technical aspects of biological weapon proliferation - federation of

TechnicalAspects ofBiological

WeaponProliferation 3

B iological and toxin warfare (BTW) has been termed“public health in reverse” because it involves thedeliberate use of disease and natural poisons to incapac-itate or kill people. Potential BTW agents include Living

microorganisms such as bacteria, rickettsiae, fungi, and virusesthat cause infection resulting in incapacitation or death; andtoxins, nonliving chemicals manufactured by bacteria, fungi,plants, and animals. Microbial pathogens require an incubationperiod of 24 hours to 6 weeks between infection and theappearance of symptoms. Toxins, in contrast, do not reproducewithin the host; they act relatively quickly, causing incapacita-tion or death within several minutes or hours.

The devastation that could be brought about by the military useof biological agents is suggested by the fact that throughouthistory, the inadvertent spread of infectious disease duringwartime has caused far more casualties than actual combat.1 Suchagents might also be targeted against domestic animals and stapleor cash crops to deprive an enemy of food or to cause economichardship. Even though biological warfare arouses generalrepugnance, has never been conducted on a large scale, and isbanned by an international treaty, BTW agents were stockpiledduring both world wars and continue to be developed as strategicweapons— “the poor man’s atomic bomb’’—by a small butgrowing number of countries.2

1 John P. Heggers, “Microbial Lnvasion-The Major Ally of War (NaturalBiological Warfare),” Military Medicine, vol. 143, No. 6, June 1978, pp. 390-394.

2 This study does not address the potential use of BTW agents by terrorist groups.For a discussion of this topic, see U.S. Congress, Oftlce of lkchnology Assessment,Technology Against Terrorism: The Federa/ Ej@rt, O’IX-ISC-481 (Washingto~ DC:U.S. Government Printing Office, July 1991), pp. 21-22. See also Jessica Eve Stem,“Will lkrronsts Tbrn to Poison?” Orbis, vol. 37, No. 3, summer 1993.

I 71

72 | Technologies Underlying Weapons of Mass Destruction

The Biological and Toxin Weapons Conven-tion of 1972, signed and ratified by some 130countries, bans the development, production,stockpiling, and transfer of BTW agents forwarfare purposes. This treaty was weakened fromthe start, however, by the impossibility of ban-ning research on BTW (agents, the fact that thedevelopment, production, and storage of BTWagents are permitted for defensive or peacefulpurposes, and the absence of formal mechanismsfor verification or enforcement.3 It has also beenalleged that key signatory states such as theformer Soviet Union have systematically violatedthe treaty. According to a recent White Housereport, ‘‘the Russian offensive biological warfareprogram, inherited from the Soviet Union, vio-lated the Biological Weapons Conventionthrough at least March 1992. The Soviet offensiveBW program was massive, and included produc-tion, weaponization, and stockpiling. ’

The biological disarmament regime has alsocome under growing pressure from the globalspread of biotechnologies suitable for both civiland military applications, and from the revolutionin genetic engineering, which has made it possi-ble to manipulate the genetic characteristicsencoded in the chemical structure of the DNAmolecule. Soon after the publication in 1973 oftechniques for cutting and splicing DNA mol-ecules across species lines, a few concernedscientists worried that these powerful methodsmight be applied to develop new and moredangerous biological-warfare agents. Today, somedefense planners believe that genetic engineeringand other biotechnologies may eventually removesome of the military liabilities of BTW agents,increasing the attractiveness of these weapons tostates of proliferation concern. It is not clear,however, that such techniques would signifi-

cantly alter the military utility of BW agentscompared with the numerous already knownagents.

Given the perceived need to strengthen theBiological Weapons Convention (BWC), and thefact that the Chemical Weapons Convention(CWC) includes formal verification measuressuch as onsite inspections, a number of countrieshave proposed establishing a similar verificationregime for the BWC. (See box 3-A, pp. 74-75.See also ch. 2 for discussion of procedures andtechnologies that might be used to detect theproduction of chemical weapons.) Nevertheless,verifying the nonproduction of biological weap-ons is inherently more difficult than for chemicalweapons, for three reasons.

First, since BW agents are living microorgan-isms that reproduce inside the host, they are muchmore potent per unit weight. Thus, whereas CWagents must be stockpiled in the hundreds orthousands of tons to be militarily significant, afew kilograms of a BW agent such as anthraxbacteria could cause comparable levels of casual-ties. Such a small quantity of agent would berelatively easy to hide.

Second, whereas the production of CW agentsrequires certain distinctive precursor materials,reactions, and process equipment and leavesbehind telltale chemical signatures, the produc-tion of BW agents involves materials and equip-ment that are almost entirely dual-use. As a result,it can be extremely difficult to distinguish illicitBW agent production from legitimate activitiespermitted under the BWC, such as the productionof vaccines.

Third, because of the potency of BW agentsand the exponential rate of microbial growth, amilitarily significant quantity of BW agent couldbe produced in a matter of days in a small, easily

3 Some analysts worry that the Chemical Weapons Convention which was recently opened for signature and includes stringent verificationmeasures, may create incentives for some prolifemnt countries to acquire biological rather than chemical arms-both because BTW agents canbe produced in smaller, more concealable facilities, and because the Biological Weapons Convention cumently lacks effective verifkationmechanisms.

4 George Bus& “The President’s Report to Congress on Soviet Noncompliance With Arms Control Agreements, ” Jan. 14, 1993, p. 14.

Chapter 3-Technical Aspects of Biological Weapon Proliferation | 73

concealed clandestine facility. All of these factorsmake the verification of compliance with theBWC a particularly challenging task.

This chapter provides technical background onthe difficulty and detectability of BTW produc-tion and weaponization. The discussion coversthe major technical hurdles involved in theacquisition of biological weapons and the associ-ated ‘‘signatures’ that might be monitored totrack their spread.

SUMMARYAlthough biological and toxin weapons are

often grouped together with chemical weapons,they differ in important ways. The most obviousdifference is that whereas CW agents are man-made, nonliving poisons, biological agents areinfectious microorganisms that reproduce withinthe host to cause an incapacitating or fatal illness.Toxins, being poisonous chemicals manufacturedby living organisms, have characteristics of bothchemical and biological agents.

Because of the ability of pathogenic microor-ganisms to multiply rapidly within the host, smallquantities of a biological agent—if widely dis-seminated through the air-could inflict casual-ties over a very large area. Weight-for-weight,BTW agents are hundreds to thousands of timesmore potent than the most lethal chemical-warfare agents, making them true weapons ofmass destruction with a potential for lethalmayhem that can exceed that of nuclear weapons.The lengthy incubation period of microbial patho-gens places a major limitation on their battlefieldutility, except in situations of attrition warfare,sabotage attacks against command and communic-ations facilities deep behind enemy lines, orstrikes against massed troops prior to theircommitment to battle. Moreover, the delayedeffects of biological weapons would not preventtheir covert use against crops, livestock, or peopleas a means of crippling the economy and psycho-logical morale of a targeted country.

Biological and toxin weapons potentially posegreater dangers than either chemical or nuclearweapons because BTW agents are so lethal on apound-for-pound basis, their production requiresa much smaller and cheaper industrial infrastruc-ture, and the necessary technology and know-howare almost entirely dual-use and thus widelyavailable. Despite the drawbacks of biologicalagents for tactical military use (e.g., delayedaction, the dependence on meteorological condi-tions for their effectiveness, and the difficulty ofprecise targeting), they might be attractive as astrategic weapon-particularly for small, non-nuclear nations embroiled in regional conflicts orthreatened by a nuclear-weapon state.

One technical hurdle to acquiring a militarilysignificant BTW capability is to ensure adequatecontainment and worker safety during productionand weapon handling. It is also technicallydifficult to deliver biological agents to a targetarea so as to cause infection in a reliable andpredictable manner. Although a supply of stand-ard BTW agents for strategic attacks againstwide-area civilian targets (e.g., cities) would berelatively easy to disseminate using crude deliv-ery systems such as an agricultural sprayer, thismeans of delivery would be largely uncontrolla-ble and subject to shifting atmospheric condi-tions. A more predictable-and hence moretactically useful-means of delivery against pointtargets on the battlefield would require extensiveresearch, development, and testing. In particular,the integration of BTW agents into long-rangedelivery systems such as cluster bombs posescomplex engineering hurdles-although theseproblems appear to have been solved for a fewagents by the United States and the Soviet Unionduring the 1950s and 1960s.

There are no specific indicators, or “signa-tures,” that can differentiate unambiguously be-tween the development of offensive BTW agentsand work on defensive measures such as vaccines,since both activities require the same basicknow-how and laboratory techniques at the R&Dstage. Moreover, certain types of civilian facili-


Box 3-A—The Debate Over BWC VerificationThe Biological and Toxin Weapons Convention (BWC) of 1972 bans agents and delivery systems of types

and in quantities that have no justification for prophylactic, protective or other peaceful purposes," yet the treatydoes not define permitted activities more precisely and lacks any formal mechanisms for verifying compliance. Atthe time the BWC was negotiated it was considered politically impossible to obtain international support for onsiteinspections and other intrusive verification measures. Since 1972, however, the emergence of genetic engineeringand other novel biotechnologies has led to renewed concern over the seriousness of the biological and toxinwarfare threat. Given the dual-use nature of the agents and equipment the feasibility of effective verification hasbeen widely debated.

At the Second Review Conference of the BWC in 1986, the participating countries sought to build confidencein the treaty regime through an annual exchange of information on permitted activities and facilities that could bepotentially associated with biological and toxin warfare. Additional confidence-building measures were adoptedat the Third Review Conference in 1991. None of these measures are legally binding, however, and less than halfof the parties to the treaty have participated to any extent in the data exchanges. At the Third Review Conference,several countries supported the drafting of a legally binding verification protocol to supplement the BWC that, interalia, would require each Party to declare all treaty-relevant biological research and production facilities. Thedeclarations would be confirmed by routine onsite inspections, supplemented by challenge inspections ofundeclared facilities.

Proponents of a verification protocol argued that while a BWC verification regime could not provide absoluteconfidence in a country’s compliance, it would serve to deter the proliferation of biological and toxin weapons by:

● imposing a risk of discovery and increasing the cost and difficulty of a clandestine program;■ providing opportunities for parties to demonstrate compliance, and enhancing confidence in the compliance of

others;■ decreasing the number of sites of proliferation concern; providing an opportunity to act on national intelligence information without public disclosure of sensitive sources

and methods; creating a legal framework for the conduct of challenge inspections; and■ reinforcing the international legal norm against the acquisition and use of BTW agents.1

The Bush administration, however, opposed the negotiation of a formal verification protocol on three grounds: the BWC could not be verified effectively because biological production facilities are dud-useand Iack distinctive

“signatures”;● a negotiated regime could not be sufficiently intrusive to detect clandestine facilities, generating false confidence

that a country was in compliance with the treaty when in fact it was not; and highly intrusive inspections by multinational teams could expose both government and commercial facilities to

foreign espionage. In particular, the loss of valuable trade secrets could weaken the competitive edge of the U.S.biotechnology and pharmaceutical industries?

1 FederationofA~d~nS~e~i~s, MMkingGrwponBioiogical andToxinMbapons Verification, “pKWrOSSin Identifying Effective and Acceptable Measures fora compliance Protocol tithe Bio@icai Mkapons convention,”working paper, May 1993.

2 Statemnt by Ambassador Ronald F. Lehman, 11, Head of United States Delegation, Biological and TotinWeapons ConventIon Third Review Conference, Sept 10, 1991. Note that In signing the (Xemioai WeaponsConvention (CWC), the U.S. Government has determined that the highly intrusive inspections mandated in thattreaty do not pose an unacceptable risk to proprietary information or national security. However, the Inspectionsspedfied in the CWC to verify that chemical weapons are not being prochced or stored wwid not necessarily besufflcientforthe purposes of verifying the Bioiogicai Mbapons Convention. Therefore, the factthat CWC Inspectionshave been judged worthwhile despite their potentiaiforespionage does notautomatioaliy mean that proposed BWCinspections wwid be also be seen as acceptable. For a discussion of measures by which industry can protect itselffrom the ioss of proprietary information due to Chemical Weapons Convention declarations and inspections, seeU.S. Congress, Office of T~noiogy Assessment lhe Chemka/ i4@apons ConWnfkw: Effecfs on the U.S.Chem/ca/ /ndusfry, OTA-BP-ISC-1O6 (Washington, DC: U.S. Government Printing Office, August 1993).


While the controversy over BWC verification has focused largely on technical issues, it is fundamentally apolitical debate over whether the burden of uncertainty associated with BWC verification would hamper moreseverely the verifier or the violator. Proponents of BWC verification argue that even imperfect monitoring measureswould create a finite probability of detection that would have a significant deterrent effect on potential proliferants.Furthermore, a verification regime based on mandatory declarations of treaty-related sites and activities woulddeter the use of known facilities for BTW production, driving any violations into Clandestine facilities and thusmaking them more difficult and costly. Verification opponents counter, however, that an ineffective verificationregime would create false confidence and hence would be worse than none at all.

There is also a semantic difference over the meaning of the term “verification.” The U.S. Government usesthis word in a narrow technical sense to mean the ability to detect violations within a specified regime with a highdegree of confidence. In contrast, proponents of verification see it as the cumulative result of many sources ofinformation, only some of which would be explicitly contained in a negotiated regime. Indeed, verificationproponents admit that no negotiated inspection regime could detect clandestine facilities with a high degree ofconfidence. Instead, they argue, a formal verification regime would provide a Iegal instrument to permit inspectionsof suspicious facilities that have been detected by covert intelligence means. While many countries could bedeterred from violating the treaty by a low probability of detection, some determined proliferants would require moreintrusive measures.

Despite the Bush administration’s opposition to a formal verification protocol for the BWC, it did agree to theestablishment of an Ad Hoc Group of Government Experts to identify and evaluate various monitoring approachesfrom a scientific and technical standpoint. This verification experts (VEREX) group met twice in Geneva during1992, from March 30 to April 10 and from November 23 to December 4. The focus of its activities was to preparea list of 21 potential BTW verification technologies, divided into onsite and offsite categories. The onsite measureswere exchange visits, inspections, and continuous monitoring; the offsite measures were information monitoringdeclarations, remote sensing, and inspections. Each of these verification measures was evaluated in terms of 6criteria:

1.2.3.4.5.6.

To

technical strengths and weaknesses, including the amount and quality of information provided;ability to distinguish between prohibited and nonprohibited activities;ability to resolve ambiguities about activities;technology, material, manpower, and equipment requirements;financial, legal, safety, and organizational implications; andimpact on scientific research, cooperation, industrial development, and other permitted activities, andimplications for the protection of commercial proprietary information.3

determine whether combining some measures would result in synergistic effects, a methodology wasdeveloped for assessing measures in combination. The results indicate that the interaction of two or moremeasures may yield synergistic capabilities or limitations that are not present when the measures are evaluatedin isolation.

Between the first and second VEREX meetings, the U.S. position on BWC verification softened noticeably,and the new Clinton administration initiated a thorough review of its BTW nonproliferation policy. The VEREX groupmet again on May 24- June 4, 1993 to evaluate the proposed verification measures. The group met a final timeon September 13-24,1993 to prepare and adopt by consensus a final report to be forwarded to the States Partiesto the BWC. This final report will provide the basis for a decision by a majority of the participating countries onwhether to proceed with the negotiation of a legally binding verification protocol for the BWC. If such a decisionis made in the affirmative, a Preparatory Conference could take place in late 1994 followed by a SpecialConference in early 1995.

3 Conferenm on Disarmament, Flna/ Deckvatkw of the Third Rev/ew Conference of the BWC, part 11,document no. BWC/CONF.111/23, p. 17.


ties will inevitably have the capacity to engage inillegal military production activities. Since exces-sive secrecy might be indicative of offensiveintent, however, greater opemmess and transpar-ency would tend to build confidence in a coun-try’s defensive intentions.

Advances in biotechnology have made it possi-ble to produce militarily significant quantities ofpathogens and toxins rapidly and in small, easilyconcealable facilities, greatly complicating thetask of detecting BTW programs with nationaltechnical means of surveillance. To monitorclandestine programs, it is necessary to integratedata from many sources, with a particular empha-sis on human intelligence: (agents and defectors).

Even though much of the equipment used toproduce BTW agents is dual-use, this is notnecessarily true of the agents themselves. Mostmicrobial agents produced for peaceful purposeshave no military utility, while those that do aremade in very few places and in small quantities.Legitimate applications of dangerous pathogensand toxins (e.g., vaccine production and the use oftoxins to treat neurological disorders and forexperimental anticancer therapy) are relativelyfew at present, and are largely confinied to sophis-ticated biomedical facilities not normally foundin developing countries (with the exception of afew vaccine production plants). Moreover, giventhe fact that the biotechnology industry is still inits infancy around the globe, the background oflegitimate activities is still relatively small.

The weaponization of BTW agents entails fieldtesting of biological aerosols, munitions, anddelivery systems, as well as troop exercises,which might be detectable by satellite or othertechnical means of verification. Nevertheless,testing of microbial aerosols might be concededor carried out at night or under the cover oflegitimate dual-use activities, such as the applica-tion of biopesticides.

Despite growing concern over the militaryimplications of genetic engineering, this technol-ogy is unlikely to result in ‘supergerms” signifi-cantly more lethal or controllable than existingBW agents or capable of eliminating many of theuncertainties associated with the use of microbialpathogens in warfare. At the same time, however,gene-splicing techniques might facilitate the weap-onization of microorganisms and toxins andenhance their operational effectiveness by render-ing them more stable during dissemination (e.g.,more resistant to heat, ultraviolet radiation, andshear forces) and insusceptible to standard vac-cines and antibiotics. Moreover, genetic engi-neering techniques could be used to develop andproduce more effective protective vaccines for theattacking forces.

In the past, most plant and animal toxins had tobe extracted from biological materials in a costlyand labor-intensive operation, but the ability to‘‘clone’ protein toxin genes in bacteria has madeit possible to produce formerly rare toxins inkilogram quantities. For the forseeable future,however, toxin-warfare agents are unlikely toprovide dramatic military advantages over exist-ing chemical weapons, although their greaterpotency makes it easier to transport and delivermilitarily significant quantities. While it is possi-ble that bioregulators and other natural bodychemicals (or synthetic analogues thereof) mightbe developed into powerful incapacitants, thenontrivial problem of delivering such agents in amilitarily effective manner would first have to besolved.

BIOLOGICAL AND TOXIN AGENTSJust because a microorganism causes a serious

disease does not make it a potential warfare agent.Of the several hundred pathogenic microbes thatdirectly or indirectly afflict humans, only about30 have been considered as likely warfare agents.5

s Department of the Army, U.S. Army Medical Researeh and Development Command, Final Programma tic Enw”ronmental ImpactStatement: Biological D#ense Research Program, RCS DD-M (AR) 1327 (Fort Detric~ MD: USAMRDC, 1989), p. A7-2.

