Introduction
History of Risk Assessment and Regulation of GEO's
Biotechnology Risk Assessment Background
Biotechnology Regulatory Background
U.S.
U.K.
EC
Biotechnology Risk Assessment
Information Requirements For Risk Assessment
Summary
References Available Electronically
Table 1:	Summary of Information Needed to Estimate Survival and Persistence of Released Microorganisms
Table 2:	Scientific Considerations For "Points to Consider For Environmental Testing of Microorganisms"
Table 3:	Data Points For Hazard Assessment
Table 4:	Data Points For Exposure Assessment
Table 5:	Genetic Characteristics of Organism (Parent and Recipient)
Table 6:	Introduction of Genetic Material
Table 7:	Environmental Considerations (Parent and Recipient)

Introduction

This chapter discusses the development of risk assessment in the field of biotechnology and examines risk assessment protocols accepted for use. As emphasized in the introduction, the primary focus of this guide is on the ecological effects and their implications for regulatory risk assessment. This chapter, therefore, gives primary consideration to risk assessment procedures as they are used to evaluate potential adverse ecological effects, and does not attempt a full discussion of human health issues. Human health effects are considered in depth when resulting from or related to environmental impacts.

Risk Assessment cannot be considered in the abstract. The activities of the risk assessor and manager are strongly affected by the relevant legislation and guidelines which have established as a result of the legislation. One can compare the risk assessment process to a filter system and risk management to devices to control flow rate. Each layer of filtration material increases product purity but reduces output. Too few layers allow unwanted contaminants to pass, resulting in adverse effect on the product. Too many layers result in lowering the rate of flow, perhaps to the point where the process is not economically feasible. A balance must be struck between purity and supply. The role of risk assessment is to determine the balance point. Risk management attempts to insure safety by use of procedural stratagems without preventing the development of useful products.

Risk assessment and management also must factor in the type of risk and an individual=s acceptance of the particular risk. Thus, one will accept a higher degree of risk to accomplish an important (to oneself) function. It is acceptable to drive at high speed even though it increases the risk of accident and greatly increases the risk of serious injury while it may not be acceptable to inhale second hand smoke even though the inhalation poses a much lower risk of adverse effect (perhaps one in 10,000 vs one in 1 million). Risk assessors (or statisticians) have calculated the probability of adverse effect on humans resulting from a diverse list of activities. Thus, it has been calculated that the risk of harm (bodily injury or disease) resulting from, for example, drinking a glass of wine a day, spending six minutes in a canoe, riding ten miles on a bicycle or flying 1000 miles is approximately equal. It is an individual judgement (i.e. risk acceptability) as to whether to accept exposure to these hazards.

The fundamental data required to evaluate ecological risk are fundamentally the same for any introduction of nonindigenous organisms into a new environment. This includes all micro- and macro organisms, whether engineered (GEOs) or natural. The risk assessor must select for emphasis data which are most pertinent to the particular environment or situation. The challenge for assessors is to obtain and interpret enough data to produce a credible assessment. However, since the assessor is strongly affected by legislation, we will first look at the historical basis for risk assessment and then consider how risk assessments are conducted.

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History of Risk Assessment and Regulation of GEOS

Biotechnology Risk Assessment Background

Risk Assessment did not begin with GEOs or biotechnology. Governments all over the world have historically assumed the responsibility of assuring various aspects of safety for their citizenry. Examples include drinking water, food, and recreation (eg swimming). This responsibility carried with it the requirement for determining the risk associated with specific activities or products, such as driving speed or newly invented chemicals. The first major application of risk assessment to environmental and public health concerns involved assessing the effects of hazardous chemicals. Over many decades, specialized procedures were developed to determine and manage chemical risks.

