Recombinant DNA Research: Actions Under the Guidelines

Federal RegisterJan 19, 1996

Ask Donna

What actually matters in this document.

Text

SUMMARY: This notice sets forth an action to be taken by the Director,

National Institutes of Health (NIH), under the NIH Guidelines for

Research Involving Recombinant DNA Molecules.

FOR FURTHER INFORMATION CONTACT:

Additional information can be obtained from Dr. Nelson A. Wivel,

Director, Office of Recombinant DNA Activities (ORDA), Office of

Science Policy and Technology Transfer, National Institutes of Health,

Suite 302, 6000 Executive Boulevard, MSC 7010, Bethesda, Maryland

20892-7010, (301) 496-9838.

SUPPLEMENTARY INFORMATION: Today's action is being promulgated under

the NIH Guidelines for Research Involving Recombinant DNA Molecules.

This proposed action was published for comment in the Federal Register

of August 18, 1994 (58 FR 44098), November 8, 1994 (59 FR 55796),

February 8, 1995 (60 FR 7630), and May 22, 1995 (60 FR 27207), and

reviewed and recommended for approval by the NIH Recombinant DNA

Advisory Committee (RAC) at its meeting on June 8-9, 1995.

I. Background Information and Decisions on Actions Under the NIH

Guidelines

A. Amendments to Sections II, III, IV, V, Appendices B, C, H, and Q of

the NIH Guidelines Regarding Updating the Classification of

Microorganisms

In a letter dated June 24, 1993, Dr. Diane Fleming, President of

the Mid-Atlantic Biological Safety Association requested the revision

and updating of Appendix B, Classification of Microorganisms on the

Basis of Hazard. The Mid-Atlantic Biological Safety Association

submitted an updated list of the classification of microorganisms for

the Recombinant DNA Advisory Committee to review which included the

latest taxonomy and agent risk group classifications as defined by the

Centers for Disease Control and Prevention.

During the September 9-10, 1993, meeting, the Recombinant DNA

Advisory Committee recommended by consensus that the current

classification of etiological agents described in the Biosafety in

Microbiological and Biomedical Laboratories, 3rd edition, May 1993,

U.S. Department of Health and Human Services, should be endorsed by the

Committee. The Committee retained the option to adopt any modifications

to the Centers for Disease Control and Prevention listing. The

Committee recommended that the revised Appendix B, Classification of

Microorganisms on the Basis of Hazard, submitted by Dr. Fleming should

not be adopted until the Committee received letters of concurrence from

both the Centers for Disease Control and Prevention and the NIH

Division of Safety.

In a telephone call on October 20, 1994, Dr. Fleming stated that

Appendix B, Classification of Microorganisms on the Basis of Hazard,

would be reviewed by experts from the Centers for Disease Control and

Prevention and the American Society for Microbiology. The revised

Appendix B was submitted to the Recombinant DNA Advisory Committee

December 1-2, 1994, meeting for review and discussion. During the

December 1994 meeting, the Committee recommended publishing the revised

Appendix B in the Federal Register for public comment, with further

review of this proposal and possible approval during the March 6-7,

1995, meeting.

During the March 6-7, 1995 meeting, the Recombinant DNA Advisory

Committee deferred approval of the proposed amendments to Appendix B

pending additional revisions to the remaining sections and appendices

of the NIH Guidelines that are required to adequately accommodate the

revised Appendix B (Sections II, III, IV, V, Appendices C, H, and Q).

The motion for deferral included a recommendation that a subcommittee

consisting of Dr. Stephen Straus (Chair of the Subcommittee), ad hoc

experts, and Office of Recombinant DNA Activities staff would meet to

develop the required modifications. The motion passed by a vote of 17

in favor, 0 opposed, and no abstentions.

On May 5, 1995, the Appendix B Subcommittee met to finalize the

document in terms of its listing of pathogens and the text of the NIH

Guidelines related to Appendix B in other sections and appendices

(Sections II, III, IV, V, Appendices C, H, and Q). During the June 8-9,

1995 meeting, the Recombinant DNA Advisory Committee reviewed the

document. There was a concurrence that the Risk Group classification

serves as an initial guidance to assign an appropriate containment

level for a particular experiment by the Institutional Biosafety

Committees and the investigators. Since the new Appendix B is primarily

concerned with human pathogenicity, it addresses only the human

etiologic agents and omits all animal agents. The Committee observed

that this omission created a problem because some of the animal agents,

particularly the group of viruses known as oncogenic viruses are

frequently used as vectors for gene transfer in the laboratories or in

human studies. The Recombinant DNA Advisory Committee approved a motion

to: (1) establish a working group to recommend exemption of additional

vector systems in Appendix C (exempt host-vector systems), and (2)

accept the proposed amendments to Appendix B with the provision to

develop a new Appendix B-V relating to animal viruses relevant to human

studies, and to list specific examples of agents under Appendix B-I,

Risk Group 1 (RG1) Agents. The motion was approved by a vote of 17 in

favor, 0 opposed, and no abstentions.