Chapter 3-Technical Aspects of Biological Weapon Proliferation! 77

Desirable characteristics of a biological agentdeveloped for military use include:

the ability to infect reliably in small doses;high virulence, or capacity to cause acuteillness resulting in incapacitation or death,without experiencing an undue loss of po-tency during production, storage, and trans-port;a short incubation period between infectionand the onset of symptoms;minimal contagiousness of the disease fromone individual to another, to avoid triggeringan uncontrolled epidemic that could boomer-ang against the attacker’s population;6

no widespread immunity-either natural oracquired-to the disease in the population tobe attacked;insusceptibility to common medical treat-ments, such as generally available antibiot-ics;suitability for economic production in mili-tarily significant quantities from availableraw materials;ease of transport, and stability under war-time field conditions of storage and delivery;ease of dissemination (e.g., as an aerosolcloud transmitted through the air);ability to survive environmental stressesduring dissemination (e.g., heat, light, desic-cation, and shear forces) long enough toinfect; andavailability of protection against the agentfor the attacking troops, such as a vaccine,antibiotics, and/or protective clothing andrespirators. 7

Figure 3-1—Toxicity of CBW Agents

Estimated lethal dose(mg/person) a

[

- 103

I

$

102CW agents

10

L 10-2 Toxin agents

i i ’ \

,0-3

104~- 10-5

i }BW agents

104

t, (--7

I

al paper clip weighs about 500 mg

SOURCE: Office of Technology Assessment, 1993,

BTW agents differ widely in infectiousness,length of incubation period, and lethality (seefigure 3-l). A variety of them, including bacteria,rickettsiae, viruses, and toxins, were weaponizedduring the U.S. offensive BTW program, whichwas terminated in 1969. Brief descriptions ofsome typical BTW agents follow.

| BacteriaBacteria are single-cell organisms that are the

causative agents of anthrax, brucellosis, tulare-mia, plague, and numerous other diseases. Theyvary considerably in infectivity and lethality. Thebacterium that causes tularemia, for example, ishighly infectious. Inhalation of as few as 10organisms causes disease after an incubationperiod of 3 to 5 days; if not treated, tularemiaresults in deep-seated pneumonia from which 30

s Some analysrs have suggested that a coun~ might deliberately develop contagious BW agents, which might be rendered insusceptibleto any drugs that could be used to combat them. Japan, for example, developed plague-a highly contagious dis~ a BW agent duringWorld War II. Wbile contagious agents are commonly dismissed as too dangerous to use, they might convey a dedsive military advantage onthe attacker if (1) he could give an antidote or vaccine to his own population, (2) the agent was designed to attack crops or livestock specificto the target country, or (3) the agent could be delivered to a distant target by a long-range delivery system such as a ballistic missile. Althoughmass vaccination of the attacker’s own population appears unlikely for logistical reasons, a ruthless aggressor-state might be willing to put itsown population at risk.

7 The effectiveness of defenses cannot be guaranteed, however. No vaccine is 100 percent effective, since even a strong immunity can beoverwhelmed by inhaling a heavy dose of agent.


Box 3-B-Anthrax as a Biological-Warfare Agent

Anthrax, a severe illness caused by the bacterium Bacillus anthracis, is considered the prototypioalbiological-warfare agent. In nature, anthrax is primarily a disease of cattle and sheep but can also infect humans.it can survive for long periods in soil in a dormant (spore) phase; after infection, it reverts to an active phase inwhich it mulitipiies rapidly in the body and secretes fatal toxins. Natural human infection can result either from skincontact with infected animals, ingestion of contaminated meat or inhalation of anthrax spores, usually fromcontaminated hides. Cases of pulmonary-and in some outbreaks gastrointestinal--anthrax are almost invariablyfatal if not treated immediately with antibiotics. Inhalation of aerosolized spores would be the primary route ofinfection if the bacteria were used deliberately as a biological-warfare agent. As extrapolated from animal studies,inhalation of about 1,000 spores or less can produce fatal pulmonary anthrax in some members of an exposedpopulation, while inhalation of about 8,000 spores-weighing about 0.08 microgram-is fatal within less than aweek to a large proportion of those exposed.l

After inhalation into the lungs, anthrax spores travel to the lymph nodes of the chest, where they becomeactive, multiplying and releasing three proteins-edema factor, lethal factor, and protective antigen. in specificcombinations, these proteins function as potent toxins, enabling the bacteria to resist host defenses and to invadeand damage host tissues via the bloodstream, resulting in uncontrollable hemorrhaging. in this manner, anthraxbacteria travel to the intestines and other areas, where they cause severe tissue damage. Initial signs of pulmonaryanthrax infection include a high fever, labored breathing, choking cough, and vomiting; it is usually fatal within 4days.2 Although infections may respond to immediate antibiotic therapy, it is relatively easy to developantibiotic-resistant anthrax strains.

in addition to its lethality, anthrax has other characteristics that make it an effective BW agent. First thedisease is not contagious from one individual to another. As a result, anthrax would not spread far beyond theintended target or boomerang against the attacker’s troops or civilian Population, assuming they do not enter acontaminated area Second, anthrax is easy to produce. The organism and its spores can be readily produced

1 Te~jmonY by Barry J. Erii~ Bioio@ai VWapons Analyst, Department of the Amy, in U.S. *nate,Committee on Governmental Affairs, Global @read of Chernkxdati B/o/o@caJ Mapons: Assessing Challengestmdl?esponsesj IOlst Congress, First Session, Feb. 9, 1989 (Washington, DC: U.S. Government Printing Offioe,1990), p. 32.

z Phiiip J. Hiits, “U.S. and Russian Researchers Tie Anthrax Deaths to %viets,” N8w Yom ~mes Mar. 15,1993, p. A6.

to 60 percent of victims die within 30 days.8 Under certain environmental conditions, anthraxBrucellosis, another bacterial disease, has a lowmortality rate-about 2 percent-but an enormouscapacity to inflict casualties. Infection gives riseto fever and chills, headache, loss of appetite,mental depression, extreme fatigue, aching joints,and sweating.9 The bacterial agent that hasreceived the most attention is anthrax, whosepulmonary form is highly lethal. (See box 3-B,)

bacteria will transform themselves into ruggedspores that are stable under a wide range ofconditions of temperature, pressure, and mois-ture. One gram of dried anthrax spores containsmore than 1011 particles; since the lethal dose byinhalation in monkeys is between l@ and l@spores, a gram of anthrax theoretically containssome 10 million lethal doses.

s TXimony byBany J, ErlicQ Biological Weapons Auilyst, Department of the Army, in U.S. Senate, Comrnitt~ on Oovernrnental Affairs,Global Spread of Chem”cal and Biological Weapons: Assessing Challenges and ReWonses, IOlst Cong., 1st sess., Feb. 9, 1989 (WashingtonDC: U.S. Government Printing Office, 1990), p. 32,

9 J. H. Rothschild, Tomorrows Weapons: Chemical and Biological (New York NY: McGraw-Hill, 19W), P. 212.

Chapter 3-Technical Aspects of Biological Weapon Proliferation 179

in the laboratory in almost unlimited quantities, and antibiotic-resistant strains have been developed with standard

selection techniques. 3

Third, when anthrax bacteria are incubated under particular conditions, they transform themselves into therugged spore form, which has along shelf-life. Although most spores can be killed by boiling for 10 minutes, theycan survive for up to 20 years or longer in soil and animal hides.4 This spore-forming ability makes anthraxparticularity well suited for delivery by missiles or bombs. The spores are stable when suspended in air, can surviveexplosive dissemination from a bomb or shell, and-unlike most pathogens-will live for several days if directsunlight is avoided. Indeed, fieldtest data have shown that anthrax spores decay at a rate of Iess than 0.1 percentper minute, which is very slow for a microorganism.5

Nevertheless, anthrax has certain liabilities as a tactical weapon. First, at lower doses there is a wide spreadin incubation times, ranging from a few days to several weeks, suggesting that the spore germinations that resultin infection can be delayed for considerable periods.6 This variability greatly reduces the predictability and hencethe military utility of the agent. Second, anthrax spores are so persistent that they can contaminate an area forlong periods, denying it both to defender and attacker. During World War II, for example, Britain detonatedexperimental anthrax bombs on Gruinard Island off the coast of Scotland, releasing spores that remained in thetop 6 to 8 inches of soil for more than 40 years.7 By infecting livestock, anthrax bacteria might also create newreservoirs of disease that could result in occasional outbreaks, making it impossible to use the affected areaproductively for long periods.8 That might be the desired intent, however, were anthrax to be used as a strategicweapon.

3 wo~d Health Organi=tion, /+ea/th @e~fs ~fchem~~/ and Bjo/ogj@/ IMgapons (Geneva: WHO, 1970),

p. 74.4 Donald Kaye and Ro~rt G. petersdorf, “Anthrax,” Eugene Br~nwalci et at., eds., HaffiSOn’S ~fil?C@/8S Of

/nferna/ Medicine, 1 Ith ed. (New Yo~ NY: McGraw Hill, 1987), p. 557.

5 Wotld Health Organization, op. dt., fOOtnOtO 3, p. 94.6 presentation ~ Matthew Meselson, Harvard University, at Smlnar on BioIo@cd VWapons in the 1990S,

sponsored by the Center for Strategic and International Studies, Washington, DC, Nov. 4, 1992.7 ~ese explosive anthr~ bombs were crude and ineffi~ent in creating an aerosol cloud COmpOSOd d Small

particles. Instead, the bombs compacted the spores into the ground. Effective BW munitions would not do this.William C. Patrick Ill, former program analysis officer, U.S. Army Medical Research Institute of Infectious Diseases,Fort Detric~ MD, personal communication, 1992.

B Wofld Health Organization, op. cit., fOOtllOM 3, p. 75.

| RickettsiaeRickettsiae are microorganisms that resemble

bacteria in form and structure but differ in thatthey are intracellular parasites that can onlyreproduce inside animal cells. Examples of rick-ettsial diseases that might be used for biologicalwarfare include typhus, Rocky Mountain spottedfever, Tsutsugamuchi disease, and Q fever. Rick-ettsiae have a wide variety of natural hosts,including mammals and arthropods such as ticks,fleas, and lice. If used as BW agents, however,they would probably be disseminated directlythrough the air.

| VirusesViruses are intracellular parasites that are about

100 times smaller than bacteria. They can infecthumans, crops, or domestic animals. Virusesconsist of a strand of genetic material (DNA orRNA) surrounded by a protective coat thatfacilitates transmission from one cell to another.The Venezuelan equine encephalitis (VEE) viruscauses a highly infectious disease that incapaci-tates but rarely kills. In contrast, some hemor-rhagic fever viruses, such as Lassa or Ebola fever,are exceedingly virulent, killing 70 out of every100 victims. The AIDS virus, despite its lethality,would not be an effective warfare agent because

80 I Technologies Underlying Weapons of Mass Destruction

its mean incubation period of 10 years is too slowto give it any tactical or strategic value in warfare,and because it cannot be transmitted through theair.

FungiFungi do not generally cause disease in healthy

humans, although the fungus Aspergillus, whichinfects by inhalation, can cause serious opportun-istic infections in people with a weakened im-mune system. A few other fungi, such as Coccidi-oides immitis and Histoplasma capsulatum, alsoinfect naturally by inhalation and can causesevere pulmonary infections in susceptible indi-viduals, but they have never been considered aspotential BW agents. Fungal diseases are, how-ever, devastating to plants and might be used todestroy staple crops and cause widespread hungerand economic hardship. Examples of plant fungalpathogens include rice blast, cereal rust, andpotato blight, which can cause crop losses of 70to 80 percent.

| ToxinsA toxin is a poisonous substance made by a

living system, or a synthetic analogue of anaturally occurring poison. An enormous varietyof toxins are manufactured by bacteria, fungi,marine organisms, plants, insects, spiders, andanimals, and more than 400 have been character-ized to date. Such toxins can exert their effects bythree different routes of exposure-injection,ingestion, and inhalation-and their potencyderives from their high specificity for cellulartargets. For example, many toxins bind to specificsites in nerve membranes, disrupting the trans-mission of nerve impulses and causing fatalrespiratory paralysis. Other toxins selectivelyblock cellular protein synthesis or other vitalphysiological functions.

From a chemical standpoint, there are twocategories of toxins: protein toxins, which consistof long folded chains of amino acids; andnonprotein toxins, which tend to be small butcomplex molecules.

PROTEIN TOXINSMost bacterial toxins, including those associ-

ated with cholera, tetanus, diphtheria, and botu-lism, are large proteins. For example, variousstrains of Staphylococcus aureus, a major bacte-rial pathogen, secrete protein toxins that causesevere nausea, vomiting, and diarrhea lastingfrom 1 to 2 days. The United States developed oneof these toxins, Staphylococcus enterotoxin B(SEB), as a warfare agent in the 1960s. Spray-dried SEB, when disseminated through the air inaerosol form, causes a chemical pneumonia thatis more debilitating than the toxin’s gastrointesti-nal effects when ingested; it can incapacitateexposed troops within hours, with recovery in 4 to6 days.l0 Botulinal toxin, secreted by the soilbacterium Clostridium botulinum, is the mostpoisonous substance known. The fatal dose ofbotulinal toxin by injection or inhalation is about1 nanogram (billionth of a gram) per kilogram, orabout 70 nanograms for an adult male.11 The toxinis also relatively fast-acting, producing deathbetween 1 and 3 days in 80 percent of victims.The U.N. inspections of Iraq after the Gulf Warconfirmed that the microbiological research facil-ity at Salman Pak had done development work onbotulinum toxin as a potential warfare agent.Nevertheless, attempts to weaponize botulinaltoxin have in the past failed to prevent extensiveloss of toxicity that accompanies dispersion.

Ricin, a plant toxin derived from castor beans,irreversibly blocks cellular protein synthesis andis lethal when inhaled in a dose of about 10micrograms (millionths of a gram) .12 Castor

10 WiU~ C. ntrick III, fo~e:rpm~ analysis officer, U.S. Army Medical Research Institute for Infectious Diseases, Fort ~tridc+ MD,personal communicatio~ 1993.

11 D. M. Gill, “Bacteri~ ‘1’bxins: A ‘Ihble of Lethal Amounts,” Microbiological Reviews, March 1982, pp. 86-94.

12 G. A. B~@ “wc~ me Toxic Protein of Castor Oil Seeds,” Tom”cology, VO1. 2, 1974, p. 80.


beans are widely cultivated as a source of castor

oil, which has numerous legitimate industrialapplications. The paste remaining after the oil hasbeen pressed out contains about 5 percent ricin,which can be purified by biochemical means.During World War II, several countries studiedricin as a potential chemical-warfare agent, andthe British developed and tested an experimentalricin weapon known as the ‘‘W bomb, ” althoughit was not ultimately deployed.13 In September1978, the Bulgarian secret police (with technical

assistance from the Soviet KGB) assassinatedGeorgi Markov, an exiled Bulgarian dissidentliving in London, by firing a tiny metal ball filledwith ricin into his thigh from a pellet-gunconcealed inside an umbrella; Markov died twodays later.14 According to published reports, Iranhas acquired 120 tons of castor beans and isallegedly purifying ricin in pharmaceuticalplants. l5

NONPROTEIN TOXINSNonprotein toxins are small organic molecules

that often have a complex chemical structure.They include tetrodotoxin @educed by a pufferfish), saxitoxin (made by marine algae known asdinoflagellates, which are taken up and concen-trated by clams and mussels), ciguatoxin andmicrocystin (synthesized by microscopic algae),palytoxin (made by a soft red Hawaiian coral),and batrachotoxin (secreted by poisonous frogsindigenous to western Colombia). Typical char-acteristics of nonprotein toxins are high toxicity,the absence of antidotes, heat stability (unlike

most protein toxins), resistance to other environ-mental factors, and speed of action. Saxitoxin, forexample, produces initial symptoms within 30seconds after ingestion and can cause laboredbreathing and paralysis in as little as 12 minutes.There is no known prophylaxis or therapy, and thelethal dose in 50 percent of those exposed maybeas low as 50 micrograms, a potency 1,000 times

greater than the chemical nerve agent VX.l6

Trichothecene mycotoxins are a family ofabout 100 poisonous compounds manufacturedby certain strains of the mold Fusarium that growon wheat, millet, and barley. When ingested bypeople or livestock, these toxins kill rapidlydividing cells such as those of the bone marrow,skin, and the lining of the gastrointestinal tract;they also block certain clotting factors in theblood, causing severe bleeding after injury. Inaerosol form, about 35 milligrams of the trichoth-ecene mycotoxin T-2 can kill a 75-kilogram man;unlike most other toxins, it is also absorbedthrough the skin. Although mycotoxins are signif-icantly less potent than chemical-warfare agentssuch as VX, they are relatively easy to produceand are highly stable.

In 1982, the Reagan administration alleged thatthe Soviet Union and its allies were using atoxin-warfare agent in Southeast Asia known as“yellow rain” whose active ingredients weretrichothecene mycotoxins.17 The Soviets deniedthe allegations, and the United States was unableto provide convincing public evidence to back upits charges in the face of criticism on the part of

13 Stwkholm htemtio~ pe~e Research Institute, The Problem of Chemical and Biological Weapons, Vol. I: The Rise of CB Weapons

(Stockholm: Almqvist & WikseU 1971), p. 123.

14 Rob@ Harris and Jeremy PaxmmL A Higher Form of Killing: The Secret Story of Gas and Germ Waglare (1-mdon: Chatto & Windus,1982), pp. 197-198; David Wise, ‘‘Was Oswald a Spy, and Other Cold War Mysteries,” New York Times Magazine, Dec. 6, 1992, p, 44.

15 Douglas Wailer, “Sneaking in the Scuds, ” Newsweek, June 22, 1992, p. 42.

16 Erlic~ op. cit., footnote 8, p. 32, See also B.J. Benton and F.C.T. Chang, ‘‘Reversal of Saxitoxin-Induced Ctiio-Resptitov F~~e byBurro XgG Antibody and Oxygen Therapy,’ Proceedings of the 1991 Medical De$ense Bioscience Review (Fort lletric~ MD: U.S. ArmyMedical Research Institute of Chemical Defense, Aug. 7-8, 1991), p. 176.

17 U.S. Department of State, Chem”cal Wag2are in Southeast Asia and Afghanistan, Special Report No. 98, w. 22, 1982, P. 30.


many in the U.S. scientific community.18 Follow-ing early reports of the presence of trichothecenesin samples from alleged attacks, the U.S. Armyand the U.K. Ministry of Defense initiated largeanalytical studies but were unable to confirm theearly findings.19 Nonetheless, U.S. intelligenceofficials, based on all the: information available tothe U.S. intelligence community, remain confi-dent in the yellow rain allegations and have notretracted them.

Compared to microbial pathogens, toxins offerthe following tactical advantages:

The most toxic toxins (e.g., botulinal toxin)are exceedingly potent, so that small, easilytransportable quantities would be militarilysignificant for certain missions.Toxins tend to deteriorate rapidly oncereleased into the environment, whereas an-thrax spores and persistent chemical agentscan centaminate soil for months or years. Forthis reason, territory attacked with toxinagents could be occupied more rapidly byattacking forces.Toxins are well suited to covert warfare.Whereas chemical agents leave telltale deg-radation byproducts that persist for longperiods in the environment, some toxinagents break down completely over a periodof weeks or months, leaving no traces.Moreover, even fresh samples of toxin mightnot provide conclusive evidence of militaryuse if the agent occurred naturally in theregion where it was employed.

