Potential Threat: Bioterrorism

Deniz Akyüz^A. , Elif Duymaz^E.

Since the dawn of humanity, societies and nascent states have often found themselves in contention with neighboring states and primitive communities. In those early years, the most formidable and influential tools wielded as leverage were geographical positioning and population size. States and communities boasting strategic locations and substantial military forces frequently emerged as dominant powers. This relentless pursuit of supremacy eventually gave rise to a concept devised by humanity: terrorism. Terrorism encompasses acts of violence and aggression directed against states, governments, or specific communities, motivated by various factors such as religion, ideology, or economic interests.

With the advent of technological revolutions, the term “terrorism” as conceived by humanity, has acquired new dimensions and roles. The concept of bioterrorism emerged with the utilization of biological weapons. The mere mention of “biological weapons and bioterrorism” can evoke profound fear¹. Microorganisms used as bioweapons (biological agents) under the umbrella of bioterrorism are not only genetically amenable to development but also practical in their application and manipulation. If these microbes are engineered and deployed against another country, it constitutes biowarfare. Biowarfare refers to the scenario arising from the use of biological weapons, which are scrutinized across a broad spectrum of areas. Genetically modified bacteria, capable of being weaponized to cause death or other biological harm to living beings in a region, underscore the looming threat of biological attacks. A biological attack involves the intentional deployment of living microorganisms or their toxic products (toxins) to induce disease and death in humans, animals, and plants². However, the primary target of biological attacks is invariably humans. In this review, the history of biological weapons, their intrinsic link to bioterrorism and classification criteria, the epidemiological investigation of bioterrorism events (supported by data), and the measures taken against biological agents will be discussed³.

Historical Context

Numerous countries, including the United States and Soviet Russia, have initiated biological warfare (BW) programs. The use of biological weapons dates back to as early as the sixth century B.C., when the Assyrians reportedly contaminated water supplies with the fungus Claviceps purpurea⁴. More recently, mycotoxins (fungal toxins) have been reported to have been used in Afghanistan in the form of what became known as “yellow rain.” In September 1981, the U.S. The Secretary of State accused the Soviet Union of supplying mycotoxins to its Vietnamese and Laotian communist allies for military use against resistance forces in Laos and Cambodia, and of deploying the same agents in combat operations in Afghanistan⁵. If these allegations were true, it would mark the first instance of toxins being used in biowarfare. The U.S. further suggested that the Soviet Union was violating the 1925 Geneva Protocol and the 1972 Biological and Toxin Weapons Convention. These accusations and the evidence presented sparked considerable debate, with some researchers proposing alternative explanations for the so-called “yellow rain.”

The American BW program was clandestinely initiated in 1942 under President Franklin Roosevelt, following suspicions of BW agent use by Germany and Japan⁶. In 1947, President Harry Truman, with Senate approval, withdrew the U.S. from the Geneva Protocol and commenced open-air field testing of non-pathogenic bacteria (such as Bacillus globigii and Serratia marcescens) on naval vessels near the Virginia Coast and San Francisco Bay⁷. This also included the dissemination of bacterial aerosols in bus stations and airports. At its peak, the American BW program engaged nearly 3,400 personnel and involved several research and production facilities. The program investigated over 30 agents, including bacteria (e.g., Bacillus anthracis, Brucella suis, Coxiella burnetii, Francisella tularensis), toxins (e.g., botulinum toxin, staphylococcal enterotoxin B), and viruses (e.g., Venezuelan equine encephalitis virus, yellow fever virus)⁶.

Bioterrorism Agents 
A biological weapon differs by causing disease and death by reproduction disease-causing organisms in the infected organism of humans, animals or plants³. The other used terminology is ‘’biological agent’’. Furthermore these agents are separated to 4 depending on types which are viruses, bacterial, toxins and chemicals. As seen in the Table 1 the biological agents can be considered not only biological based, but also the chemicals can be used as weapons.

Table 1. Comparison of biological and chemical agents used as weapons.