In 1973 a major scientific event, the expression of genetic material transferred between two living organisms in the host organism gave birth to what is now known as the Biotechnology Industry. Scientists involved in the discovery, and biologists in general, were immediately concerned that if extended, this experimental result could result in adverse effects on humans and the environment. This feeling was so strong that a major international scientific conference to discuss the ramifications of gene exchange experiments was convened in 1976 in Asilomar, California, USA. The majority of attendees were laboratory or human health oriented. Ecologists and general biologists were a small minority. The conference produced a call for voluntary guidelines for scientists working in the new area of gene exchange. The guidelines were to assure the safety of the laboratory workers, the public and the environment.

The result was a set of guidelines produced by a new committee, the Recombinant Advisory Committee, (RAC) formed at the US National Institutes of Health. The guidelines were based on containment. All experiments were to be conducted in a manner designed to prevent exposure of workers and the public to the microbes being used. Release to the environment was not acceptable. The principle of maintaining barriers around the experiment to prevent exposure was the core of the guidelines. Because the filed was new, it was made clear that the guidelines would have to be easily modified as experience was gained. The guidelines were voluntary: they had no legal standing. The guidelines have been successful: there have been no adverse effects to workers or the public when the guidelines were followed. Over the years many government agencies and countries have adopted them, or a derivation of the guidelines, as legal requirements.

Ultimately, there was concern that those with responsibility for modification of the guidelines -- the RAC -- would be knowledgeable in the field of molecular biology but would not be as experienced in biology, microbiology, ecology or other life sciences related to uncontained environmental trials. In response to this concern, RAC agreed to bring in representatives from relevant fields to broaden the risk assessment expertise.

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Biotechnology Regulatory Background

The US, UK, and EC approaches to regulation of biotechnology products are examples of three different processes to achieve similar effects. All are concerned with safety. While the EC. approach stresses harmonization of methodology to insure regional safety and exchange within the EC, the US and UK also seek to assure regional safety and encourage international trade.

EC procedures seek to harmonize those already in place in the member countries, UK and US procedures do not have this problem.

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US

In 1986, the US Office of Science and Technology Policy (OSTP) published its Coordinated Framework for the Regulation of Biotechnology (OSTP 1985), which stated that existing statutes were sufficiently broad to regulate biotechnology products. 107 existing laws, regulations and guidelines and determined which acts and agencies would have jurisdiction over genetically engineered products were reviewed and specific responsibilities identified.

Though initially controversial, the finding that existing laws were adequate to provide oversight of recombinant DNA (rDNA) technology became the basis of regulatory policy in this area in the US. Currently, most countries regulate biotechnology under existing statutes, assigning departments (or ministries or agencies) to the specific tasks based on existing statutory authority. These findings placed oversight responsibility in four US agencies: the Environmental Protection Agency (USEPA), the Department of Agriculture (USDA), the National Institutes of Health (NIH), and the Food and Drug Administration (FDA). In the US, the determination that there was no need to seek new legislation was met with skepticism by parts of the scientific community, various public interest groups, and some members of Congress. A decade of experience of field applications without apparent adverse effects gives considerable confidence in the processes in place. Although there are still those who advocate omnibus legislation regulating GEOs, the US Congress has thus far declined to take such action.

Those federal government agencies with statutory authority for regulating field trials and biotech products were placed in a position to regulate biotechnology products. Thus, the USDA and EPA which have authority over agricultural biotechnology activities were told to share responsibility for agricultural biotechnology products. The USDA's regulatory powers are exercised by the Animal and Plant Health Inspection Service (APHIS) (Plant Pest Act 7 USC 150) USDA published a guide to applicants (USDA 1990). Under this authority, USEPA has reviewed small-scale field test proposals (USDA 1987, 1993). USEPA regulatory authority derives from two specific laws: the Federal Insecticide, Fungicide and Rodenticide Act (FIFRA; 7 U.S.C.), and the Toxic Substances Control Act (TSCA; 5 U.S.C.). FIFRA regulates all pesticides, and the agency has used this law to register a number of naturally occurring microbes and viruses. The agency instituted a policy (USEPA 1987, 1988, 1989) under FIFRA (49CFR 50886) to formally extend its authority to certain small-scale field tests of genetically engineered bio-pesticides. The procedures for toxic substances were finalized in 1997 (USEPA 1977).