On June 13, 1995, the Office of Recombinant DNA Activities

forwarded two versions of the Appendix B-V, Animal Viral Etiologic

Agents in Common Use to the Appendix B Subcommittee. Most of these

agents were previously listed as Class 2 oncogenic viruses in two

separate categories of low and moderate risk agents in the original

Appendix B. Since none of these animal etiologic agents are associated

with disease in healthy human adults, one version of Appendix B-V

listed these agents as a single group recommended for Biosafety Level 1

containment and another version listed them in a two-tier system for

either Biosafety Level 1 or Biosafety Level 2 containment. Subsequent

discussion with the members of the Appendix B Subcommittee concluded

that while there was no reason to have a separate group of ``moderate''

risk agents in this list, it was prudent to recommend conducting

experiments under a Biosafety Level 2 containment with several agents

that are capable of infecting human cells, e.g., amphotropic and

xenotropic murine leukemia virus.

During the September 11-12, 1995, meeting, the Recombinant DNA

Advisory Committee reviewed the updated Appendix B along with other

sections and appendices of the NIH Guidelines (Sections II, III, IV, V,

Appendices C, H, and Q) relating to classification of microorganisms.

It was observed that some viruses in the moderate risk group could

infect human cells but their replication was largely restricted to

their animal hosts. Some Committee members pointed out that

[[Page 1483]]

some viruses with oncogenes such as SV40 have been treated more

cautiously than viruses without oncogenes; therefore, a two-tier list

should be used. Dr. Wivel explained that listing a group of animal

viruses as ``moderate risk'' agents introduces an inconsistency into

Appendix B. Some strains of these viruses, although capable of

infecting human cells, have not been shown to be associated with any

disease in healthy human adults. They fall within the definition of

Risk Group 1 agents, i.e., agents that are not associated with disease

in healthy adult humans. Two committee members inquired why several

viruses in the original Appendix B are not listed in the new version.

Dr. Thomas Shih (Executive Secretary, Appendix B Subcommittee)

explained that several rarely used viruses such as chick embryo lethal

orphan virus are deleted from the new list. The list includes commonly

used organisms, and it is not intended to be inclusive since many other

animal agents are not listed. Dr. Walters (Chair, Recombinant DNA

Advisory Committee) stated that the consensus of the committee is to

accept the list of animal viruses in Appendix B-V as a reasonable

modification of Appendix B.

The actions are detailed in Section II--Summary of Actions. I

accept these recommendations, and the NIH Guidelines will be amended

accordingly.

II. Summary of Actions

A. Amendments to Section II, Safety Considerations (Previously the

Entire Section II was Entitled Containment)

Section II is amended to read:

Section II. Safety Considerations

Section II-A. Risk Assessment

Section II-A-1. Risk Groups

Risk assessment requires the exercise of sound judgment by the

investigator. The investigator must make an initial risk assessment

based on the Risk Group (RG) of an agent (see Appendix B,

Classification of Human Etiologic Agents on the Basis of Hazard).

Agents are classified into four Risk Groups (RGs) according to their

relative pathogenicity for healthy adult humans by the following

criteria: (1) Risk Group 1 (RG1) agents are not associated with disease

in healthy adult humans. (2) Risk Group 2 (RG2) agents are associated

with human disease which is rarely serious and for which preventive or

therapeutic interventions are often available. (3) Risk Group 3 (RG3)

agents are associated with serious or lethal human disease for which

preventive or therapeutic interventions may be available. (4) Risk

Group 4 (RG4) agents are likely to cause serious or lethal human

disease for which preventive or therapeutic interventions are not

usually available.

Section II-A-2. Criteria for Risk Groups

Classification of agents is based on the potential effect of a

biological agent on a healthy human adult and does not account for

instances in which an individual may have increased susceptibility to

such agents, e.g., preexisting diseases, medications, compromised

immunity, pregnancy or breast feeding (which may increase exposure of

infants to some agents) (see Appendix B, Classification of Human

Etiologic Agents on the Basis of Hazard).

Personnel may need periodic medical surveillance to ascertain

fitness to perform certain activities; they may also need to be offered

prophylactic vaccines and boosters (see Section IV-B-1-f,

Responsibilities of the Institution, General Information).

