Ebola Virus Term Paper

Ebola virus 1

Ebola virus: Risk assessment and countermeasure options Katy Lunger BSBD 640 Term Paper Draft 1 April 5, 2009

Ebola virus 2 Table of Contents

Abstract……………………………………………………………………………………………3 Introduction………………………………………………………………………………………..4 Background………………………………………………………………………………………..5 Molecular Mechanisms for Pathogenesis…………………………………………………………6 History of Weaponization…………………………………………………………………………9 Weaponization Potential…………………………………………………………………………..9 Security and Defense Options……………………………………………………………………10 Detection and Diagnosis…………………………………………………………………………13 Medical Response to an Attack with Ebola Virus……………………………………………….15 Countermeasures…………………………………………………………………………………15 Risk Assessment…………………………………………………………………………………18 Conclusion……………………………………………………………………………………….21 References……………………………………………………………………………………….24

Ebola virus 3 Abstract Ebola virus gained widespread public attention after gruesome reports began to surface detailing outbreaks in Africa in the 1990s. Ebola virus, a member of the Filoviridae family, has a high mortality rate (50-90%), and can result in hemorrhaging and necrosis of various biological tissues. There are unconfirmed reports that Ebola virus has been weaponized, despite the lack of knowledge of the virus’s natural reservoir or a clear understanding of how the virus is spread. Several countermeasures are being studied for protection from Ebola virus infection, and though none are yet approved for human use, there are a few that are showing promise in nonhuman primates. In light of the “age of terror,” new interest in the potentially devastating effects of biological and chemical weapons has directed governmental resources toward protection from these types of attacks.

Ebola virus 4 Introduction The events surrounding September 11, 2001 and the subsequent anthrax mailings have spawned enormous efforts in the United States with regard to national defense from various threats, including those of biological weapons. Reports from various sources have also shed light on the reality of the looming threat of biological weapons. According to Alibek and Handelman (2000), the former Soviet Union conducted massive biological weapons research and production programs beginning in the 1970s until it dissolved in 1991. Dr. Kenneth Alibek, a former director of the covert program who defected to the United States in 1992, believes Russia may still be involved in illegal production and storage of biological weapons, while the vast quantities of previous weapons stocks have likely been sold to rogue nations and other private consumers (Alibek & Handelman, 2000). Various cults, terrorist organizations, and enemies of the United States, such as Aum Shinrikyo, Al Quaeda, and Iraq, are believed to have access to or research programs for biological weapons (Oehler, 1996). Much of the knowledge the United States has acquired regarding biological weapons has come from scientists who defected from the former Soviet Union (Alibek & Handelman, 2000; Fong & Alibek, 2005; PBS, 2001). According to a PBS (2001) report with Sergei Popov, a former top soviet scientist and defector, the Former Soviet Union researched weaponization for both Ebola virus and Marburg virus, among other pathogens, and his claims have been corroborated by Ken Alibek (2000). Ebola virus, especially if in a weaponized aerosol form, could be disseminated in an attack, producing high mortality rates with very low dosage, mass panic, and economic disruption. There is no treatment or immunization for Ebola virus, it is highly contagious, has a high mortality rate, and it has been classified as a Category A agent by the Centers for Disease Control and Prevention (CDC, n.d.).

Ebola virus 5

Background The first descriptions of the effects of viral hemorrhagic fevers (VHF) were reported in 1967 when 31 people in Germany and Yugoslavia became infected with Ebola’s close relative, Marburg virus. The infections were believed to have occurred when individuals came into direct contact with blood, organs or other tissues of African green monkeys that had been caught in Uganda (Simpson, 1978). After the Marburg incident, the virus seemed to disappear, until 1975 when a traveler, likely exposed in Zimbabwe, presented with Marburg in Johannesburg, South Africa (CDC, 2004). According to the Centers for Disease Control and Prevention (2004), Marburg hemorrhagic fever has appeared sporadically since then in Africa. The Ebola and Marburg viruses are the sole members of the Filoviridae family, which appears to be endemic to Central Africa in an area between the 10th parallel north and the 10th parallel south of the equator (Feldmann, Wahl-Jensen, Jones & Stroher, 2004). Ebola, like Marburg, is a highly lethal pathogen that causes hemorrhagic fever syndrome in humans and nonhuman primates (Sullivan, Yang, & Nabel, 2003). Ebola was first identified in Zaire in 1976, near the Ebola River during an outbreak. There have been a few sporadic, confirmed outbreaks in Africa since then, including large epidemics that occurred in Kikwit, Zaire in 1995 and in Gulu, Uganda in 2000 (CDC, 2004). Research has yet to determine the natural hosts of filoviruses. Because both nonhuman primates and humans can become infected with filoviruses, they are regarded as zoonotic, yet a species that can act as a life-sustaining host and vector for Ebola is elusive. Some current research has suggested that strains collected during outbreaks have indicated evolution of those strains in an as-yet unconfirmed reservoir. Zaire Ebola virus (ZEBOV) RNA has been reported