FDA reviewed its authority under the Food, Drug and Cosmetic Act and found that it could assume its component of the responsibility for assessing the effects of biotechnology products without having to change its regulations. USDA and EPA, on the other hand, elected to develop new guidelines for uncontained applications of GEOs. Final policy statements for these agencies have been published. All three agencies focused primarily on terrestrial products. No oversight policies were developed specifically for aquatic engineered organisms. <

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UK

The UK initially opted to establish health and safety standards in biotechnology under its Health and Safety at Work act by forming an advisory committee. The committee is now known as the Advisory Committee on Genetic Modification (ACGM) and has produced regulations for activities with GMOs (covering both for laboratory and field work), although a separate subcommittee (ACRE: Advisory Committee on Release to the Environment, UK1993) has been formed to focus on environmental issues. The UK did enact an Environmental Protection Act (1990) aimed at minimizing environmental damage which might result from GMOs. Thus, the UK is operating under existing laws and has enacted new statutes affecting biotechnology (UkDOE/Acre 1993 and UKHSE 1996). Contained uses are covered under the Contained Use Act (UK1996)

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EC

Operating to produce directives which affect all member nations, the EC attempts to integrate the health and safety concerns of all members as expressed in their respective statues and guidelines. Thus the EC, much as any country, concerns itself with the protection of health and safety. However, the EC must also reflect the international trade requirements of its members and therefore must also seek to harmonize procedures and for export and import of products. Harmonization (i.e. the process of developing national regulations which are serviceable between countries and facilitate rather than impede international trade) is one of the most important issues affecting international trade. The need for harmonization is clear: facilitation of import and export of products. But harmonization goes far beyond simply doing away with conflicting requirements. It involves assurances that a product is sufficiently well characterized so that the importing country can feel assured that use of the product creates only a minimal possibility of an adverse effect. The EC has promulgated two directives which seek to achieve this objective. 90/219 deals with contained use and 90/220 deals with environmental release. In keeping with EC policy, the two directives require risk assessment of products in accordance with the guidelines promulgated by the member nations.

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Biotechnology Risk Assessment

The biotechnology risk assessment and risk management process, as it has evolved over the past decade to regulate GEOs, is complex. That is, procedures vary on a case-by-case basis depending on the country, the agency charged with oversight, the mandates of the relevant statute, and the specifics of the proposed trial. An understanding of the basic elements of risk assessment science is necessary to understand its relation to the development of regulations for biotechnology products.

The US National Academy of Science (NAS) has published two documents dealing with risk assessment of biotechnology products (NAS 1987, 1989). preceded by a well received volume on risk assessment (NAS 1983). The study was oriented toward human health. Figure 1, taken from the study, has been slightly modified to emphasize the environmental component.

On a conceptual level risk assessment is very simple, consisting of the bringing together of two components, hazard and exposure. Each activity or product has a certain degree of hazard associated with it. This is the first element of risk assessment. Activities such as swimming, driving a car, or plowing a field, may lead to harm to those involved; similarly, the use of any product, from commercial foodstuff to a computer, has a probability of adverse effect on humans or the environment. Thus, step one is identification of the specific adverse effect(s) associated with a product or activity - the hazard.

The hazard component of risk assessment is then related to the level of exposure. Some activities - such as crossing a street- are more common, individualized, and short lived than others. In some cases the product is highly labile and will not persist in the environment; other products may have a longer lifespan, resulting in greater probability of larger populations coming in contact with the product. These parameters define the exposure factor. Exposure factors are based on how often one engages in the particular activity or comes in contact with a particular product. Central to exposure is identification of the exposed population.

Risk assessment is the bringing together the probabilities of hazard and exposure to produce an understanding of the likelihood of some type of adverse outcome. The process involves three components: Research (data gathering to estimate the hazard and exposure), assessment and characterization (to combine the factors) and, finally, decision making (based on the scientific and legal options available).