Section II-A-3. Comprehensive Risk Assessment

In deciding on the appropriate containment for an experiment, the

initial risk assessment from Appendix B, Classification of Human

Etiologic Agents on the Basis of Hazard, should be followed by a

thorough consideration of the agent itself and how it is to be

manipulated. Factors to be considered in determining the level of

containment include agent factors such as: virulence, pathogenicity,

infectious dose, environmental stability, route of spread,

communicability, operations, quantity, availability of vaccine or

treatment, and gene product effects such as toxicity, physiological

activity, and allergenicity. Any strain that is known to be more

hazardous than the parent (wild-type) strain should be considered for

handling at a higher containment level. Certain attenuated strains or

strains that have been demonstrated to have irreversibly lost known

virulence factors may qualify for a reduction of the containment level

compared to the Risk Group assigned to the parent strain (see Section

V-B, Footnotes and References of Sections I through IV).

A final assessment of risk based on these considerations is then

used to set the appropriate containment conditions for the experiment

(see Section II-B, Containment). The containment level required may be

equivalent to the Risk Group classification of the agent or it may be

raised or lowered as a result of the above considerations. The

Institutional Biosafety Committee must approve the risk assessment and

the biosafety containment level for recombinant DNA experiments

described in Sections III-A, Experiments that Require Institutional

Biosafety Committee Approval, RAC Review, and NIH Director Approval

Before Initiation, III-B, Experiments that Require NIH/ORDA and

Institutional Biosafety Committee Approval Before Initiation, and III-

C, Experiments that Require Institutional Biosafety Committee Approval

Before Initiation.

Careful consideration should be given to the types of manipulation

planned for some higher Risk Group agents. For example, the RG2 dengue

viruses may be cultured under the Biosafety Level (BL) 2 containment

(see Section II-B); however, when such agents are used for animal

inoculation or transmission studies, a higher containment level is

recommended. Similarly, RG3 agents such as Venezuelan equine

encephalomyelitis and yellow fever viruses should be handled at a

higher containment level for animal inoculation and transmission

experiments.

Individuals working with human immunodeficiency virus (HIV),

hepatitis B virus (HBV) or other bloodborne pathogens should consult

Occupational Exposure to Bloodborne Pathogens; Final Rule (56 FR 64175-

64182). BL2 containment is recommended for activities involving all

blood-contaminated clinical specimens, body fluids, and tissues from

all humans, or from HIV- or HBV-infected or inoculated laboratory

animals. Activities such as the production of research-laboratory scale

quantities of HIV or other bloodborne pathogens, manipulating

concentrated virus preparations, or conducting procedures that may

produce droplets or aerosols, are performed in a BL2 facility using the

additional practices and containment equipment recommended for BL3.

Activities involving industrial scale volumes or preparations of

concentrated HIV are conducted in a BL3 facility, or BL3 Large Scale if

appropriate, using BL3 practices and containment equipment.

Exotic plant pathogens and animal pathogens of domestic livestock

and poultry are restricted and may require special laboratory design,

operation and containment features not addressed in Biosafety in

Microbiological and Biomedical Laboratories (see Section V-C, Footnotes

and References of Sections I through IV). For information regarding the

importation, possession, or use of these agents see Sections V-G and V-

H, Footnotes and References of Sections I through IV.

[[Page 1484]]

Section II-B. Containment

Effective biological safety programs * * *

[Rest of Section II remains unchanged.]

B. Amendments to Section III, Experiments Covered by the NIH Guidelines

Section III-C is amended to read:

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) an indication of what protein will be produced if an attempt is to

be made to obtain expression of a foreign gene; 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), Minor Actions).

Section III-C-1. Experiments Using Risk Group 2, Risk Group 3, Risk

Group 4, or Restricted Agents as Host-Vector Systems (see Section II-A,

Risk Assessment).

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

recombinant DNA into Risk Group 2 agents will usually be conducted at

Biosafety Level (BL) 2 containment. Experiments with such agents will

usually 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 Risk Group 3 agents will usually be conducted at

BL3 containment. Experiments with such agents will usually be conducted

with whole animals at BL3 or BL3-N containment.

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

recombinant DNA into Risk Group 4 agents shall be conducted at BL4

containment. Experiments with such agents will usually 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 restricted 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 plant or animal pathogens

(see Section V-G and V-L, Footnotes and References of Sections I

through IV). 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 Risk Group 2, Risk

Group 3, Risk Group 4, or Restricted Agents (see Section V-A, Footnotes

and References of Sections I through IV) is Cloned into Nonpathogenic

Prokaryotic or Lower Eukaryotic Host-Vector Systems.

