Biological Weapons and Bio-Terrorism : a review of History and Biological Agents.

ABSTRACT Biological weapons are a thorny issue, for their destructive capabilities and for the potential to generate panic and terror among the affected people. Used since pre-Christian times, biological weapons have resulted in the decimation of whole populations and have changed the geopolitics of several places. In this work, a summary of the main war and terrorist activities carried out by biological weapons over the time is presented. Moreover, main biological warfare agents and related pathology are considered, according to American Centers for Disease Control and Prevention (CDC) priority classification. At last, the emerging more destructive potential of biological agents, due to the widespread introduction of biotechnology, is analyzed. 1. INTRODUCTION DaSilva, in his work published in 1999, defines the Biological warfare as the intentional use of microorganisms, and toxins, generally of microbial, plant or animal origin to produce disease and death in humans, livestock and crops (DaSilva, 1999). Biological warfare and bioterrorism are very complex subjects, mainly due to the many agents that can be used as a weapon and for the wide range ways for the dissemination in the environment and population. A biological event provides for the presence of at least two actors: one or more pathogens (bacteria, viruses, or toxins) and a vehicle for their dissemination. What makes biological threat so troubling? In addition to high spread capacity and lethality of biological agents, the invisibility and the extremely difficult short-term revelation makes impossible an immediate contrast with the subsequent increase of the affected. In fact, most of biological weapons (unless of toxins) have a peculiar quality that other non-conventional weapons (chemical and radiological ones) do not show: biological agents are able to multiply in the host organism and be transmitted in turn to new hosts, generating in this way not predictable effects on the population, both in terms of number of victims and in terms of spread geography. Among the reasons which make biological weapons attractive, there is their very low cost when compared to both conventional and unconventional ones. NATO sources, according to data processed in 1969 by U.S. experts, reported the following costs for an attack on an area of 1 km2 to civilian populations with different weapons: a) $1/Km2 for biological weapons, b) $600/Km2 for chemical, c) $800/Km2 for nuclear, d) $2,000/Km2 for conventional armaments (NATO, 1996). Therefore, it seems clear therefore that, in financial terms, an attack with biological weapons would be strongly cheaper when compared to any other one. In the second decade of the 21st century, to make the situation more troubling, there is the simplicity with which it is possible to produce large quantities of biological agents with facilities and expertise available to everyone, even to terrorist and paramilitary groups. The aim of this work is to present an overview on biological warfare and bioweapons, as well as to assess the state of the art on the actual offensive capability through this unconventional weapon. 2. HISTORYCAL 2.1 Pre Word Wars age The use of biological agents as war weapons is not a modern era novelty. Although it isn’t easy to identify a definite time when the use of bioweapons began, ancient information reported that in pre-Christian era, around 300 B.C., Greeks were using animal cadavers to contaminate water wells of the opponent. This strategy was also used the by Romans and the Persians (SIPRI, 1971). In a later period, during the battle of Tortona (Italy) in 1155, bodies of dead soldiers and animals were used to contaminate water wells by Barbarossa (Clarke, 1968). In the 14th century, during the siege of Kaffa by Tartar (now Feidisuja, Ukraine, a city on the Black Sea at that time under the control of the Genoese), a biological attack allowed Tartars to win the siege. During the siege, among the Tatar army, an epidemic of plague was spread: the besiegers thought to shoot the cadavers of their dead comrades within the walls of the city of Caffa (Wheelis, 2002). This resulted in a turning point in the war: the Genoese fled from Caffa, carrying with them the sick. On the return trip to Genoa, they landed several ports in the Mediterranean sea; although some sources believe a possible correlation between the epidemic of plague in Caffa and the pandemic that decimated the population of Europe in the following decades (Black Death), most of the authors share the view of two events independent (Wheelis, 2002). In 1422, during the siege of Carolstein, Lithuanian soldiers catapulted within the city cadavers of death soldiers and 2,000 chariots of excrement (Newark, 1988), frightening the population affected and spreading lethal fevers in many cases. We will have to wait for more than three centuries to find a new documented use of biological agents as a war weapon: during the French-Indian War (1754-1767), the British commander Sir. Jeffrey Amherst ordered the distribution of blankets infected with smallpox to decimate the population of Indian tribes hostile to the British (Bhalla and Warheit, 2004). The distribution of infected blankets occurred in the summer of 1763, and the resurgence of the virus among the indigenous lasted for over 200 years (Riedel, 2004). 