Ebola virus 6 in organ tissues of rodents from Central Africa, however, this possible reservoir has not yet been confirmed due to difficulty isolating the virus or because of difficulties detecting the virus with antigens (Feldmann et al., 2004). Because of the nature of the pathology and lack of treatment of Ebola virus, it is only approved for study in a Biosafety Level 4 (BSL) 4 facility; currently, there are only 5 BSL 4 labs in the United States, which is one possible limitation that may hamper the progress of Ebola virus research (McCarter, 2008). In the lab, researchers have inoculated various species with Ebola virus and found that fruit and insectivorous bats can essentially act as incubators without showing deleterious health effects (Feldmann et al., 2004), however, bats haven’t been shown to be actual reservoirs in the field. Feldmann and colleagues (2004) suggest that the difficulty in locating Ebola virus reservoirs indicates that reservoirs are either rare species or that transmission within the reservoir species is not efficient. Molecular Mechanisms for Pathogenesis The Ebola virion is generally tubular, and as is common with Filoviridae, it may appear in many different shapes, including long and branched filaments or short “U” or “6” shaped configurations (CDC, 2004). The virion is made up of a nucleocapsid, a viral envelope, and an intracellular matrix between the nucleocapsid and the viral envelope; the viral envelope is derived from host-cell lipid membranes (Puskoor & Zubay, 2000). There are four known strains of Ebola, which include the Zaire (ZEBOV), Ivory Coast (EBO-CI), Sudan (EBO-S), and Reston (EBOR) strains. All but the Reston strain can cause disease in humans, and all are known to cause disease in nonhuman primates (Sullivan et al., 2003). The Ebola virus is a negative-sense RNA virus (Sullivan et al., 2003) with a 19 kb long

Ebola virus 7 genome encoding eight proteins (Puskoor & Zubay, 2000). Sanchez and colleagues (1998) reported a single coding region responsible for two protein products, secreted glycoprotein (sGP), and transmembrane virion envelope glycoprotein (GP). GP is an edited version of sGP, which forms a trimeric complex and binds to endothelial cells (Sanchez et al., 1998), and is thought to play a key role in Ebola virus’ pathogenesis (Sullivan et al., 2003). Research has indicated that GP transmembrane proteins bind to monocytes and/or macrophages, thereby allowing the Ebola virus to spill its contents into the cell (Sullivan et al., 2003). sGP may be involved in silencing the immune response of the cells while GP, which dot the surface of the Ebola virus membrane envelope, may be responsible for the hemorrhagic fever symptoms because it binds to reticuloendothelial cells that line blood vessels (Puskoor & Zubay, 2000; Sullivan et al., 2003). In addition to GP and sGP, the Ebola virus genome encodes the following proteins: nucleoprotein (NP), which is responsible for packing the RNA genome; viral protein 35 (VP35), which is a cofactor in transcription and replication; viral protein 40 (VP40), which is associated with the membrane, viral assembly and viral budding; viral protein 30 (VP30), which appears to be involved in transcription and the addition of poly-A tails to mRNA; viral protein 24 (VP24), which is not homologous to any other known viral proteins; and an RNA polymerase; the numbers indicate approximate molecular weight of the proteins in kilodaltons (Puskoor & Zubay, 2000). Ebola virus infection usually occurs after contact with bodily fluids of infected individuals or accidental needle stick injections, though airborne transmission of viral particles is a suspected mode of communicability (Groseth, Jones, Artsob, & Feldmann, 2005). There has also been a case in which a recovered patient transmitted Marburg virus via semen three months

Ebola virus 8 after infection (Leffel & Reed, 2004). Contact between contaminated fingers and the mouth or eyes may also lead to infection as conjunctivitis has been correlated with Ebola infection (Puskoor & Zubay, 2000). Aerosolization of Ebola, as would be probable in a weaponized attack, could infect individuals through the inhalational pathway. There is some evidence indicating natural spread of Ebola virus via aerosol particles as well (Groseth et al., 2005; Puskoor & Zubay, 2000). After infection, the virus enters an incubation period lasting 2 to 21 days. During this phase, the virus infects endothelial cells, hepatocytes, monocytes and macrophages, is transported to the lymph nodes, and begins to replicate at a high rate (Sullivan et al., 2003). The infected cells leave the lymph nodes to travel throughout the body via the bloodstream. Ebola’s high rate of viral replication is thought to overwhelm the infected cell and impair its ability to synthesize proteins to ward off infection (Sullivan et al., 2003). While GP downregulates proteins associated with cell adhesion, thereby weakening the cellular network, the cell wall breaks down and releases the replicated Ebola viruses. Cytokines that were expressed in response to the invasion of the cell are also released and go on to interact with other cells in a “cytokine storm,” with the result of an induced state of vascular shock on the cells (Sullivan et al., 2003). The resulting pathology consists of necrosis of various tissues, including liver, lymph nodes, kidneys, testes, and ovaries due to the replication of the virus (Puskoor & Zubay, 2000). The individual displays an abrupt onset of fever after the initial incubation period with a variety of other symptoms such as skin rash, chills, muscle pain, and/or nausea (Groseth et al., 2005). Abnormal blood coagulation usually occurs which can lead to bleeding from orifices and bruising; necrosis of several organs may also occur (Groseth et al., 2005). External hemorrhaging is usually a rare result of infection, however, internal bleeding is very common