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Information Requirements For Risk Assessment

How can one approach assessing risk from a science perspective? One can compare the risk posed by the introduction of transgenic organisms to previous introductions of similar organisms in similar target/test environments. In this way if an introduction can be shown to be similar to a previous introduction, one which was known to present little or no risk, then the level of risk would be acceptable. For certain introductions there may be insufficient knowledge that existing practices or regulations adequately address the possible risk posed by the introduction.

However, the range of information potentially required for a successful risk assessment (simply to determine the degree of similarity to other releases) is large, and expands as the difference(s) between the Anew@ organism and the comparison organism expand. One must be able to identify and quantitate the hazard, the population at risk, the exposure dose and have the ability to measure the effect in the field. The reliability and credibility of the assessment and the efficacy of the management technique depend on the accuracy and completeness of the assessment information. Numerous groups and agencies have attempted to identify the basic requirements for individual purposes -IE Health effects (NAS, 1987,89) and ecological effects (table 1). A consensus as to what parameters are essential has emerged (Table 2).

The unifying factor for risk assessors in the US is to ensure consideration of the relevant items in "Points to Consider," a document developed by an interagency committee charged with identifying requirements for risk assessment of biotechnology products. EPA published its version of Points to Consider in 1990 (USEPA 1990) and provided an updated version (USEPA June, 1997) to meet requirements in the recent Final Rule for Microbial Biotechnology Products (USEPA 1997). The requirements identified in table 2 are universal. All risk assessments require some or all of the Points to Consider data points for producing a credible risk assessment. They are not intended to be all inclusive- i.e. not all points will be required in each assessment. Thus, the possible effect in aquatic environments would not be considered if the product is intended for use in mountainous crop areas (e.g. coffee plantations) and is biodegradable. Many of the points to consider are supported by an extensive and detailed documentation of specific assay procedures to develop data for the risk assessor. (Tables 3-7 ) (Source: Subdivision M, USEPA 199X). These tables indicate the detail required by the US EPA for developing risk assessment. Not all countries or all agencies within a country (i.e. EPA TSCA) provide detailed description of test methods.

The Points to Consider document, although not officially used by all agencies in the US or by other countries, is a complete list of information requirements for any possible risk assessment. A survey of requirements listed as possibly required by countries in and out of the EC indicates that the list is universally applicable, i.e. all countries list the same information requirements and recognize that not all assessments require information about all of the parameters identified on the list. The US has the most well developed set of specific protocols for data development.

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Summary

The objective of risk assessment is to produce a description of the magnitude and likelihood of an adverse effect related to a particular activity. The credibility and value of the assessment is directly related to the quality and quantity of data available about the product, the environment in which it will be used and the population involved. Most nations have regulations aimed at protecting citizens from adverse effects of commercial products. In most countries, these have been judged to be sufficient for providing protection vis a vie biotechnology products. In most cases, new guidelines or specific provisions to existing have been added to existing guidelines to assure safety.

Safe use of biotechnology products can be assured by adherence to risk assessment principles and development of required data using sound, science based protocols and measurement techniques. Figure Two summarizes the assessment problems and information needs in basic terms. As can be seen, the assessor requires identification of the problem: hazard and exposure, in risk assessment terminology. Hazard is identified as effect in scientific terminology: exposure is seen as initial establishment and possible transport to new locations for secondary establishment. The needs of the risk assessor are clear: Ability to enumerate the test organism and to acquire information about stability and effect of the environment on the organism. Coupled with this need is the requirement for ability to extrapolate to other environmental situations and to other exposed populations. These needs generate the requirement for specific methods, some specified by the reviewing authorities, some available in open literature and some which still require development.

Especially lacking are models which provide the ability to generalize. It is this which limits risk assessment to a case-by-case level. Only in a few situations (i.e. Bacillus thuringiensis) is sufficient background information available to permit generalization and hence relaxation of information and notification requirements for field testing engineered organisms. However, the successful conduct of over 2000 field trials over the past decade demonstrates that risk assessment is possible and has been successful, although some may argue that the procedures are overly conservative.

