Hazard Identification and Assessment
Optimal decision making about decontamination procedures requires an accurate assessment of the problem. One of the first steps is proper identification of the agent or agents used in an attack. Initially, that can be done either by measurements at the putative site of release or by retrospective analysis that identifies the source using information gleaned from records of a medical case or cluster of cases. Additional information, such as the nature of the preparation and the extent to which contamination has spread, also helps with assessment. A critical parameter in any remediation effort is accurate characterization of the amount of contaminant present at the start of the decontamination.
At the Hart Senate Office Building in Washington, D.C., the source of the contamination was evident and localized: A white powder had fallen out of an envelope, and the amount was relatively easy to characterize. In contrast, the initial source of the Bacillus anthracis at the American Media, Inc. (AMI) building in Boca Raton, Florida, has never been identified, and those who carried out the decontamination had to sample the building extensively to identify areas of contamination. The B. anthracis found at the various postal facilities resulted from cross-contamination and was widely dispersed, not localized to a specific area.
The absence of a clearly identifiable source of contamination makes the process of containment and cleanup more complicated because the area to be surveyed must be more extensive. Identification of an actual amount of contaminant provides valuable information for the selection of a method of remediation. With the knowledge that one gram (g) of dried B. anthracis spores can contain up to 1012 spores, a facility contaminated with 10 g requires substantially more than
a six-log kill. (Six-log kill is also known as 1 × 106 kill rate, which means reducing the number of live organisms by 6 orders of magnitude). Bulk material can be physically removed by cleanup methods that will leave a residuum that could be destroyed with a six-log kill. If one can not precisely identify the amount of contaminating material present at the start of a cleanup, then defining a specific level of remediation in terms of a log kill rate becomes difficult.
IDENTIFICATION OF THE AGENT
The biological agent used in an attack might become known as a result of a perpetrator’s announcement, or it could be identified from physical recognition by trained personnel, from early presumptive test kit results, or from human symptoms. A rapid overt (announced) release will give rise to identification by physical and microbial analysis of substances obtained from obviously exposed surfaces. Health monitoring of exposed people is not likely to be necessary for identification. In the case of a covert release, environmental monitors, such as those that have been deployed in major cities as part of the BioWatch program or health monitoring of exposed people, might offer the first clues. The more likely scenario for detection of a covert release—based on past experience—would be the alarm raised by health professionals who would see an unusual disease such as anthrax or smallpox, or who might see several patients who are seriously and inexplicably ill.
Surveillance for increased incidence of common symptoms in targeted patient populations, known as syndromic surveillance, is one way to identify unusual clusters of disease that could result from an act of bioterrorism. Syndromic surveillance monitors the frequency of symptom complexes identified in patients before the confirmation of a medical diagnosis. The surveillance systems complement routine public health surveillance, and they commonly provide the advantage of near-real-time data entry, analysis, and reporting. The objective is to identify an attack as quickly as possible to allow for a rapid response and effective public health intervention. The alerts or warnings provided by the systems can initiate an epidemiological investigation to determine the source and extent of the exposure in the shortest possible time. Syndromic surveillance can be used to detect increases in influenza-like illness during periods of peak influenza A and B activity and of diarrhea and vomiting during periods of suspected norovirus and rotavirus transmission (Hefferman et al., 2004), but its ability to detect a bioterrorist attack has not yet been evaluated.
Substantial information can be obtained from microbial analysis of samples of serum, pus, scabs, and stools, as well as from environmental air and surface samples. A delay in identifying a decontaminating agent would afford the possibility of sustained agent viability and growth in mechanical spaces, crevices, and so on. Many types of sampling can be done, and different approaches are appropriate in different situations. The issues of cross contamination also must be
considered, including background suppression, cell desiccation, substance stabilization, viability after impaction, and other factors that can be affected by the choice of sampling approach. Sampling is discussed in detail in Chapter 8, which outlines presumptive identification made possible with the kits used by first responders and with other early microbial analysis methods, confirmatory identification methods, and the Laboratory Response Network.
The earlier contamination is detected the easier it will be to confine the contamination and limit the number of people exposed. Environmental monitoring and syndromic surveillance systems should be evaluated for the ability to provide information that can be used to detect and limit the spread of biothreat agents in a cost-effective manner.
