Guidelines for Research Involving Recombinant DNA Molecules (NIH Guidelines); Notice DEPARTMENT OF HEALTH AND HUMAN SERVICES

Federal RegisterJul 5, 1994

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Part IV

Department of Health and Human Services

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National Institutes of Health

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Guidelines for Research Involving Recombinant DNA Molecules (NIH

Guidelines); Notice

DEPARTMENT OF HEALTH AND HUMAN SERVICES

National Institutes of Health

Guidelines for Research Involving Recombinant DNA Molecules (NIH

Guidelines)

June 1994.

These NIH Guidelines supersede all earlier versions and shall be in

effect until further notice.

Table of Contents

Section I. Scope of the NIH Guidelines

Section I-A. Purpose

Section I-B. Definition of Recombinant DNA Molecules

Section I-C. General Applicability

Section I-D. General Definitions

Section II. Containment

Section III. Experiments Covered by the NIH Guidelines

Section III-A. Experiments that Require Institutional Biosafety

Committee Approval, RAC Review, and NIH Approval Before Initiation

Section III-B. Experiments that Require NIH/ORDA and Institutional

Biosafety Committee Approval Before Initiation

Section III-B-1. Experiments Involving the Cloning of Toxin

Molecules with LD50 of Less than 100 Nanograms Per Kilogram

Body Weight

Section III-B-2. Accelerated Review of Human Gene Transfer

Experiments

Section III-B-3. Minor Modifications to Human Gene Transfer

Experiments

Section III-C. Experiments that Require Institutional Biosafety

Committee Approval Before Initiation

Section III-C-1. Experiments Using Human or Animal Pathogens (Class

2, Class 3, Class 4, or Class 5) Agents as Host-Vector Systems

Section III-C-2. Experiments in which DNA from Human or Animal

Pathogens (Class 2, Class 3, Class 4, or Class 5) Agents is Cloned

into Nonpathogenic Prokaryotic or Lower Eukaryotic Host-Vector

Systems

Section III-C-3. Experiments Involving the Use of Infectious Animal

or Plant DNA or RNA Viruses or Defective Animal or Plant DNA or RNA

Viruses in the Presence of Helper Virus in Tissue Culture Systems

Section III-C-4. Experiments Involving Whole Animals

Section III-C-5. Experiments Involving Whole Plants

Section III-C-6. Experiments Involving More than 10 Liters of

Culture

Section III-C-7. Human Gene Transfer Experiments Not Covered by

Section III-A-2, III-B-2, III-B-3, and Not Considered Exempt under

Section V-U

Section III-D. Experiments that Require Institutional Biosafety

Committee Notice Simultaneous with Initiation

Section III-D-1. Experiments Involving the Formation of Recombinant

DNA Molecules Containing No More than Two-Thirds of the Genome of

any Eukaryotic Virus

Section III-D-2. Experiments Involving Whole Plants

Section III-E. Exempt Experiments

Section IV. Roles and Responsibilities

Section IV-A. Policy

Section IV-B. Responsibilities of the Institution

Section IV-B-1. General Information

Section IV-B-2. Institutional Biosafety Committee (IBC)

Section IV-B-3. Biological Safety Officer (BSO)

Section IV-B-4. Principal Investigator (PI)

Section IV-C. Responsibilities of the National Institutes of Health

(NIH)

Section IV-C-1. NIH Director

Section IV-C-1-a. General Responsibilities

Section IV-C-1-b. Specific Responsibilities

Section IV-C-1-b-(1). Major Actions

Section IV-C-1-b-(2). Minor Actions

Section IV-C-2. Recombinant DNA Advisory Committee (RAC)

Section IV-C-3. Office of Recombinant DNA Activities (ORDA)

Section IV-C-4. Other NIH Components

Section IV-D. Compliance with the NIH Guidelines

Section IV-E. Voluntary Compliance

Section V. Footnotes and References of Sections I-IV

Appendix A. Exemptions under Section III-E-5--Sublists of Natural

Exchangers

Appendix B. Classification of Etiologic Agents and Oncogenic Viruses

on the Basis of Hazard

Appendix B-I. Class 1 Agents

Appendix B-II. Class 2 Agents

Appendix B-III. Class 3 Agents

Appendix B-IV. Class 4 Agents

Appendix B-V. Class 5 Agents

Appendix B-VI. Footnotes and References of Appendix B

Appendix C. Exemptions under Section III-E-6

Appendix C-I. Recombinant DNA in Tissue Culture

Appendix C-II. Escherichia coli K-12 Host-Vector Systems

Appendix C-III. Saccharomyces Host-Vector Systems

Appendix C-IV. Bacillus subtilis or Bacillus licheniformis Host-

Vector Systems

Appendix C-V. Extrachromosomal Elements of Gram Positive Organisms

Appendix C-VI. Footnotes and References of Appendix C

Appendix D. Major Actions Taken under the NIH Guidelines

Appendix E. Certified Host-Vector Systems

Appendix E-I. Bacillus subtilis

Appendix E-II. Saccharomyces cerevisiae

Appendix E-III. Escherichia coli

Appendix E-IV. Neurospora crassa

Appendix E-V. Streptomyces

Appendix E-VI. Pseudomonas putida

Appendix F. Containment Conditions for Cloning of Genes Coding for

the Biosynthesis of Molecules Toxic for Vertebrates

Appendix F-I. General Information

Appendix F-II. Cloning of Toxin Molecule Genes in Escherichia coli

K-12

Appendix F-III. Cloning of Toxic Molecule Genes in Organisms other

than Escherichia coli K-12

Appendix F-IV. Specific Approvals

Appendix G. Physical Containment

Appendix G-I. Standard Practices and Training

Appendix G-II. Physical Containment Levels

Appendix G-II-A. Biosafety Level 1 (BL1)

Appendix G-II-B. Biosafety Level 2 (BL2)

Appendix G-II-C. Biosafety Level 3 (BL3)

Appendix G-II-D. Biosafety Level 4 (BL4)

Appendix G-III. Footnotes and References of Appendix G

Appendix H. Shipment

Appendix I. Biological Containment

Appendix I-I. Levels of Biological Containment

Appendix I-I-A. Host-Vector 1 Systems

Appendix I-I-B. Host-Vector 2 Systems

Appendix I-II. Certification of Host-Vector Systems

Appendix I-III. Footnotes and References of Appendix I

Appendix J. Biotechnology Research Subcommittee

Appendix K. Physical Containment for Large Scale Uses of Organisms

Containing Recombinant DNA Molecules

Appendix K-I. Selection of Physical Containment Levels

Appendix K-II. Good Large Scale Practices (GLSP)

Appendix K-III. Biosafety Level 1 (BL1)--Large Scale

Appendix K-IV. Biosafety Level 2 (BL2)--Large Scale

Appendix K-V. Biosafety Level 3 (BL3)--Large Scale

Appendix K-VI. Footnotes of Appendix K

Appendix K-VII. Definitions to Accompany Containment Grid and

Appendix K

Appendix L. Release into the Environment of Certain Plants

Appendix M. Points to Consider in the Design and Submission of

Protocols for the Transfer of Recombinant DNA Molecules into the

Genome of One or More Human Subjects

Appendix M-I. Description of Proposal

Appendix M-I-A. Objectives and Rationale of the Proposed Research

Appendix M-I-B. Research Design, Anticipated Risks and Benefits

Appendix M-I-C. Selection of the Patients

Appendix M-I-D. Informed Consent

Appendix M-I-E. Privacy and Confidentiality

Appendix M-II. Special Issues

Appendix M-III. Guidelines for the Submission of Human Gene Transfer

Protocols

Appendix M-III-A. Principal Investigator-Submitted Material

Appendix M-III-B. Time Frame for Submissions

Appendix M-III-C. Oral Responses to the RAC

Appendix M-III-D. Primary Reviewers' Responses

Appendix M-IV. Reporting Requirements

Appendix M-V. Procedures to be Followed for Accelerated Review of

Human Gene Transfer Experiments by NIH/ORDA under Section III-B-2

Appendix M-VI. Procedures to be Followed for Expedited Review of

Single Patient Human Gene Transfer Experiments by the NIH Director

Under Section III-A-2

Appendix M-VII. Footnotes of Appendix M

Appendix P. Physical and Biological Containment for Recombinant DNA

Research Involving Plants

Appendix P-I. General Plant Biosafety Levels

Appendix P-II. Physical Containment Levels

Appendix P-II-A. Biosafety Level 1--Plants (BL1-P)

Appendix P-II-B. Biosafety Level 2--Plants (BL2-P)

Appendix P-II-C. Biosafety Level 3--Plants (BL3-P)

Appendix P-II-D. Biosafety Level 4--Plants (BL4-P)

Appendix P-III. Biological Containment Practices

Appendix P-III-A. Biological Containment Practices (Plants)

Appendix P-III-B. Biological Containment Practices (Microorganisms)

Appendix P-III-C. Biological Containment Practices (Macroorganisms)

Appendix Q. Physical and Biological Containment for Recombinant DNA

Research Involving Animals

Appendix Q-I. General Considerations

Appendix Q-I-A. Containment Levels

Appendix Q-I-B. Disposal of Animals (BL1-N through BL4-N)

Appendix Q-II. Physical and Biological Containment Levels

Appendix Q-II-A. Biosafety Level 1--Animals (BL1-N)

Appendix Q-II-B. Biosafety Level 2--Animals (BL2-N)

Appendix Q-II-C. Biosafety Level 3--Animals (BL3-N)

Appendix Q-II-D. Biosafety Level 4--Animals (BL4-N)

Appendix Q-III. Footnotes and References for Appendix Q

Section I. Scope of the NIH Guidelines

Section I-A. Purpose

The purpose of the NIH Guidelines is to specify practices for

constructing and handling: (i) Recombinant deoxyribonucleic acid (DNA)

molecules, and (ii) organisms and viruses containing recombinant DNA

molecules.