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References Available Electronically

EC Directive 90/219

EC Directive 90/220

UKEPA 1990. Environmental Protection Act 1990

UKDOE/ACRE. 1993. Guidance Note 1. London, UK: Department of the Environment/Advisory Committee on Release to the Environment.

UK,HSE. 1996. Guide to Genetically Modified Organisms, Contained the (as amended 1996). London, UK: Health and Safety Executive.

USDA. 1990. A Users Guide: Biotechnology Permits USDA/APHIS/BBED

Hyattsville, MD: USDA.

USDA. 1993a. Genetically Engineered Organisms and Products; Final Rule. Federal Register Mar. 31 1993 17044-17059.

USEPA. 1987. Disclosure of Reviews of Pesticide Test Data. USEPA/OPTS. Federal Register 50 #229 18833-18835.

USEPA. 1988. Pesticide Registration Procedures; Pesticide Data Requirements; Final Rule. 40 CFR parts 153, 156, 158, 162, and 163. FR 53 #88. 15952- 15999.

USEPA. 1989. General Information on Applying for Registration of Pesticides in the United States. Washington, DC: USEPA/OPP.

USEPA. 1990. Points To Consider in the Preparation and Submission of TSCA Premanufacture Notices for Microorganisms. Program Development Branch.Washington, DC: USEPA.

USEPA. 1992. Pesticide Assessment Guidelines Subdivision M. USEPA/OPPTS EPA 540/9-82028. Washington, DC: USEPA.

USEPA. 1997. Microbial Products of Biotechnology; Final Rule. Federal Register 62:2-45.

USEPA, 1997. Points to Consider in the Preparation of TSCA Biotechnology Submissions for Microorganisms. USEPA/OPPTS, Washington DC.

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Table 1:	Summary of Information Needed to Estimate Survival and Persistence of Released Microorganisms
CONTAINMENT/MITIGATION FACTORS
MITIGATING	ENHANCING
Adverse Conditions	Acceptable Conditions
Starvation	Adaptation
Competition	Competitiveness
Predation	Avoidance
Parasitism	Resistance
OUTCOME
Death	Revitalization
Debilitation	Establishment Dispersal

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Table 2:

Scientific Considerations For "Points to Consider For Environmental Testing of Microorganisms" (Adapted From USEPA 1990 and 1997)

Summary Summary of the proposed trial - objectives and significance of the proposed trial. Genetic Considerations of Modified Organisms to be Tested Characteristics of the nonmodified Parental Organism Information on identification, taxonomy, source and strain Information on organisms reproductive cycle and capacity for genetic transfer Molecular Biology of the Transgenic Organism Introduced Genes Source and function of the DNA sequence used to modify the organism to be tested in the environment. Identification, taxonomy, source and strain of organism donating the DNA. Construction of the Modified Organism Describe the method(s) by which the vector with insert(s) has been constructed include diagrams as appropriate) Describe the method of introduction of the vector used and the procedure for selection of the modified organism Specify the amount and nature of any vector and/or donor DNA remaining in the modified organism. Give the laboratory containment conditions specified by the NIH guidelines for the modified organism. Genetic Stability and Expression Present results and interpretation of preliminary tests designed to measure genetic stability and expression of the introduced DNA. Environmental Considerations The intent of gathering ecological information is to assess the effects on survival reproduction and/or dispersal of the modified organism. For this purpose, information should be provided where possible and appropriate on: relevant ecological characteristics of the nonmodified organism the corresponding characteristics of the modified organism the physiological and ecological role of donated genetic sequences in the donor and in the modified organism(s). For the following points, provide information where possible and appropriate on the nonmodified organism and a prediction of any change that may be elicited by the modification: Habitat and Geographical Distribution Physical and Chemical Factors that can affect survival, reproduction and dispersal. Biological Interactions Host range Interactions with effects on other organisms in the environment including competitors, prey, hosts, symbionts, predators, parasites and pathogens. Pathogenicity, infectivity, toxicity, virulence or as a carrier (vector) of pathogens. Involvement in biogeochemical or in biological cycling processes (e.g. mineral cycling, cellulose, lignin degradation, nitrogen fixation, pesticide degradation). Frequency with which populations undergo shifts in important ecological characteristics such as those listed in 3C, I through 4. Likelihood of exchange of genetic information between the modified organism and other organisms in nature. Proposed Field Trials Pre-Trial Considerations: Provide data related to any anticipated effects of the modified microorganism on target and nontarget organisms from microcosm, greenhouse and/or growth chamber experiments that simulate; trial conditions. The methods of detection and sensitivity of sampling techniques and periodicity of sampling should be indicated. These studies should include survival of the modified organism, replication of the modified organism dissemination of the modified organism by wind, water, soil mobile organisms and other means. Trial Conditions: Describe the trial involving release of the modified organism. Specify number(s) of organisms and methods of application Specify number(s) of organisms and methods of application Describe characteristics of the site that would influence containment or dispersal. If the modified organism has a target organism, provide the following Identification and taxonomy The anticipated mechanism and result of the interaction between the released microorganism and the target organism.