2.2 World War I and II Several biological warfare actions carried out during the World War are not sufficiently confirmed in the literature. However is frequently reported that Germans inoculated cattle with Bacillus anthracis and Pseudomonas mallei, responsible to cause severe diseases such as anthrax and glanders, before sending them in enemy states (SIPRI, 1971; Poupard and Miller, 1992; Hugh-Jones, 1992). It should be considered that the First World War saw the large-scale use of non-conventional chemical weapons, therefore it must be expected until the Second World War to find a more extensive use of biological weapons. During World War II many countries conducted research programs on the development of bioweapons; the Japanese program, conducted under the direction of Captain Shiro Ishii, was certainly most ambitious (1892-1959). The research in this direction started in 1928; during this year Captain Ishii visited many European and American countries in order to learn useful techniques and information about the possible uses of biological weapons. Returned to homeland, Captain Ishii was substantial granted in order to constitute a massive bioweapons research center, called the 731 units, located at Beiyinhe, in Manchuria. The research center could count on a staff of over 3,000 scientists, mainly microbiologists. The experiments were conducted on pow, principally Koreans, Chinese and Russian soldiers. Prisoners were used to test numerous bioweapons, including Yersinia pestis, Vibrio cholera, Neisseria meningitidis and Bacillus anthracis (Leitenberg, 2001). Christopher and colleagues report that during this research, several thousand prisoners died as a result of the experiments conducted on them (Christopher et al., 1997). However, the mortality rate around the area 731 remains very high for several years. If we consider for the total count these deaths also, we reach the considerable sum of 200,000 deaths as a result of the activities carried out by Captain Ishii (Harris, 2002). In 1942, the wrong control of the infection spread resulted in the death of 1,700 Japanese soldiers (Sokolski and Ludes, 2001). Many other nations carried out experiments on potential biological agents, but information reported in the literature are rather skinny. It is also important to remember the experiments conducted in 1942 by the British army on the Island of Gruinard, off the Scotland coast, where anthrax dirty bombs were tested (Manchee et al., 1981). The island was contaminated and uninhabitable until 1990, when an extensive land decontamination was carried out (Aldhous, 1990). 2.3 Post World Was age Until World War II, United States remained considerably behind respect the other nations in the research fields on biological weapons. In the United States, the golden age for both the test and development of bioweapons in the United States was immediately after the conclusion of World War II, when U.S. received the results of the experiments performed by the Japanese unit 731. The U.S. could also rely on working directly with Captain Ishii, the former director of the unit 731 (Christopher et al., 1997). In September of 1950, the U.S. Navy conducted an experiment on civilians in order to assess the vulnerability of a large American coastal town to a biological attack: in the San Francisco Bay a cloud of Serratia marcescens (a low pathogenic bacterium mainly responsible for infections of skin and respiratory tract) was spread by boat. The infection struck, as a result of subsequent checks, almost the entire population (1 million people). Even though the bacterium had to be almost harmless, several individuals showed the effects of respiratory diseases and some of them died (Christopher et al., 1997). Few years later (1956-1958), in Georgia and Florida, swarms of mosquitoes, probably carriers of yellow fever, were released in order to verify the vulnerability to an air attack. Even though the documents are still top secret, several sources report that some individuals died for the bite of insects. A last large scale experiment which is documented, consists in the dissemination of Bacillus subtilis in the New York subway in the summer 1966. The experiment resulted in the infection, although without consequences, of more than one million people. It demonstrated that the spread of a pathogen in the whole subway network from a single station, due to the displacement of air in the tunnels, was possible (Zygmunt, 2006). In the seventies, USSR conducted an ambitious research program on biological weapons; however, unlike the United States programs, on which the secrecy has been partially removed, an aura of mystery on Russian research programs still remains. Some information about the research carried out in the USSR are provided in the paper published by Davis in 1999 on Emerging Infectious Disease. The USSR, between 1973 and 1974, formed an organization called "Chief Directorate for Biological Preparation" (Biopreparat), with the purpose to develop and produce biological weapons. Although there are no unambiguous data about the number of individuals employed by the Biopreparat, it is believed that more than 50,000 people were working in the whole system connected to the structure, including scientists and technicians, who could count on 52 research and producing factories. In these structures, high amounts of etiologic agents of plague, tularemia, anthrax, glanders, smallpox