Section III-C-2-a. Experiments in which DNA from Risk Group 2 or

Risk Group 3 agents (see Section II-A, Risk Assessment) is transferred

into nonpathogenic prokaryotes or lower eukaryotes may be performed

under BL2 containment. Experiments in which DNA from Risk Group 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, Exempt

Experiments). 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, Experiments Involving the

Cloning of Toxin Molecules With LD50 of Less than 100 Nanograms

Per Kilogram Body Weight) or shall be conducted under NIH specified

conditions as described in Appendix F, Containment Conditions for

Cloning of Genes Coding for the Biosynthesis of Molecules Toxic for

Vertebrates.

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

DNA from restricted agents is transferred into nonpathogenic

prokaryotes or lower eukaryotes shall be determined by NIH/ORDA

following a case-by-case review (see Section V-L, Footnotes and

References of Sections I through IV). A U.S. Department of Agriculture

permit is required for work with plant or animal pathogens (see Section

V-G, Footnotes and References of Sections I through IV).

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

RNA Viruses or Defective 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-J, Footnotes and

References of Sections I through IV) being considered identical (see

Section V-K, Footnotes and References of Sections I through IV), are

considered defective and may be used in the absence of helper virus

under the conditions specified in 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-C-3-a. Experiments involving the use of infectious or

defective Risk Group 2 viruses (see Section V-A, Footnotes and

References of Sections I through IV, and Appendix B-II, Risk Group 2

Agents) in the presence of helper virus may be conducted at BL2.

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

defective Risk Group 3 viruses (see Section V-A, Footnotes and

References of Sections I through IV, and Appendix B-III-D, Risk Group 3

(RG3)--Viruses and Prions) in the presence of helper virus may be

conducted at BL3.

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

defective Risk Group 4 viruses (see Section V-A, Footnotes and

References of Sections I through IV, and Appendix B-IV-D, Risk Group 4

(RG4)--Viral Agents) in the presence of helper virus may be conducted

at BL4.

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

defective restricted poxviruses (see Section V-A and V-L, Footnotes and

References of Sections I through IV) in the presence of helper virus

shall be determined on a case-by-case basis following NIH/ORDA review.

A U.S.

[[Page 1485]]

Department of Agriculture permit is required for work with plant or

animal pathogens (see Section V-G, Footnotes and References of Sections

I through IV).

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

defective viruses in the presence of helper virus which are not covered

in Sections III-E-3-a through III-C-3-d 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-like

(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 nonhuman 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, Footnotes and References of

Sections I through IV). 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, Experiments Involving Whole Animals. For experiments involving

recombinant DNA-modified Risk Groups, 2, 3, 4, or restricted organisms,

see Sections V-A, V-G, and V-L, Footnotes and References of Sections I

through IV. It is important that the investigator demonstrate that the

fraction of the viral genome being utilized does not lead to productive

infections. A U.S. Department of Agriculture permit is required for

work with plant or animal pathogens (see Section V-G, Footnotes and

References of Sections I through IV).

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, Experiments

Using Risk Group 2, Risk Group 3, Risk Group 4, or Restricted Agents as

Host-Vector Systems, or III-C-4-a, Experiments Involving Whole Animals,

the appropriate containment shall be determined by the Institutional

Biosafety Committee.

[The rest of the Section III-C remains unchanged.]

C. Amendments to Section IV, Roles and Responsibilities

Section IV-C-1-b-(2)-(e) is amended to read:

Section IV-C-1-b-(2)-(e). Setting containment under Sections III-C-

1-d, Experiments Using Risk Group 2, Risk Group 3, Risk Group 4, or

Restricted Agents as Host-Vector Systems, and III-C-2-b, Experiments in

which DNA from Risk Group 2, Risk Group 3, Risk Group 4, or Restricted

Agents is Cloned into Nonpathogenic Prokaryotic or Lower Eukaryotic

Host-Vector Systems;

[The rest of the Section IV-C-1-b-(2) remains unchanged.]

D. Amendments to Section V, Footnotes and References of Sections I

Through IV

Section V is amended to read:

Section V. Footnotes and References of Sections I through IV

Section V-A. 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), Minor Actions). The revised list of organisms in each

risk group is reprinted in Appendix B, Classification of Human

Etiologic Agents on the Basis of Hazard.

Section V-B. Section III, Experiments Covered by the NIH

Guidelines, 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), Principal Investigator). For cases falling under

Sections III-A through III-D, Experiments Covered by the NIH

Guidelines, 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),

Institutional Biosafety Committee). 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, Office of Recombinant DNA Activities). 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), Minor Actions).