Virus	Bacteria	Toxin	Chemical
Variola virus(small pox)	Bacillus anthrax (anthrax)	Staph entero-B	Mustard Gas
Viral hemorrhagic fever (ebola)	Francisella tularensis (tularemia)	Botulinium neurotoxin	Tabun Gas(GA)
Viral encephalitis	Brucella spp (Brucellosis)	T-2Mycotoxin	Sarin Gas(GB)
Crimean-Congo hemorrhagic fever	Vibrioc cholerea (cholerea)	Shiga toxin	Soman(GD)
Rift vailey fever	Burkhoideria pseudomail ei (glanders)	Ricin	Choking-agents:chlorine gas,phosgene,diphosgene and chloropicrin
Tick-borne encephalitis complex	Coxiella burnetti (Q fever)	Clostridium perfringens epsilon toxin	Cyandies
Yellow fever		Abrin	Vesicants: Sulphur mustard, Lewisite[dichloro(2-chlorovinyl) arsine] and nitrogen mustard

Viral Hemorrhagic Fever Virus

Viralhemorrhagic fever virus (VHF) refers to severe febrile illness with abnormal vascular regulation, vascular damage, and hemorrhagic manifastation that are caused by several viruses of different familias⁹. They are all lipid-enveloped RNA viruses and they require an animal or insect host reservoir¹⁰. Human disease is sporadic and usually follows accidental exposure to contaminated saliva, urine or feces of infected animals, insect bites or occasionally from human to human due to exposure to contaminated tissue or body fluids¹⁰. While humans are considered to be accidental hosts, many arthropods and rodents serve as effective reservoirs for the virus transmission⁹.

Venezulen Equine Encephalitis VirusVenezuelan equine encephalitis virus complex (VEE) refers to mosquito-borne α-viruses that cause human disease in Central America, Mexico and occasionly in the United States¹⁰.The virus is transmitted by the bites of  mosquitoes, which is infected by feeding on resorvoirs that contain high levels of viremia. The viremia process is produced in the salivary glands, and then, the viruses’ passage to new reservoirs during the feeding of the vector¹¹. Infection with Western equine  encephalitis  or Eastern equine encephalitis virus is clinically  indistinguishable  from VEE; however, VEE is more likely to be candidate because of  the lower human infective dose¹⁰.In humans early symptoms of VEEV infection include flu-like symptoms, such as fever, chills, maleise, severe headache, myalgia in the legs and lower back, tachycardia, and in some cases, nausea, vomiting, and diarrhea. If there are neurological signs, they can include seizures, drowsiness, confusion and photophobia¹¹.

Smallpox (variola)

Smallpox (variola) virus is one of the most dangerous viral potential bio-weapons. Smallpox is a severe viral infection that, until recently, caused significant human disease. So, it was a highly feared, disfiguring disease known to humans for thousands of  years ,until its eradication in 1980¹². Smallpox virus is known  to infect only humans. Therefore, it can be transmitted human to human however, it can not be observed that transmission from animal to human. Smallpox is a highly  contagious infection: it is only slightly  inferior to  measles and chickenpox in this parameter^12,13. Human neurological symptoms of  smallpox are headache, backache (prodrome) and viral encephalitis.Headache is quite commonly observed. Smallboxes have double-stranded DNA and large,asymmetric , brick-shaped  virions that encode unique enzymes that allow viral replication in the cytoplasm  of infected cells¹⁰.

Stealth Viruses

These consist of cryptic viral agents bearing potential human oncogenes that can be illicitly or secretly transferred to human genomes. Usually, they remain dormant for many years but exposure to a single natural stimulus can activate oncogenic determinants present on the stealth viruses and could cause vast destruction in the human population. For example, human herpes virus can cause oral and genital lesions after induction. Similarly, people who have contracted chicken pox previously present a natural reservoir of varicella virus that sometimes rejuvenates in the form of herpes zoster virus causing shingles disease in some people¹⁴. 

Bacteria
During the 19^th century, when modern microbiology saw great advancements, ironically it was the same time when the use of infectious bacteria to terrorize populations was first observed and predominantly established as a warfare agent. Hence, the lack of concern at that time, along with simultaneous advancements in the field of bacteriology, made bacteria a preferred and unique choice among biological warfare agents¹⁵. Bacteria commonly live in dense, multispecies,communities where there is competition over scarce source¹⁶. As shown in Table 1, there are various bacteria which are used as biological weapons. Furthermore some of the important bacteria from Table 1 will be mentioned in this part such as; Bacillus anthracis, T.pestis and Brucellas suis.

Bacillus anthracis (B.anthracis)

A German physician Robert Koch discovered Bacillus anthracis (B.anthracis) in 1876⁸. It is  the one of the most dangerous species of the bacteria class, a potential biological agent.In the fashion World, there has been an increasing demand for animal products like wool, leathers, hides and hair products. These biological agents infect the environment by contamination of the mentioned areas. The bacteria spor enters through abraded skin during handling of the abovementioned contaminated products and causes infection by cutaneous anthrax. B. anthracis is the main causative factor for this zoonotic disease¹⁵. In the human, When B. anthracis spores are inhaled, alveolar macrophages and dendritic cells transport the organism to the proximal lymph nodes. Bacteremia begins after an asymptomatic incubation period of 1 to 6 days¹⁶. Following the onset of bacteremia, patients can experience flu-like symptoms for 1 to 5 days prior to an acute disease stage that lasts 1 to 2 days¹⁷. Initially there is acute inflammation in the intestinal tract characterized by nausea, vomiting, loss of appetite, and fever, which may be followed by severe diarrhea and vomiting blood¹⁵.