Section I-A-1. Any recombinant DNA experiment, which according to

the NIH Guidelines requires approval by the NIH, must be submitted to

the NIH or to another Federal agency that has jurisdiction for review

and approval. Once approval, or other applicable clearances, has been

obtained from a Federal agency other than the NIH (whether the

experiment is referred to that agency by the NIH or sent directly there

by the submitter), the experiment may proceed without the necessity for

NIH review or approval (see exceptions in Sections I-A-2 and I-A-3).

Section I-A-2. Certain experiments that involve the deliberate

transfer of recombinant DNA or DNA or RNA derived from recombinant DNA

into one or more human subjects (see Section V-U) shall be considered

Major Actions (see Section IV-C-1-b-(1)), and shall require RAC review

and NIH Director approval, if determined by NIH/ORDA in consultation

with the RAC Chair and/or one or more RAC members, as necessary, to:

(i) Represent novel characteristics (e.g., target disease or vector),

(ii) represent an uncertain degree of risk to human health or the

environment, or (iii) contain information determined to require further

public review (see Section III-A-2).

Section I-A-3. Experiments involving the transfer of recombinant

DNA to one or more human subjects that are not considered under Section

III-A-2 may qualify for Accelerated Review (see Section III-B-2 and

Appendix M-V) and will be considered as Minor Actions (see Section IV-

C-1-b-(2)-(a)). Actions that qualify for Accelerated Review will be

reviewed and approved by NIH/ORDA in consultation with the RAC Chair

and/or one or more RAC members, as necessary.

Certain experiments involving the transfer of recombinant DNA or

DNA or RNA derived from recombinant DNA into one or more human subjects

(see Section V-U) may be considered exempt from RAC and/or NIH/ORDA

review and/or NIH Director approval and only require registration with

NIH/ORDA (see Section III-C-7).

Section I-B. Definition of Recombinant DNA Molecules

In the context of the NIH Guidelines, recombinant DNA molecules are

defined as either: (i) Molecules that are constructed outside living

cells by joining natural or synthetic DNA segments to DNA molecules

that can replicate in a living cell, or (ii) molecules that result from

the replication of those described in (i) above.

Synthetic DNA segments which are likely to yield a potentially

harmful polynucleotide or polypeptide (e.g., a toxin or a

pharmacologically active agent) are considered as equivalent to their

natural DNA counterpart. If the synthetic DNA segment is not expressed

in vivo as a biologically active polynucleotide or polypeptide product,

it is exempt from the NIH Guidelines.

Genomic DNA of plants and bacteria that have acquired a

transposable element, even if the latter was donated from a recombinant

vector no longer present, are not subject to the NIH Guidelines unless

the transposon itself contains recombinant DNA.

Section I-C. General Applicability

Section I-C-1. The NIH Guidelines are applicable to:

Section I-C-1-a. All recombinant DNA research within the United

States (U.S.) or its territories that is conducted at or sponsored by

an institution that receives any support for recombinant DNA research

from the NIH, including research performed directly by the NIH. An

individual who receives support for research involving recombinant DNA

must be associated with or sponsored by an institution that assumes the

responsibilities assigned in the NIH Guidelines.

Section I-C-1-b. All recombinant DNA research performed abroad:

Specifically:

Section I-C-1-b-(1). Research supported by NIH funds.

Section I-C-1-b-(2). If they involve testing in humans of materials

containing recombinant DNA developed with NIH funds and if the

institution that developed those materials sponsors or participates in

those projects. Participation includes research collaboration or

contractual agreements, not mere provision of research materials.

Section I-C-1-b-(3). If the host country has established rules for

the conduct of recombinant DNA research, then the research must be in

compliance with those rules. If the host country does not have such

rules, the proposed research must be reviewed and approved by an NIH-

approved Institutional Biosafety Committee or equivalent review body

and accepted in writing by an appropriate national governmental

authority of the host country. The safety practices that are employed

abroad must be reasonably consistent with the NIH Guidelines.

Section I-D. General Definitions

The following terms, which are used throughout the NIH Guidelines,

are defined as follows:

Section I-D-1. An ``institution'' is any public or private entity

(including Federal, state, and local government agencies).

Section I-D-2. An ``Institutional Biosafety Committee'' is a

committee that: (i) Meets the requirements for membership specified in

Section IV-B-2, and (ii) reviews, approves, and oversees projects in

accordance with the responsibilities defined in Section IV-B-2.

Section I-D-3. The ``Office of Recombinant DNA Activities (ORDA)''

is the office within the NIH that is responsible for: (i) Reviewing and

coordinating all activities relating to the NIH Guidelines, and (ii)

performing other duties as defined in Section IV-C-3.

Section I-D-4. The ``Recombinant DNA Advisory Committee'' is the

public advisory committee that advises the Department of Health and

Human Services (DHHS) Secretary, the DHHS Assistant Secretary for

Health, and the NIH Director concerning recombinant DNA research. The

RAC shall be constituted as specified in Section IV-C-2.

Section I-D-5. The ``NIH Director'' is the Director of the National

Institutes of Health, or any other officer or employee of NIH to whom

authority has been delegated.

Section I-D-6. ``Deliberate release'' is defined as a planned

introduction of recombinant DNA-containing microorganisms, plants, or

animals into the environment.

Section II. Containment

Effective biological safety programs have been operative in a

variety of laboratories for many years. Considerable information

already exists about the design of physical containment facilities and

selection of laboratory procedures applicable to organisms carrying

recombinant DNA (see section V-A). The existing programs rely upon

mechanisms that can be divided into two categories: (i) A set of

standard practices that are generally used in microbiological

laboratories; and (ii) special procedures, equipment, and laboratory

installations that provide physical barriers that are applied in

varying degrees according to the estimated biohazard. Four biosafety

levels are described in Appendix G. These biosafety levels consist of

combinations of laboratory practices and techniques, safety equipment,

and laboratory facilities appropriate for the operations performed and

are based on the potential hazards imposed by the agents used and for

the laboratory function and activity. Biosafety Level 4 provides the

most stringent containment conditions, Biosafety Level 1 the least

stringent.

Experiments involving recombinant DNA lend themselves to a third

containment mechanism, namely, the application of highly specific

biological barriers. Natural barriers exist that limit either: (i) The

infectivity of a vector or vehicle (plasmid or virus) for specific

hosts, or (ii) its dissemination and survival in the environment.

Vectors, which provide the means for recombinant DNA and/or host cell

replication, can be genetically designed to decrease, by many orders of

magnitude, the probability of dissemination of recombinant DNA outside

the laboratory (see Appendix I).

Since these three means of containment are complementary, different

levels of containment can be established that apply various

combinations of the physical and biological barriers along with a

constant use of standard practices. Categories of containment are

considered separately in order that such combinations can be

conveniently expressed in the NIH Guidelines.

Physical containment conditions within laboratories, described in

Appendix G, may not always be appropriate for all organisms because of

their physical size, the number of organisms needed for an experiment,

or the particular growth requirements of the organism. Likewise,

biological containment for microorganisms described in Appendix I may

not be appropriate for all organisms, particularly higher eukaryotic

organisms. However, significant information exists about the design of

research facilities and experimental procedures that are applicable to

organisms containing recombinant DNA that is either integrated into the

genome or into microorganisms associated with the higher organism as a

symbiont, pathogen, or other relationship. This information describes

facilities for physical containment of organisms used in non-

traditional laboratory settings and special practices for limiting or

excluding the unwanted establishment, transfer of genetic information,

and dissemination of organisms beyond the intended location, based on

both physical and biological containment principles. Research conducted

in accordance with these conditions effectively confines the organism.