Ebola virus 9 (Groseth et al., 2005). Currently, there are no treatment options for Ebola virus; one only has hope that their immune system will mount an adequate defense. The mortality rate associated with Ebola virus infection is between 50-90% (CDC, 2004). History of Weaponization According to a statement made by Dr. Kenneth Alibek before the US Congress Joint Economic Committee (1998), the Former Soviet Union extensively studied the weaponization of several viral hemorrhagic fevers, including Ebola virus, both before and after signing the Biological and Toxin Weapons Convention of 1972. Alibek has divulged that the Russian government developed techniques and equipment for large-scale cultivation and concentration of many pathogens (Alibek, 1998). He also claims that extensive research was done on the smallpox vaccine virus, vaccinia, and that many different experiments were conducted in which genes from different viruses were inserted into vaccinia’s genome, including genes from Ebola virus (Alibek, 1998). Aum Shinrikyo has reportedly attempted to obtain Ebola virus during a natural outbreak in Zaire, however, this report has never been confirmed and there is no evidence of the sect actually using it (Olson, 1999). Thus far, only faint rumors of weaponized Ebola virus exist; however, it is highly possible that the virus may be weaponized in the future if it hasn’t already. Weaponization Potential Ebola virus has a high potential for weaponization because it is stable at neutral pH, and can survive for long periods in blood and other body fluids; in proper laboratory media, the virus could be cultivated (Leffel & Reed, 2004). Leffel and Reed (2004) report that a review of research on filoviruses shows that they are stable in aerosols and can retain virulence after lyophilization for long periods of time, however, it should be noted that the referenced studies

Ebola virus 10 were done with Marburg virus, and not with Ebola virus. Experiments have shown aerosolized Ebola to be lethal in rhesus monkeys (Johnson, Jaax, White, & Jahrling, 1995). The fact that virtually no cure, antidote, or vaccine exists for Ebola infection makes its use as a weapon highly viable. Further, public panic and social disruption would likely be high in the even of an Ebola virus attack. The lack of scientific understanding of reservoirs for Ebola and other viral hemorrhagic fevers could further enhance public terror. Aerosolization of Ebola virus could inflict a massive death toll if dispersed over a large population of individuals. With a mortality rate between 50-90%, Ebola would pack quite a punch as a biological weapon. Moreover, it is known to be highly contagious and the longer the disease progresses the more communicability increases. One slight drawback for someone interested in weaponizing Ebola might be that by the time an individual was highly contagious they may have already sought treatment (Borio et al., 2002). On the other hand, individuals who have mounted an immune defense against the virus have been known to spread active virus through sexual intercourse months later, infecting unsuspecting individuals, further increasing the latent power of a weaponized form of Ebola virus. Other concerns have been raised about the possible spread of the disease via sweat and other body fluids (Borio et al., 2002), however, more data is required to confirm these suspicions. Besides being aerosolized, Ebola virus could be used as a weapon if it could be spread through the food or water supply, or even through infecting individuals in order to create an epidemic. Security and Defense Options Current defensive procedures against biological weapons in the United States are still in their infancy. The Department of Homeland Security (DHS) was created in 2002 under the Bush

Ebola virus 11 administration in response to the terror attacks on September 11, 2001 and the subsequent anthrax mailings (Center for Democracy & Technology, 2002). The DHS consolidated 22 agencies, and formed four directorates: Information Analysis and Infrastructure Protection, Science and Technology, Border and Transportation Security, and Emergency and Preparedness Response (Center for Democracy & Technology, 2002). The Science and Technology (S&T) Directorate is concerned with “threat awareness, surveillance and detection, response and recovery, and agro-defense, particularly against foreign animal diseases” (Cohen, 2007). The S&T collaborates with the Department of Defense (DoD), Department of Health and Human Services (HHS), the United States Department of Agriculture (USDA), the Department of Justice (DOJ), the Environmental Protection Agency (EPA), and the Department of State (Cohen, 2007). The S&T Directorate is required to play the “lead role in conducting assessments of the evolving biological threat” in order to guide prioritization of investments and research for biosecurity and biodefense (Cohen, 2007). The S&T Directorate has played a major role in employing the “BioWatch” aerosol monitoring system (Cohen, 2007). BioWatch is a program in which at least 31 cities have been equipped with aerosol samplers; the samplers are actually filters attached to EPA air sampling stations (Shea & Lister, 2003). The filters are tested every 24 hours for the presence of pathogens via DNA extraction and PCR by lab technicians (Shea & Lister, 2003). Ideally, PCR results would alert authorities to a biological attack before affected individuals presented with symptoms at health care facilities. BioWatch has been criticized for several reasons, including the placement of sensors with EPA stations instead of probable attack areas, the fact that sensor installation and PCR testing of filters is labor intensive and costly, and that scientific studies have not concluded the efficiency or usefulness of the sensors (Shea & Lister, 2003).