Using Epidemiology to Identify the Agent
Efforts to identify a biological agent following an act of bioterrorism can be both difficult and time consuming. If there are no witnesses and no group claims responsibility for a deliberate release, nobody other than the perpetrator might be aware that an event has occurred, particularly if there are no real-time environmental monitoring systems at the site. In some instances, identification could only occur as a result of epidemiological monitoring or through medical diagnosis, as was the case at the AMI building in 2001. In such situations, it is difficult to determine whether the symptoms are the result of a natural outbreak or an intentional attack. That problem could be compounded by a lack of timely communication between epidemiologists and forensics experts.
Epidemiological Investigation Leading to Source Identification
A major bioterrorist attack could be unannounced or covert, and the source of the release might need to be identified through an extensive epidemiological investigation. Finding a confirmed case of the suspected disease is critical to many of those investigations. The definition may be clinical, with laboratory confirmation, or it could be done on the basis of laboratory evidence confirmed by one or two supportive laboratory tests. Suspected cases or clinically compatible cases linked to a confirmed environmental exposure, but without corroborative laboratory evidence of exposure or infection, may also be defined in an epidemiological investigation. Laboratory criteria for diagnosis must be defined as well. Follow-up includes enhanced case finding; retrospective and prospective surveillance systems; and environmental assessments and sampling of patients’ homes, work sites, and travel destinations over the period preceding symptom onset and consistent with the incubation period of the suspected disease. Investigations can take weeks, during which time the released agent could be widely disseminated, in the case of spores, or transmitted, in the case of communicable diseases.
Epidemiological Factors Affecting Decontamination Efforts
Exposure reconstruction and risk characterization are important to epidemiological investigation and to decontamination. In the case of the letters tainted with B. anthracis spores, it was important to understand that exposure can be associated with the passage of powder-containing letters through the mail and to validate the model using empirical outcome data (CDC, 2001). Other research priorities include analysis of reaerosolization of settled spores and identification of risk for disease among secondarily exposed individuals; follow-up surveillance in those potentially exposed; the effects of long-term, low-level exposures; quantification of background contamination by potential agents in urban and rural environments; and identification of the occurrence of sporadic cases of zoonotic diseases that are considered possible threats. It also is necessary to decide how much sampling and decontamination will be done at satellite locations to which agents could have been transported.
EVALUATING THE STATE OF THE AGENT
Specific knowledge about the harmful biological agent used in an attack is important for emergency response, and it is essential for proper cleanup. Unlike the spores of B. anthracis, Yersinia pestis cells are sensitive to extremes in environmental conditions and therefore should not pose the same long-term hazard to the general population after a release (Inglesby et al., 2000). Naturally occurring Y. pestis is unlikely to remain viable for more than a few days after release, so its detection and identification can be troublesome. Recovery of viable organisms is unlikely unless samples are obtained and tested immediately after a release. Culturing the organism takes several days so PCR identification would be most timely, despite the fact that PCR cannot answer questions about viability. Like Y. pestis, variola major is sensitive to environmental conditions and in its natural form would not persist in droplets for long outside a human host.
For the case of biological agents, there also is the possibility of weaponization—engineering of the organism to improve its stability or other properties. In general, the weaponization begins with the growth of the agent (lag, log, and stationary phases each have unique properties mixed in with the culture media), then fermentation; centrifuging and separation; drying; milling for respirable particle size; additives to prevent aggregation and clumping, neutralize electrical charge, and increase survival in air; and microencapsulation for stability and viability. Each phase leaves physical and chemical clues that can help investigators to distinguish the agent substance from a normal background presence. Expertly prepared weapons are likely to be more resistant to natural attenuation and may be more resistant to decontamination.
Characteristics of Biological Agents That May Affect Hazard Assessment
The type of processing done before an agent is used as a weapon can alter how hazardous it is to humans and its persistence in the environment. This processing might be termed weaponization if it increases the ability of the agent to cause harm by making the agent more stable, more infectious, or better able to penetrate the human body. For agents that cause harm via inhalation, the size of the particles is crucial. Particle size also affects the ability of the agent to be aerosolized or reaerosolized.