For research involving plants, four biosafety levels (BL1-P through

BL4-P) are described in Appendix P. BL1-P is designed to provide a

moderate level of containment for experiments for which there is

convincing biological evidence that precludes the possibility of

survival, transfer, or dissemination of recombinant DNA into the

environment, or in which there is no recognizable and predictable risk

to the environment in the event of accidental release. BL2-P is

designed to provide a greater level of containment for experiments

involving plants and certain associated organisms in which there is a

recognized possibility of survival, transmission, or dissemination of

recombinant DNA containing organisms, but the consequence of such an

inadvertent release has a predictably minimal biological impact. BL3-P

and BL4-P describe additional containment conditions for research with

plants and certain pathogens and other organisms that require special

containment because of their recognized potential for significant

detrimental impact on managed or natural ecosystems. BL1-P relies upon

accepted scientific practices for conducting research in most ordinary

greenhouse or growth chamber facilities and incorporates accepted

procedures for good pest control and cultural practices. BL1-P

facilities and procedures provide a modified and protected environment

for the propagation of plants and microorganisms associated with the

plants and a degree of containment that adequately controls the

potential for release of biologically viable plants, plant parts, and

microorganisms associated with them. BL2-P and BL3-P rely upon accepted

scientific practices for conducting research in greenhouses with

organisms infecting or infesting plants in a manner that minimizes or

prevents inadvertent contamination of plants within or surrounding the

greenhouse. BL4-P describes facilities and practices known to provide

containment of certain exotic plant pathogens.

For research involving animals, which are of a size or have growth

requirements that preclude the use of conventional primary containment

systems used for small laboratory animals, four biosafety levels (BL1-N

through BL4-N) are described in Appendix Q. BL1-N describes containment

for animals that have been modified by stable introduction of

recombinant DNA, or DNA derived therefrom, into the germ-line

(transgenic animals) and experiments involving viable recombinant DNA-

modified microorganisms and is designed to eliminate the possibility of

sexual transmission of the modified genome or transmission of

recombinant DNA-derived viruses known to be transmitted from animal

parent to offspring only by sexual reproduction. Procedures, practices,

and facilities follow classical methods of avoiding genetic exchange

between animals. BL2-N describes containment which is used for

transgenic animals associated with recombinant DNA-derived organisms

and is designed to eliminate the possibility of vertical or horizontal

transmission. Procedures, practices, and facilities follow classical

methods of avoiding genetic exchange between animals or controlling

arthropod transmission. BL3-N and BL4-N describe higher levels of

containment for research with certain transgenic animals involving

agents which pose recognized hazard.

In constructing the NIH Guidelines, it was necessary to define

boundary conditions for the different levels of physical and biological

containment and for the classes of experiments to which they apply.

These definitions do not take into account all existing and anticipated

information on special procedures that will allow particular

experiments to be conducted under different conditions than indicated

here without affecting risk. Individual investigators and Institutional

Biosafety Committees are urged to devise simple and more effective

containment procedures and to submit recommended changes in the NIH

Guidelines to permit the use of these procedures.

Section III. Experiments Covered by the NIH Guidelines

This section describes five categories of experiments involving

recombinant DNA: (i) Those that require RAC review and NIH and

Institutional Biosafety Committee approval before initiation (see

section III-A), (ii) those that require NIH/ORDA and Institutional

Biosafety Committee approval before initiation (see section III-B);

(iii) those that require Institutional Biosafety Committee approval

before initiation (see section III-C), (iv) those that require

Institutional Biosafety Committee notification simultaneous with

initiation (see section III-D), and (v) those that are exempt from the

NIH Guidelines (see section III-E).

Note: If an experiment falls into either section III-A or

section III-B and one of the other categories, the rules pertaining

to section III-A or section III-B shall be followed. If an

experiment falls into section III-E and into either sections III-C

or III-D categories as well, the experiment is considered exempt

from the NIH Guidelines.

Any change in containment level, which is different from those

specified in the NIH Guidelines, may not be initiated without the

express approval of NIH/ORDA (see Minor Actions, section IV-C-1-b-(2)

and its subsections).

Section III-A. Experiments that Require Institutional Biosafety

Committee Approval, RAC Review, and NIH Approval Before Initiation

Experiments in this category are considered Major Actions (see

section IV-C-1-b-(1)) and cannot be initiated without submission of

relevant information on the proposed experiment to the Office of

Recombinant DNA Activities, National Institutes of Health, Building 31,

room 4B11, Bethesda, Maryland 20892, (301) 496-9838, the publication of

the proposal in the Federal Register for 15 days of comment, reviewed

by the RAC, and specific approval by the NIH (not applicable for

Expedited Review single patient human gene transfer experiments

considered under Appendix M-VI). The containment conditions for such

experiments will be recommended by the RAC and set by the NIH at the

time of approval. Such experiments require Institutional Biosafety

Committee approval before initiation. Specific experiments already

approved are included in Appendix D which may be obtained from the

Office of Recombinant DNA Activities, National Institutes of Health,

Building 31, room 4B11, Bethesda, Maryland 20892, (301) 496-9838.

Section III-A-1. Deliberate transfer of a drug resistance trait to

microorganisms that are not known to acquire the trait naturally (see

section V-B), if such acquisition could compromise the use of the drug

to control disease agents in humans, veterinary medicine, or

agriculture.

Section III-A-2. Certain experiments involving the deliberate

transfer of recombinant DNA or DNA or RNA derived from recombinant DNA

into one or more human subjects (see section V-U) shall be considered

Major Actions (see section IV-C-1-b-(1) and Appendix M-III), and shall

require RAC review and NIH Director approval, if determined by NIH/

ORDA, in consultation with the RAC Chair and one or more RAC members,

as necessary, to: (i) Represent novel characteristics (e.g., target

disease or vector), (ii) represent an uncertain degree of risk to human

health or the environment, or (iii) contain information determined to

require further public review. The requirement for RAC review shall not

be considered to preempt any other required review or approval of

experiments with one or more human subjects. Relevant Institutional

Biosafety Committee and Institutional Review Board reviews and

approvals of the proposal should be completed before submission to NIH.

Certain experiments involving deliberate transfer of recombinant DNA or

DNA or RNA derived from recombinant DNA into one or more human subjects

may qualify for the Accelerated Review process (see section III-B-2).

Certain categories of experiments involving the deliberate transfer of

recombinant DNA or DNA or RNA derived from recombinant DNA into one or

more human subjects and that are not covered by section V-U, may be

considered exempt from RAC and/or NIH/ORDA review and/or NIH Director

approval and only require registration with NIH/ORDA (see section III-

C-7).

Section III-B. Experiments That Require NIH/ORDA and Institutional

Biosafety Committee Approval Before Initiation

Section III-B-1. Experiments Involving the Cloning of Toxin Molecules

with LD50 of Less than 100 Nanograms per Kilogram Body Weight

Deliberate formation of recombinant DNA containing genes for the

biosynthesis of toxin molecules lethal for vertebrates at an LD50

of less than 100 nanograms per kilogram body weight (e.g., microbial

toxins such as the botulinum toxins, tetanus toxin, diphtheria toxin,

and Shigella dysenteriae neurotoxin). Specific approval has been given

for the cloning in Escherichia coli K-12 of DNA containing genes coding

for the biosynthesis of toxic molecules which are lethal to vertebrates

at 100 nanograms to 100 micrograms per kilogram body weight. Specific

experiments already approved under this section may be obtained from

the Office of Recombinant DNA Activities, National Institutes of

Health, Building 31, room 4B11, Bethesda, Maryland 20892, (301) 496-

9838.

Section III-B-1-(a). Experiments in this category cannot be

initiated without submission of relevant information on the proposed

experiment to NIH/ORDA. The containment conditions for such experiments

will be determined by NIH/ORDA in consultation with ad hoc experts.

Such experiments require Institutional Biosafety Committee approval

before initiation (see section IV-B-2-b-(1)).

Section III-B-2. Accelerated Review of Human Gene Transfer Experiments

As determined by NIH/ORDA, in consultation with the RAC Chair and

one or more RAC members, as necessary, certain categories of human gene

transfer experiments may be considered as Minor Actions and qualify for

Accelerated Review and approval (see section IV-C-1-b-(2)-(a), Appendix

M-III-A, and Appendix M-V). The RAC Chair will present a report of all

NIH/ORDA approved human gene transfer protocols at the next regularly

scheduled RAC meeting. If NIH/ORDA determines that an experiment does

not qualify for the Accelerated Review process, the Principal

Investigator must submit the proposal for full RAC review 8

weeks prior to the next scheduled RAC meeting (See section III-A-2).

Section III-B-3. Minor Modifications to Human Gene Transfer Experiments

A minor modification in a human gene transfer protocol is a

modification that does not significantly alter the basic design of the

protocol and that does not increase risk to human subjects or the

environment. After approval has been obtained by the relevant

Institutional Biosafety Committee and Institutional Review Board, NIH/

ORDA will consider the change in consultation with the RAC Chair and

one or more RAC members, as necessary. Submit minor modifications to

the Office of Recombinant DNA Activities, National Institutes of

Health, Building 31, Room 4B11, Bethesda, Maryland 20892, (301) 496-

9838. The RAC Chair will provide a report on any such approvals at the

next regularly scheduled RAC meeting.