Ebola virus 12 President Bush signed the Project BioShield Act into law in 2004 with the aim of integrating national medical countermeasures into governmental preparedness programs against weapons of mass destruction (HHS, n.d.). Project BioShield grants the National Institues of Health the authority to streamline funding awards for projects that will aid in biodefense (HHS, n.d.). The Biomedical Advanced Research and Development Authority (BARDA) manages Project BioShield and is directly responsible for the management and integration of vaccines, drugs, and therapies into the biological defense system of the United States (HHS, n. d.). The “Select Agent Program” emerged out of the USA Patriot Act after the anthrax mailings in 2001, and was fully implemented in 2005 (Center for Biosecurity of UPMC, 2009). The program enforces accountability for those who work with certain pathogens and is run by the CDC, the USDA’s Animal and Plant Health Inspection Service (APHIS), and the Department of Justice, which performs background checks on the scientists involved in the program (Center for Biosecurity of UPMC, 2009). “Select” agents (more than 70 viruses, bacteria, and other pathogens that are deemed as biological threats) are under regulation of these agencies; and therefore only certain scientists are legally allowed to study them (Center for Biosecurity of UPMC, 2009). While APHIS regulates agents that relate to agriculture, the CDC regulates the human pathogens like Ebola virus (Center for Biosecurity of UPMC, 2009). As of April 2008, about 14,000 people from 324 government, private, and academic laboratories had been given clearance by the Department of Justice to work with the special pathogens (Center for Biosecurity of UPMC, 2009). The natural occurrence of “select” agents, including Ebola virus, limits the control the US government can have over the acquisition and transfer of them; rumors of Aum Shinrikyo’s attempt to acquire Ebola virus during a natural outbreak highlight the case in point, and this

Ebola virus 13 methodology of culturing pathogens is a very possible one. According to the Center for Biosecurity at the University of Pittsburgh Medical Center (2009), the Select Agent Program has resulted in negative consequences as relates to public health, science, and international collaborations. For example, clinical laboratories frequently destroy samples immediately in order to stay within governmental regulations when further research on the samples could prove fruitful for understanding how the pathogen harms its host, or for work on vaccines. Early-warning systems for biological attacks could potentially improve response time and recognition of a biological attack, even before physicians know what disease they are dealing with in the clinic or hospital. Several states already have systems where clinicians input symptomatic data into an electronic system that sends the information to the CDC; the CDC’s version is called Early Aberration Reporting System (EARS), and the tool began in New York City after the September 11, 2001 terror attacks (CDC, 2006). Data in this system is gathered through a variety of ways, including 911 emergency calls, school and work absenteeism, and over-the-counter drug sales (CDC, 2006). According to the CDC (2006), there are several US cities that are currently using EARS as a tool for biological terrorism preparedness. Some drawbacks, however, include data input that may not actually correlate to an outbreak of a serious pathogen in the United States. For example, over-the-counter sales of a drug may increase if the drug is on sale and not because of an epidemic; also, accidents in input coding of symptomatic data could possibly lead to expensive and unnecessary responses by the CDC, for example if someone accidentally coded an asthma attack as an Ebola virus infection. Detection and Diagnosis Infection with viral hemorrhagic fevers in North America is quite rare, and therefore physicians may not initially suspect Ebola virus infection if a patient presents with flu-like

Ebola virus 14 symptoms and rash. In the case of an epidemic outbreak, as could be the scenario in the event of a bioterrorist attack, many severely ill individuals reporting to hospitals and clinics will likely be what sets off warning bells. According to Puskoor and Zubay (2000), elevation of cytokines, interleukin-2, interleukin-10, TNF-alpha, interferon-alpha, and interferon-gamma, were all found in individuals who died from Ebola infection. Patients infected with Ebola usually complain about sudden onset of fever, headache, myalgia, bloody diarrhea, and vomiting; full-blown disease can lead to shock and bleeding, and eventually death (Leffel & Reed, 2004). Once a clinician suspects Ebola infection, it must be reported to local and state health departments and to the CDC’s Special Pathogens Branch (CDC, 2005). Samples should not be taken from the patient without first consulting the Special Pathogens Branch (CDC, 2005). Because of the high level of toxicity of Ebola, and the fact that there are currently no treatments available for infection, samples must be handled extremely carefully, and clinicians should follow all CDC guidelines, including the use of personal protective equipment when working near infected individuals. According to the CDC (2005), viral hemorrhagic fever should be suspected in patients that have a fever and that have either 1) traveled to a location where there has been a recent outbreak; 2) had contact with blood or other body fluids from an infected person or animal; or 3) were possibly exposed in a laboratory or clinical setting in which viral hemorrhagic fever was present. There are only a few public health laboratories that are properly equipped to identify viral hemorrhagic fever infection (Groseth et al., 2005), and therefore samples must be sent to one identified by the CDC Special Pathogens Branch. Enzyme-linked ImmunoSorbent Assay (ELISA) and reverse transcription polymerase chain reaction (RT-PCR) are techniques capable

Ebola virus 15 of identifying Ebola virus infection; also, staining of formalin-fixed tissues with immunoperoxidase has been used for filovirus identification in infected individuals (Groseth et al., 2005). Medical Response to an Attack with Ebola Virus Currently, the United States is ill equipped to handle a biological attack, especially one with Ebola virus since treatment is mostly supportive and includes very strict isolation and containment protocols. Once a quarantine facility was arranged to handle a large number of infected or potentially infected patients, they would likely follow military-type protocols that include providing supportive medical care in an intensive care facility, the administration of oxygen, an intravenous route of hydration, ventilation support for severe cases of Ebola infection, management of pain, and the avoidance of blood-thinning drugs like aspirin (US Army Office of the Surgeon General, 2000). Countermeasures Currently there is no cure, vaccine, or antidote for Ebola virus infection. Investigations into the molecular biology of Ebola virus have offered insight into how the virus attacks its host, and therefore research has focused on ways in which to defend against the virus’ pathological offenses. Rodent and nonhuman primate models have been utilized for Ebola virus infection research, including injection, inhalation, or conjunctivial routes of infection (Reed & Mohamadzadeh, 2007). Mouse models of Ebola infection are of limited use because mice do not develop coagulopathy or lymphocyte apoptosis like humans do; guinea pigs show coagulopathy but not to the level of nonhuman primates (Reed & Mohamadzadeh, 2007). Vaccines that protect guinea pigs or mice, however, don’t always protect nonhuman primates either (Reed & Mohamadzadeh,

