHS is in charge during a biological 
attack, EPA is primarily responsible for the coordination of the 
recovery process. So, DHS will coordinate with EPA to ensure 
appropriate investments are made to explore improved sampling." With 
respect to our recommendation that DHS develop probability-based 
sampling strategies, DHS said that it must first define the necessary 
requirements for the sampling process and then evaluate targeted and 
probability-based sampling strategies against those requirements. DHS 
said that targeted sampling may be beneficial for some applications. We 
agree with DHS on the need to define the requirements for the sampling 
process and to evaluate sampling approaches against those requirements. 
On the basis of the work we have done on this review, we believe that 
(1) DHS will find that targeted sampling will not always meet all the 
requirements to answer the question of whether a facility is 
contaminated and (2) probability-based sampling will be necessary when 
information on the source and path of potential contamination is not 
definitive. In our view, probability sampling will be necessary in 
order for DHS to achieve its goal of having a "scientifically 
defensible sampling strategy and plan."

Mr. Chariman, this concludes my prepared statement. I will be happy to 
answer any questions you or Members of the Subcommittee may have.

Contacts and Acknowledgments:

If you or your staff have any questions about this report or would like 
additional information, please contact me at (202) 512-6412, or Sushil 
Sharma, PhD., DrPH, at (202) 512-3460. We can also be reached by e-mail 
at and

Other staff that contributed to this report include Hazel Bailey, 
Heather Balent, Venkareddy Chennareddy, Crystal Jones, Jack Melling, 
Penny Pickett, Laurel Rabin, Mark Ramage, and Bernard Ungar.

[End of section]

Appendix I: Scope and Methodology:

To respond to your request, we interviewed officials from federal 
agencies involved in sampling the postal facilities. The federal 
agencies included the Centers for Disease Control and Prevention (CDC), 
the Environmental Protection Agency (EPA). We also interviewed U.S. 
Postal Service (USPS), Association of Public Health Laboratories 
(APHL), public health and private sector laboratories, and experts on 
microbial detection in indoor environments.

We reviewed documentation provided or developed by CDC, EPA, and USPS, 
including sample collection strategies, guidance, environmental 
collection and analytical methods and protocols. In addition, we 
reviewed and analyzed test results data, that is, sample collection and 
analytical data collected by federal agencies, their contractors, and 
public health laboratories. We did not independently verify these data.

We conducted site visits to some postal facilities affected by anthrax 
and some public health and private sector laboratories that were 
involved in analyzing samples. We also reviewed studies on sampling 
methods for detecting biological substances, including anthrax, on 
surfaces and in the air. We conducted our review from May 2003 through 
November 2004 in accordance with generally accepted government auditing 

Although our study focused on anthrax testing relating to 2001 anthrax 
incident, we believe that the issue we identified concerning the need 
for validated methods and sound sampling strategies would apply to 
similar incidents in future. This is particularly evident given the 
consequences arising from the March 2005 incident involving facility 
closures following preliminary anthrax testing in the Washington, D.C. 


[1] GAO, Anthrax Detection: Agencies Need to Validate Sampling 
Activities in Order to Increase Confidence in Negative Results, GAO-05-
251 (Washington, D.C.: March 31, 2005).

[2] "Anthrax" in this testimony reflects commonly used terminology. 
Technically, the term refers only to the disease caused by the 
microorganism Bacillus anthracis, not the bacterium itself or its 

[3] Anthrax contamination had been found earlier in several Florida 
postal facilities that processed mail for the American Media 
Incorporated building there. However, no letter containing anthrax was 
ever found.

[4] According to the head of the Postal Inspection Service, more than 
7,000 hoaxes, threats, and suspicious letters and packages--an average 
of almost 600 a day--were reported to his agency in the weeks following 
the first anthrax incident. As a result, nearly 300 postal facilities 
had to be evacuated.

[5] We use "sampling event" to refer to initial sample collection by a 
specific agency on a specific day and at a specific time in a specific 
facility. Multiple agencies collected samples on the same day in some 
of the same facilities; therefore, each agency's sample collection is 
considered a separate sampling event. As a result, there were more 
sampling events than the total number of facilities sampled.

[6] The published literature provided some information on the 
efficiency of a few sample collection methods. In all the methods 
studied, swabs were always premoistened before samples were collected. 
However, according to one study, the most efficient method caused 
problems when used with certain analytic methods.

[7] Department of Transportation, 49 C.F.R. subchapter C--Hazardous 
Materials Regulation. The USPS regulations mirror the Department of 
Transportation regulations. However, to be transported as mail, 
material must be classified as mailable. By statute, infectious 
materials, such as anthrax spores, that are "disease germs or scabs, 
[or] other natural or artificial articles, compositions, or material 
which may kill or injure another" cannot be mailed. Such materials are 
termed "nonmailable matter." Knowingly mailing such material is a 
criminal offense, and doing so with the intent to kill or injure is a 
felony. When an etiologic material is not "outwardly or of [its] own 
force dangerous or injurious to life, health, or property," USPS may 
allow it to be mailed, subject to appropriate rules and regulations 
governing its preparation and packing. As a result, USPS allows the 
mailing of small quantities of appropriately packaged infectious 
material, but only if it is intended for medical or veterinary use, 
research, or laboratory certification related to public health.

[8] Using synthetic swabs and a particular type of buffer could lead to 
70 to 75 percent extraction. However, repeating the test with the same 
type of buffer made by different companies yielded different results. 
The official said that this test showed that there were too many 
variables. Even when analysts followed the same procedure, the results 
were not always reproducible, casting doubt on the reliability of the 
test results.

[9] When bacteria stained with Gram's stain retained the color of the 
primary stain (crystal violet), they were considered gram-positive, a 
characteristic of anthrax. Hemolysis, a procedure involving culturing, 
identified whether the colonies gave no evidence of red blood cell 
lysis, a characteristic of anthrax. Motility refers to whether the 
colonies showed no movement in microscopic observation, another 
characteristic of anthrax.

[10] See GAO, U.S. Postal Service: Better Guidance Is Needed to Ensure 
an Appropriate Response to Anthrax Contamination, GAO-04-239 
(Washington D.C.: Sept. 9, 2004).