Section V-C. U.S. Department of Health and Human Services, Public

Health Service, Centers for Disease Control and Prevention and the

National Institutes of Health. Biosafety in Microbiological and

Biomedical Laboratories, 3rd edition, 1993. Copies are available from:

Superintendent of Documents, U.S. Government Printing Office,

Washington, DC 20402 (stock # 017-040-00523-7), Phone (202)-512-2356.

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. Benenson, Abram S. ed., Control of Communicable

Diseases in Man, 15th edition. 1990. American Public Health

Association, Washington, DC.

Section V-F. World Health Organization Laboratory Biosafety Manual,

2nd edition. 1993. WHO Albany, NY. Copies are available from: WHO

Publication Centre, USA, (Q Corp) 49 Sheridan Avenue, Albany, New York

12210; Phone: (518)-436-9686 (Order # 1152213).

Section V-G. A U.S. Department of Agriculture permit, required for

import and interstate transport of plant and animal pathogens, may be

obtained from the U.S. Department of Agriculture, ATTN: Animal and

Plant Health Inspection Service (APHIS), Veterinary Services, National

Center for Import-Export, Products Program, 4700 River Road, Unit 40,

Riverdale, MD 20737. Phone: (301)-734-8499; Fax: (301)-734-8226.

[[Page 1486]]

Section V-H. American Type Culture Collection Catalogues of plant

viruses, animal viruses, cells, bacteria, fungi, etc. are available

from American Type Culture Collection, 12301 Parklawn Drive, Rockville,

Maryland 20852-1776. Phone: (800)-638-6597; Fax: (301)-231-5826.

Section V-I. U.S. Department of Labor, Occupational Safety and

Health Administration. 1991. Occupational Exposure to Bloodborne

Pathogens, Final Rule (56 FR 64175-64182).

Section V-J. As classified in the 6th Report on the International

Committee on Taxonomy of Viruses: Classification and Nomenclature of

Viruses, F.A. Murphy et al., Archives of Virology/Supplement 10, 1995,

Springer-Verlag, New York, New York.

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

not exceed two-thirds of the genome.

Section V-L. Organisms including alastrim, smallpox (variola) and

whitepox may not be studied in the United States except at specified

facilities. 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-M. 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-G, Footnotes and References of

Sections I through IV). 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.

E. Amendments to Appendix B, Classification of Human Etiologic Agents

on the Basis of Hazard

Appendix B is amended to read:

Appendix B. Classification of Human Etiologic Agents on the Basis of

Hazard

Appendix B includes those biological agents known to infect humans,

as well as selected animal agents, that may pose theoretical risks if

inoculated into humans. Included in the lists are species known to be

pathogenic, mutated, or recombined; non-pathogenic species and strains

are not considered. Non-infectious life cycle stages of parasites are

excluded.

This appendix reflects the current state of knowledge and should be

considered a resource document. The more commonly encountered agents

are included; however, this appendix is not meant to be all inclusive.

Information on agent risk assessment may be found in the Agent Summary

Statements of the Centers for Disease Control and Prevention/National

Institutes of Health publications, Biosafety in Microbiological and

Biomedical Laboratories (see Sections V-C, V-D, V-E, and V-F, Footnotes

and References of Sections I through IV). Further guidance on agents

not listed in Appendix B may be obtained through: Centers for Disease

Control and Prevention, Biosafety Branch, Atlanta, Georgia 30333,

Phone: (404)-639-3883, Fax: (404)-639-2294; National Institutes of

Health, Division of Safety, Bethesda, Maryland 20892, Phone: (301)-496-

1357; National Animal Disease Center, U.S. Department of Agriculture,

Ames, Iowa 50010, Phone: (515)-862-8258.

A special committee of the American Society for Microbiology will

conduct an annual review of this appendix and its recommendation for

changes will be presented to the Recombinant DNA Advisory Committee as

proposed amendments to the NIH Guidelines.

Appendix B--Table 1.--Basis for the Classification of Biohazardous

Agents by Risk Group (RG)

Risk Group 1 (RG1)..................... Agents that are not associated

with disease in healthy adult

humans.

Risk Group 2 (RG2)..................... Agents that are associated with

human disease which is rarely

serious and for which

preventive or therapeutic

interventions are often

available.

Risk Group 3 (RG3)..................... Agents that are associated with

serious or lethal disease for

which preventive or

therapeutic interventions may

be available (high individual

risk but low community risk).

Risk Group 4 (RG4)..................... Agents that are likely to cause

serious or lethal human

disease for which preventive

or therapeutic interventions

are not usually available

(high individual risk and high

community risk).

Appendix B-I. Risk Group 1 (RG1) Agents

RG1 agents are not associated with disease in healthy adult humans.