Despite these operational advantages, how-ever, toxins have drawbacks for battlefield use:

Protein toxins such as botulinal toxin decom-pose rapidly on exposure to sunlight, air, andheat, and thus would have to be used at night.Protein toxins may be inactivated by themechanical shear forces caused by passagethrough an aerosol sprayer.20 While low-molecular-weight toxins such as saxitoxinand trichothecene mycotoxins are more sta-ble than protein toxins, they are less stablethan chemical-warfare agents.Most toxins (with the exception of trichoth-ecene mycotoxin T-2) do not penetrate theskin, nor would toxin lying on the groundcreate a vapor hazard.21 Weaponization there-fore requires the creation of a small-particleaerosol cloud, in which the toxin mustremain airborne to be effective.The inhalation threat posed by protein toxinssuch as botulinal toxin can be counteredeffectively with modern gas masks (althougha surprise or covert attack might exposepersonnel to lethal concentrations beforethey could don their masks).

For conventional battlefield use, toxins offerfew military advantages over chemical nerveagents. Toxins would, however, probably besuperior for small-scale clandestine operations.

ACQUIRING A BTW CAPABILITYThe acquisition of a militarily significant BTW

capability would probably involve the followingsteps (see figure 3-2):

la Jfi~RobinsoG Je~e Guille~ and Matthew Meselso~ “Yellow Rain in Southeast Asia: ‘he StOry COlkipSeS,” SUS~ wrigh~ cd.,Preventing a Biological Arms Race (Cambridge, MA: MIT Press, 1990), pp. 220-238. See also U.S. Senate, Cornrnittee on Foreign Relations,Subcommittee on Arms Control, Oceans, International Operations, and Environment, 98th Cong., 1st sess., YelZ~w Rain: The Arms ConrrolImplications, Feb. 24, 1983 (Washington DC: U.S. Government Printing Off@ 1983).

19 Robinson et al., op. cit., footnote 18, pp. **8-**9.

20A fw prote~ tox~ we quite s~ble: SEB ~ ~~ for e~ple, is not &s~oy~ ~t~ boiling for 30 minutes. III additio~ the stibility

of protein toxins can be increased through a technique known as microeneapsulation (see production section below), David S. Huxsoll, formerdirector, U.S. Army Medical Research Institute for Infectious Diseases, personal comrnunicatiom 1992.

21 Shirley Freeman, ‘‘Disease as a Weapon of War, ” Pacific Research, vol. 3, February 1990, p. 5.


R&D

v’or novel agent

– — v — – - -

Figure 3-2—Biological Weapon Acquisition

Produce agent

, ;;;>~~~;~>E-->F ->?, Test suitability

A >’ for weapon

I~ Mampulate genehc ~

characteristics I

II

(optional)

Design, test, and build munitions

EMicro-> encapsulate

agent

I Area delwery: I vI sprayer system I I ~ ~ ‘-~L.––—.:–-–L— - )

~—.--.—iField- ‘ MaSS-

‘ > Filltest produce munitions < -

I

Point dellvery: ‘cluster bomb or warhead ,~— —- . -–. J

Acquire delivery system

r?=fl

r%%=”!E!!Z2EE!IL JDevelop strategic and

tactical BW battle plans

EiiiiaaSOURCE: Office of Technology Assessment, 1993.

delivery system

Acquire operational capability I

1. Establishment of one or more facilities and 3.associated personnel with organizationaland physical provisions for the conduct ofwork in secret; 4.

2. Research on microbial pathogens and tox-ins, including the isolation or procurementof virulent or drug-resistant strains;

>

~Integrate

weapon systemsinto military ,

df o r c e s

@ockpile>1 f i l l e d

Lm!!!’ti!!s

mOperational> capability

Pilot production of small quantities of agentin flasks or small fermenter systems;

Characterization and military assessment ofthe agent, including its stability, infectivity,course of infection, dosage, and the feasibil-ity of aerosol dissemination;


5.

6.

7.

8.

Research, design, development, and testingof munitions and/or other disseminationequipment;

Scaled-up production of agent (possibly inseveral stages) and freeze-drying;

Stabilization of agent (e.g., through micro-encapsulation) and loading into spray tanks,munitions, or other delivery systems; and

Stockpiling of filled or unfilled munitionsand delivery vehicles, possibly accompa-nied by troop training, exercises, and doc-trinal development. (In some but not allcases, a country planning the offensive useof BTW agents would take measures toprotect its own troops, such as immuniza-tion, the acquisition of respirators, andtraining in self-protective measures.)

The key steps in this acquisition sequence areexamined below in terms of their technicaldifficulty.

| Research, Development, andWeaponization

Countries seeking a BTW capability are likelyto start with the development of standard agentsthat have been weaponized in the past, such asanthrax, tularemia, and botulinum toxin. Nearlyall proliferant states lack the sophisticated scien-tific and technological infrastructure needed todevelop novel agents such as exotic viruses,whose military characteristics are poorly under-stood.

BTW agents are widely accessible. Pathogenicmicroorganisms are indigenous to many countriesand can be cultured from infected wild animals(e.g., plague in rodents), living domestic animalsor infected remains (e.g., Q fever in sheep,anthrax in cattle), soil in endemic areas (whichmay contain trace amounts of anthrax bacteriaand other pathogens), and spoiled food. Certainbiological supply houses also ship strains of

microbial pathogens to scientists throughout theworld. For example, American Type CultureCollection (ATCC), a nonprofit company inRockville, MD, acts as a clearinghouse forresearch institutions around the world, shippingeach year approximately 130,000 cultures ofweakened (’‘attenuated”) pathogens to 60 na-tions. 22 While such attenuated strains are notvirulent and hence could not be converted directlyinto biological weapons, they would be useful forBTW research and development and for preparingself-protective vaccines. Methods for culturingorganisms and for inducing spore formation arealso described in the open scientific literature, andstandard microbiological procedures can be usedto produce more virulent or antibiotic-resistantstrains of microbial pathogens.

Once a proliferant had acquired BTW agents,they might be modified genetically throughsimple selection techniques to increase theirvirulence or effectiveness. For example, incubat-ing microbial pathogens in the presence ofstandard antibiotics can induce the emergence ofdrug-resistant strains, which can then be subcul-ture and mass-produced. Agent developmentwould also involve “weaponization," or a thor-ough assessment of the agent military potential,including its stability, infectivity, course of infec-tion, and effective dosage. This step wouldinclude the testing of candidate agents to deter-mine their effectiveness, including the feasibilityand reliability of aerosol dissemination. Suchtests might be carried out either in a sealed aerosolchamber or in field studies of simulant microorg-anisms at a remote testing range.

THE DUAL-USE DILEMMAA fundamental problem in countering the

proliferation of biological and toxin weapons isthe fact that much of the necessary know-how andtechnology is dual-use, with legitimate applica-tions in the commercial fermentation and biotech-nology industries. Many developing countries

n Wc N~Im d RoM WiIIdreW, “&adly Contagio~” The New Republic, vol. 204, No. 5, Feb. 4, 1991, p. 18.


have acquired industrial microbiology plants forthe production of fermented beverages, vaccines,

antibiotics, ethanol (from corn or sugar cane),enzymes, yeast, vitamins, food colors and flavor-ings, amino acids, and single-cell protein as asupplement for animal feeds.23 This global expan-sion of the civilian biotechnology industry, com-bined with the growing number of molecularbiotechnologists trained in the West, has createdmuch broader access to the expertise and equip-ment needed for the development of BTW agents.Sophisticated laboratories that might be used forthe design of novel BW agents are inexpensivecompared with nuclear weapon plants. Moreover,biotechnology is information-intensive ratherthan capital-intensive, and much of the rele-vant data are available in the published scien-tific literature. For these reasons, it is virtuallyimpossible for industrialized states to preventthe diffusion of weapon-relevant informationto states of proliferation concern.

It has been estimated that more than 100countries have the capability-if not necessarilythe intent-to develop at least crude biologicalweapons based on standard microbial and toxinagents. 24 In addition to the United States, Russia,Western Europe, and Japan, countries with anadvanced commercial biotechnology infrastruc-ture include Argentina, Brazil, Chile, Cuba, India,Israel, the People’s Republic of China, Taiwan,and Thailand. Cuba, in particular, has an ad-

vanced biotechnology industry that exports vac-

cines and reagents to other Latin Americancountries .25 While Iraq lags somewhat behind thisgroup of countries, Baghdad established a na-

tional Center for Genetic Engineering and Bio-technology in the late 1980s, initially staffed withonly four scientists.26 As an increasing number ofdeveloping countries become involved in com-mercial biotechnology, they may be tempted toexplore its military potential.

In addition, the legitimate use of toxins formedical therapy and biomedical research is in-creasingly widespread. Botulinal toxin, for exam-ple, is used to treat abnormal muscle spasmsknown as dystonias by selectively paralyzing thespastic muscles; it has also been applied cosmeti-cally to smooth wrinkles.27 Toxins such as ricin,when linked to antibodies that selectively targetcancer cells, have shown promise in clinical trialsas an anticancer therapy.28 Furthermore, saxitoxin

and other exotic toxins that bind specifically tochannels or receptors in nerve-cell membranes arevaluable research tools in neuroscience. Theinherently dual-use nature of many pathogens andtoxins makes the prevention of BTW-relevantresearch extremely difficult. Consumption oftoxins for medical therapy and research hasalready expanded to the current level of hundredsof grams per year, and the anticipated furthergrowth of such therapies will eventually blur the

23 For an overview, see U.S. Congr=s, OffIce of ‘lkchnology Assessment, Biotechnology in a Global Economy, OTA-BA-494 (w~mto~DC: U.S. Government Printing Office, October 1991).

u ~s~ony of moms Welcb D~~ty Assistant Smre- of Defense (~emic~ Matters), reported in D#ense Week, May 9, 1988.

2S ~wond A. Zilinska% “Biological Warfare and the Third World” Politics and the L~e Sciences, vol. 9, No. 1, August 1990, p. 61.

26 presentation by Raymond Zilinskas, Washington Sh’ategy s~, Washington, DC, July 14, 1992.27 Tom Waters, ‘me F~e M of M.alcing Poisou” Discover, vol. 13, No. 8, August 1992, p. 32; and Anna Evangeli, ‘ ‘Botulism Gives Fac~

New base of Life, ” New Scien~ist, vol. 137, No. 1859, Feb. 6, 1993, p. 18. See also Fritz P. Gluckstein and Mark Hallet, Clinical Use ofBotulinum Toxin: January 1987 through September 1990, 318 Citations (Washington DC: National Library of Medicine/U.S. GovernmentPrinting Office, 1990).

28 David Fitzgerald and Isa Pastaq ‘‘Targeted Toxin Therapy for the Treatment Of Cancer, ” Journal of the National Cancer Institute, vol.81, No. 19, October 1989, pp. 1455-1463; Andrew A. Hertler and Arthur E. Frankel, “Imrnunotoxins: A Clinical Review of Their Use in theTreatment of Malignancies,” Journal of Clinical Oncology, vol. 7, No. 12, December 1989, pp. 1932-1942; Lee H. Pai and Ira PastarL‘‘Immunotoxin Therapy for Cancer,’ Journal of the Amen’can Medical Association, vol. 269, No. 1, Jan. 6, 1993, pp. 78-81.


distinction between medically useful and militar-ily significant quantities of toxins. 29 The legiti-mate applications of toxins will therefore have toremain relatively open to preclude their use forillicit purposes.

Finally, the development of defenses againstBTW attack-an activity explicitly permitted bythe Biological Weapons Convention-draws onmuch of the same knowledge base needed todevelop offensive BTW agents. Indeed, “threatassessment, an important aspect of some biolo-gical-defense programs, includes the evaluationof defenses under simulated warfare conditionsand may be indistinguishable from the develop-ment of offensive BTW agents .30 Furthermore,certain defensive activities may have offensiveapplications. According to one assessment, ‘‘thevirulence of micro-organisms is studied both forits relevance to the field of natural infections andin order to produce living, attenuated vaccines.Such knowledge can obviously be used more orless directly to make a BW agent more viru-lent.”31 For these reasons, biological-defenseactivities such as the development of vaccinesmay arouse concerns about offensive intentionsunless they are conducted openly and in anunclassified environment..

In sum, research on potential BTW agents doesnot necessarily imply an offensive weapon pro-gram because much of the relevant knowledge ismultiuse. This inherent ambiguity means that atthe R&D stage, the only difference betweenoffensive and defensive activities is one of intent.The policy dilemma is that progress in controllinginfectious diseases requires the free and openflow of information, so that researchers can buildon and validate the work of others; imposingcontrols on the publication of results with poten-

tial military implications would seriously impedelegitimate scientific research worldwide. Never-theless, openness may impose some limits on themisuse of biomedical research for maliciouspurposes.

| Large-Scale ProductionBTW agents would be relatively easy and

inexpensive to produce for any nation that has amodestly sophisticated pharmaceutical or fer-mentation industry. Indeed, mass-productionmethods for growing pure cultures are widelyused in the commercial production of yogurt,yeast, beer, antibiotics, and vaccines. Nearly allthe equipment needed for the production ofpathogens and toxins is dual-use and widelyavailable on the international market, increasingthe potential for concealing illicit activities underthe cover of legitimate production. Whereas atypical vaccine production facility costs a mini-mum of $50 million, a much less elaborateindustrial fermentation plant suitable for conver-sion to BTW agent production could be built forabout $10 million.32 In such a‘ ‘no-frills’ facility,bacteria could be grown in standard dairy tanks,brewery fermenters, or even in the fiberglasstanks used by gas stations.

In contrast to chemical-warfare (CW) agents,no specialized starting materials are required forthe production of biological and toxin agentsexcept for a small seed stock of a disease-producing organism. Nutrients such as fermenta-tion medium, glucose, phosphates, peptone, anda protein source (e.g., casein, electrodialyzedwhey, or beef bouillon) are widely available andare routinely imported by developing countriesthat have commercial fermentation industries. Astate seeking a CW capability, in contrast, re-

29& p. ~licoff, Senior Me~~, ‘lMmical Staff, Sandia National Laboratories, personal COmlll@catiolL 1992.

30 s= su~m Wn@t ~d SW: Kc@@ ‘‘~ probl~ of ~terpre@ tie U.S. BiOIOgic~ D~e~ llese~ch program,’ SUSan Wrigh~

cd., Preventing a Biological Arms Race (Cambridge, MA: MIT Press, 1990), pp. 167-196.

31 stw~o~ htemtio~ Pea:e Re~~h lnsti~te (SIPIU), The Problem of Chemical and Biological Wa~are, Vol. VI: Technical Aspects

of Early Warning and Verification (Stockholm: Almqvist & Wiksell, 1975), p. 24.

32 ~temiew tit.b Dr. Ron l“hibeauto~ Wyeth-Ayerst vaccine production pkm~ Marietta, pA, &t. 22, 1992.


Figure 3-3-Production of Biological Agents by Fermentation

,~’y /’~

Freeze-driedseed culture fl

T

Propagation vessels ~LY@l.

Production-scalefermenter

c--—

C2

.

‘\

“3.

‘7.—. .

/’”

Final processing— >

SOURCE: U.S. Senate, Committee on Governmental Affairs, Global Spread of Chemical and Biological k$bapons,IOlst Cong., 1st sess., Feb. 9, 1989 (S. Hrg. 101-794), p. 241.

quires hundreds or thousands of tons of unusualprecursor chemicals that may be difficult toobtain.

PRODUCTION OF BACTERIAL AGENTSA biological-warfare plant would contain fer-

menters and the means to sterilize and dispose ofhazardous biological wastes on a large scale. Asmall vial of freeze-dried seed culture, grown ina fermenter in a nutrient medium kept at constant

temperature, can result in kilograms of product(e.g., anthrax bacteria) in as little as 96 hours.33

(See figure 3-3.) Microbial pathogens such asplague bacteria can also be cultivated in livinganimals, ranging from rats to horses.

Fermentation can be carried out on a batchbasis or in a continuous culture from whichorganisms are constantly removed and an equalvolume of new culture medium is added. Anadvantage of continuous culture is shorter turna-

33 Er~c~ op. cit., foomote 8, P. 32.


round time, increasing the productivity of eachfermenter. Indeed, if nutrients are supplied con-tinuously and natural growth-inhibitors are re-moved as soon as they are formed, the bacterialculture can be maintained indefinitely in a phasein which it multiples exponentially. A continuousculture can therefore yield nearly 10 times asmuch product per volume of culture medium asthe batch approach.34 Nevertheless, batch culturehas generally been used to cultivate BW agents inthe past because continuous culture is technicallymore complex and sometimes results in a loss ofpotency. High levels of purity are not required forBW agents; 60 to 70 percent purity will sufficeand is easy to obtain. The main technical hurdlesin bacterial production are:

the danger of infecting production workers;genetic mutations that may lead to a loss ofagent potency; andthe contamination of bacterial cultures withother microbes (e.g., bacterial viruses) thatmay kill them or interfere with their effects.

Although biological agents can be grown inordinary laboratory flasks, an efficient productioncapability would require the use of specializedfermenters. Until fairly recently, large-scale pro-duction of bacteria for commercial or militarypurposes required tank-type bioreactors contain-ing thousands of liters of culture, with mechanicalstirring or a flow of air to oxygenate the culturemedium. During World War II, for example, theJapanese Army ran a top-secret BTW facility inoccupied Manchuria at which more than 3,000workers grew kilogram quantities of pathogenic

bacteria (including the agents of anthrax, brucel-losis, plague, and typhus) in giant vats.35

Also during World War II, the United Statesand Britain planned to produce anthrax bacteria inlarge quantities for use in a strategic bombingcampaign against Germany. In 1943, a pilotanthrax production plant became operational atCamp Detrick, MD, staffed with about 500bacteriologists, lab assistants, chemical engi-neers, and skilled technicians.36 Based on thisexperience, the decision was made to build afill-scale plant at Vigo, Indiana, at a cost of $8million, where 1,000 workers would manufacturemore than 500,000 anthrax bombs a month (or,alternatively, 250,000 bombs filled with botulinaltoxin). Since both agents store well, they could bestockpiled in large quantities. The Vigo plant wascompleted in early 1945 but never actually wentinto production.37 Although it is far from certain

the anthrax bombs would have worked as de-signed, it is possible that large areas of Germanycould have been rendered uninhabitable for dec-ades.

In 1950, the U.S. Congress voted $90 millionto build another BTW plant called X-201 at arenovated arsenal near Pine Bluff, Arkansas. Thenew production facility had 10 stories, 3 of themunderground, and was equipped with 10 fermen-ters for the mass-production of bacterial patho-gens on short notice.38 To give some idea of thescale involved, the Pine Bluff facility and itsassociated munitions-falling plant required a watersupply of 2 million gallons per day, an electricalpower supply of 5 megawatts, and an initialworkforce of 858 people.39 Production of BW

~ SIPM, VO1. VI, op. cit., footnote 31, p. 43.

N me Jap~~B~ fmfi~ WaS c~e.~~ Unit 731. John W. Powe~ “A Hidden Chapter in History,” Bulletin ojtheAro?nic ~denti$t$,vol. 37, No. 8, October 1981, pp. 44-52. For a more detailed deseriptio~ see Petes Williams and David Wallace, Unit 731: Japan’s SecretBiological Warjare in World War II (New Yorlq NY: Free Press, 1989).

3 6 = ~d p~, op. cit., footnote 14, pp. 1(X L101.

37 Ibid,, p. 103.

SE Ibid., p. 160.