Yersine pestisOne of the common hazardous bacteria affecting human beings,Y. pestis, is transmitted with flea bites and eventually causes plague-it even sometimes causes fatal bubonic plague^18,19.The clinical symptoms of plague include fever, chills, weakness, headache, tender lymph nodes etc¹⁵. Other types of plague include pneumonic plague, infected via respiratory droplets from infected individuals or animals¹⁵. Pneumonic plague is the most likely target for bioterrorists in the form of aerosolized Y. pestis, which may cause sudden severe pneumonia and sepsis¹⁵. Symptoms of primary pneumonic plague typically begin between 2 and 4 days after inoculation and include a productive cough with bloody sputum, fever, headache, malaise, nausea, and vomiting¹⁹. Though rare, multidrug resistant strains of Y. pestis have been isolated, highlighting the need for research focused on identifying novel therapeutic targets^20,21.

Brucellas suis

One of the neglected zoonotic bacterial disease is caused by  Brucella suis¹⁵. Brucella suis is an emerging, zoonotic disease predominantly affecting dogs and humans that engage in feral pig hunting in Australia and other countries. Although B. suis infection in dogs shares some clinical similarities to the host-adapted species (B. canis), B. suis remains an incompletely understood pathogen in dogs with limited published data on its pathogenesis and clinical featurest²².Other livings that it generally chooses as hosts are cattle, buffola, goats and horses. It is a facultative intracellular, gramnegative pathogen, transmitted to the human population via dairy products. General symptoms of brucellosis include fever, body pain, especially muscle and joint pain, splenomegaly, etc¹⁵. In Australia, dogs and humans become infected with B. suis following contact with body fluids and/or tissues from infected feral pigs, making hunting and butchering of feral pigs and feeding of uncooked pig meat the most common risk factors for infection^23,24. Being a neglected disease nowadays, it is an emerging target for the bioterrorist²⁵. Even an aerosol form of brucellosis is presently considered to be a dangerous biothreat as it causes respiratory brucellosis²⁶.

Biological Toxins

Toxins are extremely poisonous products of the metabolism of living organisms like bacteria, plants, animals, and fungi²⁷. Most toxins are based biological.These molecules induce detrimental effects in other organisms by inhalation, injection, ingestion or absorption²⁸. Many of them affect the nervous system, disrupting the conduction of nerve impulses, for example by blocking the release of the acetylcholine neurotransmitter and ultimately the muscle contraction (botulinum toxin). At sublethal doses, the effect that this causes is temporary. Others toxins cause damage to cell membranes, and as a result lead to disturbances in the functioning of tissues and organs. The effects of such toxins are often irreversible and cause permanent damage to health²⁹.In this part, some of the biological toxins will be mentioned with their characteristics; botulinum neurotoxins, ricin and abrin.

Botulinum neurotoxins

Botulinum toxin is called the most toxic substance in the known world. Botulinum neurotoxins (BoNTs) are produced by the spore-forming, anaerobic Gram-positive bacteria of the genus Clostridium which consists more than 150 species. They lead to a disease called botulism. BoNTs comprise 7 serotypes (A to G) and more than 40 subtypes. However, A, B, C, E and F serotypes are responsible for inducing human botulism²⁹.

The mechanism of the toxic action of botulin toxin inhibits the release of neurotransmitters, including acetylcholine, within neuromuscular junctions, resulting in relaxation and paralysis of skeletal muscles. The first symptoms occur within a few to 36 h of poisoning with the toxin. Irrespective of the type, there are similar clinical manifestations of poisoning. Initially these are speech and swallowing difficulties, double and blurred vision, anxiety, lack of saliva and tears. There then follows a loss of control over the body, and atrophy of the throat reflex appears. Respiratory muscle paralysis then causes respiratory failure, and this is the main cause of death of infected patients ^30-32.

During a bioterrorist attack, botulinum toxin may be deployed in aerosol form or by contaminating water and food. The efficacy of the neurotoxin is equally high regardless of route of entry, due to having almost identical disease symptoms. A bioterrorist attack with botulinum neurotoxin is difficult to identify. Essentially, only an increase in the number of people with symptoms of toxin poisoning could indicate its use in a bioterrorist attack^14,33.