and Venezuelan equine encephalomyelitis were studied and produced. Not only biological agents from natural sources were studied in the Soviet Union. The Soviets studied and applied technologies of genetic engineering in order to increase the aggressiveness of biological agents through biotechnology The aim of this work was the production of a new more dangerous, more easily spread and more difficult to identify and combat generation of biological weapons (Davis, 1999). Among the countries that developed a massive program on bioweapons research, in the post-war era, we should quote Iraq. Iraq started its research and development program in the field of biological warfare in 1974, contextualizing it in an organization called "State Organization for Trade and Industry" (Davis, 1999). The program consisted on the study and production of botulinum toxin, anthrax, aflatoxin and ricin, as well as antiplants and viral agents (Rotavirus, Infectious Hemorrhagic Conjunctivitis, Camel Pox) (Leitenberg, 2001). The program involved about 300 scientists, who completed their training in Western Europe countries (Leitenberg, 2001). 2.4 International treaties The first measures against the use of biological weapons were taken in the 19th century during the Hague Conference in 1899, and then confirmed in the same place in 1907: with the document "Laws and Customs of War on Land", signed by 24 countries, the prohibition against the use of poisoned arms was ratified (Leitenberg, 2001). In 1925, with the awareness of the horrors of World War I, especially as regards the use of chemical weapons, the Geneva Protocol was signed. It "Prohibited the use in war of Asphyxiating, Poisonous or Other Gases, and of Bacteriological methods of Warfare". Although this treaty was signed by a considerable number of nations (even though it was ratified by the U.S. not until the mid-seventies), it prohibited the use of biological agents as weapons only, but not their development or their stocking (Christopher et al., 1997). In view of the limited effectiveness of the Protocol of Geneva in the control of the bioweapons development and proliferation, in 1972 the "Conventions on the Prohibition of the Development, Production and Stockpiling of Bacteriological (Biological) and Toxin Weapons and on Their Desctruction"was initialed. Initially signed by over 100 nations, the Convention became effective in 1975; however this convention, such as that Geneve one, has several loopholes: first of all, it does not provide guidelines for the protocol compliance verification; moreover it prohibits the use and development of bioweapons in “quantities that have no justification for prophylactic, protective or other peaceful purposes” (Riedel, 2004). However, it is evident how this assertion is open to interpretation; de facto it does not define threshold quantities or substantial limitations to the development and production of bioweapons (SIPRI 1971 b, 1973). The events consequent to the ratification of the Biological Weapons Convention of 1972, up to the most recent, have confirmed this observation. 2.5 Last decades: bioterrorism and biotechnology advent Even after the entry into force of the "Conventions on the Prohibition of the Development, Production and Stockpiling of Bacteriological (Biological) and Toxin Weapons and on Their Destruction", a large number of countries went on to develop, produce and test biological agents for military purposes. Since the 80s, increasingly terrorist groups have begun to consider the bioweapon as an economic and highly destabilizing way for civil society. It must be considered that the large scale advent of biotechnology, and the reduced difficulty in genetically modified organisms producing, has made the potential creation of multi-drug resistant pathogens, or with enhanced virulence factors possible. The use of biological agents in the last decades is mainly attributable to terrorist groups, more or less isolated, who used the bioweapons as a strategy to defend extremist religious ideas striking the civilian population or sensible government targets. In 1984 in The Dalles, Oregon, United States, a group of extremist followers of Bhagwan Shree Rajneesh (also known as Osho), contaminated the salad in 10 different salad bar with the pathogen of salmonellosis, Salmonella thyphimurium, in order to disable the population. 751 people contracted the disease and several of them were hospitalized; although there were no fatalities, this terrorist act is considered the largest bioterrorist attack in the history of the United States for involved people (Török et al., 1997). In the nineties the Japanese cult of Aum Shinrikyo tested different biological weapons, including botulin toxin, anthrax, cholera, and Q fever; in 1993, during a humanitarian mission in Africa, it tried to obtain samples of the Ebola virus. Between 1990 and 1995, the cult attempted to carry out several bioterrorist actions in Tokyo using vaporized biological agents including botulinum toxin and anthrax spores. Fortunately, the attacks were unsuccessful (Olson, 1999). A significant bioterrorist event happened in the United States contextually to the dramatic attacks to the World Trade Center in New York in September 2001. The release of Bacillus anthracis spores through the U.S. postal system was carried out with letters addressed to the press and to government officials. There were 22 confirmed cases of anthrax contamination, 12 cutaneous and 10 inhalational cases. The 12 cutaneous responded positively to the antibiotic treatment, while of the 10 inhalational cases, 4 were fatal (McCarthy, 2001). In 2002, in Manchester, UK, six terrorists were arrested for having been found in possession of ricin and in 2004, traces of the same toxin were found at the Dirksen Senate Office Building in Washington DC. So, it appears evident then that the use of biological agents has moved, in recent times, to terrorist groups. This creates very strong concerns: the use of biological weapons by terrorists can generate unexpected scenarios characterized by massive destructive potential. 