Section III-C. Experiments that Require Institutional Biosafety

Committee Approval Before Initiation

Prior to the initiation of an experiment that falls into this

category, the Principal Investigator must submit a registration

document to the Institutional Biosafety Committee which contains the

following information: (i) The source(s) of DNA; (ii) the nature of the

inserted DNA sequences; (iii) the host(s) and vector(s) to be used;

(iv) if an attempt will be made to obtain expression of a foreign gene,

and if so, indicate the protein that will be produced; and (v) the

containment conditions that will be implemented as specified in the NIH

Guidelines. For experiments in this category, the registration document

shall be dated, signed by the Principal Investigator, and filed with

the Institutional Biosafety Committee. The Institutional Biosafety

Committee shall review and approve all experiments in this category

prior to their initiation. Requests to decrease the level of

containment specified for experiments in this category will be

considered by NIH (see Section IV-C-1-b-(2)-(c)).

Section III-C-1. Experiments Using Human or Animal Pathogens (Class 2,

Class 3, Class 4, or Class 5 Agents (see Section V-A) as Host-Vector

Systems

Section III-C-1-a. Experiments involving the introduction of

recombinant DNA into Class 2 agents shall be conducted at Biosafety

Level (BL) 2 containment. Experiments with such agents shall be

conducted with whole animals at BL2 or BL2-N (Animals) containment.

Section III-C-1-b. Experiments involving the introduction of

recombinant DNA into Class 3 agents shall be conducted at BL3

containment. Experiments with such agents shall be conducted with whole

animals at BL3 or BL3-N containment.

Section III-C-1-c. Experiments involving the introduction of

recombinant DNA into Class 4 agents shall be conducted at BL4

containment. Experiments with such agents shall be conducted with whole

animals at BL4 or BL4-N containment.

Section III-C-1-d. Containment conditions for experiments involving

the introduction of recombinant DNA into Class 5 agents shall be set on

a case-by-case basis following NIH/ORDA review. A U.S. Department of

Agriculture permit is required for work with Class 5 agents (see

Sections V-R and V-T). Experiments with such agents shall be conducted

with whole animals at BL4 or BL4-N containment.

Section III-C-2. Experiments in Which DNA From Human or Animal

Pathogens (Class 2, Class 3, Class 4, or Class 5 Agents (see Section V-

A) is Cloned Into Nonpathogenic Prokaryotic or Lower Eukaryotic Host-

Vector Systems

Section III-C-2-a. Experiments in which DNA from Class 2 or Class 3

agents (see Section V-A) is transferred into nonpathogenic prokaryotes

or lower eukaryotes may be performed under BL2 containment. Experiments

in which DNA from Class 4 agents is transferred into nonpathogenic

prokaryotes or lower eukaryotes may be performed under BL2 containment

after demonstration that only a totally and irreversibly defective

fraction of the agent's genome is present in a given recombinant. In

the absence of such a demonstration, BL4 containment shall be used. The

Institutional Biosafety Committee may approve the specific lowering of

containment for particular experiments to BL1. Many experiments in this

category are exempt from the NIH Guidelines (see Section III-E).

Experiments involving the formation of recombinant DNA for certain

genes coding for molecules toxic for vertebrates require NIH/ORDA

approval (see Section III-B-1) or shall be conducted under NIH

specified conditions as described in Appendix F.

Section III-C-2-b. Containment conditions for experiments in which

DNA from Class 5 agents is transferred into nonpathogenic prokaryotes

or lower eukaryotes shall be determined by NIH/ORDA following a case-

by-case review. A U.S. Department of Agriculture permit is required for

work with Class 5 agents (see Sections V-R and V-T).

Section III-C-3. Experiments Involving the Use of Infectious Animal or

Plant DNA or RNA Viruses or Defective Animal or Plant DNA or RNA

Viruses in the Presence of Helper Virus in Tissue Culture Systems

Caution: Special care should be used in the evaluation of

containment levels for experiments which are likely to either enhance

the pathogenicity (e.g., insertion of a host oncogene) or to extend the

host range (e.g., introduction of novel control elements) of viral

vectors under conditions that permit a productive infection. In such

cases, serious consideration should be given to increasing physical

containment by at least one level.

Note: Recombinant DNA or RNA molecules derived therefrom, which

contain less than two-thirds of the genome of any eukaryotic virus

(all viruses from a single Family (see Section V-Q) being considered

identical (see Section V-S), are considered defective and may be

used in the absence of helper under the conditions specified in

Section III-D-1.

Section III-C-3-a. Experiments involving the use of infectious or

defective Class 2 animal viruses (see Section V-A, Appendix B-II, and

Appendix B-II-E) in the presence of helper virus may be conducted at

BL2.

Section III-C-3-b. Experiments involving the use of infectious or

defective Class 3 animal viruses (see Section V-A and Appendix B-III-D)

in the presence of helper virus may be conducted at BL3.

Section III-C-3-c. Experiments involving the use of infectious or

defective Class 4 animal viruses (see Section V-A and Appendix B-IV-D)

in the presence of helper virus may be conducted at BL4.

Section III-C-3-d. Experiments involving the use of infectious or

defective Class 5 viruses (see Section V-A and Appendix B-V) in the

presence of helper virus shall be determined on a case-by-case basis

following NIH/ORDA review. A U.S. Department of Agriculture permit is

required for work with Class 5 agents (see Sections V-R and V-T).

Section III-C-3-e. Experiments involving the use of infectious or

defective animal or plant viruses in the presence of helper virus are

not covered in Sections III-C-3-a through III-C-3-d and may be

conducted at BL1.

Section III-C-4. Experiments Involving Whole Animals

This section covers experiments involving whole animals in which

the animal's genome has been altered by stable introduction of

recombinant DNA, or DNA derived therefrom, into the germ-line

(transgenic animals) and experiments involving viable recombinant DNA-

modified microorganisms tested on whole animals. For the latter, other

than viruses which are only vertically transmitted, the experiments may

not be conducted at BL1-N containment. A minimum containment of BL2 or

BL2-N is required.

Caution--Special care should be used in the evaluation of

containment conditions for some experiments with transgenic animals.

For example, such experiments might lead to the creation of novel

mechanisms or increased transmission of a recombinant pathogen or

production of undesirable traits in the host animal. In such cases,

serious consideration should be given to increasing the containment

conditions.

Section III-C-4-a. Recombinant DNA, or DNA or RNA molecules derived

therefrom, from any source except for greater than two-thirds of

eukaryotic viral genome may be transferred to any non-human vertebrate

or any invertebrate organism and propagated under conditions of

physical containment comparable to BL1 or BL1-N and appropriate to the

organism under study (see Section V-B). Animals that contain sequences

from viral vectors, which do not lead to transmissible infection either

directly or indirectly as a result of complementation or recombination

in animals, may be propagated under conditions of physical containment

comparable to BL1 or BL1-N and appropriate to the organism under study.

Experiments involving the introduction of other sequences from

eukaryotic viral genomes into animals are covered under Section III-C-

4-b. For experiments involving recombinant DNA-modified Class 2, 3, 4,

or 5 organisms, see Section V-A. It is important that the investigator

demonstrate that the fraction of the viral genome being utilized does

not lead to productive infection. A U.S. Department of Agriculture

permit is required for work with Class 5 agents (see Section V-R and V-

T).

Section III-C-4-b. For experiments involving recombinant DNA, or

DNA or RNA derived therefrom, involving whole animals, including

transgenic animals, and not covered by Sections III-C-1 or III-C-4-a,

the appropriate containment shall be determined by the Institutional

Biosafety Committee.

Section III-C-5. Experiments Involving Whole Plants

Experiments to genetically engineer plants by recombinant DNA

methods, to use such plants for other experimental purposes (e.g.,

response to stress), to propagate such plants, or to use plants

together with microorganisms or insects containing recombinant DNA, may

be conducted under the containment conditions described in Sections

III-C-5-a through III-C-5-e. If experiments involving whole plants are

not described in Section III-C-5 and do not fall under Sections III-A,

III-B, or III-E, they are included in Section III-D.

Note: For recombinant DNA experiments falling under Sections

III-C-5-a through III-C-5-d, physical containment requirements may

be reduced to the next lower level by appropriate biological

containment practices, such as conducting experiments on a virus

with an obligate insect vector in the absence of that vector or

using a genetically attenuated strain.

Section III-C-5-a. BL3-P (Plants) or BL2-P + biological containment

is recommended for experiments involving most exotic (see Section V-W)

infectious agents with recognized potential for serious detrimental

impact on managed or natural ecosystems when recombinant DNA techniques

are associated with whole plants.