Examples of RG1 agents include asporogenic Bacillus subtilis or

Bacillus licheniformis (see Appendix C-IV-A, Bacillus subtilis or

Bacillus licheniformis Host-Vector Systems, Exceptions), Eschenrichia

coli-K12 (see Appendix C-II-A, Escherichia coli K-12 Host-Vector

Systems, Exceptions), and adeno-associated virus types 1-4.

Those agents not listed in Risk Groups (RGs) 2, 3 and 4 are not

automatically or implicitly classified in RG1; a risk assessment must

be conducted based on the known and potential properties of the agents

and their relationship to agents that are listed.

Appendix B-II. Risk Group 2 (RG2) Agents

RG2 agents are associated with human disease which is rarely

serious and for which preventive or therapeutic interventions are often

available.

Appendix B-II-A. Risk Group 2 (RG2)--Bacterial Agents Including

Chlamydia

--Acinetobacter baumannii (formerly Acinetobacter calcoaceticus)

--Actinobacillus

--Actinomyces pyogenes (formerly Corynebacterium pyogenes)

--Aeromonas hydrophila

--Amycolata autotrophica

--Archanobacterium haemolyticum (formerly Corynebacterium haemolyticum)

--Arizona hinshawii--all serotypes

--Bacillus anthracis

--Bartonella henselae, B. quintana, B. vinsonii

--Bordetella including B. pertussis

--Borrelia recurrentis, B. burgdorferi

--Burkholderia (formerly Pseudomonas species) except those listed in

Appendix B-III-A (RG3))

--Campylobacter coli, C. fetus, C. jejuni

--Chlamydia psittaci, C. trachomatis, C. pneumoniae

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

histolyticum, Cl. novyi, Cl. septicum, Cl. tetani

--Corynebacterium diphtheriae, C. pseudotuberculosis, C. renale

--Dermatophilus congolensis

--Edwardsiella tarda

--Erysipelothrix rhusiopathiae

--Escherichia coli--all enteropathogenic, enterotoxigenic,

enteroinvasive and strains bearing K1 antigen, including E. coli

O157:H7

--Haemophilus ducreyi, H. influenzae

--Helicobacter pylori

--Klebsiella--all species except K. oxytoca (RG1)

--Legionella including L. pneumophila

--Leptospira interrogans--all serotypes

--Listeria

--Moraxella

--Mycobacterium (except those listed in Appendix B-III-A (RG3))

including M. avium complex, M. asiaticum, M. bovis BCG vaccine strain,

M. chelonei,

[[Page 1487]]

M. fortuitum, M. kansasii, M. leprae, M. malmoense, M. marinum, M.

paratuberculosis, M. scrofulaceum, M. simiae, M. szulgai, M. ulcerans,

M. xenopi

--Mycoplasma, except M. mycoides and M. agalactiae which are restricted

animal pathogens

--Neisseria gonorrhoea, N. meningitidis

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

transvalensis

--Rhodococcus equi

--Salmonella including S. arizonae, S. cholerasuis, S. enteritidis, S.

gallinarum-pullorum, S. meleagridis, S. paratyphi, A, B, C, S. typhi,

S. typhimurium

--Shigella including S. boydii, S. dysenteriae, type 1, S. flexneri, S.

sonnei

--Sphaerophorus necrophorus

--Staphylococcus aureus

--Streptobacillus moniliformis

--Streptococcus including S. pneumoniae, S. pyogenes

--Treponema pallidum, T. carateum

--Vibrio cholerae, V. parahemolyticus, V. vulnificus

--Yersinia enterocolitica

Appendix B-II-B. Risk Group 2 (RG2)--Fungal Agents

--Blastomyces dematitidis

--Cladosporium bantianum, C. (xylohypha) trichoides

--Cryptococcus neofomans

--Dactylaria galopava (Ochroconis gallopavum)

--Epidermophyton

--Exophiala (Wangiella) dermatitidis

--Fonsecaea pedrosoi

--Microsporum

--Paracoccidioides braziliensis

--Penicillium marneffei

--Sporothrix schenckii

--Trichophyton

Appendix B-II-C. Risk Group 2 (RG2)--Parasitic Agents

--Ancylostoma human hookworms including A. duodenale, A. ceylanicum

--Ascaris including Ascaris lumbricoides suum

--Babesia including B. divergens, B. microti

--Brugia filaria worms including B. malayi, B. timori

--Coccidia

--Cryptosporidium including C. parvum

--Cysticercus cellulosae (hydatid cyst, larva of T. solium)