39 ~~ew Me~lsO~ hlartin M. Kapl~ and Mark A. Mokukk-y, ‘ ‘Vcrifkat.ionof BiologieaJ and ‘Rm.in WMpOrM Di saxmarnen4° Science& Global Security, vol. 2, Nos. 2-3, 1991, p. 237.

Chapter 3–Technical Aspects of Biological Weapon Proliferation | 89

agents on such a vast scale is far in excess of whata country would need to wreak enormous destruc-tion on an adversary.

Over the past decade, technological ad-vances associated with the commercial bio-technology industry have made it possible toproduce large quantities of microorganisms inmuch smaller facilities. The introduction ofcomputer-controlled, continuous-flow fermentersand compact ultrafiltration methods has vastlyincreased productivity, making it possible toreduce the size of a fermenter to about 1,000-foldless than conventional batch fermenters that giveequivalent production.40 Real-time sensors and

feedback loops under microprocessor controlhave also optimized culture conditions, resultingin much higher yields and better quality productsthan in the past. The resulting increase in produc-tivity has made it possible to reduce the amountof trained manpower needed to operate large-scale fermenters and to use smaller, more con-cealable production equipment. Of course, adeveloping country could produce many small-scale batches of BTW agents in laboratoryglassware without the need for high-technologyfermenters.

PRODUCTION OF VIRAL AND RICKETTSIALAGENTS

Pathogenic viruses and rickettsiae are intracell-ular parasites that can only reproduce insideliving cells. There are two approaches to cultivat-ing these agents: in intact living tissue (e.g., chickembryos or mouse brains) or in isolated cellsgrowing in tissue culture. The latter approach is

technically simpler because it requires only flasksand nutrient medium, but certain viruses (e.g.,influenza) do not grow well in tissue culture andmust be cultivated in fertilized eggs. In 1962, FortDetrick used more than 800,000 eggs for thecultivation of pathogenic viruses.41

Growing viruses and rickettsiae in culturedmammalian cells offers greater control but in-volves certain technical hurdles. The cells mustadhere to a surface to grow and also require acomplex culture medium based on blood serumobtained from horses and cows. Until recently,cultured mammalian cells were grown on theinner surface of rotating glass bottles, whichlimited the volume of production. Over the pastdecade, however, new methods for cultivatingmammalian cells have been developed that permithigher concentrations of cells and greater recov-ery of product. For example, allowing the cells togrow on surface of beads suspended in culturemedium has permitted the scaling-up of produc-tion. Yield has been improved further by replac-ing the beads with microcarriers, which have aporous internal structure into which animal cellscan grow.42

Hollow-fiber technology offers an even moreefficient method of growing anchorage-dependent mammalian cells in high concentra-tions for the cultivation of viruses or rickettsiae.The cells are grown on the outer surface of thinfibers that are immersed in the growth medium;air is pumped through the fibers and diffusesthrough the fiber wall to reach the cells.43 Sincea single hollow-fiber bioreactor is equivalent toseveral thousand one-liter roller bottles, it occu-

~ Gove~ent of Aus~~ia, ‘Impact of Recent Advances in Science and ‘Ikdmology on the Biological Weapons Convention’ BackgroundDocument on New Scientific and Technological Developments Relevant to the Convention on the Prohibition of the Development, Productionand Stockpiling ofBacten’ological (Biological) and Tom’n Weapons and on Their Destruction, Third Review Conference of the BWC (Genev%Switzerland), Document No. BWC/CONF.111/4, Aug. 26, 1991, p. 3.

41 SePOW M. Her~~ Chemical ~nd Biological Wa$are: Am~ca’~ Hidden Arsenal @I&uMpofis, IN: Bobbs-M@l, 1%8), p. 78.

42 S.B. Wose, Molecular Biotechnology, 2d ed. (Oxford, England: Blackwell Scientitlc Publications, 1991), P. 116.

43 Government of the U.S.S.R, “Selected Scientilc and Technological Developments of Relevance to the BW Conventio~” BackgroundDocument on New Scientific and Technological Developments Relevant to the Convention on the Prohibition of the Development, Productionand Stockpiling of Bacteriological (Biological) and Tom”n Weapons and on Their Destruction, Third Review Conference of the BWC, Genev~Switzerland, Document No. BWC/CONF.111/4/Add.1, Sept. 10, 1991, p. Al 1.


pies less than one-twentieth the volume of theprevious technology.

44 Advantages include econ-

omy and the high concentration and purity of theend-product, which reaches 98 percent on leavingthe reactor. In sum, the new cell-culture tech-niques greatly simplify the production of vi-ruses and rickettsiae and allow large-scaleyields from very small facilities.

PRODUCTION OF TOXINSThe most efficient way to produce bacterial

toxins is through fermentation. Botulinal toxin,for example, is derived from a culture of Clostrid-ium botulinum bacteria, which multiply rapidlyunder the right conditions of temperature, acidity,and the absence of oxygen. It takes only about 3days to grow up a dense culture of the bacterialcells, which extrude botulinal toxin into thesurrounding culture medium. (Purification of thetoxin is neither necessary nor desirable, since ittends to reduce stability.) During World War II,Japan’s Unit 731 produced kilogram quantities ofbotulinal toxin in a fermenter approximately 10feet high and 5 feet wide.45 A crude preparation oftoxin can be freeze-dried down to a solid cake,which is then milled into a fine powder suitablefor dissemination through the air. The millingoperation is exceedingly hazardous, however, andmust be carried out under high-containmentconditions. Plant toxins such as ricin, whose rawmaterial is widely available, could easily beproduced in the hundreds of kilograms.

With recombinant-DNA techniques, rareanimal toxins—formerly available only in mil-

ligram amounts-can be prepared in signifi-cant quantities in microorganisms. Althoughthese techniques are still largely restricted to theadvanced industrial countries, they are spreadingrapidly around the world. One method, known asthe “cloning” of toxin genes, involves identify-ing DNA sequences in plants and animals thatgovern the production of protein toxins, transfer-ring these genes to a suitable microbial host, andmass-producing the toxin in a fermenter. In thisway, ordinary bacterial cells can be transformedinto miniature toxin factories.% Production ofanimal toxins in bacteria involves certain techni-cal hurdles, however. Bacteria typically produceand secrete toxins only under special conditions,which may not be met in an artificial environ-ment; and bacteria may be unable to performcertain biochemical “processing” steps neededto convert a protein toxin to its active form.47 Forthese reasons, it maybe necessary to clone plantand animal toxins in yeast or mammalian cells, atechnically more challenging task.

Nonprotein toxins are considerably harderthan protein toxins to produce in militarilysignificant quantities. Until recently, even smallamounts of nonprotein toxins such as saxitoxinhad to be extracted from large quantities ofbiological material with costly and labor-intensive purification methods. For example, 270kilograms of toxin-containing clam siphons yieldedless than 5 grams of saxitoxin.

48 Although somenonprotein toxins such as saxitoxin and tetro-dotoxin can be synthesized in the test tube withmultistep procedures, the overall yield is only

44 @vement of the United States, “lkchnological Developments of Relevance to the Biological and ‘Ibxin Weapons Convention”BackgroundDocument on New Sciennj?c and Techno[ogica[Developments Relevant to the Convention on the Prohibition of the Development,Production and Stockpiling of Bacteriological (Biological) and Toxin Weapons and on Their Destruction, Third Review Conference of theBWC (Genev& Switzerland), Document No, BWC/COITF.111/4, Aug. 26, 1991, p. 30.

45 Wflh ~d JVmce, op. cit., fmtnote 35, p. 124.

46 Akm Wiseman, “The Organization of Production of Genetically-Engineered Proteins in Yeast,” Endeavor (new series), vol. 16, No. 4,1992, pp. 190-193,

4T Mc~d Novick ~d Seth S,h- “New FOMM of Biological Warfare?’ Susan Wrigh4 cd., Preventing A Biological Arms Race

(Cambridge, MA: MIT Press, 1990), p. 115.

48 Edwmd J. SChantZ et al., ‘‘Paralytic Shellfish Poison. IV. A Procedure for the Isolation and PurMcation of Poison from Toxic Clam andMussel Tissues,” Journal of the American Chemical Society, vol. 79, Oct. 5, 1957, pp. 5230-5235,

..— ———

Chapter 3–Technical Aspects of Biological Weapon Proliferation I 91

Table 3-l—Key Production Techniques for BTW Agents

Type of agent Low-tech production High-tech production

Bacteria Batch fermentation, production in Genetically engineered strains, con-animals tinuous-flow fermentation

Rickettsiae and viruses Cultivation in eggs, mouse brains, Culture in mammalian cells grownor tissue culture (roller bottles) on beads, microcarriers, or hollow

fibers

Protein toxins Batch fermentation and purification Cloning of toxin gene in microbialof a bacterial toxin, or extraction of host, extractiontoxin from a plant or animal source

Nonprotein toxins Extraction from plant or animal Cloning of a series of genes, eachsource governing production of one of the

enzymes needed to complete astep In the biosynthetic pathway

SOURCE: Office of Technology Assessment, 1993.

about 0.1 percent, making it unlikely that militar-ily significant quantities of toxin could be pro-duced by chemical synthesis.49 Biotechnologicalapproaches are possible but technically challeng-ing, involving the synthesis not of a single proteinbut of an entire series of enzymes, each necessaryto catalyze one step in a complex series ofreactions .50

Production techniques for the various types ofBTW agents are summarized in table 3-1. Withadvanced fermentation techniques availabletoday, a militarily significant supply of BTWagents could be produced over a period ofseveral days, obviating the need for the long-term stockpiling of agents. As a result, a BTWproduction facility might remain largely quies-cent in peacetime. After completing R&D,weaponization, and pilot-production tests onBTW agents, a proliferant could build productionand storage facilities and either keep them moth-balled or in use for legitimate commercial pur-poses. Clandestine production facilities might bekept in reserve, ready to be diverted to the rapidmanufacture of BTW agents in the event a major

conflict breaks out. Alternatively, a commercialproduction facility could be kept in operation andconverted to BTW production in wartime. Theadvantage of the latter option is that it would beeasier to retain the necessary trained staff andup-to-date equipment, albeit at some cost insecrecy.

CONTAINMENT MEASURESSince working with pathogenic microorgan-

isms is extremely hazardous, specialized physical-containment or ‘‘barrier’ measures are needed toprotect plant workers and the surrounding popula-tion from infection. Great care must be taken toprevent BTW agents from escaping from aproduction facility and causing a devastatingplague in the country producing the weapons.

In advanced industrial countries, work onhighly infectious microbial agents is carried outin high-centainment (Biosafety Level 3 or 4)facilities. In a BL3 facility, all personnel havebeen immunized against the infectious agentsthey work with. Since no vaccine is 100 percenteffective, however, they also wear protective

@ Muel L. Sanches et al., Chemical Weapons Convention (CWC) Signatures Analysis (Ml figto% VA: System plm g Corp, FinalTechnical Report No. 1396, August 1991), p. 89.

so ~05e, op. cit., footnote 42, p. 84.


clothing, goggles, and face masks. Microorgan-isms are manipulated in special biohazard safetycabinets maintained under “negative’ pressure(lower than the outside atmosphere), so that airflows into the work area. In addition to theseprimary barriers, secondary barriers to the spreadof infectious materials include the use of high-efficiency particulate air filters and the inciner-ation of exhaust.51 Because viral particles areabout a hundredth the size of bacteria, they aremore difficult to contain with filters and othermeans. Moreover, spore-forming bacteria (e.g.,the agents of anthrax and tetanus) foul theair-handling system with long-lived spores, whichcan easily contaminate other products. As a result,these bacteria are normally produced in separatefacilities. 52

A BL4 facility, the highest level of contain-ment, is designed to isolate the human operatorfrom the infectious agents. Since research ondangerous microbial pathogens requires handlingmuch lesser quantities of” hazardous material thandoes production, it is generally performed insmall BL4 enclosures inside a less stringent BL3facility .53 Such enclosures generally consist ofsealed boxes with rubber-glove ports that provideabsolute containment, and ‘hoodlines’ that makeit possible to move hazardous cultures directlyfrom a glove-box to an autoclave, which destroysthe infectious microorganisms with superheatedsteam. In a larger BL4 research or productionfacility, the human workers are isolated from themicrobes, rather than vice-versa. Each operatorwears a self-contained “space suit” and iscompletely isolated from the surrounding room,which is contaminated with infectious agents. Heor she enters the laboratory through an air-lockand hooks up to a supply of compressed air. Thesuit is kept under positive pressure so that if there

is any loss of physical integrity, the leaking airwill blow outwards, reducing the risk of infection.

A developing country seeking to developbiological weapons would probably use muchless elaborate containment measures. DuringWorld War II, for example, the Japanese Army’sUnit 731 produced vast quantities of highlyinfectious agents, yet the workers were protectedonly by wearing rubberized suits, masks, surgicalgloves, and rubber boots, and by receivingvaccinations against the agents they were work-ing with. The United Nations inspections of Iraqafter the 1991 Gulf War revealed that BTWresearchers in that country’s BTW program usedsurprisingly rudimentary containment measures,at the level of a BL2 facility. Laboratory techni-cians were vaccinated against the infectiousagents they worked with and used simple labora-tory hoods, but they did not wear masks orprotective clothing.

Commercially available containment systemsused for vaccine production might be suitable forcultivating highly pathogenic organisms. To com-ply with environmental and occupational-healthstandards and to ensure the purity of products forhuman use, many pharmaceutical plants carry outthe microbial production of antibiotics and otherdrugs in a “clean room” that is comparable to aformally designated BL4 facility. Clean rooms fordrug production are normally kept Under positivepressure to keep contaminants out, whereas areasof a vaccine plant used for the culture of infectious microorganisms are kept under negativepressure to prevent the dangerous microbes fromescaping. In principle, the direction of air flowcould be reversed, albeit with some difficulty.

Methods for sterilizing equipment after usewith hazardous microorganisms include physicalmeasures such as dry heat or pressurized, super-heated steam; ionizing radiation such as x-rays

‘1 Department of the Army, U.S. Army Medical Resetirch and Development Coremand, op. cit., footnote 5, p. 7-15.52 G- E&fi vice pr~ident for operations, Connaught Laboratories (Svfifhvater, IA), pUSO@ Commticatio% 1~.

53 Ha&fl M~covi@ “verification of High-Containment Facil.itiea,” S.J. Lund@ cd., Views on Possible Verification Measures for theBiological Weapons Convention (Oxford, England: SIPRI/Oxford University Preas, 1991), p. 55.


and high-energy ultraviolet radiation; and chemi-cal treatment with formaldehyde or bleach.54

Sophisticated biotechnology plants often haveself-sterilizing fermenters and process equipment,whether or not they are handling hazardous micro-organisms. Such plants would therefore have aninherent capability to work with BTW agents.

| Stabilization of BTW AgentsOnce BTW agents have been produced, it is

necessary to process them into a form thatenhances their stability in storage and afterdissemination, so that they remain viable longenough to infect. Since microbial pathogens areliving organisms, they will eventually deteriorateand die unless their metabolism is slowed downor stopped. Such a process of suspended anima-tion occurs naturally in the case of spore-formingmicroorganisms such as anthrax, which cansurvive for decades in the dorman t spore form.Nonspore-forming microbes and most toxins,however, tend to break down rapidly in theenvironment if not protected. For this reason,BTW agents are generally most effective ifdisseminated within a few days after production.If rapid use is not feasible, the live agents must beconverted into a more stable form so that they cansurvive the stresses of storage, transport, anddissemination.

FREEZE-DRYINGOne method for enhancing the stability of

BTW agents is rapid freezing and subsequentdehydration under a high vacuum, a processknown as freeze-drying or lyophilization. In a fewhours, a lyophilizer, a device mainly used in thepharmaceutical industry, reduces a solution ofbacteria and a sugar stabilizer to a small cake of

dried material that can then be milled into anydesired state of freeness. Lyophilization avoidsthe need to maintain microorganisms in incon-venient and dangerous liquid suspensions duringstorage and transportation. It also makes possiblea significant increase in agent potency by directinhalation of particles of dried agent into thelungs. 55 This technique is also applicable totoxins; a fine dust of dried toxin, if inhaled, can bedeadly in extremely small quantities.

If kept in cold storage, the desiccated orga-nisms will remain viable for long periods, al-though they still deteriorate. For example, freeze-dried brucellosis bacteria can be stored for severalmonths, and Q-fever rickettsiae for up to 8years. 56 Lyophilization alSO extends the shelf-life

of protein toxins: freeze-dried Staphylococcusenterotoxin B (SEB) can be stored for up to a year.Even so, the virulence and viability of lyophilizedBTW agents decays over time: there is a loss ofpotency of a factor of 10 to 100 over a period of1 to 5 years, so that much larger quantities of olderagent are required to produce the same militaryeffect. 57

CHEMICAL ADDITIVESThe stability of a microbial aerosol can be

increased by adding a variety of compounds to thespray material.58 Moreover, antiagglomerants suchas colloidal silica help prevent the clumping offreeze-dried microbial agents and toxins that havebeen milled into a fine powder. Agriculturalresearch on biological pesticides, such as theinsect-killing bacterium Bacillus thuringiensis,has provided much information on methods forstabilizing bacterial agents in the field. Forexample, new formulations of B. thuringiensishave been developed that extend the life of the

~ DW~ent of my, U.S, AI-My MedicaJ Research and Development Co remand, op. cit., footnote 5, pp. A-132, A-13-3.55 WIiH~ ad Wfice, op. cit., footnote 35, p. 72.

SC Rotichild, op. cit., foo~ote 9, pp. 206-219.

57 SPRI, VO1. VI, op. cit., foomote 31, p. 50.

5 8 Row J. Goodlo~ ~d F~~c A. ~~, “Viability and I.nfcctivity of Microorganisms in Experimental Airborne InfectiorL”Bacteriological Reviews, vol. 25, 1%1, p. 185.


disseminated bacteria by means of ultravioletprotestants and other additives that ensure com-patibility with existing agricultural sprayers.59

MICROENCAPSULATIONAnother approach to stabilization, known as

microencapsulation, emulates natural spore for-mation by coating droplets of pathogens orparticles of toxin with a thin coat of gelatin,sodium alginate, cellulose, or some other protec-tive material. (An industrial example of microen-capsulation is the production of carbonless carbonpaper, in which ink droplets are coated in thismanner.) Microencapsulation can be performedwith physical or chemical methods.60

Micromcapsulation production methods canbe set up to generate particles of a selected sizerange (e.g., 5 to 10 microns).6l The polymercoating protects the infectious agent againstenvironmental stresses such as desiccation, sun-light, freezing, and the mechanical stresses ofdissemination, and permits cold-storage of micro-bial pathogens for several months. Microcapsulescan be charged electrostatically to reduce particleclumping during dissemination, or ultraviolet-light blocking pigments can be added to themicrocapsule to protect microorganisms againstdegradation by sunlight. Once in the targetenvironment, such as the interior of the lung, thepolymer coating dissolves, releasing the agent.Microencapsulation can also be applied to toxins,making them more stable, predictable, and saferto handle.

| Integration With Delivery SystemsA biological or toxin agent is of little military

utility if it does not produce consistent andreliable effects and cannot be delivered to a target.BTW agents are all nonvolatile solids that wouldbe disseminated either as a liquid slurry or a drypowder of freeze-dried organisms or toxin.62

Possible delivery systems range in complexityand effectiveness from an agricultural sprayermounted on a truck to a specialized clusterwarhead carried on a ballistic missile. T h edifficulty of delivery-system development de-pends on the proliferant’s military objectives.It is not hard to spread BTW agents in anindiscriminate way for the purpose of produc-ing large numbers of casualties over a widearea. It is much more difficult, however, todevelop BTW munitions that have predictableor controllable military effects against pointtargets, such as troop concentrations on thebattlefield.