Section III-C-5-b. BL3-P or BL2-P + biological containment is

recommended for experiments involving plants containing cloned genomes

of readily transmissible exotic (see Section V-W) infectious agents

with recognized potential for serious detrimental effects on managed or

natural ecosystems in which there exists the possibility of

reconstituting the complete and functional genome of the infectious

agent by genomic complementation in planta.

Section III-C-5-c. BL4-P containment is recommended for experiments

with a small number of readily transmissible exotic (see Section V-W)

infectious agents, such as the soybean rust fungus (Phakospora

pachyrhizi) and maize streak or other viruses in the presence of their

specific arthropod vectors, that have the potential of being serious

pathogens of major U.S. crops.

Section III-C-5-d. BL3-P containment is recommended for experiments

involving sequences encoding potent vertebrate toxins introduced into

plants or associated organisms. Recombinant DNA containing genes for

the biosynthesis of toxin molecules lethal for vertebrates at an

LD50 of 50

100 nanograms per kilogram body weight in organisms other

than Escherichia coli K-12 (see Section III-B-1 and Appendices F-I and

F-II);

Section IV-C-3-b. Reviewing and approving certain experiments

involving the deliberate transfer of recombinant DNA or DNA or RNA

derived from recombinant DNA into one or more human subjects, in

consultation with the RAC Chair and one or more RAC members, as

necessary, that qualify for the Accelerated Review process (see Section

III-B-2);

Section IV-C-3-c. Reviewing and approving minor changes to human

gene transfer protocols approved under Sections III-A-2 and III-B-2, in

consultation with the RAC Chair and one or more RAC members, as

necessary;

Section IV-C-3-d. Reviewing and approving the membership of an

institution's Institutional Biosafety Committee, and where it finds the

Institutional Biosafety Committee meets the requirements set forth in

Section IV-B-2 will give its approval to the Institutional Biosafety

Committee membership;

Section IV-C-3-e. Publishing in the Federal Register:

Section IV-C-3-e-(1). Announcements of RAC meetings and agendas at

least 15 days in advance (NOTE--If the agenda for a RAC meeting is

modified, ORDA shall make the revised agenda available to anyone upon

request at least 72 hours in advance of the meeting);

Section IV-C-3-e-(2). Proposed Major Actions to the NIH Guidelines

(see Section IV-C-1-b-(1)) at least 15 days prior to the RAC meeting;

Section IV-C-3-f. Serve as the focal point for data management of

NIH-approved human gene transfer protocols approved under Sections III-

A-2 and III-B-2 and registered with NIH/ORDA as required under Section

III-C-7;

Section IV-C-3-g. Serve as the executive secretary of the RAC; and

Section IV-C-3-h. Maintain a list of Major and Minor Actions

approved under Section III-A-2 and III-B-3 and a list of experiments

registered with NIH/ORDA as described in Section III-C-7.

Section IV-C-4. Other NIH Components

Other NIH components shall be responsible for certifying maximum

containment (BL4) facilities, inspecting them periodically, and

inspecting other recombinant DNA facilities as deemed necessary.

Section IV-D. Compliance with the NIH Guidelines

As a condition for NIH funding of recombinant DNA research,

institutions shall ensure that such research conducted at or sponsored

by the institution, irrespective of the source of funding, shall comply

with the NIH Guidelines. The policies on noncompliance are as follows:

All NIH-funded projects involving recombinant DNA techniques must

comply with the NIH Guidelines. Non-compliance may result in: (i)

suspension, limitation, or termination of financial assistance for such

projects and of NIH funds for other recombinant DNA research at the

institution, or (ii) a requirement for prior NIH approval of any or all

recombinant DNA projects at the institution.

All non-NIH funded projects involving recombinant DNA techniques

conducted at or sponsored by an institution that receives NIH funds for

projects involving such techniques must comply with the NIH Guidelines.

Noncompliance may result in: (i) suspension, limitation, or termination

of NIH funds for recombinant DNA research at the institution, or (ii) a

requirement for prior NIH approval of any or all recombinant DNA

projects at the institution.

Information concerning noncompliance with the NIH Guidelines may be

brought forward by any person. It should be delivered to both NIH/ORDA

and the relevant institution. The institution, generally through the

Institutional Biosafety Committee, shall take appropriate action. The

institution shall forward a complete report of the incident

recommending any further action to the Office of Recombinant DNA

Activities, National Institutes of Health, Building 31, Room 4B11,

Bethesda, Maryland 20892, (301) 496-9838.

In cases where NIH proposes to suspend, limit, or terminate

financial assistance because of noncompliance with the NIH Guidelines,

applicable DHHS and Public Health Service procedures shall govern.

Section IV-E. Voluntary Compliance

Section IV-E-1. Basic Policy

Individuals, corporations, and institutions not otherwise covered

by the NIH Guidelines are encouraged to do so by following the

standards and procedures set forth in Sections I through IV. In order

to simplify discussion, references hereafter to ``institutions'' are

intended to encompass corporations and individuals who have no

organizational affiliation. For purposes of complying with the NIH

Guidelines, an individual intending to carry out research involving

recombinant DNA is encouraged to affiliate with an institution that has

an Institutional Biosafety Committee approved under the NIH Guidelines.

Since commercial organizations have special concerns, such as

protection of proprietary data, some modifications and explanations of

the procedures are provided in Sections IV-E-2 through IV-E-5-b in

order to address these concerns.

Section IV-E-2. Institutional Biosafety Committee Approval

It should be emphasized that employment of an Institutional

Biosafety Committee member solely for purposes of membership on the

Institutional Biosafety Committee does not itself make the member an

institutionally affiliated member. Except for the unaffiliated members,

a member of an Institutional Biosafety Committee for an institution not

otherwise covered by the NIH Guidelines may participate in the review

and approval of a project in which the member has a direct financial

interest so long as the member has not been, and does not expect to be,

engaged in the project. Section IV-B-2-a-(4) is modified to that extent

for purposes of these institutions.

Section IV-E-3. Certification of Host-Vector Systems

A host-vector system may be proposed for certification by the NIH

Director in accordance with the procedures set forth in Appendix I-II.

In order to ensure protection for proprietary data, any public notice

regarding a host-vector system which is designated by the institution

as proprietary under Section IV-E-5-a will be issued only after

consultation with the institution as to the content of the notice.

Section IV-E-4. Requests for Exemptions and Approvals

Requests for exemptions or other approvals as required by the NIH

Guidelines should be submitted based on the procedures set forth in

Sections I through IV. In order to ensure protection for proprietary

data, any public notice regarding a request for an exemption or other

approval which is designated by the institution as proprietary under

Section IV-E-5-a will be issued only after consultation with the

institution as to the content of the notice.

Section IV-E-5. Protection of Proprietary Data

Section IV-E-5-a. General

In general, the Freedom of Information Act requires Federal

agencies to make their records available to the public upon request.

However, this requirement does not apply to, among other things,

``trade secrets and commercial or financial information that is

obtained from a person and that is privileged or confidential.'' Under

18 U.S.C. 1905, it is a criminal offense for an officer or employee of

the U.S. or any Federal department or agency to publish, divulge,

disclose, or make known ``in any manner or to any extent not authorized

by law any information coming to him in the course of his employment or

official duties or by reason of any examination or investigation made

by, or return, report or record made to or filed with, such department

or agency or officer or employee thereof, which information concerns or

relates to the trade secrets, (or) processes--of any person, firm,

partnership, corporation, or association.'' This provision applies to

all employees of the Federal Government, including special Government

employees. Members of the RAC are ``special Government employees.''

In submitting to NIH for purposes of voluntary compliance with the

NIH Guidelines, an institution may designate those items of information

which the institution believes constitute trade secrets, privileged,

confidential, commercial, or financial information. If NIH receives a

request under the Freedom of Information Act for information so

designated, NIH will promptly contact the institution to secure its

views as to whether the information (or some portion) should be

released. If the NIH decides to release this information (or some

portion) in response to a Freedom of Information request or otherwise,

the institution will be advised; and the actual release will not be

made until the expiration of 15 days after the institution is so

advised except to the extent that earlier release in the judgment of

the NIH Director is necessary to protect against an imminent hazard to

the public or the environment.

Section IV-E-5-b. Presubmission Review

Any institution not otherwise covered by the NIH Guidelines, which

is considering submission of data or information voluntarily to NIH,

may request presubmission review of the records involved to determine

if NIH will make all or part of the records available upon request

under the Freedom of Information Act.