Ebola virus 16 2007). Therefore, nonhuman primates are the most relevant models for human infection with Ebola; various primates have been used including African green monkeys, rhesus macaques, cynomolgus macaques, and baboons (Reed & Mohamadzadeh, 2007). While each show variations of pathology when infected with Ebola, the cynomolgus macaques are thought to be the most stringent model since the Zaire strain of Ebola is extremely virulent with this species as compared to others (Reed & Mohamadzadeh, 2007). According to Reed & Mohamadzadeh (2007), little is known about the pathogenesis of inhaled filovirus aerosol, and it isn’t thought to be the common route of exposure in natural outbreaks, though it has been a suspected route of communication. But experimental studies have shown filoviruses to be stable in aerosol, and therefore more research is necessary in order to understand the health effects of breathing in weaponized Ebola aerosol. Early experimental countermeasures for Ebola were vaccines made with formalin or gamma-irradiation inactivated virus; but often such a vaccine worked in one animal model but not another, and therefore that approach has been abandoned (Reed & Mohamadzadeh, 2007). Several new avenues of research exist for Ebola virus protection. Exploration of DNA vaccines that express the GP of Marburg and Ebola viruses have shown some promise in guinea pigs when it was boosted with baculovirus-expressed GP lacking the transmembrane domain (Reed & Mohamadzadeh, 2007). In 2004, a DNA vaccine construct was successful in protecting guinea pigs and nonhuman primates from the Zaire strain of Ebola, and began phase I clinical trials (Reimenschnieder et al., 2003). Vector-based vaccines have been utilized in research as well. Genes of interest are mutated to remove toxicity and then inserted into a vector; ideally the recipient’s immune system will build antibodies to the gene so that when the real live virus infects, the immune system will

Ebola virus 17 be prepared to attack it. Various issues exist that make the use of vector-based vaccines problematic. Live viral vectors can be harmful to immunosuppressed individuals, and people may also be immune to the vector, thereby blocking any potential immunity formation against the inserted gene (Reed & Mohamadzadeh, 2007). Adenovirus-based vectors have been successful in protecting nonhuman primates against Zaire strain Ebola. Sullivan and colleagues (2003) reported a single immunization with adenovirus vector containing the GP gene from the Zaire strain of Ebola protected cynomolgus macaques from lethal dose injection of the virus. Reed and Mohamadzadeh (2007) pointed out that this study used a dose that is much lower than a common accidental needlestick exposure; still, all control animals were killed. This study used a nonreplicating adenovirus, making it necessary to receive a very high dose of vaccine; further, pre-existing immunity to the adenovirus vector in the general population may be an issue; there are reports that as much as 50% of the human population are immune to the adenovirus used by Sullivan and colleagues (2003) (Reed & Mohamadzadeh, 2007). Vesicular stomatitis virus (VSV) engineered to express GP of Ebola (Zaire strain) has protected cynomolgus macaques from challenge, as well as from rechallenge with different strains of Ebola (Reed & Mohamadzadeh, 2007). Because VSV is a live virus, it was necessary to test it in immunocompromised models, as live virus vaccines can be dangerous for such individuals. Geisbert and colleagues (2008) reported a VSV for Ebola that protected against lethal Ebola challenge and was well-tolerated in nonhuman primates. Virus-like particles (VLP) are viral protein aggregates; they mimic the native protein’s conformation but do not confer safety concerns as do live viruses. By transfecting 293T cells with GP and VP40 genes from Ebola, Bavari and colleagues (2002) discovered the cells

Ebola virus 18 produced VLPs that were virtually indistinguishable from Ebola virus particles; therefore the introduced genes were building VLPs using materials available within the cell, including the cells lipid membrane. These VLPs may be useful for eliciting immune responses, and therefore as vaccines for Ebola virus. Guinea pigs that were vaccinated with VLPs have been fully protected from Zaire strain Ebola as well as Marburg virus (Reed & Mohamadzadeh, 2007). Risk Assessment Proof of previous weaponization of Ebola virus has been elusive, despite reports of soviet defectors (Alibek & Handelman, 2000; PBS, 2001). Because Russian defectors have claimed such weaponization, and because former Russian President Boris Yeltsin admitted that Russia covered up biological weapons research performed even after signing the Biological Weapons Convention treaty in 1972 (Alibek & Handelman, 2000; Fong & Alibek, 2005; PBS, 2001; PBS, 2006), the potential threat of weaponized Ebola virus has been taken seriously by the United States. Further, the United States has not visited Russian biological research facilities since 1994 (Moodie, 2001), leaving 15 years with which possible weapons could have been created and/or sold during the collapsed soviet economy. Aerosolization of biological weapons is considered one of the most efficient methods of delivery for a variety of reasons, including the potential to infect large numbers of people (or other targets, such as livestock or crops) with one delivery of agent. Ebola virus is a highly lethal agent, with a mortality rate between 50-90% (Groseth et al., 2005). Because there is no treatment or vaccine available, an attack with Ebola virus would create mass panic and likely overwhelm health care facilities that are not equipped to contain a BSL 4 virus. It is likely that infected individuals may present at hospitals with flu-like symptoms, and diagnosis of Ebola infection may not occur until obvious symptoms arise, at which time the individual may have