Knowing the particle size of the pathogenic agent is critical in determining its potential for dispersal, reaerosolization, and infectivity—especially if the agent is released and spread as an aerosol. The particle size distribution depends on the agent (e.g., spore, vegetative cell, viron), the degree of weaponization sophistication (e.g., electrically neutralized, finely milled, encapsulated), and aerosol transport mechanism (e.g., dry cells, wet aerosol). A crudely weaponized agent is likely to have a large particle size distribution that varies from single particles of 0.2-2 µm to clumps of many particles or liquid droplets as large as 30 µm. Variola major virons can have complex shapes from 0.2-0.4 µm, Y. pestis cells are rod shaped and range from 0.5 × 1 µm to 1 × 2 µm, B. anthacis vegetative cells are rod shaped from 0.25 × 1 µm, and B. anthracis spores are spherical and 1-1.5 µm. There are many routes for hazardous insult by the threat agent ranging from contact with eyes or broken skin to inhalation into the respiratory tract. Infection is promoted by the growth of threat agent cells in local macrophages or by the proliferation of cells into the bloodstream. Most morbid infections stem from inhalation of aerosols though the nose or mouth. Large particle clumps or droplets (10-20 µm) can lodge in the mucosa of the nasal cavity or the pharynx, causing infection by local macrophages or gastrointestinal infection by ingestion. Particle clumps or droplets in the range 5-15 µm can lodge in the trachea. The most dangerous infections are caused by 0.1-10 µm particles lodged in the lungs, where they may be retained in the upper bronchiole region (5-10 µm particles) or in the lower alveolar region (0.1-5 µm particles).
The dynamics of particle size retention depend on the flow rate, mass impaction, diffusion, and gravitational settling which are, in turn, related to the activity of the person, tidal volume, and oral versus nasal inhalation. Several modeling efforts have helped to explain those dynamics. Calculations by Yu and Diu (1983) for spherical uncharged particles in the lung showed good agreement with experimental data. Yeh and Schum (1980) performed detailed in vitro measurements on lung molds created from human cadavers to validate deposition equations, again with spheres. Harvey and Hamby (2002) presented a model for deposition differences by age and sex. Generally the experiments show a retention rate of about 20-30% for 0.1-0.2 µm diameter spheres, which drops to about 10% for spheres in the 0.3-0.5 µm range and then rises to 90% or more at diameters 6 µm and greater. All of the models and data clearly reflect the partial clearance (exhala-
tion) of particles in the 0.3-3 µm range and the marked retention of larger diameter particles.
Retention of the actual organisms depends on other factors, such as the particle shape, the particle charge, and the hydrophilic or hydrophobic character. Goodlow and Leonard (1961) determined that the LD50 for Francisella tularensis aerosol in guinea pigs increased by nearly 4 orders of magnitude when the particle size of the organism was increased from 1 µm to 12 µm. Fothergill (1957) had published concordant work on the effects of particle size on the LD50 of 6 aerosolized pathogens in guinea pigs. More research is needed to explain the particle retention dynamics in the lung and the infectivity of real organisms in healthy people and also immunocompromised subjects.
One approach to characterizing an aerosol biological agent is called the agent-containing-particles per liter of air (ACPLA) method. That technology combines sample collection with a slit sampler, dichotomous sampler, or all-glass impingers with statistical analysis to determine numbers of a viable agent, such as B. anthracis spores, in a single particle. Studies that use liquid suspensions of B. subtilis spores have shown that not all spores in a suspension will be viable (Ho et al., 2001). It can be assumed that the same holds true for an aerosol of biological material. To test this hypothesis, ACPLA has been used in field trials for testing biological aerosol detectors. Particles in a bioaerosol may be of varying sizes and may contain a mixture of viable and nonviable particles. ACPLA determinations are important because a fundamental characteristic of a biological aerosol threat is that agent particles are linked to infectivity. Research that used B. globigii spores has shown ACPLA values of about 4.5 viable spores in a typical particle of 2.5-4 µm (Ho et al., 2001).