A request for presubmission review should be submitted to NIH/ORDA

along with the records involved to the Office of Recombinant DNA

Activities, National Institutes of Health, Building 31, Room 4B11,

Bethesda, Maryland 20892, (301) 496-9838. These records shall be

clearly marked as being the property of the institution on loan to NIH

solely for the purpose of making a determination under the Freedom of

Information Act. NIH/ORDA will seek a determination from the

responsible official under DHHS regulations (45 Code of Federal

Regulations, Part 5) as to whether the records involved, (or some

portion) will be made available to members of the public under the

Freedom of Information Act. Pending such a determination, the records

will be kept separate from NIH/ORDA files, will be considered records

of the institution and not NIH/ORDA, and will not be received as part

of NIH/ORDA files. No copies will be made of such records.

NIH/ORDA will inform the institution of the DHHS Freedom of

Information Officer's determination and follow the institution's

instructions as to whether some or all of the records involved are to

be returned to the institution or to become a part of NIH/ORDA files.

If the institution instructs NIH/ORDA to return the records, no copies

or summaries of the records will be made or retained by DHHS, NIH, or

ORDA. The DHHS Freedom of Information Officer's determination will

represent that official's judgment at the time of the determination as

to whether the records involved (or some portion) would be exempt from

disclosure under the Freedom of Information Act if at the time of the

determination the records were in NIH/ORDA files and a request was

received for such files under the Freedom of Information Act.

Section V. Footnotes and References of Sections I Through IV

Section V-A. The original reference to organisms as Class 1, 2, 3,

4, or 5 refers to the classification in the publication Classification

of Etiologic Agents on the Basis of Hazard, 4th Edition, July 1974,

U.S. Department of Health, Education, and Welfare, Public Health

Services, Centers for Disease Control and Prevention, Office of

Biosafety, Atlanta, Georgia 30333. The NIH Director, with advice of the

RAC, may revise the classification for the purposes of the NIH

Guidelines (see Section IV-C-1-b-(2)-(e)). The revised list of

organisms in each class is reprinted in Appendix B.

Section V-B. Section III describes a number of places where

judgments are to be made. In all these cases, the Principal

Investigator shall make the judgment on these matters as part of his/

her responsibility to ``make the initial determination of the required

levels of physical and biological containment in accordance with the

NIH Guidelines'' (see Section IV-B-4-c-(1)). For cases falling under

Sections III-A through III-D, this judgment is to be reviewed and

approved by the Institutional Biosafety Committee as part of its

responsibility to make an ``independent assessment of the containment

levels required by the NIH Guidelines for the proposed research'' (see

Section IV-B-2-b-(1)). The Institutional Biosafety Committee may refer

specific cases to NIH/ORDA as part of NIH/ORDA's functions to ``provide

advice to all within and outside NIH'' (see Section IV-C-3). NIH/ORDA

may request advice from the RAC as part of the RAC's responsibility for

``interpreting the NIH Guidelines for experiments to which the NIH

Guidelines do not specifically assign containment levels'' (see Section

IV-C-1-b-(2)-(f)).

Section V-C. Laboratory Safety at the Centers for Disease Control,

September 1974, U.S. Department of Health, Education, and Welfare

Publication No. CDC 75-8118.

Section V-D. Classification of Etiologic Agents on the Basis of

Hazard, 4th Edition, July 1974, U.S. Department of Health, Education,

and Welfare, Public Health Service, Centers for Disease Control, Office

of Biosafety, Atlanta, Georgia 30333.

Section V-E. National Cancer Institute Safety Standards for

Research Involving Oncogenic Viruses, October 1974, U.S. Department of

Health, Education, and Welfare, Publication No. (NIH) 75-790.

Section V-F. National Institutes of Health Biohazards Safety Guide,

1974, U.S. Department of Health, Education, and Welfare, Public Health

Service, NIH, U.S. Government Printing Office, Stock No. 1740-00383.

Section V-G. A. Hellman, M. N. Oxman, and R. Pollack (eds.), 1973,

Biohazards in Biological Research, Cold Spring Harbor Laboratory, Cold

Spring Harbor, NY.

Section V-H. Furr, A. K., Handbook of Laboratory Safety, 2nd ed,

The Chemical Rubber Co., Boca Raton, Florida, 1990.

Section V-I. American Public Health Association, Bodily, J. L.,

General Administration of the Laboratory, 6th ed., ``Diagnostic

Procedures for Bacterial, Mycotic, and Parasitic Infections,'' New

York, 1981.

Section V-J. H. M. Darlow, Safety in the Microbiological

Laboratory, in J. R. Norris and D. W. Robbins (eds.), Methods in

Microbiology, Academic Press, Inc, New York, New York, 1969, pp. 169-

204.

Section V-K. C. M. Collins, E. G. Hartley, and R. Pilsworth, The

Prevention of Laboratory Acquired Infection, Public Health Laboratory

Service, Monograph Series No. 6, 1974.

Section V-L. Chatigny, M. A, ``Protection Against Infection in the

Microbiological Laboratory: Devices and Procedures,'' in W.W. Umbreit

(ed.), Advances in Applied Microbiology, Academic Press, New York, New

York, 1961, 3:131-192.

Section V-M. Design Criteria for Viral Oncology Research

Facilities, U.S. Department of Health, Education, and Welfare, Public

Health Service, NIH, DHEW Publication No. (NIH) 75-891, 1975.

Section V-N. Kuehne, R. W., Biological Containment Facility for

Studying Infectious Disease, Appl. Microbiol. 26:239-243, 1973.

Section V-O. Runkle, R. B., and G. B. Phillips, Microbial

Containment Control Facilities, Van Nostrand Reinhold, New York, 1969.

Section V-P. Chatigny, M. A., and D. I. Clinger, ``Contamination

Control in Aerobiology,'' in R. L. Dimmick and A. B. Akers (eds.), An

Introduction to Experimental Aerobiology, John Wiley & Sons, New York,

1969, pp. 194-263.

Section V-Q. As classified in the Third Report of the International

Committee on Taxonomy of Viruses: Classification and Nomenclature of

Viruses, R. E. F. Matthews (ed.), Intervirology 12 (129-296), 1979.

Section V-R. A U.S. Department of Agriculture permit is required

for the importation, interstate movement, and release into the

environment of certain organisms that are plant or animal pathogens,

whether genetically engineered or not. Permits are required for

veterinary biologics and for certain plants or microorganisms derived

through genetic engineering using genetic sequences from plant pests

(pathogens). Specific information about regulated organisms and

procedures for obtaining a permit for regulated organisms may be

obtained from the Director, Biotechnology, Biologics, and Environmental

Protection, Animal and Plant Health Inspection Service, U.S. Department

of Agriculture, 6505 Belcrest Road, Room 850, Hyattsville, Maryland

20782, (301) 436-7601.

Section V-S. i.e., the total of all genomes within a family shall

not exceed two-thirds of the genome.

Section V-T. All activities, including storage of variola and

whitepox, are restricted to the single national facility (World Health

Organization Collaborating Center for Smallpox Research, Centers for

Disease Control and Prevention, Atlanta, Georgia).

Section V-U. Human studies in which the induction or enhancement of

an immune response to a vector-encoded microbial immunogen is the major

goal, such an immune response has been demonstrated in model systems,

and the persistence of the vector-encoded immunogen is not expected,

are not covered under Sections III-A-2, III-B-2, or III-B-3. Such

studies may be initiated without RAC review and NIH approval if

approved by another Federal agency.

Section V-V. For recombinant DNA experiments in which the intent is

to modify stably the genome of cells of one or more human subjects (see

Sections III-A-2, III-B-2, and III-B-3).

Section V-W. In accordance with accepted scientific and regulatory

practices of the discipline of plant pathology, an exotic plant

pathogen (e.g., virus, bacteria, or fungus) is one that is unknown to

occur within the U.S. (see Section V-R). Determination of whether a

pathogen has a potential for serious detrimental impact on managed

(agricultural, forest, grassland) or natural ecosystems should be made

by the Principal Investigator and the Institutional Biosafety

Committee, in consultation with scientists knowledgeable of plant

diseases, crops, and ecosystems in the geographic area of the research.

Appendix A. Exemptions Under Section III-E-5--Sublists of Natural

Exchangers

Certain specified recombinant DNA molecules that ``consist entirely

of DNA segments from different species that exchange DNA by known

physiological processes, though one or more of the segments may be a

synthetic equivalent are exempt from these NIH Guidelines (see Section

III-E-5). Institutional Biosafety Committee registration is not

required for these exempt experiments. A list of such exchangers will

be prepared and periodically revised by the NIH Director with advice

from the RAC after appropriate notice and opportunity for public

comment (see Section IV-C-1-b-(1)-(c)). See Appendices A-I through A-VI

for a list of natural exchangers that are exempt from the NIH

Guidelines.'' Section III-E-5 describes recombinant DNA molecules that

are: (1) composed entirely of DNA segments from one or more of the

organisms within a sublist, and (2) to be propagated in any of the

organisms within a sublist (see Classification of Bergey's Manual of

Determinative Bacteriology; 8th edition, R. E. Buchanan and N. E.