Ebola virus 19 already infected hospital staff. Hospitals would only be able to offer supportive care for victims of Ebola, and it is likely that severe disruption of the healthcare system in the affected area would occur. Hospitals would likely not accept patients known to be infected with Ebola virus in an effort to protect the uninfected as quarantine quarters would probably not be available. A separate quarantine facility would have to be set up as quickly as possible after an attack, and other potentially infected individuals would need to be identified for quarantine in an effort to contain spread. Humans infected with Ebola virus have been found to contain high titers of virus in their skin and sweat glands (Borio et al., 2002). While evidence is lacking, it is possible that Ebola could be spread from person-to-person via touch. More probable would be infection of small abrasions or cuts from touching an infected person, as several people in the Democratic Republic of the Congo have contracted Ebola after washing the bodies and cutting the hair and nails of dead, Ebola infected bodies (Borio et al., 2002). There has been some evidence for the possibility of transmission of Ebola virus through small drops of airborne particles, such as those ejected during coughing. Though most evidence suggests that the virus is not contagious during the incubation period, the evidence is not conclusive (Groseth et al., 2005). It is possible, though the probability not completely understood, that if an attack occurred, unsuspecting individuals could spread the virus to others before the onset of symptoms. In the event that an Ebola virus attack occurred without warning or suspicion, individuals may not seek help even after the onset of symptoms, increasing the risk of spreading the disease to others, as the risk of transmission seems to increase with time after infection (Groseth et al., 2005). While some individuals infected will mount an effective immune response and overcome

Ebola virus 20 Ebola infection, there is evidence that the virus can persist in body fluids such as semen and ocular fluids for 12 weeks after the onset of symptoms (Groseth et al, 2005). Individuals, therefore, may appear quite healthy and active yet still transmit the disease to others. Not enough evidence exists yet to determine possible vectors, such as mosquitoes or rodents indigenous to the United States, for transmission of Ebola. Pigs have been infected with Ebola Reston, however, and therefore infection of livestock as a weapon of terror is possible (Pig Progress, 2008). Rumors have circulated that the Japanese cult, Aum Shinrikyo, once sent a group to Zaire during an Ebola outbreak to gather viral samples in order to culture them for biological weapons (Olson, 1999). It appears, therefore, that the acquisition of virus is highly possible; weaponization has probably already occurred in Russia; and aerosolization has proven to be highly lethal in monkeys, with very low doses (400 viral particles) (Leffel & Reed, 2004).

Table 1. Risk Assessment Rubric Description Effectiveness of Ebola virus as a weapon Lethality 23 Availability 10 Likelihood of 3 misdiagnosis Ability to spread 4 (natural vectors) Contagious 5 Ability to weaponize 5 Ability to employ 5 Ability to 15 terrorize/panic Economic toll 10 Ability to infect 10 livestock and/or other agricultural resources Difficulty in treating 15 Total usefulness 105

Standard 25 20 5 5 5 10 10 15 10 15 15 135

Ebola virus 21 Conclusion While Ebola virus can cause devastating and gruesome disease in infected individuals, proof of its weaponization has not been attained. Dr. Kenneth Alibek, former deputy director of the Russian agency Biopreparat, is the most notable proponent of the idea of Ebola weaponization. His credibility is questionable, and therefore, so is the notion that Ebola has been weaponized. According to an article in the Los Angeles Times, some of Alibek’s research findings have not withstood peer review; he promotes a nonprescription pill that allegedly bolsters the immune system; his claims that Iraq definitely had stores of smallpox was never proven, even after the US invaded after September 11, 2001 (Willman, 2007). Alibek’s claims have been part of the driving force behind the US government’s biodefense agenda, which has thus far cost billions of dollars. Nevertheless, the possibility of Ebola virus being used as a weapon exists and defensive plans have begun to take shape in the United States. Because of claims that Russia has experimented with deadly pathogens that defy the Biological and Chemical Weapons Convention that they are party to, inspections of Russian facilities should be conducted by the United States. Not only should this be done for the safety of the world, but also as a way to determine Alibek’s credibility. There is virtually no evidence thus far to prove that Ebola virus has ever been used as a weapon, and no concrete evidence that it has been weaponized. Project BioWatch is underway in the United States. The project is ambitious, and it is just beginning. While it has been criticized for falling short of the ideal of complete protection of biological and chemical attacks, the project has been a giant stride forward in terms of biosecurity and biodefense. Refining the detection factors is necessary in order to ensure that Project BioWatch will move closer and closer toward the goal of complete protection; however,