Many pathogens infect man naturally by meansof an intermediary organism (“vector’ ‘), such asa mosquito or tick.63 Military microbiologistsdiscovered during World War II, however, thatBTW agents can be disseminated through the air,making it possible to infect large numbers ofpeople simultaneously. Many microbial patho-gens and toxins-even those normally transmitt-ed by vectors or in food-can invade the bodythrough the lungs, giving rise to foci of infectionor traveling through the bloodstream to otherparts of the body. The key to producing large-

59 ~Ve~nt of the United Kingdom, ‘‘General Developments Relevant to the BWC,’ in Background Document on New Scientific andTechnological DevelopmentsRelevant to the Convention on the Prohibition of the Development, Production and Stockpiling ofBacteriological(Biological) and Toxin Weapons and on Their Destruction, Third Review Conference of the BWC (Genev& Switzerland), Document No.BWC/CONF.111/4, Aug. 26, 1991, p. 25.

@ -U OSO1 et al,, eds., Remington’s Pharmuceutica2 Sciences, 15th ed. (Eastou PA: M@ mbl- CO., 1975), P. 1604.

61 A ~c.n iS ~ ~ou~th of a fi~eter.

62 C.V. Chester and G. P. Ziummnaq ‘‘Civil Defense hnplieat.ions Of Biological WMPOIIS,” Journal of Civil De$ense, vol. 17, No, 6,December 1984, p. 6.

63 me disme v~~r is usually some type of arthropod: mosquitoes ~“t yellow fever and dengue fevw, fleas transmit plague; and tickstransmit tularernia and Q fever, During 1932-45, the Japan~e BW facility known as Unit 731 set up flea “nurseries” for the production of135 miUion plague-infested fleas every 4 months. As a delivexy systenA porcelain bombs were developed that could contain about 30,000infected fleas. See Williams and Wallace, op. cit., footnote 35, p. 27.

Chapter 3–Technical Aspects of Biological Weapon Proliferation | 95

scale respiratory infections is to generate abiological “aerosol”: a stable cloud of sus-pended microscopic droplets, each containingfrom one to thousands of bacterial or virusparticles. (Fogs and smokes are examples ofvisible aerosols.) Biological aerosols can beproduced with a relatively simple piece of ma-chinery, analogous to a home vaporizer, thatsprays a suspension of microorganisms throughfine nozzles, converting about 85 percent of theirstarting material into droplets in the desired sizerange. 64 The concentration of organisms in the

starting solution influences the distribution oforganisms among the aerosol particles.65

Aerosol dissemination of many vector-bornediseases, such as yellow fever, Rocky Mountainspotted fever, tularemia, and tick-borne encepha-litis, can produce atypical infections of therespiratory tract. Respiratory infection with suchagents bypasses normal protective mechanismssuch as local inflammatory processes and in-creases the virulence of pathogens that normallyhave a low lethality, such as Venezuelan equineencephalitis (VEE).66 In the case of microbialpathogens that can be transmitted by differentroutes, such as anthrax bacteria, respiratory infec-tion results in by far the most virulent form of thedisease. For example, whereas untreated skinanthrax is fatal in only about 5 percent of cases,pulmonary anthrax is fatal in more than 90percent of cases.67

Freeze-dried toxins can also be disseminated inthe form of an aerosol. Recent studies have shownthat saxitoxin and T-2 trichothecene mycotoxinare at least 10 times more toxic when adminis-

tered by aerosol than by intravenous injection.68

Because protein toxins are large organic mol-ecules, however, they are susceptible to environ-mental stresses such as heat, oxidation, and shearforces. As a result, attempts to aerosolize toxinshave encountered problems in maintainingg thestability of the agent before and after dissemina-tion. It is also difficult to formulate protein toxinscapable of penetrating the skin.

The primary challenge in weaponizing BTWagents for long-range delivery is to keep themalive long enough to infect enemy troops. Theagent must be capable of withstanding thephysical stresses involved in the disseminationprocess without losing activity. Technical hur-dles involved in the design of self-dispensingbiological weapons are as follows:

■

the munition or delivery system must gener-ate a cloud of aerosol particles with dimen-sions that allow them to be inhaled deep intothe lungs of the target personnel;the agent must be physically stabilized sothat it can survive the process of dissemina-tion long enough to infect the target person-nel;the agent must disseminated slowly to permitaerosolization while avoiding loss of viabil-ity or toxicity; andthe overall size and shape of the aerosolcloud and the concentration of agent withinit must be reasonably predictable, so that thedispersion pattern can be matched to thetarget.69

These technical hurdles are discussed below.

64 Wton Lcitenberg, “Biological Weapons, ’ Scientist and Citizen, vol. 9, No. 7, August-September 1%7, p. 157.

65 Goodlow and Leonard, op. cit., footnote 58, p. 1*4.

66 World He~~ org~=tiom Hea/~h A~pecr~ ~~C~em”ca/ and Biological Weapons (Geneva: W-lo, 1970), p. 61.

ST ~llcoff, op. cit., footnote 29.

68 Gove~ent of the U. S. S. R., “Selected Scientific and Technological Developments of Relevance to the BW Convention” op. cit.,footnote 43, p. A lO.

69 stoc~o~ Intemtio~ pe~e Re~em~h Imti~te (SrpRr), The Problem of chemical andBiOIOgical wa~are, VOI. II: CB wt?clpO?lS TOC@I

(Stockholm: Alrnqvist & WikeH, 1973), pp. 72-73.


EFFECT OF PARTICLE SIZEThe particle size of an aerosol is critical to both

its atmospheric stability and its military effective-ness. Whereas larger particles tend to settle out ofthe air, microscopic particles between one andfive microns in diameter form a stable aerosol inwhich the particles remain airborne for a longtime. The very low settling velocity of theparticles will by itself keep a biological aerosolcloud suspended in the air for long periods. Sucha cloud may therefore be transported by the windover long distances. Moreover, losses resultingfrom fallout and washout are negligible and donot significantly reduce the concentration of anaerosol cloud. Particles less than 5 microns indiameter generally do not collide with smoothsurfaces in their path but are carried over them byair currents. In contrast, transport over roughsurfaces for distances of more than a kilometercan result in significant deposition.

Aerosolized BTW agents generally do notpenetrate the skin and thus do not represent asignificant contact hazard; instead, they infectindividuals only if inhaled into the lungs .70Particle size is also critical for respiratory infec-tion. Almost all particles larger than 5 microns indiameter are trapped in the phlegm and passagesof the upper respiratory tract, while particlessmaller than 1 micron diameter are exhaledwithout being retained in the deep lung tissue.Only particles between 1 and 5 microns indiameter are small enough to reach the tinyterminal air sacs (alveoli) of the lung, bypassingthe body’s natural filtering and defense mecha-nisms. In one set of experiments on the effect ofparticle size on respiratory infection, tularemiabacteria were administered to guinea pigs as anaerosol. When the aerosol. particles were 1 micron

in diameter, only 3 bacterial cells per animal wereneeded to kill 50 percent of the guinea pigs, butwhen the particle size was increased to 7 microns,the number of bacteria per animal required to killhalf of the guinea pigs rose to 6,500.71

PHYSICAL STABILIZATIONThe use of mechanical devices to generate

aerosols from a bulk storage tank places a varietyof mechanical stresses on microorganisms, reduc-ing the number of viable, infectious cells. Rela-tively few microbial pathogens can meet thestability requirements of bulk dissemination.72

Those agents best suited for long-range attack caninfect with a small number of microorganismsand are hardy enough to survive for a fairly longperiod floating in the air. Once released, however,the aerosol cloud “decays” over time as themicroorganisms die as a result of exposure tooxygen, atmospheric pollutants, sunlight, anddesiccation, resulting in a loss of viability (abilityto survive and multiply) and virulence (ability tocause disease and injury). A BW agent dissemi-nated into a given environment may also retain itsviability while losing its virulence.73

Decay of an aerosol cloud occurs in two stages.Initial dissemination is followed by a period ofvery rapid cell death during the first severalseconds after the cloud has been released. Indeed,producing a liquid aerosol by explosive disper-sion or passage through a spray device may kill asmany as 95 percent of the microorganisms. Thisinitial stage is followed by a much slower rate ofdecay, so that the aerosol cloud may persist forlong periods of time. A relative humidity of over70 percent promotes microbial survival.74 Large-particle clouds are more resistant to the lethaleffects of solar radiation than small-particle

70 Ibid., p. 29.

71 ~j. WiU~ D. Sawy=, “Airborne rnfectiom” Military Medicine, February 1%3, pp. 90-92.

72 SIPRI, VOI. II, op. cit., footnote 69, p. 30.

73 *UP of com~~t ~p~ on ~efic~ ~d Bacteriolo@~ @iolo@@ WqXMM, Chem”cal ati Bacteriological (Biological)

Weapons and the Z2’ecrs of Their /Dossible Use, United Nations Report No. E.69.I.24 (New York NY: Ballantine Books, 1970), p. 13.74 r)q~ent of the AIIIy, U.S. Army Medical Research md Development CO mman~ op. cit., footnote 5, p. A7-16.

Chapter 3-Technical Aspects of Biological Weapon Proliferation! 97

clouds, and dry disseminated aerosols are moreresistant than wet aerosols.75 As the plumedisperses, long-lived particles (e.g., anthraxspores) may be deposited on the ground, wherethey may then adhere to large particles of surfacesoil and dust. If the surface is disturbed, either bythe wind or by human activities, the spores canagain be resuspended, potentially causing addi-tional infections.76 The inhalation hazard is muchreduced, however, owing to the large particle size.

TYPES OF AEROSOL ATTACKSThere are two types of aerosol dissemination of

BTW agents. “Area” attack involves releasingan aerosol cloud upwind and allowing it to driftover the target area. In contrast, “point” attackinvolves projecting the agent in a cannister thatreleases the agent immediately over the target.77

Area attackA BTW weapon designed for area attack would

disseminate its payload as an aerosol cloudcontaining a sufficient concentration of viablemicroorganisms to infect the targeted personnelwith particles in the 1 to 5 micron size range. Thesimplest means of area delivery is with spraytanks mounted on manned aircraft, unmannedremotely piloted vehicles (RPVs), or cruisemissiles, which can release a large quantity ofagent over a controlled line of flight. A slow-flying aircraft such as a crop duster coulddischarge a line of agent that, as it travelleddownwind, would reach the ground as a vast,elongated infective cloud. Such a linear cloud ofagent, known as a ‘‘line source, can cover alarger area than a cloud released from a singlespot, or ‘‘point source. ’

Air rushing past the spray tank can be used toforce out its contents; alternatively, compressedair or carbon dioxide may be used to disseminatethe agent. Aerosol generators might also beoperated from offshore ships or submarines paral-lel to a coastline, producing an invisible cloud ofBTW agents that could be carried by the prevail-ing winds over key coastal cities or militarybases. 78 The discharge rate must be slow enoughto generate a stable aerosol, yet slow-flyingaircraft and RPVs are extremely vulnerable to airdefenses.

Area attack with a biological aerosol dependsheavily on atmospheric diffusion and wind cur-rents to dilute and spread the agent over the areabeing attacked. The most stable atmosphericconditions occur on cold, clear nights or early inthe morning, when the ground and the layer of airabove it are cooler than the next higher 1ayer ofair. This phenomenon, called an inversion, isideal for the delivery of BTW agents because thestable interface of warm and cold air prevents thevertical mixing of the cloud and causes it to hugthe ground, keeping the organisms at a lowaltitude where they can be inhaled. In contrast,bright sunlight causes atmospheric turbulencethat breaks up the aerosol cloud, and also containsultraviolet rays that kill many microorganisms.For these reasons, a BTW attack would be mostlikely to come at dusk or at night.79

The effectiveness of an area attack also re-quires detailed knowledge of the prevailing winddirection and speed. Under favorable wind condi-tions, an aerosol cloud could contaminate the airover large areas, but if the wind is erratic orexcessively strong, the agent might fail to reachthe target or might be dissipated too rapidly to be

75 G@ow and bonard, op. cit., footnote 58, p. 184.

76 ~onB~Wige,~n~a/ation HUardfiom Reaero$o/izedBio/ogicalAgent$:A Review, Repod No. ~EC-l_’R~13 (A~rd@nprov@

Ground, MD: Chemical Research, Development and Engineering Center, September 1992).

77 SIPRI, vol. II, op. cit., footnote 69, p. 28.78 W, Seth cm, “The poor Man’s Atomic Bo&?”: Biological Weapons in the Middle East, Washington IDstitute Policy Papers No. 23

(WashingtorL DC: Washington Institute for Near East Policy, 1991), p. 11.79 ~ester ~d zimme= op. cit., footnote 62, P. 7.


effective. Assuming the target is nearby, theattackers will know the wind direction and canplan the attack at a favorable time. If the target isdeep inside enemy territory, local meteorologicalconditions would be harder to assess withoutaccess to current weather data. Nevertheless,hundreds of airports worldwide broadcast winddirection, speed, cloud cover, and temperatureevery 3 hours according to World MeteorologicalOrganization guidelines.

A remotely piloted vehicle or subsonic cruisemissile flying at low altitude might reduce suchproblems by disseminating the toxic cloudclose to the ground just upwind of the target—assuming, of course, that the attacker had themeans of knowing which way the wind wasblowing over the target area. In 1960, the U.S.Army began developing a drone aircraft thatcould be used to deliver chemical and biologicalweapons. The pilotless plane was designed tocarry 200 pounds of germ agents as far as 115miles. 80 Even relatively unsophisticated cruisemissiles might be capable of generating line-source aerosols for off-target attacks.81 For thisreason, the simultaneous proliferation of bio-logical weapons and cruise-missile capabilitiesmay become a major security threat in thefuture.

Point attackA point BTW attack would be performed with

munitions delivered by artillery, rockets, mis-siles, or aircraft. Although the targeted personnelwould be warned of the attack by the arrival of themunition, the rapid formation of a concentratedaerosol (within 15 to 30 seconds) means thatmany soldiers would inhale an incapacitating orlethal dose of agent before they had time to put ontheir gas masks properly, assuming they wereavailable. 82 A point attack that dropped the agent

directly over the targeted personnel would also bemuch less dependent on meteorological condi-tions, although it would require a much higherpayload of munitions per area covered.

MUNITIONS FOR POINT ATTACKMunitions developed for chemical-warfare agents

are generally unsuited for biological warfarebecause of the lower stability of BTW agents andtheir susceptibility to environmental factors suchas ultraviolet radiation and air pollution. Thereare two basic methods for disseminating BTWagents from a munition: explosive and pressur-ized. Whereas explosive dissemination producesan almost instantaneous build-up of aerosolconcentration over the target, it destroys a largeportion of the infectious agents and tends toproduce drops that are considerably larger thanthe optimal droplet size for inhalation.83 I ncontrast, pressurized munitions do not disperseagent as rapidly as explosive munitions butprovide better control of particle size, are gentleron the microorganisms, and produce an aerosolcloud that is visible for a shorter period of time.One method of pressurized delivery is to force aliquid suspension of agent through a fine nozzle,which breaks up the material into droplets of theappropriate size.

A BTW bomb or warhead may either befilled with bulk agent or with numerousself-dispensing cluster-bomb units (CBUs). Acluster bomb has a casing that breaks open duringdelivery to scatter a large number of smallersubmunitions over a wide area. The submunitionsthen fall to earth and are triggered to go off at analtitude of about 15 to 20 feet off the ground. Eachof these bomblets generates a small aerosol cloud;these multiple point sources are then coalesced byair currents into a single large cloud. DuringWorld War II, the United States and Great Britain

80 I-k@ op. cit., foo~ote 41, .P. 71.

al ~, op. cit., foomote 78, PP. 1o11.

82 Rowhild, op. cit., footnote 9, p. 76.

83 Ibid., p. 60.


jointly developed a 500-pound bomb for thedelivery of anthrax spores. Each bomb casingcontained 106 four-pound bomblets designed toburst in midair, producing dense aerosols ofspores. Twenty of these bomblets could cause ahigh fatality rate among humans and livestockover a one-square-mile area, and British war planscalled for using as many as 40,000 anthrax bombsagainst six German cities.84

Nevertheless, the technical difficulty of dis-pensing submunitions should not be underesti-mated. Missile delivery would place severe envi-ronmental stresses on microbial agents, includingfreezing temperatures prevailing at high altitudesand friction-heating of the missile noseconeduring reentry through the atmosphere, whichcould be fatal to microbes without sufficientinsulation. The timing of agent disseminationwould also be critical, since if it occurred at thewrong altitude, the agent would not form amilitarily effective aerosol. Releasing the agenttoo high would cause it to dissipate before it couldbe inhaled by the targeted troops; releasing it toolow would merely produce a puddle of toxicmaterial on the ground. In sum, effective agentdissemination requires a series of mechanicalsteps to work perfectly, and atmospheric con-ditions to cooperate.

Other problems associated with weaponizationinclude the hazards of loading munitions withagent, and corrosion and seepage from filledmunitions. Despite these technical hurdles, how-ever, effective biological munitions have beendeveloped and deployed in the past. In 1951, thefirst anticrop cluster bombs were placed inproduction for the U.S. Air Force; each bombletcontained turkey feathers centaminated with ce-

real rust spores.85 Also during the 1950s, theUnited States developed small, self-dispersingBTW bomblets for the Honest John missile.86

In conclusion, strategic BTW attacks againstcities might be carried out with relatively simpleoff-target delivery systems, such as a spray tankcarried by an aircraft. After dissemination, theaerosol cloud might behave in unexpected waysin response to changes in the wind and weather,potentially boomeranging against the attacker’sown troops or population.87 More controlled pointattacks with BTW agents for military purposeswould require the use of missiles or bombs,possibly equipped with cluster munitions, butsuch attacks would be technically more difficultto carry out.

Some analysts contend that the most probableuse of BTW agents would be for covert warfareagainst crops, livestock, or human populations forpurposes of economic destabilization. In thiscase, relatively small quantities of BTW agentsmight be introduced by human saboteurs directlyinto the air or water supply of a city or militaryinstallation.

INDICATORS OF BTW AGENTPRODUCTION

Detection and monitoring of BTW agent pro-duction is an extremely challenging task for thefollowing reasons:

■ All equipment and feedstock materials usedto make BTW agents is dual-use and couldbe used for legitimate purposes in the bio-technology or pharmaceutical industries.

■ Utilizing new biotechnologies, productioncould take place in facilities that are muchsmaller and less conspicuous than in the past,

84 Barton J. Berstein, “Churchill’s Secret Biological Weapons,” Bulletin of the Atomic Scientists, January/February 1987, pp. 4&50.

85 SIPRI, VO1. II, op. cit., footnote 69, p. 160.

86 Rothschild, op. cit., footnote 9, p. 78.

87 h 1942, dtig World War II, special Japanese troops spread diseases such as cholera, typhoi~ plague, ~d ~~ ~ -.Subsequently, more than 10,000 Japanese soldiers fell ill after they overran a contamma“ ted are% presuma bly because regular soldiers had notbeen informed about the use of biological weapons. Barend ter Haar, The Future of Biological Weapons, The Washington Papers No. 151 (NewYork NY: Praeger, 1991), p. 5.


with no obvious signs to indicate illicitactivity.