Ricin

Ricin toxin isolated from the castor bean (Ricinus communis) is one of the most potent and lethal molecules known³⁴. The castor plant grows wild in tropical and subtropical climates and the plant is cultivated in large scale for the commercial production of castor oil, a major component of the castor bean, with almost two million tons of seeds harvested annually²⁹. Ricin, a byproduct of castor oil, is a real threat for bioterrorism and for biological warfare, especially when dispersed by aerosol³⁴. Ricin can be used in acts of bioterrorism in several forms, as it has many ways to penetrate the organism. In addition,it can be used as an aerosol and by poisoning water and food. The toxicity of ricin depends on different factors, such as the route of entry into the body, the dose and species²⁹. Ricin inhalation causes acute lung injury characterized by a massive inflammatory response³⁴.

Ricin is also characterized by its stability in the environment and relative ease of extraction. Despite having a well-understood mechanism of action, treatment is still based on supporting the organism, which in a bioterrorist attack with a high casualty count, can cause chaos in the local healthcare system. Therefore, more research is needed to be more accurate. Due to its ease of production, good stability, availability and high lethality, ricin exhibits potential as a biological weapon. Furthermore, ricin is accepted according to a report of US Centers for Disease Contral (CDD) as the second highest priority agent²⁹. 

Tetrodotoxin

Tetrodotoxin (TTX) is one of the most lethal toxins in the marine environment. It is a naturally occurring toxin responsible for human fatalities and intoxication. The name of TTX was established after the Tetraodontidae family of fish. In Japan fugu, or puffer fish, is known for its potential for TTX toxicity²⁷. It is considered that more than 20 species of puffer fish is housed to the toxin.Besides puffer fish, there are several livestock species which are gastropods, clab, blue-ringed,octopuses, ringworms. Moreover, tetrodotoxication is still an important health problem today, as TTX has no known antidote. TTX poisonings were most commonly reported from Japan, Thailand, and China, but today the risk of TTX poisoning is spreading around the world. Recent studies have shown that TTX-containing fish are being found in other regions of the Pacific and in the Indian Ocean, as well as the Mediterranean Sea³⁵.  Bacteria are responsible for production of TTX. However, the quantity of TTX produced by these bacteria is very low. Despite low toxin production by bacterial strains in laboratory conditions, even minimal amounts of TTX produced by intestinal microflora of an animal can contribute to its toxicity²⁷. 

TTX poisoning results in inhibition of neurotransmission as it blocks the voltage gated sodium channels and thus effects both action potential generation and impulse conduction, resulting in a blockade of the neuron action potential and in muscle paralysis. Little information is available about the absorption and excretion of TTX and its analogs in humans. Raw or boiled tissue extracts of L. lagocephalus exhibited hepatotoxic and nephrotoxic effects in rats. TTX also effects the respiratory system. An i.v. injection of TTX in rats leads to respiratory failure, due to paralysis of the respiratory muscles, which were apparently more susceptible to the action of tetrodotoxin than the respiratory and other motor nerves²⁷.

 Chemicals 

The use of toxic chemicals in warfare is as old a srecorded history. Chemical Weapons (CWs) have recently come under the international spotlight.They have been repeatedly used, to tragic humanitarian effect, in the Syrian Civil War. In February 2017, a nerve agent was used to assassinate the half-brotherof North Korean leader Kim Jong-unina Malaysian airport. In March 2018, a chemical agent was used in anassassination attempt targeting Sergei Skripal, a Russian defector living in the UK, and inadvertently poisoned several others.These incidents highlighted the risk posed by CWs and fuelled fears that these weapons are making a come back on the international stage³⁶. Cyanide substance is one of the common used chemicals as weapons. It is mentioned in this review;symptoms that it causes, signs.

Cyanide 

Cyanide refers to a highly toxic chemical compound containing one atom of carbon and nitrogen each³⁷. Cyanide poisoning, whether it be accidental or intentional, remains a major threat to civilians and military personnel worldwide. It is readily available, highly lethal, and easily weaponized. These low-dose exposures frequently cause headache, dizziness, mild confusion, abdominal cramping, nausea, and vomiting. Exposure to even  small amounts of cyanide can cause some symptoms. of cyanide Large-dose exposures eventually lead to dyspnea, respiratory depression, apnea, hypotension, arrhythmias, coma, and seizure³⁸. These large-dose effects can result in irreversible injury and death within minutes of the onset of symptoms³⁹.

Classification of Biological Agents

Many microorganisms can be considered as potential biological weapons (bio-agents). The biological weapons as per the CDC classification are classified into three categories, category A, B, C is given in the Table 2; based on agents’ features and  risk factors that they could make.