Ebola virus 22 that goal may be unattainable. The best possible protection will be in prevention of an Ebola virus attack; however, because that is not completely possible, vaccines and antidotes will be key in ensuring the survival of victims of an attack. Understanding aspects of a biological attack, such as wind currents, background levels of pathogens, likelihood of attack at location where sensor is placed, and the re-evaluation of placing BioWatch sensors with EPA sensor sites will all need to be taken into account. More information must be gathered regarding the weaponization potential of Ebola virus, including Ebola-specific aerosolization studies to include the ease with which Ebola can be weaponized as an aerosol. Though some information has shown that Marburg virus can be aerosolized and can survive for long periods of time on contaminated surfaces, this has not been proven for Ebola (Leffel & Reed, 2004). There has been a report of the spread of Ebola virus to uninfected macaques that were housed in the same room as experimentally infected macaques, and Leffel and Reed (2004) suggest that it was possibility that the spread occurred through aerosolized particles infected with Ebola during cage cleaning. This theory has not yet been examined under scientific scrutiny. Much is known about the pathogenesis of Ebola virus infection via injection, but not much is known about inhalational infection because of a lack of research in this area. An aerosolized form of Ebola virus would arguably be the most effective method of dissemination on a target; therefore more research on aerosolized Ebola virus needs to be done. Moreover, more research should be done on individuals who have mounted immunity to Ebola virus, as their biochemistry may provide valuable information to scientists in search of protection from the virus. Another aspect of an attack with Ebola virus, or any other pathogen, that should be

Ebola virus 23 studied is the possibility of genetic modification which may be done in order to increase virulence, to add layers of disease by combining genes from another pathogen, for example, adding a gene from Venezuelan equine encephalitis (VEE) with Ebola virus, or to mask the identity of the pathogen. Dr. Alibek has made claims that Biopreparat experimented with chimeras while he was employed there (Alibek & Handelman, 2000). By determining sequences within the Ebola virus’ genome that can be modified in order for it to maintain its virulence could help in designing a robust vaccine that could still defend against a modified virus. Enormous strides have been made in regard to developing protective countermeasures against Ebola virus. While there is not yet an approved vaccine for humans, one is surely on the horizon. With Geisbert and colleagues (2008) report detailing a well-tolerated vaccine in immunocompromised nonhuman primates, future clinical trials are possible. Improvements in all aspects of biological terror preparedness in the United States are underway, and, while currently little evidence exists for the use of Ebola virus as a weapon, perhaps governmental vigilance for protecting its citizens may benefit Ebola research. Because Ebola virus has thus far rarely shown itself in the United States, it has not been considered an imminent threat; the fact that is must be studied in a BSL-4 facility makes fast progress for Ebola virus protection difficult when there are only 5 such facilities in the US.

Ebola virus 24 References Alibek, K. (1998). Terrorist and intelligence operations: Potential impact on the US economy. Washington, DC: Joint Economic Committee, United States Congress. Retrieved March 21, 2009, from http://www.house.gov/jec/hearings/intell/alibek.htm Alibek, K., & Handelman, S. (2000). BIOHAZARD. New York: Random House. Bavari, S., Bosio, C. M., Wiegand, E., Ruthel, G., Will, A. B., & Geisbert, T. W., et al. (2002). Lipid raft microdomains: A gateway for compartmentalized trafficking of Ebola and Marburg viruses. The Journal of Experimental Medicine, 195(5), 593-602. Borio, L., Inglesby, T., Peters, C. J., Schmaljohn, A. L., Hughes, J. M., & Jahrling, P. B., et al. (2002). Hemorrhagic fever viruses as biological weapons. Journal of the American Medical Association, 287(18), 2391-2405. CDC. (n.d.). Bioterrorism agents/diseases by category. Retrieved March 10, 2009, from http://www.bt.cdc.gov/agent/agentlist-category.asp CDC. (2002). Ebola hemorrhagic fever information packet. Retrieved March 2, 2009, from http://www.cdc.gov/ncidod/dvrd/spb/mnpages/dispages/Fact_Sheets/Ebola_Fact_Booklet .pdf CDC. (2004). Filoviruses. Retrieved March 12, 2009, from http://www.cdc.gov/Ncidod/dvrd/spb/mnpages/dispages/filoviruses.htm CDC. (2005). Interim guidance for managing patients with suspected viral hemorrhagic fever in US hospitals. Retrieved March 22, 2009, from http://www.cdc.gov/ncidod/dhqp/bp_vhf_interimGuidance.html CDC. (2006). CDC early aberration reporting system (EARS). Retrieved March 28, 2009, from http://www.bt.cdc.gov/surveillance/ears/