■ Legitimate facilities might be diverted to theproduction of BTW agents in a relativelyshort time.

■ Since microbial agents can be grown in anadvanced fermenter from a few cells to manykilograms of agent over a period of days orweeks, it would not be necessary to maintainlarge stockpiles (although filled or emptymunitions would need to be stored).

The extreme potency of BTW agents meansthat as little as a few kilograms can bemilitarily significant.

■ BTW agents would be hard to distinguishfrom naturally occurring pathogens, particu-larly if the agents are endemic to the affectedarea.

There is an enormous variety of potentialBTW agents, each requiring a specific detec-tion method.88

According to a State Department official, “Inmany ways, recent progress in biological technol-ogy increases the ease of concealment of illicitmanufacturing plants, particularly for biologi-cally derived chemicals such as toxins. . . . Notonly has the time from basic research to massproduction decreased but the ability to createagents and toxins with more optimal weaponpotential has increased. Simply put, the potentialfor undetected breakout from treaty constraintshas increased significantly.”89 Thus, while thecharacteristics of a given facility may be consist-ent with an offensive BTW program, the odds offinding a “smoking gun’’ -such as a munitionfilled with BTW agent—are quite low.

Despite these difficulties, however, a fewfactors constrain the detection problem. Micro-bial pathogens and toxins of military concern

have relatively few civilian uses in scientificresearch and medical therapy, and such applica-tions are generally confined to sophisticatedbiomedical research centers not often found indeveloping countries. As a result, there are someproduction and weaponization signatures that-ifintegrated effectively with national intelligencecollection and declarations of activities andfacilities relevant to the Biological and ToxinWeapons Convention-might provide strong cir-cumstantial evidence of a clandestine BTWprogram.

| Research and Development SignaturesMany research and development activities

related to BTW agents are inherently ambiguousin that they can support both defensive andoffensive purposes. It is therefore essential toevaluate the evidence in the context of a countryoverall behavior and the openness and transpar-ency of its nominally defensive program. Telltaleindicators of a BTW program might include theexistence of biological research facilities oper-ated under military control, the large-scale pro-duction of vaccines in excess of legitimatedomestic needs, or the purchase of dual-usebiological materials and equipment. Analystssearching for indicators of foreign BTW activitiesshould avoid “mirror-imaging’ ‘—the temptationto judge other countries by U.S. standards. Onlyby first understanding a country’s commercialstandards for biological containment and goodmanufacturing practice is it possible to identifyanomalies that do not fit this basic profile.90

SCIENTIFIC PUBLICATIONSThe collection and systematic analysis of

scientific and technical information published bya country of proliferation concern can help to

S6 stwhen S, Morse, ‘‘Strategies for Biological Weapons Verification” Proceedings of the Arms Control and Verification TechnologyConference, Williamsburg, VA, June 1992 (Washingtoxq DC: Defense Nuclear Ageney, in press.)

69 H. Allen Ho~es, “Biolo@c~ WM~ns proliferatio~” Department of Sfate Bufkrin, VO1. 89, JulY 1989, p. 43.

w Graham S. Pearsoq ‘Biological Weapons: ~ British View,’ p=sentation at a Seminar on Biological Weapons in the 1990s, sponsoredby the Center for Strategic and International Studies, Washington DC, Nov. 4, 1992.

Chapter 3-Technical Aspects of Biological Weapon Proliferation I 101

monitor research trends, identify institutions andscientists associated with such research activities(including cooperation with foreign states), andidentify gaps or abrupt halts in open research onparticular topics that may be suggestive ofmilitary censorship. Although such literatureanalysis is unlikely to reveal BTW activities thathave been deliberately concealed, it can raisequestions about the capabilities and activities ofvarious facilities and is useful when combinedwith other sources of information. For example,useful intelligence on Soviet BW activities wasreportedly gleaned from clues picked up in theSoviet scientific literature. By tracking the awardof academic honors and by noticing obvious gapsin a series of published papers, Western intelli-gence analysts could judge which fields ofbiological research Soviet military scientists hadentered.91 During the 1970s and ‘80s, for exam-ple, the Soviet literature contained a remarkablylarge number of publications on toxin research.92

Nevertheless, publication tracking is not areliable indicator of a BTW program. First, itrequires the existence of a preliminary scientificresearch effort before a country undertakes pro-duction of BTW agents. In the past, countries withoffensive BTW programs, such as Japan, Ger-many, the United States, and the Soviet Union,launched a major scientific research effort inrelevant areas of microbiology before they coulddevelop biological weapons. Today, however,much of the basic science is already well under-stood, and explicit weapon-development pro-grams could be undertaken with relatively littlepreliminary research.

Second, because many variables affect scien-tific productivity and publication rates, publica-

tion tracking can only provide a broad indicationof a state’s BTW activities. The scientific culturein many developing countries does not demandlarge numbers of publications, so there will befewer to monitor. Finally, the biomedical data-bases available for tracking (e.g., MEDLINE) donot have representative numbers of scientificabstracts from the countries of greatest prolifera-tion concern, particularly for papers not publishedin English.93 For this reason, the number ofabstracts in the database dealing with microbialand toxin agents may not correlate either posi-tively or negatively with a country’s BTWactivities. For all these reasons, publication analy-sis is an unreliable-although potentially useful—indicator of BTW activity.

HUMAN INTELLIGENCEBecause of the limited value of technical

collection systems such as satellites for monitor-ing BTW activities, human intelligence is vital inthis area. For example, Vladimir Pasechnik, aRussian microbiologist with first-hand knowl-edge of the Soviet BTW program, defected toBritain in 1989 while attending a scientificconference in London and provided extensiveinformation on Soviet BTW activities that wasunobtainable by other means.94 Human agentsand defectors can also confirm suspicions aboutsites and activities based on other sources ofintelligence. Nevertheless, the UNSCOM biolog-ical inspections of Iraq have shown that humanintelligence can be unreliable or misleading if theindividual reporting does not have direct knowl-edge or is unfamiliar with the technical details ofwhat he is describing.95

91 Harris and Paxman, op. cit., foomote 14, p. 114.92 David B&r, “~emi~~ ~d Biologic~ w~~e Agen~_AFre~App~~~’ Ja~e’~J~re//~ge~ceReview, VO]. s, ~0, ~, ~a13~ 199s,

p. 44.

w ~licoff, op. cit., foomote 29.

94 Bin Gem, “Defecting Russian Scientist Revealed Biological - ~ofis,” The Washington Times, July 4, 1992, p. A4.95 ~lqhone fitwiw ~~ David H~soll, de- S~hOol of Vete- Medicine, ~~.s@ Swte Un.iversiv, Baton Rouge, ~, Aug. 17,

1992.


| Weaponization and Testing SignaturesWeaponization involves the determination of

whether a candidate BTW agent is militarilyeffective and how it would be used. Theseactivities have no obvious civilian counterpart,and hence would be indicative of a clandestineBTW development program. Signatures associ-ated with the testing of candidate BTW agentsmight be easier to detect than agent productionsignatures, and inspections focusing on weaponi-zation activities would also be more acceptable tothe biotechnology industry than production moni-toring, since they would not compromise tradesecrets.

Examples of weaponization and testing signa-tures that might be observable with overheadphotography include field tests of aerosol disper-sal patterns, tests of effectiveness against largeanimals, and the surreptitious burial of deadanimals from weaponization tests .96 Other weap-onization signatures would only be visibleduring an onsite inspection. For example, spe-cialized aerosol test chambers might be used tostudy the behavior of biological aerosols in theenvironment or the detonation of BTW muni-tions. If such a test facility were found in anominally civilian vaccine plant, it would beextremely suspicious. Indeed, the secret SovietBW facility at the All-Union Research Institute ofApplied Microbiology in Oblensk reportedlycontained two such test chambers. The “aerosol-dissemination test chamber’ consisted of a steelcube, roughly 50 feet on a side, in whichexperimental animals were tethered to the floorand exposed to BW aerosols released from ceilingvents. There was also a reinforced ‘‘explosive-test chamber’ in which the detonation of BWmunitions was simulated.97

Although some analysts contend that weap-onization could be carried out in enclosed,unobtrusive facilities, others argue that theintegration of signatures from a variety ofsources would make a militarily significantweaponization program difficult to hide. Nev-ertheless, there are a number of potential conceal-ment strategies:

Weaponization studies short of actual fieldtests could be performed inside productionfacilities.Open-air testing of BTW weapons would bedifficult to detect if the test grid weremasked, or if there were no distinctivedelivery systems or advance indications ofwhere to look. In addition, since many BTWagents are sensitive to sunlight, they wouldbe tested at night.98

Certain legitimate activities, such as thedissemination of biopesticides on crops, orthe use of conventional smoke bombs, mightbe used as a cover for BTW weaponizationtesting.

growing number of countries, for example,are replacing chemical pesticides with certainmicroorganisms that are natural insect-killers.Although over 100 bacteria, fungi, and virusesinfect insects, only a very few are in commercialproduction. The most widely used is Bacillusthuringiensis, a bacterium that produces a crystal-line protein toxic to insects and is applied to cropsas an aerosol from agricultural sprayers.99 Al-though B. thuringiensis does not have the charac-teristics of a BW agent and would be a poorsimulant for weaponization studies, field testswith bacteria more closely resembling BW agentscould be disguised as biopesticide application.Thus, if biopesticides are already in use in the

96 Rst rmges will Mely be at remote locations. The former Soviet UnioW for example, operated a BTW test site on isolated VO~fi-deniye Island in the Aral Sea.

97 John Bamy, “P1 arming a Plague?” Newsweek, Feb. 1, 1993, p. 40.

‘3S J~es -s, Engines of Wlzr: Merchants of Death and the New Arms Race (New Yor& NY: Atlantic Monthly press, 1990), P. 221.

99 primrose, op. cit., footnote 42, P. 76+


local agricultural sector, a covert proliferant coulduse them to test the open-air dissemination ofmicrobial aerosols in preparation for germ war-fare, and also to justify the acquisition of hard-ware needed to disperse the agent. Nevertheless,substantial field tests using grams or kilograms ofmicron-sized particles could be detected in sam-ples taken at great distances downwind if theorganism had distinctive DNA sequences andthose doing the detection knew what to look for.

| Production SignaturesProduction of BTW agents is nearly impossible

to detect by visual inspection alone, although thisapproach may add some useful pieces to thepuzzle.

EXTERNAL VISUAL SIGNATURESBTW production facilities may sometimes be

detected or monitored with overhead photogra-phy, although the evidence is nearly alwaysambiguous. For example, the Institute of Microbi-ology and Virology in the Russian city ofSverdlovsk, 850 miles east of Moscow, report-edly aroused the suspicions of U.S. intelligenceanalysts in the 1970s because of certain character-istics observed by satellite. Photointerpretersidentified tall incinerator stacks, large cold-storage facilities, animal pens, sentries, anddouble barbed-wire fences. These features, notunlike those at the former U.S. offensive BTWfacility at Fort Detrick, suggested that theSverdlovsk facility might serve military pur-poses. 100 In Much 1980, the U.S. Governmentattributed a serious outbreak of pulmonary an-thrax in Sverdlovsk the previous year to SovietBW activities at the microbiology institute. Al-though Soviet officials denied the allegations atthe time, in May 1992 Russian President Boris

Yeltsin finally admitted that the anthrax epidemichad been caused by an accident at the militaryfacility. Russian military spokesmen have in-sisted, however, that the anthrax work atSverdlovsk was “defensive” in nature, and thisquestion has yet to be resolved. l0l

Recent innovations in biotechnological pro-duction technology aimed at increasing produc-tivity, cutting costs, and improving safety havefurther blurred distinctions important for verifica-tion, such as between a laboratory and a produc-tion facility. In the past, large batch fermentersand refrigerated storage vaults provided signa-tures of BTW production; today they are beingreplaced in advanced industrial countries withsmall, continuous-flow fermenters that can pro-duce large quantities of highly infectious materi-als rapidly in a laboratory-scale facility. Suchadvances in production technology have greatlyincreased the difficulty of detection by reduc-ing the size of plants needed to producemilitarily significant quantities of BTW agentsand the amount of time needed to break out ofdisarmed status. With the new production tech-nologies, a clandestine BTW plant might be moreeasily camouflaged, buried underground, or hid-den within a larger complex that produces legiti-mate commercial products.

Still, satellite or aerial photography might helpto monitor sites judged suspicious on the basis ofother sources of intelligence. The followingsignatures of BTW agent production might bedetected or monitored with overhead imaging:

“Excessive” secrecy and security surround-ing a nominally civilian microbiologicalfacility, such as a brewery, sugar refinery,infant-formula plant, or single-cell proteinplant. Telltale security measures might in-clude double or triple fencing, watch towers,

IW Elisa D. Harris, “Sverdlovsk and Yellow Rain: Two Cases of Soviet Noncompliance?” International Secun”fy, vol. 11, No. 4, spring1987, pp. 41-95.

Iol See Milton Leitenberg, “A Return to Sverdlovsk: Allegations of Soviet Activities Related to Biological Weapons,’ Arms Control, vol.12, No. 2, September 1991 ($mblished April 1992), pp. 161-190; and Milton Leitenberg, “Anthraz in Sverdlovsk: New Pieees to the Puzzle, ”Arms Control Toa2zy, vol. 22, No. 3, April 1992, pp. 1013.


and air-defense missile batteries-althoughconcealment, camouflage, and deception op-erations are possible.Elaborate microbiological production facili-ties inconsistent with the level of sophistica-tion of other, clearly civilian plants.Facilities for housing large numbers ofprimates, horses, rats, mice, rabbits, sheep,goats, or chickens (for producing eggs),when such animals are not clearly associatedwith vaccine production.Changes in activity at nominally civilianproduction facilities.

PLANT DESIGN AND LAYOUTOther signatures of BTW production would not

be visible from outside a suspect facility, and thuscould only be detected during an intrusive onsiteinspection. The basic equipment in a BTWproduction facility would be much the same asthat in a vaccine plant, including equipment andmaterials for microbial fermentation, cell culture,or egg incubation, followed by harvesting, purifi-cation, and lyophilization. Both a vaccine plantand a BTW production facility would require asource of pharmaceutical-grade distilled water toremove bacterial contaminants in tap water thatwould interfere with the growth of desired micro-bial agents. And both types of facilities wouldrequire autoclaves to sterilize the growth mediaand decontaminate the equipment after produc-tion.

It is also important to evaluate BTW-relatedactivities in their socioeconomic context. Indeveloped countries, civilian production facilitiesthat utilize microorganisms, such as pharmaceuti-cal plants and even breweries, now incorporatesafety and environmental equipment that wereonce unique to BTW production facilities. Thisfact has made it easier to use commercial produc-tion as a cover for illicit military work, although

the presence in a vaccine-production plant ofprocesses that cannot be justified on technical oreconomic grounds may provide indicators ofpossible conversion or diversion to BTW produc-tion. Nevertheless, a clandestine BTW productionfacility could be so small that it could be easilyhidden.

PHYSICAL CONTAINMENTIn developed countries, an important difference

between a vaccine plant and a BTW productionfacility may be the level and type of physicalcontainment measures that are employed. Threeaspects of physical containment might be sugges-tive of a clandestine BTW program:

First, production of vaccines involves the useof living, attenuated microbial strains that areeither further weakened to produce a live vaccineor killed immediately after cultivation. As aresult, stringent containment measures are re-quired only during the initial phase of vaccineproduction, which involves the cultivation ofagents before they are attenuated or killed.102

According to one assessment, “Extensive safetyprecautions during the whole production cycle fora vaccine are hardly defensible economically andwould hence be suspect. ’ ’103

A second difference between a vaccine plantand a BTW production facility is the presence inthe former of costly measures to protect thepurity, sterility, and reproducibility of the prod-ucts so that they are suitable for human use. Thus,an indicator of illicit BTW production in apharmaceutical facility might be the absence ofmeasures to purify the product and ensure itssterility.

Although BTW agents would probably becultivated under negative pressure to preventdangerous microbes from escaping from theplant, this would not be a reliable signaturebecause negative pressure would also be needed

In David L, H~ol~ ~@ El. &ott, ~d Wik C. RI@icIK III, “Medicine in Defense Agtit Biological Wtime,’ yOIUd Of theAmerican Medical Association, vol. 262, No. 5, Aug. 4, 1989, p. 679.

103 SH, VOI. VI, op. cit., footnote 31, p. 45.


in a legitimate pharmaceutical plant that isproducing live, attenuated vaccines. It would alsobe possible to reverse the pressurization of afacility before an inspection by changing thedirection of flow of the faltered air, but only if thesystem were engineered for this purpose inadvance. Moreover, governments engaged in thecovert production of BTW agents would probablynot hesitate to cut comers on containment andworker safety in order to avoid signaling theirintentions.

In addition to the type and level of physicalcontainment, several other potential signatures ofclandestine BTW production might be detectedduring an onsite inspection. l04 The utility of suchsignatures is highly controversial, however, andeach one is open to criticism. These signaturesmight include:

Bad odors associated with microbial fermen-tation, since multiplying bacteria produce avariety of volatile and odiferous gases.However, odors do not travel far and arenonspecific.Seed stocks and cell lines inappropriate fordeclared activities such as production ofvaccines or single-cell protein, or in amountsexceeding immediate research needs. How-ever, such materials would probably not bedeclared and could be easily hidden.Activities related to microorganisms andtoxins that cannot be explained by civilianneeds, such as development of vaccinesagainst rare, nonindigenous disease agents.However, such activities could be easilyhidden.Production capacity greatly in excess ofdemand for the plant’s legitimate products,such as vaccines. However, such excesscapacity would not be required for a BTWcapability.

A discrepancy between a small quantity ofoutput product (e.g., packaged vials of vac-cine) and a large quantity of input materials(e.g., fermentation media). However, calcu-lation of a precise material balance is proba-bly impossible.Air compressors, air tanks, or lines forair-supplied protective suits as a means ofenhancing physical containment. However,compressors are easily hidden.Facilities for rapid decontamination andcleaning of the production line, or evidenceof recent large-scale decontamination opera-tions, fumigation, or removal of materials orequipment. However, decontamination rou-tinely occurs in pharmaceutical facilities.Large stockpiles of bleach (sodium hy-pochlorite or sodium hydroxide) for use as adecontaminating agent. However, such agentsare also widely used in legitimate commer-cial facilities.Specialized equipment for the lyophiliza-tion, milling, or microencapsulation of BTWagents. However, lyophilization machinesare ubiquitous in the pharmaceutical indus-

W.Refrigerated storage bunkers, freezers, orlarge quantities of liquid nitrogen for storingstockpiles of live or freeze-dried pathogens.However, since significant quantities ofbiological agents can be grown relativelyquickly, long-term stockpiling of agents inrefrigerated bunkers is unnecessary.Anomalous transport of microbial productsor wastes off-site. However, microbial wastecan be steam-sterilized.Incomplete or anomalous plant productionrecords. However, a proliferant engaged inillicit production is unlikely to provide suchrecords voluntarily. Records might also be

1~ F@emtion of herican Scientists, Working Group on Biological and lbxin Weapons Verii3catioQ ‘‘A hgally BiIMI@ ComPl~ceRegime for the Biological Weapons Convention: Reftiment of Proposed Measures Through Trial Facility Visits,” draft manuscrip~ March1992, p. 12. See also SIPRI, vol. VI, op. cit., footnote 31, p. 35.


forged, although it is hard to do so convinc-ingly.