Gibbons, editors, Williams and Wilkins Company; Baltimore, Maryland

1984). Although these experiments are exempt, it is recommended that

they be performed at the appropriate biosafety level for the host or

recombinant organism (see Biosafety in Microbiological and Biomedical

Laboratories, 3rd edition, May 1993, U.S. DHHS, Public Health Service,

Centers for Disease Control, Atlanta, Georgia, and NIH Office of

Biosafety, Bethesda, Maryland).

Appendix A-I. Sublist A

Genus Escherichia

Genus Shigella

Genus Salmonella--including Arizona

Genus Enterobacter

Genus Citrobacter--including Levinea

Genus Klebsiella--including oxytoca

Genus Erwinia

Pseudomonas aeruginosa, Pseudomonas putida, Pseudomonas fluorescens,

and Pseudomonas mendocina

Serratia marcescens

Yersinia enterocolitica

Appendix A-II. Sublist B

Bacillus subtilis

Bacillus licheniformis

Bacillus pumilus

Bacillus globigii

Bacillus niger

Bacillus nato

Bacillus amyloliquefaciens

Bacillus aterrimus

Appendix A-III. Sublist C

Streptomyces aureofaciens

Streptomyces rimosus

Streptomyces coelicolor

Appendix A-IV. Sublist D

Streptomyces griseus

Streptomyces cyaneus

Streptomyces venezuelae

Appendix A-V. Sublist E

One way transfer of Streptococcus mutans or Streptococcus lactis DNA

into Streptococcus sanguis

Appendix A-VI. Sublist F

Streptococcus sanguis

Streptococcus pneumoniae

Streptococcus faecalis

Streptococcus pyogenes

Streptococcus mutans

Appendix B. Classification of Etiologic Agents and Oncogenic Viruses on

the Basis of Hazard (See Appendix B-VI-A).

Appendix B-I. Class 1 Agents

All bacterial, parasitic, fungal, viral, rickettsial, and

chlamydial agents not included in higher classes shall be considered

Class 1 agents.

Appendix B-II. Class 2 Agents

Appendix B-II-A. Class 2 Bacterial Agents

Acinetobacter calcoaceticus

Actinobacillus--all species

Aeromonas hydrophila

Amycolata autotrophica

Arizona hinshawii--all serotypes

Bacillus anthracis

Bordetella--all species

Borrelia recurrentis, B. vincenti

Campylobacter fetus

Campylobacter jejuni

Chlamydia psittaci

Chlamydia trachomatis

Clostridium botulinum, Cl. chauvoei, Cl. haemolyticum, Cl.

histolyticum, Cl. novyi, Cl.

septicum, Cl. tetani

Corynebacterium diphtheriae, C. equi, C. haemolyticum, C.

pseudotuberculosis, C.

pyogenes, C. renale

Dermatophilus congolensis

Edwardsiella tarda

Erysipelothrix insidiosa

Escherichia coli--all enteropathogenic, enterotoxigenic, enteroinvasive

and strains

bearing K1 antigen

Haemophilus ducreyi, H. influenzae

Klebsiella--all species except oxytoca

Legionella pneumophila

Leptospira interrogans--all serotypes

Listeria--all species

Moraxella--all species

Mycobacteria--all species except those listed in Class 3

Mycobacterium avium

Mycoplasma--all species except Mycoplasma mycoides and Mycoplasma

agalactiae, which are in Class 5

Neisseria gonorrhoea, N. meningitides

Nocardia asteroides, N. brasiliensis, N. otitidiscaviarum, N.

transvalensis

Pasteurella--all species except those listed in Class 3

Rhodococcus equi

Salmonella--all species and all serotypes

Shigella-all species and all serotypes

Sphaerophorus necrophorus

Staphylococcus aureus

Streptobacillus moniliformis

Streptococcus pneumoniae, S. pyogenes

Treponema carateum, T. pallidum, and T. pertenue

Vibrio cholerae, V. parahemolyticus

Yersinia enterocolitica

Appendix B-II-B. Class 2 Fungal Agents

Blastomyces dermatitidis

Cryptococcus neoformans

Paracoccidioides braziliensis

Appendix B-II-C. Class 2 Parasitic Agents

Endamoeba histolytica

Leishmania sp.

Naegleria gruberi

Schistosoma mansoni

Toxocara canis

Toxoplasma gondii

Trichinella spiralis

Trypanosoma cruzi

Appendix B-II-D. Class 2 Viral, Rickettsial, and Chlamydial Agents

Adenoviruses--human--all types

Cache Valley virus

Coronaviruses

Coxsackie A and B viruses

Cytomegaloviruses

Echoviruses--all types

Encephalomyocarditis virus (EMC)

Flanders virus

Hart Park virus

Hepatitis viruses--associated antigen material

Herpesviruses--except Herpesvirus simiae (Monkey B virus) which is in

Class 4

Influenza viruses--all types except A/PR8/34, which is in Class 1

Langat virus

Lymphogranuloma venereum agent

Measles virus

Mumps virus

Parainfluenza virus--all types except Parainfluenza virus 3, SF4

strain, which is in Class 1

Polioviruses --all types, wild and attenuated

Poxviruses--all types except Alastrim, Smallpox, and Whitepox which are

Class 5 and Monkey pox which depending on experiments is in Class 3 or

Class 4

Rabies virus--all strains except Rabies street virus which should be

classified in Class 3

Reoviruses--all types

Respiratory syncytial virus

Rhinoviruses--all types

Rubella virus

Simian viruses--all types except Herpesvirus simiae (Monkey B virus)

and Marburg virus which are in Class 4

Sindbis virus

Tensaw virus

Turlock virus

Vaccinia virus

Varicella virus

Vesicular stomatitis virus (see Appendix B-VI-B)

Vole rickettsia

Yellow fever virus, 17D vaccine strain

Appendix B-II-E. Class 2 Oncogenic Viruses (See Appendix B-VI-C)

Appendix B-II-E-1. Low-Risk Oncogenic Viruses

Adenovirus 7-Simian virus 40 (Ad7-SV40)

Adenovirus

Avian leukosis virus

Bovine leukemia virus

Bovine papilloma virus

Chick-embryo-lethal orphan (CELO) virus or fowl adenovirus 1

Dog sarcoma virus

Guinea pig herpes virus

Lucke (Frog) virus

Hamster leukemia virus

Marek's disease virus

Mason-Pfizer monkey virus

Mouse mammary tumor virus

Murine leukemia virus

Murine sarcoma virus

Polyoma virus

Rat leukemia virus

Rous sarcoma virus

Shope fibroma virus

Shope papilloma virus

Simian virus 40 (SV-40)

Appendix B-II-E-2. Moderate-Risk Oncogenic Viruses

Adenovirus 2-Simian virus 40 (Ad2-SV40)

Epstein-Barr virus (EBV)

Feline leukemia virus (FeLV)

Feline sarcoma virus (FeSV)

Gibbon leukemia virus (GaLV)

Herpesvirus (HV) ateles

Herpesvirus (HV) saimiri

Simian sarcoma virus (SSV)-1

Yaba

Appendix B-III. Class 3 Agents

Appendix B-III-A. Class 3 Bacterial Agents

Bartonella--all species

Brucella--all species

Francisella tularensis

Mycobacterium bovis, M. tuberculosis

Pasteurella multocide type--``buffalo'' and other foreign virulent

strains (see Appendix B-VI-B)

Pseudomonas mallei (see Appendix B-VI-B)

Pseudomonas pseudomallei (see Appendix B-VI-B)

Yersinia pestis

Appendix B-III-B. Class 3 Fungal Agents

Coccidioides immitis

Histoplasma capsulatum

Histoplasma capsulatum var. duboisii

Appendix B-III-C. Class 3 Parasitic Agents

None

Appendix B-III-D. Class 3 Viral, Rickettsial, and Chlamydial Agents

Monkey pox virus--when used in vitro (see Appendix B-VI-D)

Arboviruses--all strains except those in Class 2 and 4. (Arboviruses

indigenous to the United States are in Class 3 except those listed in

Class 2. West Nile and Semliki Forest viruses may be classified up or

down depending on the conditions of use and geographical location of

the laboratory).