Ebola virus 25 Center for Biosecurity of UPMC. (2009). Preventing and deterring biological attacks: Priorities that should emerge from the WMD commission report. Retrieved March 26, 2009, from http://www.upmc-biosecurity.org/website/resources/commentary/2008-12-19preventdeterbioweapons.html Center for Democracy and Technology. (2002). The new homeland security department: Challenge, potential, and risk- privacy guidelines, careful oversight required. Retrieved March 13, 2009, from http://www.cdt.org/security/homelandsecuritydept/021210cdt.shtml Cohen, J. M. (2007). Six years after the anthrax attack: Are we better prepared to respond to bioterrorism? Washington: Government Printing Office. Feldmann, H., Wahl-Jensen, V., Jones, S. M., & Stroher, U. (2004). Ebola virus ecology: a continuing mystery. TRENDS in Microbiology, 12(10), 433-437. Fong, I. W., & Alibek, K. (2005). Bioterrorism and infectious agents: A new dilemma for the 21st century. New York: Springer. Geisbert, T. W., Daddario-DiCaprio, K. M., Lewis, M. G., Geisbert, J. B., Grolla, A., & Leung, A., et al. (2008). Vesicular stomatitis virus-based Ebola vaccine is well-tolerated and protects immunocompromised nonhuman primates. Public Library of Science Pathology, 4(11), 1-16. Retrieved March 23, 2009, from http://www.plospathogens.org/article/info:doi%2f10.1371%2fjournal.ppat.1000225 Groseth, A., Jones, S., Artsob, H., & Feldmann, H. (2005). Hemorrhagic fever viruses as biological weapons. In I. W. Fong & K. Alibek (Eds.), Bioterrorism and infectious agents: A new dilemma for the 21st century (pp. 169-191). New York: Springer. HHS. (n.d.). Project BioShield. Retrieved March 13, 2009, from

Ebola virus 26 http://www.dhhs.gov/aspr/barda/bioshield/index.html Johnson, E., Jaax, N., White, J., & Jahrling, P. (1995). Lethal experimental infections of rhesus monkeys by aerosolized Ebola virus. International Journal of Experimental Pathology, 76(4), 227-236. Leffel, E. K., & Reed, D. S. (2004). Marburg and Ebola viruses as aerosol threats. Biosecurity and Bioterrorism: Biodefense Strategy, Practice, and Science, 2(3), 186-191. McCarter, M. (2008, October 17). Two of five BSL-4 labs flunk security test. Homeland Security Today. Retrieved March 28, 2009, from http://www.hstoday.us/content/view/5659/128/ Moodie, M. (2001, Spring). The Soviet Union, Russia, and the biological and toxin weapons convention. Nonproliferation Review, 59-69. Retrieved March 13, 2009, from http://cns.miis.edu/npr/pdfs/81moodie.pdf Oehler, G. C. (1996). The growing chemical and biological weapons threat. Retrieved March 12, 2009, from http://www.fas.org/irp/cia/product/go_testimony_032796.html Olson, K. B. (1999). Aum Shinrikyo: Once and future threat? Retrieved March 13, 2009, from http://www.cdc.gov/ncidod/eid/vol5no4/olson.htm PBS. (2001). Interviews with biowarriors: Sergei Popov. Retrieved March 4, 2009, from http://www.pbs.org/wgbh/nova/bioterror/biow_popov.html PBS. (2006). Timeline: Biological weapons. Retrieved March 13, 2009, from http://www.pbs.org/wgbh/amex/weapon/timeline/timeline2.html Pig Progress. (2008). Ebola detected in Philippine pigs. Retrieved March 29, 2009, from http://www.pigprogress.net/news/ebola-detected-in-philippine-pigs-2409.html Puskoor, R., & Zubay, G. (2000). Ebola viruses. In G. Zubay (Ed.), Agents of bioterrorism (59-

Ebola virus 27 78). New York: Columbia University Press. Reed, D. S., & Mohamadzadeh, M. (2007). Status and challenges of filovirus vaccines. Vaccine, 25, 1923-1934. Riemenschneider, J., Garrison, A., Giesbert, J., Jarhling, P., Hevey, M., Negley, D., et al. (2003). Comparison of individual and combination DNA vaccines for B. anthracis, Ebola virus, Marburg virus and Venezuelan equine encephalitis virus. Vaccine, 21(2526), 4071-4080. Sanchez, A., Yang, Z. Y., Xu, L., Nabel, G. J., Crews, T., & Peters, C. J. (1998). Biochemical analysis of the secreted and virion glycoproteins of Ebola virus. Journal of Virology, 72(8), 6442-6447. Shea, D. A., & Lister, S. A. (2003). The BioWatch program: Detection of bioterrorism. (Order Code RL32152). Washington, DC: CRS Report for Congress. Retrieved March 13, 2009, from http://www.fas.org/sgp/crs/terror/RL32152.html Simpson, D. I. H. (1978). Viral haemorrhagic fevers of man. Bulletin of the World Health Organization, 56(6), 819-832. Sullivan, N. J., Giesbert, T. W., Giesbert, J. B., Xu, L., Yang, Z. H., Roederer, M, et al. (2003). Accelerated vaccination for Ebola virus haemorrhagic fever in non-human primates. Nature, 424(6949), 681-684. Sullivan, N., Yang, Z. H., & Nabel, G. J. Ebola virus pathogenesis: Implications for vaccines and therapies. Journal of Virology, 77(18), 9733-9737. US Army Office of the Surgeon General. (2000). Medical NBC aspects of Ebola. Retrieved March 28, 2009, from http://209.85.173.132/search?q=cache:uHlEQuCNdMJ:https://www.cbrniac.apgea.army.mil/Documents/Presentations/ebola.ppt+medical+re

Ebola virus 28 sponse+to+ebola&cd=1&hl=en&ct=clnk&gl=us&client=safari Willman, D. (2007, July 1). Selling the threat of bioterrorism. The Los Angeles Times. Retrieved March 26, 2009, from http://www.latimes.com/news/nationworld/nation/la-nabioterror1jul01,1,1052428.story