--Echinococcus including E. granulosis, E. multilocularis, E. vogeli

--Entamoeba histolytica

--Enterobius

--Fasciola including F. gigantica, F. hepatica

--Giardia including G. lamblia

--Heterophyes

--Hymenolepis including H. diminuta, H. nana

--Isospora

--Leishmania including L. braziliensis, L. donovani, L. ethiopia, L.

major, L. mexicana, L. peruvania, L. tropica

--Loa loa filaria worms

--Microsporidium

--Naegleria fowleri

--Necator human hookworms including N. americanus

--Onchoerca filaria worms including, O. volvulus

--Plasmodium including simian species, P. cynomologi, P. falciparum, P.

malariae, P. ovale, P. vivax

--Sarcocystis including S. sui hominis

--Schistosoma including S. haematobium, S. intercalatum, S. japonicum,

S. mansoni, S. mekongi

--Strongyloides including S. stercoralis

--Taenia solium

--Toxocara including T. canis

--Toxoplasma including T. gondii

--Trichinella spiralis

--Trypanosoma including T. brucei brucei, T. brucie gambiense, T.

brucei rhodesiense, T. cruzi

--Wuchereria bancrofti filaria worms

Appendix B-II-D. Risk Group 2 (RG2)--Viruses

Adenoviruses, Human--All Types

Alphaviruses (Togaviruses)--Group A Arboviruses

--Eastern equine encephalomyelitis virus

--Venezuelan equine encephalomyelitis vaccine strain TC-83

--Western equine encephalomyelitis virus

Arenaviruses

--Lymphocytic choriomeningitis virus (non-neurotropic strains)

--Tacaribe virus complex

--Other viruses as listed in the reference source (see Section V-C,

Footnotes and References of Sections I through IV)

Bunyaviruses

--Bunyamwera virus

--Rift Valley fever virus vaccine strain MP-12

--Other viruses as listed in the reference source (see Section V-C,

Footnotes and References of Sections I through IV)

Calciviruses

Coronaviruses

Flaviviruses (Togaviruses)--Group B Arboviruses

--Dengue virus serotypes 1, 2, 3, and 4

--Yellow fever virus vaccine strain 17D

--Other viruses as listed in the reference source (see Section V-C,

Footnotes and References of Sections I through IV)

Hepatitis A, B, C, D, and E Viruses

--Herpesviruses--except Herpesvirus simiae (Monkey B virus) (see

Appendix B-IV-D, Risk Group 4 (RG4)--Viral Agents)

--Cytomegalovirus

--Epstein Barr virus

--Herpes simplex types 1 and 2

--Herpes zoster

--Human herpesvirus types 6 and 7

Orthomyxoviruses

--Influenza viruses types A, B, and C

--Other tick-borne orthomyxoviruses as listed in the reference source

(see Section V-C, Footnotes and References of Sections I through IV)

Papovaviruses

--All human papilloma viruses

Paramyxoviruses

--Newcastle disease virus

--Measles virus

--Mumps virus

--Parainfluenza viruses types 1, 2, 3, and 4

--Respiratory syncytial virus

Parvoviruses

--Human parvovirus (B19)

Picornaviruses

--Coxsackie viruses types A and B

--Echoviruses--all types

--Polioviruses--all types, wild and attenuated

--Rhinoviruses--all types

--Poxviruses--all types except Monkeypox virus (see Appendix B-III-D,

Risk Group 3 (RG3)--Viruses and Prions) and restricted poxviruses

including Alastrim, Smallpox, and White-pox (see Section V-L, Footnotes

and References of Sections I through IV)

--Reoviruses--all types including Coltivirus, human Rotavirus, and

Orbivirus (Colorado tick fever virus)

Rhabdoviruses

--Rabies virus--all strains

--Vesicular stomatitis virus--laboratory adapted strains including VSV-

Indiana, San Juan, and Glasgow

Togaviruses (see Alphaviruses and Flaviviruses)

--Rubivirus (rubella)

Appendix B-III. Risk Group 3 (RG3) Agents

RG3 agents are associated with serious or lethal human disease for

which preventive or therapeutic interventions may be available.