Still, although the indicators listed abovewould not necessarily be associated with illicitproduction activities, a pattern of them mightarouse suspicions.

BIOCHEMICAL SIGNATURESBTW agents do not possess a single common

chemical signature, such as the phosphorus-methyl bond characteristic of most nerve agents,but pathogenic microorganisms can be identifiedin minute quantities using sensitive immunologi-cal, biochemical, and genetic techniques. (Seebox 3-C, pp. 108-109.) Telltale traces of DNAfrom virulent strains of bacteria and viruses mightbe discovered in samples collected during anonsite inspection of a suspect site, even after thefacility had undergone decontamination. Suchtraces might be indicative of previous research orproduction activities at the facility. Fermentersalso generate large quantities of liquid wastes thatmight contain unusual metabolic byproducts andother telltale chemicals even after decontamina-tion treatment.

Nevertheless, the fact that certain toxins andmicrobial pathogens have defensive or medicaluses means that it can be difficult to distinguishlegitimate from illicit BTW activities. Toxinsalso differ from chemical-warfare agents in thatthey do not leave persistent traces in the environ-ment and are easily destroyed by autoclaving withsuperheated steam. The extent to which heat-neutralized protein toxins could be detected byimmunological methods is unknown. Finally, theability of modem analytical techniques to detecttrace amounts of biological organisms couldmake legitimate biotechnology facilities reluctantto submit to such intrusive inspection for fear of

losing proprietary information. Analytical instru-ments could probably be “blinded,” however, todetect only the presence or absence of knownBTW agents.

BIOMARKERSSince the workforce in a BTW production

facility would likely be immunized against theagents being produced, another approach toverification would be to determine whether theblood of workers in a suspect fermentation orvaccine facility contains antibodies against knownBTW agents. Monitoring of immunization pro-grams would involve taking blood samples fromplant workers for onsite immunological analysis.Another approach would be to take blood samplesfrom wild animals (e.g., rodents) in the vicinity ofa suspect facility to detect possible exposure tounusual infectious agents.

Although most vaccine production plants re-quire all workers to undergo initial and periodicblood collection and analysis, it would be difficultto negotiate a verification regime that requiressuch intrusive inspections. Furthermore, perform-ing such tests as part of routine onsite inspectionsmight violate U.S. constitutional protectionsagainst “unreasonable searches and seizures,”since it would be difficult to protect confidentialmedical information unrelated to the purpose ofthe inspection. According to one legal scholar,“No treaty could empower inspectors to conductrandom intrusive body searches for possibletelltale evidence of radiation or biological weap-ons. “105 Another analyst argues, however, thatbiomedical sampling might be upheld by thecourts on grounds of national security if thereis a clear connection between the objectives ofthe regime and the analysis of biochemicalindicators. l06

105 David A, Koplow, “Arms Control Inspection: Constitutional Restrictions on ‘IYeaty Verification in the United States,” New YorkUniversity Luw Review, vol. 66, May 1988, p. 355.

1~ Jq R. S@ckto~ Edward A. Tanzman, and Barry Ke- Harmonizing the Chemical Weapons Convention With the Um”ted StatesConstitution (McLeq VA: BDM International, Report No. DNA-TR-91-216, April 1992), p. 59.


| Stockpile and Delivery System SignaturesStockpiling of agent or loading into sprayers,

munitions, or other delivery systems might beassociated with a number of signatures. Thefollowing might be observable by aerial orsatellite photography:

■

■

■

cold storage of bulk BTW agents in refriger-ated bunkers or igloos, although small quan-tities of stored agent would probably not bedetected;storage depots for BTW-capable munitionsand delivery systems in proximity to possi-ble production facilities; andheavy trucks for the transport of empty orfilled munitions in the vicinity of a biologi-cal production facility.

The remaining signatures could only be de-tected during an onsite inspection:

inappropriate metal-working equipment orstock that might be used to fashion muni-tions;specialized equipment for filling BTW agentsinto munitions and warheads;breeding of insect vectors, or acquisition ofequipment for disseminating biological agentsand toxins as an aerosol cloud;munitions or parts thereof for disseminatingBTW agents; andthe training of troops in the tactical use ofBTW agents.

| Weapon Use SignaturesBTW agents might either be used deliberately

or escape accidentally from a secret militaryresearch or production facility, as happened in theSoviet city of Sverdlovsk. It is therefore impor-

tant to determine whether an outbreak of infec-tious disease in an area where it is not endemic isthe result of clandestine biological warfare activi-ties and, if possible, to identify its source. Fieldepidemiology can help investigate alleged casesof biological and toxin warfare.107 Indeed, theCenters for Disease Control’s Epidemic Intelli-gence Service was originally founded in 1951 outof concern that terrorists or foreign intelligenceagencies might launch a covert BTW attackagainst the United States.l08

As a first step, all of the likely natural causes ofan epidemic must be investigated and excluded—a difficult task given the enormous variability ofinfectious diseases. Covert attacks aimed ateconomic sabotage are most likely to involveanimal or plant pathogens. The best known caseof a suspicious epidemic took place in 1981 inCuba, which suffered a severe outbreak of denguefever, a mosquito-borne viral illness. Of theestimated 350,000 people who developed thedisease, approximately 10,000 suffered fromsevere (hemorrhagic) symptoms and 158 died, amortality rate of 1.6 percent.l09 Cuban PresidentFidel Castro blamed the epidemic on covert U.S.biological warfare, which he alleged was beingrun by the Central Intelligence Agency. Epidemi-ological analysis indicated, however, that theoutbreak was of natural origin. The Cubanepidemic occurred a few months after a majoroutbreak of hemorrhagic dengue fever in South-east Asia. Epidemiologists determined that Cubanconstruction workers building a hotel in Hanoi,Vietnam, had become infected with the disease.After returning home to Cuba, they were bitten by

WI Peter B~s, “Epid~c Fie]d hvestigation as Applied to Allegations of Chemierd, Biological, or lbxhl W@ire, ” PO/itiCS andthe Lz~eSciences, vol. 11, No. 1, February 1992, pp. 5-22.

1~ Stephen S. Morse, ‘‘Epidemiological Surveillance for Investigating Chemical or Biological Warfare and for Improving Human Heal@’ ‘Politics and the Life Sciences, vol. 11, No, 1, February 1992, pp. 28-29.

1~ Jay P. Sanford, ‘‘Arbovirus Infections,’ Eugene Braunwald et al., Harrison ’s Principles oflnternal Medicine, 1 Ith ed. (New YoX NY:McGraw-Hill, 1987), p. 727.


Box 3-C-Biochemical Detection of BTW Agents

Inspections of microbioiogical laboratories or production facilities for indications of BTW activities mightinvolve the collection and analysis of samples to detect the presence of undeclared pathogens or toxins. Suchsamples might include wipes from equipment and air filters or liquid samples from the waste stream or theenvironment near the plant Alternatively, air filters might be used to screen large volumes of air inside, or evenat a considerable distance from, a plant.

Immunological techniques. The fastest method for detecting pathogens is to use specic antibodies that havebeen Iabelled with a tag of some type, such as a fluorescent molecule or a radioactive atom. Such antibodies wouldbind to the pathogen with high specificity, providing nearly unambiguous evidence of its presence. Techniquesfor producing large quantities of “monocional” antibodies, all specific to a single marker protein on the surface ofa microorganism, permit the detection and identification of minute quantities of bacterial and viral agents (andprotein toxins) in Mood or environmental samples. Such immunological screening techniques includeradioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA). A drawback of such assays is thatthey could not identify BTW agents that had been genetically modified to alter their immunological characteristics.

Bioassays. A pathogen or toxin can be detected by measuring its physiological effects on intact organismsor on isolated cell or enzyme systems. For example, many toxins work by specifically inhibiting the enzymeacetylcholinesterase involved in nerve transmission, thereby reducing its ability to break down the messengerchemical acetylcholine. Devices known as ‘biosensors” are under development that use receptor molecules orenzymes immobilized on the surface of a chip to detect the binding of toxins or viral agents. One biosensor capableof detecting toxins, developed by engineers at Arthur D. Little, is moving into a manufacturing prototype.1

Genetic analysis. The advent of recombinant-DNA techniques has made it possible to identify minusculequantities of microorganisms in complex samples.The first step is to prepare standards by isolating single-strandDNA sequences from specific microorganisms or synthesizing them chemically and labeling them with aradioactive isotope or a fluorescent dye. These labeled DNA fragments, known as “DNA probes,” can pair up or“hybridize” with DNA in a sample if the sample contains DNA from the same microorganism. The advantage ofDNA probes is their unique specificity, which enables them to identify a single type of pathogen even in complexmixtures.

Because of background noise, it can be difficult to detect probe/target hybrids when only a small number ofmicroorganisms are present in the sample. This problem was solved in 19$5 with the development of a powerfulnew technique known as polymerase chain reaction (PCR), which can amplify a given DNA sequence as muchas 1O8 times. PCR therefore makes it posssible to use DNA probes to identify pathogens present in tracequantities-as few as tensor hundreds of microorganisms-without having to grow them into larger colonies overa period of days or weeks. Since PCR reagents are available in kit form, this technique has greatly speeded thediagnosis of infectious diseases, including potential BW agents such as anthrax bacteria.2 PCR is also useful foranalyzing biological samples in the field or during an onsite inspection of a suspect facility it has even detectedkilled bacteria in autoclaved samples?

1 Ri~a~F.Tayl~,~h~D. IJttie Corp., Ciportabie, Real-ThneB iosensorsforChemicai AgentVerifioation,”presentation at the Chemical Wafxms Convention Vedfioation Teohnoiogy Researoh and DevelopmentConference, Herndon, VA, Mar. 2-3,1993.

2 ~~~~ of ogf~~, A~~@ fkpcwf to CongraIss on the IWeamh, w-t ~- and ~-ju~t~onof ttm Chemloa~idogical LWWse Program * the Petfod October 1, 19$0 77nwgh Septemher 30, 7991,RCS:DD-USDRE(A) 1085, p. 51. See also, M. Carl et al., CIDeteotlon of spores of Bad#us anthrads using thepolymerase chain reaotion;’ Journa/of/nib@ous IXseas~ voi. 185, 1992, pp. 1145-1148.

3 T. ~rry and F. (3annon, W)irgot C3enomio DNA Arnpiifkation From Autooiaved Infeotlws -OOrgankrnsUsing PCR T~hnology,” PCR A4ethoo% and A#oatAww, VOL 1,1991, p. 75.


Nevertheless, PCR has some limitations. First, it is only suitable for identifying known organisms, since onemust decide in advance which DNA sequences to use as probes. Second, because many pathogenic microbes(e.g., anthrax bacteria) are ubiquitous in the environment in trace amounts, a probe of sufficient sensitivity mayfind “prohibited” DNA everywhere! Third, the accuracy of PCR depends on both the length of the target DNAsequence and the length of the PCR “primers,” which bind to the target DNA to initiate the amplification process.As one tests for shorter sequences (e.g., 100 instead of 1,000 DNA base-pairs), the sensitivity of the techniqueincreases but its specificity declines, since several different microbial species may have identical short DNAsequences. For this reason, two levels of detection have been proposed, depending on the characteristics of theDNA probe. The first level would identify the group of pathogenic bacteria to which a suspect agent belongs bydetecting a DNA sequence common to all species in that group; the second level would provide species-specificidentification by using longer DNA probes specific to each microorganism targeted for detection.5

Genetic fingerprinting. Known technically as restriction-fragment length polymorphism (RFLP) analysis, thistechnique involves the use of special “restriction” enzymes that cut microbial DNA at specific sites. This treatmentresults in a pattern of DNA fragments of different sizes, which can be analyzed by separating the fragments ona gel. Genetic fingerprinting can also be done with RNA viruses. The result of this technique is a characteristicpattern of spots on the gel. Since different DNA sequences will result in a different pattern of spots, comparingsuch maps will reveal the extent to which two strains of a bacterium or virus differ genetically.

All microbial pathogens can be “fingerprinted” by analyzing their genetic material (DNA or RNA). Since thereare always minor genetic differences among various strains of a pathogenic microorganism, it is very likely thata laboratory-developed strain is genetically distinct from an indigenous strain. Moreover, an indigenous strain thathas been produced in large quantities is likely to be more genetically homogeneous than the causative agent ofa natural epidemic. For many microbial pathogens, scientists have compiled a library of characterized strains t hatcan be compared with any newly discovered strain. Thus, genetic fingerprinting often provides enough informationto determine the source of a virus and whether it has been modified genetically in the laboratory. Trace amountsof genetic material can be amplified for further analysis using PCR.

Town analysis. For toxins, analytical techniques for detection and characterization include immunoassay, aswell as analytical chemistry techniques such as combined gas chromatography/mass spectrometry (GC/MS),liquid chromatography, and others. Quadruple mass spectrometry is used to analyze protein toxins, as well assamples dissolved in water.6 Very large biomolecules must be broken down into smaller components for analysis.To this end, a technique known as pyrolysis mass spectrometry involves heating complex materials in a controlledmanner to generate characteristic chemical signatures that can then be analyzed by a mass spectrometer.7 Thesesignatures are compared with a large computer database of known chemical spectra to identify the compoundspresent Finally, if a pure sample of a protein toxin or peptide bioregulator is available, it may also be possible toidentify it from its amino-acid sequence. Off-site detection of toxins is nearly impossible, however, because theylack volatility.

4 Moreover, a laboratory might detect a contaminant from past rather than current WO*, SUM as anthraxspores from earlier samples.

5 BarbaraJ. Mann, De@cfjono~Bb/o@a/ WarfareAgenfs Ush?gl the Po/ymerase Chah? l?eacfion (ResearchTriangle Park NC: Battelle Memorial Institute, September 1992), DTIC No. AD-A259391.

6 External Affajrs and International Trade Can-Verification Research Unit, V&lf&NIOn: De@opmnf ofa Porfab/e Trichothecene SensorK7f fortf?e Defecfkm of T-2 f@cofox/n In Human B/oodSampfes (Ottawa: ExternalAffairs, March 1987).

7 Diane M. Kotras, “New Detection Approaches for Chemical and Biological Defense,” ArmY Resear@,Development and~cquisltion Bu//etin, January-February 1989, p. 2.


mosquitoes, which then transmitted the disease toothers. 110

Another suspicious epidemic that still remainsto be explained took place during the civil war inRhodesia (now Zimbabwe) from 1978 to 1980.An unprecedented outbreak of cattle anthrax wasalmost entirely confined to the Tribal TrustLands-the areas then assigned to Rhodesia’sblacks and accounting for about 17 percent of thecountry’s land area. 111 Since cattle were the

primary source of wealth for black farmers, theepidemic led to the severe impoverishment of the—

ected rural populations. The outbreak of cattlethrax was accompanied by a secondary humanidemic, which resulted in more than 10,000

infections and 182 human deaths. Since anthrax isnot contagious from one individual to another, theexplosive nature of the human epidemic wasstriking: the reported incidence of human anthraxcases during the 1979-80 period was more than400 times the average incidence of the previous29 years. Some epidemiologists believe that thelosing Rhodesian government forces may haveresorted to biological warfare with anthrax againstcattle in order to impoverish the rural blackpopulation, as a desperate tactic in the finalmonths of the civil war, and that humans wereinfected secondary to contact with infected ani-mals or animal products. 112

EPIDEMIOLOGICAL ANALYSISDistinguishing natural disease outbreaks from

those produced deliberately requires careful in-vestigation and knowledge of local diseases andendemic infections. There is at present no gener-

ally accepted methodology for investigating thepossible use of BTW agents. But Dr. JackWoodall, an epidemiologist with the WorldHealth Organization in Geneva, has identified anumber of characteristics of a disease outbreak

would suggest it was not of natural origin:113

The appearance of an endemic disease faroutside its established range. A naturaldisease outbreak might be distinguishedfrom a BTW attack by determiningg whetherits source is an agent endemic to the region.Although jet travel has made it easier forinfectious agents to spread discontinuouslyfrom one continent to another, the progres-sion of an epidemic typically involves grad-ual spread to contiguous regions or alongtransportation routes.Appearance of a vector-borne disease in theabsence of natural vectors or reservoirhosts. Plague epidemics, for example, typi-cally begin in rats and are spread to man byinfected fleas. The initial form of the diseasein humans is the bubonic form affecting thelymph nodes, which later converts into themore lethal and contagious pneumonic form.Thus, the sudden appearance of pneumonicplague in humans (1) in the absence ofinfected rats and fleas, and (2) withoutprecursor cases of the bubonic form, wouldbe suggestive of a covert BW attack.Pulmonary disease in the absence of naturalmechanisms for producing high-concentra-tion biological aerosols. Since many infec-tious diseases do not naturally infect thelungs, the anomalous appearance of a respi-

110 q+leph~~e ~tmiew +~ Dy, sc~~ ~te~, ~soci~e Dir@or of tie H~th Scien@s Divisio~ Rockefeller Fourl&itiou New yOr~

NY, Aug. 6, 1992.

111 See J.C.A. Davies, “A Major Epidemic of Anthrax in Zimbabwe, Part l,” Central Afi”can Journal of Medicine, vol. 28, 1982, pp.291-298; and J.C.A. Davies, “A Major Epidemic of Anthrax in Zimbabwe, Part 2,” Central Afi”can Journal of Medicine, vol. 29, 1983, pp.8-12.

112 M~lN=, ~ c~~Epimoticin~~~e, 1978-1980: ~etoDeh~mte Spr~d?’ The PSR Q@er/y, vol. 2, No.4, December 1992,

pp. 198-209.113 Job p. w-, ~ ‘WO 1{~~ ~d Epidemic ~o~tion M a &MiS for v~~tion Activities Under the Biological W~IIS

Convention” S.J. Lundin, cd., Vievvs on Possible Verlj7cation Measures for the Biological Weapons Convention, SIPRI Chemical & BiologicalWarfare Studies No. 12 (Oxford, England: Oxford University Press, 1991), pp. 59-70.

Chapter 3-Technical Aspects of Biological Weapon Proliferation

ratory form of such a disease might beindicative of a deliberate aerosol attack.Other human activities than deliberate mili-tary attack may generate infectious aerosols,however. The outbreak of Legionnaires’ Dis-ease at a hotel in Philadelphia, for example,was traced to a natural microbial contaminationof the building’s air-conditioning system.Unusual epidemiological patterns that differfrom natural disease outbreaks. A deliberateBW attack by aerosol dissemination wouldinfect a large number of exposed individualssimultaneously, causing a majority of themto develop symptoms at approximately thesame time. Thus, instead of a gradual risefrom a smaller number of precursor cases,there would be an “explosive’ outbreak ofdisease in many thousands of people.114

While these characteristics are all plausible, therecent natural outbreak in New Mexico of ‘ ‘Na-vajo flu, ’ a virulent respiratory illness withgreater than 30 percent mortality, meets nearly allof the criteria Woodall proposes. Hanta virus,now known to be the cause of the illness, hadnever before been known to occur among humansin the Western Hemisphere, Whereas all previousreported cases around the world were hemor-rhagic fevers with shock and kidney failure, thisrecent outbreak took the form of a respiratoryillness. Finally, the epidemiology of the diseasewas extremely unusual and confused investiga-tors for months. This episode points out thedifficulty of distinguishing a highly anomalousdisease outbreak of natural origin from the

111

deliberate or accidental release of biological-warfare agents. 115

An important task of epidemiological analy-sis is to characterize the strains of disease-causing agents indigenous to the affected area,thereby making it possible to distinguish preex-isting “background” strains from BW agentsintroduced from the outside. Even if it could beascertained that a disease outbreak was of artifi-cial origin, however, it might still not be clearwho had initiated the attack. It might also bedifficult to collect the necessary data if investiga-tors were denied permission to visit the sites f

Y;* i t.the alleged attacks.