Table 2.  Classification of biological agents⁴.

Category A	Category B	Category C
High priorty agents include organisms that pose a risk to national security because they are: – Easily dissemanited  – Cause high mortality  – Cause public panic and social distruption  – Require special action for public health prepardness	Second highest priorty agents include those that are: – Moderately easy to disseminate – Cause moderate morbidity – Require enchanced disease surveillance and public health diasgnostic capacity	Third highest priorty agents include emerging pathogens                             – That could be engineered for mass dissemination in the future – Have potantial for high morbidity, mortality and major health impact

The Centers for Disease Control and Prevention (CDC) categorizes bioterrorism agents according to their threat and priority levels. Highest priority Category A agents include: Anthrax, Botulism, Plague, Smallpox, Tularemia and Viral Hemorrhagic Fevers. Category B agents include: Brucellosis, Epsilon Toxin of Clostridium Perfringens, food safety threats such as Salmonella, E.coli and Shigella, Glanders, Melioidosis, Psittacosis, Q Fever, Ricin, Staphylococcal Enterotoxin B, Typhus fever, Viral Encephalitis and water safety threats such as Vibrio Cholerae and Cryptosporidium Parvum. Category C agents, the third highest priority agents include emerging pathogens that could be engineered for mass dissemination in the future because of availability, ease of production and distribution, and potential for high morbidity and mortality rates. Emerging infectious diseases such as Nipah virus and Hantavirus are currently classified as Category C. In table 3, it is shown that Bacillus antrachis is the highest priorty to pose risk.Thus, it is in Category A. Due to its extremely pathonegic potantian and sporulation, anthrax is a top candidate for use in bioterrorist attacks. In 1970, the World Health Organisation (WHO) estimated that between 130,000 and 3 million individuals may die following the introduction of 300 kg of B. antrachis in 1993¹.This is just as deadly as a hydrogen bomb. According to research from animals,the human LD50 (a dose that kills %50 of infected people) is estimated to be between 2,500 and 55,000 inhaled spores⁸.

Table 3. Agents of bioterrorism.

Category A Agents	Category B Agents	Category C Agents
Bacillus antrachis (anthrax)	Alpha viruses	Hanta viruses
Clostridium botulinum toxin(botulism)	Eastern and western equine encephalomyelitis viruses (EEE, WEE)	Multidrug-resistant tuberculosis
Francisella tularensis (tularemia)	Venezuelan equine encephalomyelitis virus (VEE)	Nipah virus
Veriola majör (smallpox)	Brucella species (brucellosis)	Tickborne encephalitis viruses
Yersinia pestis (plague)	Burkholderia mallei (glanders)	Tickborne haemorrhagic fever viruses
Filo viruses	Coxiella burnetii(Q fever)	Yellow fever
Ebola virua (Ebola hemorrhagic fever)	Epsilon toxin of Clostridium perfringens
Marburg virüs (Marburg hemorrhagic fever)	Ricin toxin from Ricinus communis
Arena viruses	Staphylococcal enterotoxin B
Junin virüs (Argentinian hemorrhagic fever) and related viruses
Lassa virüs (Lassa fever)

Biological Attacks with Analysis

It is quite significant to understand and define the concept of biological attack. The biological attack is essentially a perverted misuse of all knowledge of nature, causes, distribution, suppression and prevention of naturally occurring infections and poisonings³. Bioterrorist attack could be caused by virtually any pathogenic microorganism. However, microorganisms (like virus, bacteria, fungi or toxins) to be effective as a bioterrorist agent should consistently produce a given effect, death or disease, at low concentration⁴. Many lethal microorganisms used in biological attacks are mentioned.These microorganisms are listed as Table 3. Twenty of the attacks involved anthrax, 5 involved salmonella, 3 involved ricin, 2 involved faecal matter, 1 involved botulinum toxin, 1 involved the use of HIV infected razor blades and 1 involved either ricin or anthrax. Seven of the recorded deaths were linked to anthrax attacks and 2 remaining deaths were related to salmonella incidents. Of the 806 injuries reported, 776 were related to 2 attacks involving salmonella, 25 were related to anthrax events and 1 was related to an event involving faecal matter as a biological agent (Fig. 1)⁴⁰.

Figure 1.  Breakdown of biological agents used in bioterrorism attacks.