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Table 3:	Data Points For Hazard Assessment
TEST PRINCIPLES
Test at multiple of anticipated dose Maximum hazard dose used if positive results obtained
SINGLE SPECIES TESTS
Avian single dose oral toxicity Avian dietary toxicity Freshwater fish acute bioassay Freshwater invertebrate test Nontarget plant Nontarget insect Honey bee Estuarine nontarget species Honey bee
FIELD SURVEY
Identify population(s) at risk

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Table 4:	Data Points For Exposure Assessment
1. SCALE OF TEST
2. CONTAINMENT PROCEDURES EMPLOYED
3. SPECIFIC DATA REQUIRED:
Biological Fate of Gene Habitat of MPCA Survival and Replication Factors Gene Flow Gene Construct, Probability of Transfer and Expression Expression Level Chemical Fate of Gene Fate of Gene/Gene Product in Soil Fate of Gene/Gene Product in Water

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Table 5:	Genetic Characteristics of Organism (Parent and Recipient)
IDENTIFICATION
Taxonomic description
GENOTYPE
Characterization of genetic material
POTENTIAL FOR GENE TRANSFER
Capability and mechanism Evidence for exchange in nature
PHENOTYPE
Rationale for selection Anticipated changes in host Culture requirements, life cycle, habitat Pathogenicity data (type, virulence) Antibiotic resistance and production Survival and persistence data Control mechanisms Natural agents, effective disinfectants

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Table 6:	Introduction of Genetic Material
HOW MODIFIED
Source and function of inserted DNA Methods used to identify, isolate and Insert DNA
VECTOR
Identification Site of gene insertion Method of introduction to host Characterization of inserted genes Location, amount, stability of vector DNA
COMPARISON OF MPCA TO PARENT
Laboratory data describing relative survival, persistence, multiplication and dissemination

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Table 7:	Environmental Considerations (Parent and Recipient)
ORGANISM
HABITAT
SURVIVAL FACTORS
Microcosm data (environmental conditions favoring or adversely affecting survival and growth)
DATA DESCRIBING SURVIVAL, REPLICATION, DISSEMINATION
POTENTIAL FOR BIOLOGICAL INTERACTION
IDENTIFICATION OF SPECIFIC POTENTIAL ADVERSE EFFECTS
TRIAL
Conditions of Trial Location Site Characteristics Probability of dissemination, description of target and nontarget population present Containment Procedures to be Employed Onsite containment procedures (physical, biological), transportation procedures, employee training, security procedures.
MONITORING
Procedures to be Employed Description of techniques (discuss sensitivity, and reliability). Discuss available data on recovery, sensitivity and reliability of techniques with MPCA and parent. Mitigation Procedures Termination procedures Disposal procedures Disinfection procedures

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