&

t

A current problem with depending on epideology to detect the use of BTW agents is that suchskills are unlikely to be available in those regionsof the world where biological warfare is mostlikely. In order to detect new infectious diseasessuch as AIDS before they reach epidemic propor-tions, the epidemiologist Donald A. Hendersonhas proposed the establishment of an interna-tional network of research centers to monitor theemergence and spread of new infectious diseases,linked to a global rapid-response system. ll6

Beyond its obvious public-health benefits, such aglobal surveillance system would make it easierto distinguish artificially induced epidemics asso-ciated with the covert use of BTW agents fromordinary background noise. ‘‘117 It would therebyhelp to deter biological warfare and also to identifyfalse claims of BW, an important objective.

Table 3-2 summarizes the various potentialsignatures associated with BTW development,

114 Nevefieless, tie anthrax epdtmic in the Soviet town of Sverdlovsk (now Yekaterinburg) in 1979-now recogniz.ed to have ~ntheredtof an accidental release of anthrax spores from a Soviet military biological facility-was associated with a gradual increase in the number ofcases over a period of several weeks, a pattern that appeared consistent with a natural epidemic. It is lmo~ however, that at low levels ofexposure, anthrax spores ge rminate at different rates in exposed primate hosts, resulting in highly variable incubation periods. MatthewMeselso~ Harvard University, personal communication 1993.

I Is m~coff, 0p. cit., footnote 29.

1ls Donald A. Henderson, “Surveillance Systems and Intergovernmental Cooperation” Stephen S. Morse, cd., Emerging Viruses (NewYork: Oxford University Press, 1992), pp. 283-289. See also Joshua Lederberg et al., Emerging Infections: Microbial Threats to Health in theUnited States (Washingto~ DC: National Academy Press, 1992), pp. 134-137.

1‘7 Mark L. Wheelis, “Strengthening Biological Weapons Control Through Global Epidemiologicat Surveillance, ” Politics and the LfeSciences, vol. 11, No. 2, August 1992, pp. 179-189.


Table 3-2—Biological Weapon Program Signatures and Concealment

Program stage Signature Detection methods (examples) Concealment methods, comment

Research &development

-. . . . . . .Scientific and technical publi-cations (presence or absence)

Literature survey and analysis 1. Manage publication activities

2. Use widely available technicalinformation rather than designnew agents or techniques

Nondeclaration of work withpotential BTW agents or withpathogen aerosols

Human intelligenoe (humint),on-site inspections

Conceal undeclared activities

Clandestineproduction plant

Security measures Overhead imaging or humint

Humint

Conceal measures, or place plantwithin other secure facilities

Large numbers of eggs orlaboratory animals for virusproduction

Storage depots for BTW-capable munitions

Use tissue culture rather thananimals for production of viruses

Overhead imaging or humint Conceal depots underground(although faciiity building wouldbe visible)

imports of dual-use equipment(fermenters, lyophilizers, mi-croencaipsulation systems)

Tracking of exports tosuspected proliferants

1. Obtain equipment from multiplesuppliers, or through interme-diaries

2. Divert equipment from legiti-mate civil activities

3. Make equipment indigenously

Converted ormultipurposepharmaceuticalplant

Security measures Overhead imaging or humint Conceal measures

Residues of virulent microbialstrains or genetically modifiedagents

Sampling of air, water, or soil inor near suspect plants; togetherwith various forms of biochemi-cal analysis (e.g., ELISA, bio-assay, DNA probes, PCR)

1. Decontaminate production linewlth bleach or superheated steamand autoclave cultures

2. Remove wastes for off-sitedisposal

3. Claim that BTW agents arebeing used for defensive acti-vities

Special safety and containmentmeasures

Onsite inspection of suspectplants

1. Sacrifice worker safety2. Modern biotech plants increas-

ingly have these features

Processes or capacity that can-not be justified on technical oreconomic grounds

Seed stocks, cell lines, andequipment (e.g., microencap-sulation) inappropriate fordeclared activities

Omission of costly measuresto ensure purity and sterilityof pharmaceuticals or toinactivate agent to makevaccine

Facilities for rapid, large-scaledecontamination

Onsite inspection of suspectplants

Such assessments are highly sub-jective

Onsite inspection and sampling Claim that material and equipmentis being used for legitimate medi-cal applications, although possibil-ities may be limited

Onsite inspection Employ measures to simulate phar-maceutical production (costly)

Onsite inspection Use legitimate vaccine productionactivities as a cover


Program stage Signature Detection methods (examples) Concealment methods, comment

Evidence of immunization toBTW agents in plant workersor evidence of infection in peo-ple or animals nearby

Blind tests Refuse permission to take bloodsampils

Weaponization andtesting

Uniquely configured arsenals(e.g. distribution of storagebunkers)

Overhead imaging Pattern facilities after conventionalarsenals

Cold storage of BTW agents Thermal infrared imageryExcess electrical capacit y

1. Produce large quantities of agentshortly before use to minimizeneed for storage

2. Mask thermal-infrared emissionsfrom refrigerators

Specialized equipment forfilling agents into munitions

Onsite inspection Conceal filling operation at someremote location

BTW testing facilities, suchas small aerosol chambers

Onsite inspection, samplingand analysis

Carry out tests inside dosedbuildings

Field testing of aerosolgenerators and delivery sys-tems

Overhead imaging, onsiteinspection, sampling andanalysis

1. Mask test grid2. Use legitimate activity such as

biopesticide dissemination as acover for illicit activities, althoughhigh security might be a give-away

Large animals for aerosol test-ing

Field training of troops

Uniquely configured test facili-ties

Overhead imaging 1. Make special features temporary2. Test on overcast days, at night,

or in absence of overhead imag-ing systems

Weapon use Anomalous characteristics of adisease outbreak (e.g., atypi-cal agent, explosive diseasespread, pulmonary disease inabsence of natural respiratory

1. Field epidemiology2. Genetic fingerprinting of

disease agent

Use a disease agent indigenous tothe area being attacked


production, weaponization, and use. Many ofthese indicators are nonspecific, since their pres-ence could be associated with other, legitimateactivities. Even so, a pattern of such signatureswould be suggestive of a clandestine BTW pro-gram that could then be confirmed by other means.

lar level. Genetic engineering involves identify-ing regions along the DNA molecules that encodedesirable genetic characteristics and cutting andsplicing these segments of DNA with enzymetools to create “recombinant” strains. Since allliving creatures contain DNA, it is also possibleto combine genes across species lines to give anorganism novel traits that do not occur in nature.MILITARY IMPLICATIONS OF GENETIC

ENGINEERINGThe past two decades have seen revolutionary

advances in the ability to manipulate the genetic| Novel Agents?

Techniques for the engineering of genes inbacteria and animal cells, and for the modificationcharacteristics of living organisms at the molecu-


of proteins, have become widely available. Al-though advanced genetic-engineering capabilitiesare still rare in the developing world, gene-splicing “kits” containing the necessary equip-ment and reagents (e.g., restriction enzymes) canbe easily obtained by mail order, and much of thenecessary know-how is openly published in thescientific literature. Some analysts have specu-lated that gene-splicing technologies could beused to develop ‘second-generation’ BW agentswith greater military utility by making the behav-ior of these agents in the environment morepredictable. 118 Toxin genes and Virulence factors

might also be transferred from one species ofmicroorganism to another. According to JohnBirkner, a foreign technology analyst for theDefense Intelligence Agency, “recombinant-DNA techniques could open up a large number ofpossibilities. Normally harmless, nondisease-producing organisms could be modified to be-come highly toxic and produce effects for whichan opponent has no known treatment. Otheragents, now considered too unstable for storage orbiological warfare applications, could be changedsufficiently to become effective. ’ ’119

Although it could theoretically add a toxingene to a harmless bacterium to render it virulent,recombinant-DNA technology is unlikely to pro-duce novel pathogens more devastating than thehighly infective and lethal agents that alreadyexist in nature. The reason is that any attempt tocombine genes from unrelated organisms is likelyto interfere with the highly developed and inte-grated pattern of genetic traits that give rise topathogenic behavior. Since a whole constellationof genes must work together for a microorganismto cause disease, altering a few genes withrecombinant-DNA techniques is unlikely to yielda novel pathogen significantly more deadly thannatural disease agents. 120 It is therefore doubtful

that genetic engineering could result in novelBW agents with greater potency than natu-rally occurring agents.

| Increased Controllability of MicrobialAgents

Nevertheless, the genetic modification of stand-ard BTW agents might, however, overcomespecific obstacles that currently limit their milit-ary utility. In particular, genetic engineering andmodern biotechnologies can facilitate microbialproduction, improve storage and delivery, createantibiotic resistance, and enhance the controlla-bility of existing pathogens. It is not clear, how-ever, that these modifications would significantlyalter the military utility of BW agents comparedwith the numerous already known agents.

SHORTER INCUBATION TIMEModifying BW agents to act more rapidly

would increase their tactical utility on the battle-field, although this is unlikely to be accomplishedanytime soon.

ENVIRONMENTAL STABILITYGenetic engineering might be able to increase

the ability of microorganisms and toxins towithstand some of the stresses associated withstorage and dissemination, for example, by insert-ing complexes of genes for resistance to inactiva-tion by temperature, ultraviolet radiation, drying,and the shear forces associated with aerosolformation. Most of these traits are geneticallycomplex, however, and are not well understood.

INCREASED VIRULENCEDevelopment of a system for the super-

expression of toxin genes has made it possible todevelop recombinant bacterial strains that pro-

118 fi~d Geisslm, “~plica~o~ of @netic I@ina@ for Chemical and Biological Wti~,” W~r/dAr~~nt.S and Disannanent:

SZPRI Yearbook 1984 (lmdon: lhylor & Francis, 1984), pp. 421451.

119 Job BhkXICX, cit~ in R. J&ffey %n.i~ “The Dark Side of Biotechnology,” Science, vol. 224, June 15, 1984, p. 1215.In Jom~ B. ~cker, ‘‘Gme: WarS, ” Foreign Policy, No. 57, winter 1984-85, p. 62.

— . . -.

duce 10 tostrains.

ANTIBIOTICInserting

Chapter 3–Technical Aspects of Biological Weapon Proliferation I 115

100 times more toxin than natural

RESISTANCEantibiotic-resistance genes into natu-

rally infectious agents can make them resistant toone or more prophylactic or therapeutic drugs,rendering such defenses useless. At the sametime, an attacker could immunize his troopsagainst the modified agent, protecting themwithout the need for antibiotics. Reportedly, theSoviet Union launched a secret program in 1984to develop a genetically engineered form ofplague that was resistant to antibiotics.121

VACCINE PRODUCTIONRecombinant-DNA techniques make it easier

and safer to produce specific vaccines to matchnovel agents, thus enabling the attacker to protecthis own forces while denying a vaccine to thedefender. In the past, the difficulty of producingeffective protective vaccines was a major obstacleto acquiring an offensive BTW capability. Never-theless, recombinant vaccines are not alwayseffective because they represent one or a fewantigens rather than the full set of antigens presentin the actual pathogen.

CONTROLLED PERSISTENCEGenetic engineering might result in more

controllable BW agents through the manipulationof genes to program the survival of a bacterialpopulation released into the environment. Forexample, it might be possible to program micro-organisms genetically to survive only under anarrow set of environmental conditions. Alterna-tively, one might design regulatory sequencesknown as ‘‘conditional suicide genes, ’ whichcause a microorganism to die off after a specified

number of cell divisions. 122 By inserting such genes

into pathogens, it might be possible to create aBW agent that would cause disease for a limitedperiod of time and then spontaneously die off.

IMMUNOLOGICAL MODIFICATIONBy means of gene transfers, it would be

possible to modify the antigenic (antibody-inducing) proteins on the outer surface of a pathogenic virus or toxin, thereby rendering themodified agents insensitive to a preexisting hostimmunity or to standard vaccines and antitoxins.(Since most toxin antigens are located on thescaffolding of the molecule rather than near thesite responsible for its toxic effects, it would bepossible to alter the immunological characteris-tics of a toxin without changing its biologicalactivity.) Further, since most diagnostic proce-dures for BTW agents rely on the detection ofcertain surface antigens with antibodies, modifl-

,%”cation of the antigens would make it harder todetect, identify, and counter the modified agents.

HOST SPECIFICITYSome analysts have raised the grotesque possi-

bility of making microbial pathogens more dis-criminate by designing ‘‘ethnic weapons’ thatexploit differences in gene frequency betweenpopulations to selectively incapacitate or kill aselected ‘‘enemy’ population to a greater extentthan a‘‘&iendly’ population. Yet human popula-tions are not uniform enough to be uniquelytargeted by a given pathogen.123

* * *

These possibilities notwithstanding, the practi-cal obstacles to developing more controllable BWagents remain enormous. Even if genetic engi-neering could produce recombinant pathogensthat survived for a predetermined length of

121 Bw, op. cit., foo~ote 97! p- 41”

122 ~qmatow Cohttee for tie ~rd Review Conference of the Parties to the BWC, Background Document on NW Scientific and

Technological Developments Relevant to the Convenh”on on the Prohibition of the Development, Production andstockpiling ofBacteriological(Biological) and Toxin Weapons and on Their Destruction, Document No. BWC/CONF.HI/4, Aug. 26, 1S91, p. 11.

In Novick and Shul~ op. cit., footnote 47, p. 114.


time in the environment, they would remainincalculable in their effects, since their dissem-ination would still rely on wind and weather,and mutations might change the behavior of agenetically modified agent after it had beenreleased. Once released,, living pathogens mightpropagate, evolve, and develop ecological rela-tionships with other living things in ways thatcannot be entirely foreseen. Furthermore, geneti-cally engineered pathogens would require exten-sive trials to verify that they would survive longenough to infect target personnel after beingdisseminated by a weapon system and exposed tothe natural environment, in which most microor-ganisms are extremely fragile. Thus, testing inhuman subjects might be required to give amilitary planner confidence in genetically engi-neered biological agents.

| Modified Toxins and BioregulatorsAnother potential threat from the biotechnol-

ogical revolution is the development of newtoxin-warfare agents. Known protein toxins, suchas botulina1 and ricin, deteriorate in response toenvironmental factors such as temperature andultraviolet radiation, and thus rapidly lose toxic-ity after dissemination. Although genetic engi-neering is unlikely to increase the potency ofnaturally available toxins, it might conceivablybe applied to modify the chemical structure oftoxins to:

● increase the stability of toxins so that theycan better be disseminated as an aerosol;

■ alter the antigenic structure of toxin mol-ecules, rendering them insusceptible to exist-ing antitoxins or antibody-based diagnostictechniques;

develop “chimaeric” toxins (combinationsof two different toxin molecules, such asricin and diphtheria toxin) that are morecapable of penetrating and killing targetcells; anddesign novel peptide toxins (possibly con-—sisting only of the biologically active regionof a protein toxin) that are as poisonous asnerve agents but are small enough to pene-trate the filters currently used in masks andprotective garments.l24

BIOREGULATORSRecombinant-DNA research may also lead to

the development of more effective incapacitants.With genetic engineering, even the body’s ownnatural substances might be utilized as warfareagents. “Bioregulators’ are small, physiologi-cally active peptides (chains of amino acidssmaller than proteins) that are normally present inthe body in minute quantities and that orchestratekey physiological and psychological processes.They are active at very low concentrations andinfluence the full spectrum of life processes, bothphysiological and mental. Bioregulators govern,for example, hormone release, control of bodytemperature, sleep, mood, consciousness, andemotions. An important subgroup of the bioregu-lators are the opioid peptides, which can induceanalgesia and euphoria. Since these naturallyoccurring peptides are active in the body in traceamounts, the application of larger quantitiesmight induce euphoria, fear, fatigue, paralysis,hallucinations, or depression, giving them somepotential as nonlethal incapacitating weapons.l25

Bioregulators might be modified chemically toenhance their physiological activity, stability, orspecificity. For example, the modification of thepeptide hormone LHRH (leutenizing hormone

124 fite~ ~fis ~d ~tematio~ T~de CaMda, VtiWtionReWh Unit, Novel Toxins and Bioregulators: The Emerging Scientific

and Technological Issues Relating to Verification of the Biological and Tom-n Weapons Convention (Ottawa: External Affairs, September1991), p. 47.

12S Sw~shNatiO~ ~eme ;Resemch~ti~te, &nefiCEnginee~”ng u~DiOJOgiCuz weupo~, @)ortNo. PB88-210869 ~m~, Swdem

National Defense Research Institute, November 1987; translated for the Office of International Affairs, National THmieal Information Semice,h’ftIy 1988), p. 58.


Table 3-3—implications of Genetic Engineering for Biological and Toxin Warfare

Capability Possible now May be possible May be possibleIn 5 years In 10 years, If ever

Shorter incubation time xTemperature stability Xa

UV stability Xb

Drying/aeosol stability xAntibiotic resistance X c

Controlled persistence xImmunological modification x c

Host specificity xCloning of toxin genes xMore stable toxins xNovel toxins xa For ~rtain Protein toxinsb For bacteriaC In some cases, not all


releasing hormone) by substituting a single aminoacid yielded a product 50 times more potent.126

Even so, it would be difficult to disseminatepeptides through the air in a militarily effectiveway. The ability of a peptide to diffuse across themucosa1 membranes of the respiratory tract de-pends on its molecular size. Although attempts todeliver the small peptide hormone ADH (antidiu-retic hormone) with a nasal aerosol have beensuccessful, similar efforts with insulin have failedbecause of the molecule’s relatively large size.127

The possible implications of genetic engineer-ing for biological and toxin warfare are summa-rized in table 3-3. Although the potential for themisuse of genetic engineering to develop new andmilitarily more effective BTW agents currentlyappears limited, this emerging threat clearlydeserves to be monitored carefully. Advanced

genetic-engineering capabilities are still rare inthe developing world, but most of the largercountries in the Middle East already have thetechnical capability to selectively breed microbialstrains with enhanced virulence, survivability,and antibiotic resistance. In According to one

analyst, “If you can identify the gene you want tomove, it is possible to do so. ’ 129

For at least the medium-term, BTW prolif-erants are likely to produce proven agents suchas anthrax and botulin toxin, rather thaninvest large amounts of time and money onexperimentation with genetically engineeredmicroorganisms. Eventually, however, techno-logically sophisticated proliferants might tryto modify standard agents to make them morestable during dissemination or more difficultto detect or to defend against.

lx Gove~ent of tie United States, op. cit., footnote 44, p. 29.

127 Zficoff, Op. cit., footnote 29.12E ~~ony H. Cordesu weapons of ~a~~ De~~ctiOn in r~e ~~/e East @ndOn: BrzI.ssey’s ~), 1991), p. 77.In ~~cot=f, Op. cit., foomote 29.

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