Clear and detailed documentation of terrorist events is further hindered by restrictions on reporting, the lack of independent corroboration and the lack of transparency within certain government sources. Infrastructure needed to report, detect and identify biological agents is likely lacking in many parts of the world leading to potential under reporting of events. The greater number reported in the United States may be due to greater scrutiny of potential events. Event descriptions as provided by the Global Terrorism Database(GTD) rely mostly on grey literature sources and as such accuracy is only limited to those sources. Bioterrorism attacks have been historically rare but have the ability to inflict large-scale, mass casualty events. Anthrax has been most commonly used in previous bioterrorism events with the vast majority of reported attacks occurring in the United States by a single suspected perpetrator. With new advances in microbiology and synthetic biology, it is becoming increasingly possible for individual or small groups of rogue actors to develop and disseminate advanced bioweapons. While the Covid19 pandemic has likely raised the awareness levels of first responders to biothreats, it has also exposed response and preparedness vulnerabilities in the healthcare sector. Counter-Terrorism Medicine and Disaster Medicine specialists need to be proactive in delivering ongoing educational sessions on biological events to first responder communities, and anticipate emerging novel biotechnology threats⁴⁰.

Environment and Public Health

Biological weapons cause catastrophic effects on public health, biodiversity and the environment¹. The technical knowledge and materials needed to produce the biological weapons is available, however the knowledge about targeting these materials is limited. However, precautionary and preventive measures need to be taken considering their chances of being used¹.

A 1970 WHO study estimated that 50 kg of Bacillus anthracis released over an urban population of 5 million would sicken 250,000 and kill 100,000 people, and a 1993 Office of Technology Assessment (OTA) study estimated that between 130,000 and 3 million deaths would follow the release of 100 kg of B. anthracis. This data reflects that bioterrorist attacks pose some major threats to the public health of any nation. Firstly, most of the countries in the world are dealing with developing their public health infrastructures. In such a situation a bioterrorist attack will add to this existing burden. Secondly, it is very difficult to identify and differentiate a bioterrorist attack from a natural disease outbreak. Thirdly, most of the drugs and vaccines have a limited shelf life and hence cannot be stocked up. In such a situation, the countries might not be in a situation to provide effective and immediate remedy to the public in case of a possible bioterrorist attack. The outcome of such attacks depends on the preparation of a country to deal with it. Lastly, unorthodox use of bioweapons may result in disease with unexpected symptoms and epidemiology. Until 2001, the possibility of cross-contamination of anthrax through the mail was discounted by most experts¹.

As mentioned, bioterrorism and its derivatives cause huge destruction to the environment and public health. So many of the instances mentioned above to you,the reader, explain this situation. Moreover, these destructions lead to irreversible consequences. Permanent and definitive solutions are essential for bioterrorism and the agents used.

Preventing the Bioterrorism Attack

In the previous part, we have mentioned about the importance of the environmental and public health against to biological attacks. Environment and public health refer to developing strategies and methods agains to potentially dangerous-biological and chemical based-attacks on each element that constitutes it(human, animal, plant etc.) The responsibilities of public health agencies are surveillance of infectious diseases, detection and investigation of outbreaks, identification of etiologic agents and their modes of transmission and the development of prevention and control strategie⁴. There are 5 essential phase that need to be followed for succesful response to a bioterrorist attack⁴.

(a)Preparedness Phase: This phase includes actions to be taken by different agencies to ensure the required state of preparedness. These include evaluation of the laboratory facilities and upgrading the same, evaluating the hospital preparedness in emergency response and case management in case of an imminent attack, conduct training of health professionals, rapid response team (RRT) and quick response medical team (QRMT) who would be the first responders, work out the legal provision and their implications, ensure that requirement of safe drinking water is met, ensure availability of adequate stocks of medicines and vaccines, coordinate with security organization, organize mock drills for health professionals, government departments, animal husbandry, security, law enforcing and other agencies so as to assess their preparedness levels to act in case of an attack, prepare contact details so that communications is unhampered during an attack. Public should be kept aware about imminent attacks so that voluntary reporting is encouraged. It is important to carry out a review of the situation based on current information of threat perception⁴.

(b) Early Warning Phase:

The early warning in the surveillance system includes activities like case definitions, notification, compilation and interpretation of epidemiological data. Early detection and rapid investigation by public health epidemiologists is critical in determining the scope and magnitude of the attack and to implement effective interventions⁴.