3. BIOTERRORISM AND BIOWEAPONS The American Center for Disease Control and Prevention (CDC) defines a bioterrorism attack as “the deliberate release of viruses, bacteria or other germs (agents) used to cause illness or death in people, animals, or plants” (CDC, 2013). CDC classifies the most important bioterrorism agents into three categories (tab. 1): a) Category A: Agents that can be easily disseminated or transmitted from person to person. They result in high mortality rates and have the potential for major public health impact. They might cause public panic and social disruption, and require special action for public health preparedness. b) Category B: Agents that are moderately easy to disseminate. They result in moderate morbidity rates and low mortality, and require specific enhanced diagnostic capacity and disease surveillance. c) Category C: Emerging agents that could be engineered for mass dissemination in the future because of their availability. They are easy to produce and disseminate. They are potentially linked to high morbidity and mortality rates, and major health impact. Table 1: Major biological agents possible used as bioweapons (CDC, 2013). Groups Diseases Agents A Anthrax Bacillus anthracis Botulism Clostridium botulinum toxin Plague Yersinia pestis Smallpox Variola major Tularemia Francisella tularensis Viral hemorrhagic fevers Filoviruses and Arenaviruses B Brucellosis Brucella spp. Epsilon toxin Clostridium perfringens Food safety threats Salmonella spp., E.coli O157:H7, Shigella Glanders Burkholderia mallei Melioidosis Burkholderia pseudomallei Psittacosis Chlamydia psittaci Q fever Coxiella burnetii Ricin toxin Ricinus communis Staphylococcal enterotoxin B Staphylococcus spp. Typhus fever Rickettsia prowazekii Viral encephalitis Alphaviruses Water safety threats Vibrio cholerae, Cryptosporidium parvum C Emerging infectious diseases Nipahvirus and Hantavirus Generally, biological agents (included those used as weapons) can be further classified according to certain characteristics that define the hazard to health (NATO, 1996): a) Infectivity: The aptitude of an agent to penetrate and multiply in the host; b) Pathogenicity: The ability of the agent to cause a disease after penetrating into the body; c) Transmissibility: The ability of the agent to be transmitted from an infected individual to a healthy one; d) Ability to neutralize: It means to have preventive tools and/or therapeutic purposes. Biological agents can be transmitted through one or more ways. The transmission modes are the following (La Placa, 2010): a) Parenteral: Agents that are transmitted through body fluids or blood; b) Airway (by droplets): Agents that are emitted by infected people, which can then be inhaled by surrounding people; c) Contact: Through which the agents present on the surface of the infected organism can infect another organism; d) Oral-fecal route: Through objects, foods or other items contaminated with the feces of infected patients, or through sexual contact. 3.1 Bioweapons There are numerous pathogens (bacteria, viruses and toxins) that cause diseases in humans, animals and plants; however, only very few possess the characteristics to be a bioweapon. In literature, in 1997 Eitzen describes the characteristics that make a biological agent a potential bioweapon. Ideally, a biological weapon should be easy to find or to produce. In order to develop a biological attack towards sensitive targets or the population, large amounts of biological agents are in fact required; it must be considered that it is necessary quite a number of biological agents (or a certain amount of toxin) to generate a disease in an target. The ideal bioweapon also must hold a high capacity to incapacitate the affected or, alternatively, be highly lethal. It is appropriate to choose an agent with an incubation period depending on whether immediate or delayed effects are required. Other important characteristics for a biological weapon are the route of transmission, and hence, the ease of dissemination with an appropriate method of delivery. Finally, the stability of the agent must be assessed, especially when large quantities must be stored for indefinite periods (Eitzen, 1997). Below some of the key features of the most relevant biological agents (included in A category by CDC) are reported, categorized according to the biological origin. 3.2 Bacteria Bacillus anthracis. Bacillus anthracis is a Gram-positive, non-motile, facultative anaerobic endospore forming bacteria, usually surrounded by a capsule; it is the etiological agent of anthrax. The disease occurs most frequently when an epizootic or enzootic of herbivores becomes infected after acquiring spores from direct contact with contam