B. anthracis spores are about 1-1.5 µm—an appropriate size for deposition in the alveoli of the lung. The spores germinate in the macrophage to produce the anthrax toxin and capsule, which in turn initiate the cascade of events that leads to disease. Studies of anthrax outbreaks in employees in New England wool mills (Brachman et al., 1960) found that workers may have inhaled 600-2150 spore particles daily without becoming ill. Some 150-700 of the spore particles were less than 5 µm in diameter (Brachman et al., 1960). Dahlgren and colleagues (1960) reported that, even in the dirtiest parts of a goat hair processing plant, employees inhaled 600-1300 spores during the work day and that only 25% to 50% of those particles were smaller than 5 µm in diameter. Although daily exposure may have served as a mechanism through which the workers became immune, the spores also could have aggregated to form particles that were too large to reach deep into the alveoli of the lungs and thus never encountered macrophages. The data substantiate earlier reports of the correlation between larger particle size and increased LD50.
Secondary aerosolization of biological agents is a subject of great debate. The agent’s characteristics—its physical state (e.g., vegetative spore), particle size, shape, electrical charge, and hydrophobicity—are important. The agent
might also be transported with and by other kinds of cells (culture media, skin particles), in environmental dust and aerosols, and in weaponization platforms (silica, beads). Several recent investigations have used computational fluid dynamics models and calculations to predict the major effects of normal air circulation, wall turbulence, and particle diffusion (Fennelly et al., 2004; Scorpio et al., 2003). Those results, using spherical particles with aerodynamic particle diameters from 0.5-10 µm as a proxy for anthrax spores, predict exposures of as much as one LD50 per breath from reaerosolization of 1 gram of material lying dormant on a desk in a normal office. The reaerosolized lethal dose exposure has been predicted to present itself in as little as 10 minutes. In another investigation, data from air samples, dust, and swab samples from a contaminated U.S. Senate office building were used to estimate the reaerosolization of anthrax. Colony-forming units were measured for semi-quiescent and active periods, and the conclusion was that secondary aerosolization was probable and problematic. Recent laboratory experiments by the Defence Research Establishment, Suffield, Canada (Defence R&D Canada) using the simulant B. globigii (B. atrophaeus) measured the rate of accumulation of spores onto slit sample agar devices in several office settings.
EVALUATING THE STATE OF THE CONTAMINATED BUILDING
To completely assess risk, the amount of material initially used in an attack (the source term) must be identified to the extent possible. Initial loading and sample volume affect the types of sampling that can be done and the extent of cleanup required. The building’s internal environment and its structure also are crucial. Humidity, air circulation, air exchange rate, mechanical complexity (HVAC system), architecture (stack effects), functional space (walls, floors, office material), and electrical complexity (lights, computers) all affect the rate of dissemination, viability, and lesion and reaerosolization. Those topics are discussed in Chapter 7.
FINDINGS AND RECOMMENDATIONS
Detailed characterization (including screening for known threat agents, genetically modified and emerging threat organisms) of a suspected biological pathogen is required for proper analysis and to inform decision making.
Research should be conducted to develop a characterization system that can inexpensively identify, or approximately characterize, all potential threat agents including genetically modified and emerging threat agents.
Identifying and characterizing the properties of an organism (or organisms), and
the amount and extent of its concentration at the time cleanup begins, are critical to making decisions about response options.
Characterizing the contaminating agent or agents should be done before selecting the approach for large-scale remediation. The remediation approach chosen should be one that can adequately destroy (or remove) the amount of agent present at the start of the procedure.
The earlier contamination is detected the easier it will be to restrict the area of contamination and the number of individuals who will be exposed. In the case of the 2001 anthrax letter mailings, the event first came to light through the observations of an astute physician. Different monitoring systems—environmental (e.g., Biowatch) and medical (e.g., syndromic surveillance) in nature—have since been put in place with the hope of obtaining the earliest possible indicator regarding the release of a biological agent.
Existing environmental monitoring systems and syndromic surveillance systems need to be evaluated for their abilities to provide information that can be used to detect and to limit the spread of biothreat agents in a cost effective manner. If those systems prove to be effective, they could be deployed in public facilities that may be likely targets for attacks.
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