(b1) Detection of Attack

Deduction phase is considered a subgroup of early warning phase.In case of any attack, biological agents’ data need to be measured and detected by unique equipment to improve new strategies and methods against agents. These equipment are separated depending on method.Sensors constitute a large part of these tools. A sensor is defined as a device that is able to detect the presence of an analyte in a sample and quantify it⁴¹. The sensor consists of a detection system called a receptor, a transducer, and a readout system⁴¹. When evaluating sensors,the following features should be taken into consideration;selectivity, sensitivity,and repeatability⁴¹. Selectivity is the ability of the sensor to distinguish between the sample and other materials. High sensitivity refers to the capacity of sensor’s detection to the minimal changes in analyte concentration. Repeatability is important to ensure accuracy. While the biological layer is responsible for the specific interaction with the analyte, the transducer converts the information from this interaction into a measurable effect^43-45. For example, mechanical transducers convert the analyte bioreceptor interaction to a change in bending or resonant frequency, optical transducers usually convert this phenomenon to a change in light frequency or intensity, and electrochemical transducers to a change in current, potential, and so on. In the result, the reading system measures these changes. A biochip also includes an array of single biosensors that can operate separately and are commonly used to detect multiple analytes simultaneously⁴¹. In this part, the electrochemical biosensors will be mentioned.

Electrochemical Biosensors

The electrochemical method utilized is a distinguishing aspect of an electrochemical biosensor. In addition to the electrochemical method, the sample handling approach and sensor signal readout format also provide distinguishing aspects of a biosensor-based approach for pathogen detection⁴⁵.

Electrochemical methods of analysis are based on measuring current changes, potential, conductivity characteristics between two electrodes, impedance and field-effect. These techniques are divided into two categories: static (with zero current) and dynamic (with current passing). Static techniques include direct potentiometry and potentiometric titration, and dynamic techniques include potentiostatic (with potential control) and galvanostatic (with controlled flow) methods⁴¹. Electrochemical measurement systems usually consist of two control electrodes and a detector. 

(c) Notification Phase:It is mandatory to report any unusual syndrome or usual syndromes in unusual numbers to appropriate authorities. The activities in this phase include rapid epidemiological investigations, quick laboratory support for confirmation of diagnosis, quarantine, isolation, keeping health care facilities geared for impending casualty management and evolving public health facilities for control⁴.

(d) Response Phase: In this phase the activities include rapid epidemiological investigation, quick laboratory support, mass casualty management and initiation of preventive, curative and specific control measures for containing the further spread of the disease.In order to achieve them, following steps can be followed⁴.

i- Assess the situation: Initiate the response by assessing the situation in terms of time, place and person distribution of those affected, routes of transmission, its impact on critical infrastructure and health facilities, the agencies and organizations involved in responding to the event, communicate to the public health responders, local, state and national level emergency operation centres for event management etc⁴.

ii- Contact key health personnel: Contact and coordinate with personnel within the health department that have emergency response roles and responsibilities. Record all contacts and follow-up actions⁴.

iii- Develop action plan: Develop initial health response objectives that are specific, measurable and achievable. Establish an action plan based on the assessment of the situation. Assign responsibilities and record all action⁴.

iv- Implementation of the action plan: The RRTs/ QRMTs investigate the outbreak /increase in the disease incidence, collect samples and send it to the identified state/national laboratory for testing. Hospitals are alerted for receiving the patients and their treatment. If necessary tented hospitals are set up. Methods to control the disease and quarantine measures are instituted. Once the disease is identified, treatment protocols are sent to all concerned by the fastest possible means. Standard operating procedures (SOP) for laboratory testing is made by the identified laboratory and the same is sent to all the hospital laboratories and district hospitals for implementation. Laboratory reagents are distributed to the concerned laboratories. Public is taken into confidence to prevent any panic. The list of ‘Do’s and Don’ts’ are circulated thorough the print and electronic media. Hospitals ensure appropriate isolation, quarantine, waste disposal and personal protective measures. All contaminated clothing and equipment are carefully disposed of by incineration. An impact assessment team assesses the impact of the attacks on humans, animals and plants⁴.

(e) Recovery Phase: The setbacks suffered as a result of the bioterrorist attack are restored and lessons learnt in this phase are incorporated in the future preparedness plans. The damage done to the public health facilities and the essential items utilized during the response phase are replenished. Public advisories are issued regarding restoration of normalcy. The RRTs compile and analyze data to identify the deficiencies experienced in the implementation of the response measures. The necessary modifications are then incorporated in the contingency plan for future⁴⁶. 

Relationship with Artificial Intelligence (AI) and Future Perspective 

It is widely known that there is no discipline that artificial intelligence has not reached and it is not involved.The future perspective of bioterrorism is infinitely to discuss. Therefore, this section focuses on looking to the future and collusion of humanity with AI in the war against bioterrorism. In this section, it will be briefly discussed how artificial intelligence technologies are used in the detection of biological agents and how strategies are applied to achieve successful results.