Dengue virus--when used for transmission or animal inoculation

experiments

Lymphocytic choriomeningitis virus (LCM)

Rickettsia--all species except Vole rickettsia when used for

transmission or animal inoculation experiments

Yellow fever virus--wild, when used in vitro

Appendix B-IV. Class 4 Agents

Appendix B-IV-A. Class 4 Bacterial Agents

None

Appendix B-IV-B. Class 4 Fungal Agents

None

Appendix B-IV-C. Class 4 Parasitic Agents

None

Appendix B-IV-D. Class 4 Viral, Rickettsial, and Chlamydial Agents

Ebola fever virus

Monkey pox virus--when used for transmission or animal inoculation

experiments (see Appendix B-VI-D)

Hemorrhagic fever agents--including Crimean hemorrhagic fever, (Congo),

Junin, and Machupo viruses, and others as yet undefined

Herpesvirus simiae (Monkey B virus)

Lassa virus

Marburg virus

Tick-borne encephalitis virus complex--including Russian spring-summer

encephalitis, Kyasanur forest disease, Omsk hemorrhagic fever, and

Central European encephalitis viruses

Venezuelan equine encephalitis virus, epidemic strains--when used for

transmission or animal inoculation experiments

Yellow fever virus-wild--when used for transmission or animal

inoculation experiments

Appendix B-V. Class 5 Agents (see Appendix B-VI-E)

Appendix B-V-A. Animal Disease Organisms which are Forbidden Entry into

the United States by Law

Foot and mouth disease virus

Appendix B-V-B. Animal Disease Organisms and Vectors which are

Forbidden Entry into the United States by U.S. Department of

Agriculture Policy

African horse sickness virus

African swine fever virus

Besnoitia besnoiti

Borna disease virus

Bovine infectious petechial fever

Camel pox virus

Ephemeral fever virus

Fowl plague virus

Goat pox virus

Hog cholera virus

Louping ill virus

Lumpy skin disease virus

Mycoplasma mycoides--contagious bovine pleuropneumonia

Mycoplasma agalactiae--contagious agalactia of sheep

Nairobi sheep disease virus

Newcastle disease virus--Asiatic strains

Rhinderpest virus

Rickettsia ruminatium--heart water

Rift valley fever virus

Sheep pox virus

Swine vesicular disease virus

Teschen disease virus

Theileria annulata

Theileria bovis

Theileria hirci

Theileria lawrencei

Theileria parva--East Coast fever

Trypanosoma evansi

Trypanosoma vivax--Nagana

Vesicular exanthema virus

Wesselsbron disease virus

Zyonema

Appendix B-V-C. Organisms which may not be Studied in the United States

Except at Specified Facilities

Alastrim (see Appendix B-VI-D)

Small pox (see Appendix B-VI-D)

White pox (see Appendix B-VI-D)

Appendix B-VI. Footnotes and References of Appendix B

Appendix B-VI-A. The original reference for this classification was

the publication Classification of Etiologic Agents on the Basis of

Hazard, 4th edition, July 1974, U.S. DHHS, Public Health Service,

Centers for Disease Control and Prevention, Office of Biosafety,

Atlanta, Georgia 30333. For the purposes of these NIH Guidelines, this

list has been revised by the NIH.

Appendix B-VI-B. A U.S. Department of Agriculture permit, required

for import and interstate transport of pathogens, may be obtained from

the U.S. Department of Agriculture, ATTN: Animal and Plant Health

Inspection Service, Import-Export Products Office, Room 756, Federal

Building, 6505 Belcrest Road, Hyattsville, Maryland 20782.

Appendix B-VI-C. National Cancer Institute Safety Standards for

Research Involving Oncogenic Viruses, U.S. Department of Health,

Education, and Welfare Publication No. (NIH) 75-790, October 1974.

Appendix B-VI-D. All activities, including storage of variola and

whitepox, are restricted to the single national facility (World Health

Organization Collaborating Center for Smallpox Research, Centers for

Disease Control and Prevention, Atlanta, Georgia).

Appendix B-VI-E. U.S. Department of Agriculture, Animal and Plant

Health Inspection Service.

Appendix C. Exemptions Under Section III-E-6

Section III-E-6 states that exempt from these NIH Guidelines are

``those that do not present a significant risk to health or the

environment (see Section IV-C-1-b-(1)-(c)), as determined by the NIH

Director, with the advice of the RAC, and following appropriate notice

and opportunity for public comment. See Appendix C for other classes of

experiments which are exempt from the NIH Guidelines.'' The following

classes of experiments are exempt under Section III-E-6:

Appendix C-I. Recombinant DNA in Tissue Culture

Recombinant DNA molecules containing less than one-half of any

eukaryotic viral genome (all viruses from a single family (see Appendix

C-VI-D) being considered identical (see Appendix C-VI-E), that are

propagated and maintained in cells in tissue culture are exempt from

these NIH Guidelines with the exceptions listed in Appendix C-I-A.

Appendix C-I-A. Exceptions

The following categories are not exempt from the NIH Guidelines:

(i) experiments described in Section III-A which require specific RAC

review and NIH and Institutional Biosafety Committee approval before

initiation, (ii) experiments described in Section III-B which require

NIH/ORDA and Institutional Biosafety Committee approval before

initiation, (iii) experiments involving DNA from Class 3, 4, or 5

organisms (see Appendix C-VI-A) or cells known to be infected with

these agents, (iv) experiments involving the deliberate introduction of

genes coding for the biosynthesis of molecules that are toxic for

vertebrates (see Appendix F), and (v) whole plants regenerated from

plant cells and tissue cultures are covered by the exemption provided

they remain axenic cultures even though they differentiate into

embryonic tissue and regenerate into plantlets.

Appendix C-II. Escherichia coli K-12 Host-Vector Systems

Experiments which use Escherichia coli K-12 host-vector systems,

with the exception of those experiments listed in Appendix C-II-A, are

exempt from the NIH Guidelines provided that: (i) the Escherichia coli

host does not contain conjugation proficient plasmids or generalized

transducing phages; or (ii) lambda or lambdoid or Ff bacteriophages or

non-conjugative plasmids (see Appendix C-VI-B) shall be used as

vectors. However, experiments involving the insertion into Escherichia

coli K-12 of DNA from prokaryotes that exchange genetic information

(see Appendix C-VI-C) with Escherichia coli may be performed with any

Escherichia coli K-12 vector (e.g., conjugative plasmid). When a non-

conjugative vector is used, the Escherichia coli K-12 host may contain

conjugation-proficient plasmids either autonomous or integrated, or

generalized transducing phages. For these exempt laboratory

experiments, Biosafety Level (BL) 1 physical containment conditions are

recommended. For large scale fermentation experiments, the appropriate

physical containment conditions need be no greater than those for the

host organism unmodified by recombinant DNA techniques; the

Institutional Biosafety Committee can specify higher containment if

deemed necessary.

Appendix C-II-A. Exceptions

The following categories of experiments are not exempt from the NIH

Guidelines: (i) experiments described in Section III-A which require

Institutional Biosafety Committee approval, RAC review, and NIH

approval before initiation, (ii) experiments described in Section III-B

which require Institutional Biosafety Committee and NIH/ORDA approval

before initiation, (iii) experiments involving DNA from Class 3, 4, or

5 organisms (see Appendix C-VI-A) or cells known to be infected with

these agents may be conducted under containment conditions specified in

Section III-C-2 with prior Institutional Biosafety Committee review and

approval, (iv) large scale experiments (e.g., more than 10 liters of

culture), and (v) experiments involving the cloning of toxin molecule

genes coding for the biosynthesis of molecules toxic for vertebrates

(see Appendix F).

Appendix C-III. Saccharomyces Host-Vector Systems

Experiments involving Saccharomyces cerevisiae and Saccharomyces

uvarum host-vector systems, with the exception of experiments listed in

Appendix C-III-A, are exempt from the NIH Guidelines. For these exempt

experiments, BL1 physical containment is recommended. For large scale

fermentation experiments, the appropriate physical containment

conditions need be no greater than those for the host organism

unmodified by recombinant DNA techniques; the Institutional Biosafety

Committee can specify higher containment if deemed necessary.

Appendix C-III-A. Exceptions

The following categories are not exempt from the NIH Guidelines:

(i) Experiments described in Section III-A which require Institutional

Biosafety Committee approval, RAC review, and NIH approval before

initiation, (ii) experiments described in Section III-B which require

Institutional Biosafety Committee and NIH/ORDA approval before

initiation, (iii) experiments involving DNA from Class 3, 4, or 5

organisms (see Appendix C-VI-A) or cells known to be infected with

these agents may be conducted under containment conditions specified in

Section III-C-2 with prior Institutional Biosafety Committee review and

approval, (iv) large scale experiments (e.g., more than 10 liters of

culture), and (v) experiments involving the deliberate cloning of genes

coding for the biosynthesis of molecules toxic for vertebrates (see

Appendix F).

Appendix C-IV. Bacillus subtilis or Bacillus licheniformis Host-Vector

Systems

Any asporogenic Bacillus subtilis or asporogenic Bacillus

licheniformis strain which does not revert to a spore-former with a

frequency greater than 10-7 may be used for cloning DNA with the

exception of those experiments listed in Appendix C-IV-A. For these

exempt laboratory experiments, BL1 physical containment conditions are

recommended. For large scale fermentation experiments, the appropriate

physical containment conditions need be no greater than those for the

host organism unmodified by recombinant DNA techniques; the

Institutional Biosafety Committee can specify higher containment if it

deems necessary.

Appendix C-IV-A. Exceptions