AI technologies that can be used to aid the planning and perpetuation of biological terrorism⁴⁷. AI has the ability to predict and track the spread of viruses. To do this, the artificially intelligent system is able to figure out where clusters of disease begin to form and how they travel. In the same way, AI can locate areas in cities or cities themselves where certain diseases are more prominent. This will help determine the kinds of biological agents that will be most effective in these areas. Information or data about the spread of disease is essential to the effective and efficient dispatching of treatment, aid allocation, and more⁴⁸. For instance, a few different COVID-19 tracking apps, hereafter referred to as “CTAs”, have been developed in an effort to curb the spread of the virus⁴⁹. It can be mentioned that two main processes to summarize the role of artificial intelligence in war against bioterrorism; the data mining process and predictive analysis. 

The Data Mining Process 

The data mining process consists of 5 processes. The first step the data mining process is data cleaning. Data cleaning refers to an indepth scrubbing of the data that has been collected from various sources. Scrubbing refers to the removal of noise from data.However, noise can be added to a data set to prevent someone from mining and misusing the collected data. This is one available method we can use to protect the sensitive information that we have stored. The second step is data integration. This step of the process is used to compile all the relevant data, note the use of the term relevant, that has been cleaned in the previous step. There is no longer any noise or irrelevant information which does not suit the purpose or intent of the process⁴⁷. The other step is data transformation. This step in the data mining process occurs after the various forms of the collected data have been stored together in the same location. There is a shift away from articles, pictures, vlogs, news reports, and recordings, to a single location where all this information can be stored.  An interesting aspect of data transformation is that, while it may seem harmless, it can pose real dangers.For this reason, the data transformation process carries a couple of risks. “The Boiling Frog” analogy specifically applies to data transformation and broadly to data mining as a whole⁴⁶. To be brief, there are two ways to boil a frog with differing levels of success⁴⁷. One option is to toss the frog into the water when it is already boiling. However, the outcome of this is that the frog will immediately jump out of the pot to save its own life⁴⁷. The second option is to put the frog in the water while it is cold and heat the pot thereafter⁴⁸. The temperature will increase slowly, alarming the frog only once it is too hot and the damage has been done. The fourth step is data mining; it is a fundamental element of the process. The step after collecting and storing data is to make sense of it.Knowledge representation is the last step. The data, at this point, has been cleaned, integrated, transformed, and mined. It is ready to be represented to highlight the results or outcome of the process⁴⁹.

Predictive Analysis

After collecting, transforming and storing data, predicting data and developing strategies is the first step to follow. The collection of medical data poses a risk should the data be mined and used to perpetrate an attack on a particular area or group of people. Biological warfare is something that can be achieved even on a small scale, even when there is no clear war going on around us. Biodefense refers to defensive strategies and methods that can be taken against the threat of a biological attack, whether it be weapons or biological agents⁴⁷. 

Predictive analysis, just as is the case with data mining, has clear benefits while posing grave risks and potential consequences. As has been stated, while we enjoy the benefits of mining data, big data, and predictive analysis, the potential implications for AI are far-reaching and pose enough of a risk to warrant further investigation. The concern here lies in the preservation of privacy within big data and data mining, so that it is possible to reap the rewards of data collection without compromising the safety of private or personally identifiable information⁴⁸.

Through AI, governments and healthcare facilities can conduct contact tracing. The AI system can “augment mobile health applications where smart devices like watches, mobile phones, cameras, and an entire range of wearable devices can be employed” states that there are a multitude of ethical concerns regarding contact tracing and the surveillance that it involves, namely a) privacy, in that people are being tracked, which leads us to the next issue, b) voluntariness, where the voluntariness of the participant must be taken into account, c) beneficence, the concern for who benefits from the data, and d) incentives, referring to the influence of incentives on citizens⁵⁰.

Conclusion 

In this review, some of the terminologies related to bioterrorism were mentioned; biological agents, biological attack, environment and public health.These terminologies help  understand the concept of bioterrorism. As we mentioned, biodefense refers to all the methods and strategies developed and applied against biological attacks. 

Bio-warfare is a concept that has been emphasized and has been at war with humanity since  ancient times.The highly unpredictable nature of any event involving biological warfare agents has given rise to the need for developing rapid and accurate detection systems. The bioterrorist events are difficult to predict and prevent; in the case of a release, accurate, easy deployable detection systems are needed to minimize the damage and to prevent further spread of these agents. By developing artificial intelligence and its technologies, humanity has a stronger trump card against biological attacks and weapons. Nevertheless, it should not be considered that artificial intelligence has a nature that will completely eliminate bioterr noorism applications. In addition to this, the individual who exists in the sociological order must be conscious in bioterrorism on behalf of itself and society. 

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