[[Page 1488]]

Appendix B-III-A. Risk Group 3 (RG3)--Bacterial Agents Including

Rickettsia

--Bartonella

--Brucella including B. abortus, B. canis, B. suis

--Burkholderia (Pseudomonas) mallei, B. pseudomallei

--Coxiella burnetii

--Francisella tularensis

--Mycobacterium bovis (except BCG strain, see Appendix B-II-A, Risk

Group 2 (RG2)--Bacterial Agents Including Chlamydia); M. tuberculosis

--Pasteurella multocida type B--``buffalo'' and other virulent strains

--Rickettsia akari, R. australis, R. canada, R. conorii, R. prowazekii,

R. rickettsii, R. siberica, R. tsutsugamushi, R. typhi (R. mooseri)

--Yersinia pestis

Appendix B-III-B. Risk Group 3 (RG3)--Fungal Agents

--Coccidioides immitis (sporulating cultures; contaminated soil)

--Histoplasma capsulatum, H. capsulatum var. duboisii

Appendix B-III-C. Risk Group 3 (RG3)--Parasitic Agents

None

Appendix B-III-D. Risk Group 3 (RG3)--Viruses and Prions

Alphaviruses (Togaviruses)--Group A Arboviruses

--Semliki Forest virus

--St. Louis encephalitis virus

--Venezuelan equine encephalomyelitis virus (except the vaccine strain

TC-83, see Appendix B-II-D, Risk Group 2 (RG2)--Viruses)

--Other viruses as listed in the reference source (see Section V-C,

Footnotes and References of Sections I through IV)

Arenaviruses

--Lymphocytic choriomeningitis virus (LCM) (neurotropic strains)

Bunyaviruses

--Hantaviruses including Hantaan virus

--Rift Valley fever virus

Flaviviruses (Togaviruses)--Group B Arboviruses

--Japanese encephalitis virus

--Yellow fever virus

--Other viruses as listed in the reference source (see Section V-C,

Footnotes and References of Sections I through IV)

Poxviruses

--Monkeypox virus

Prions

--Transmissible spongioform encephalopathies (TME) agents (Creutzfeldt-

Jakob disease and kuru agents) (for containment instruction, see

Section V-C, Footnotes and References of Sections I through IV)

Retroviruses

--Human immunodeficiency virus (HIV) types 1 and 2

--Human T cell lymphotropic virus (HTLV) types 1 and 2

--Simian immunodeficiency virus (SIV)

Rhabdoviruses

--Vesicular stomatitis virus

Appendix B-IV. Risk Group 4 (RG4) Agents

RG4 agents are likely to cause serious or lethal human disease for

which preventive or therapeutic interventions are not usually

available.

Appendix B-IV-A. Risk Group 4 (RG4)--Bacterial Agents

None

Appendix B-IV-B. Risk Group 4 (RG4)--Fungal Agents

None

Appendix B-IV-C. Risk Group 4 (RG4)--Parasitic Agents

None

Appendix B-IV-D. Risk Group 4 (RG4)--Viral Agents

Arenaviruses (Togaviruses)--Group A Arboviruses

--Guanarito virus

--Lassa virus

--Junin virus

--Machupo virus

Bunyaviruses (Nairovirus)

--Crimean-Congo hemorrhagic fever virus

Filoviruses

--Ebola virus

--Marburg virus

Flaviruses (Togaviruses)--Group B Arboviruses

--Tick-borne encephalitis virus complex including Absetterov, Central

European encephalitis, Hanzalova, Hypr, Kumlinge, Kyasanur Forest

disease, Omsk hemorrhagic fever, and Russian spring-summer encephalitis

viruses

Herpesviruses (alpha)

--Herpesvirus simiae (Herpes B or Monkey B virus)

Hemorrhagic fever agents and viruses as yet undefined.

Appendix B-V. Animal Viral Etiologic Agents in Common Use

The following list of animal etiologic agents is appended to the

list of human etiologic agents. None of these agents is associated with

disease in healthy adult humans; They are commonly used in laboratory

experimental work.

A containment level appropriate for RG1 human agents is recommended

for their use. For agents that are infectious to human cells, e.g.,

amphotropic and xenotropic strains of murine leukemia virus, a

containment level appropriate for RG2 human agents is recommended.

Baculoviruses

Herpesviruses

--Herpesvirus ateles

--Herpesvirus saimiri

--Marek's disease virus

--Murine cytomegalovirus

Papovaviruses

--Bovine papilloma virus

--Polyoma virus

--Shope papilloma virus

--Simian virus 40 (SV40)

Retroviruses

--Avian leukosis virus

--Avian sarcoma virus

--Bovine leukemia virus

--Feline leukemia virus

--Feline sarcoma virus

--Gibbon leukemia virus

--Mason-Pfizer monkey virus

--Mouse mammary tumor virus

--Murine leukemia virus

--Murine sarcoma virus

--Rat leukemia virus

F. Amendments to Appendix C, Exemptions Under Section III-E-6

Appendix C-I-A is amended to read:

Appendix C-I-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 Director 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 Risk Groups 3, 4, or

restricted organisms (see Appendix B, Classification of Human Etiologic

Agents on the Basis of Hazard, and Sections V-G and V-L, Footnotes and

References of Sections I through IV) 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

[[Page 1489]]

F, Containment Conditions for Cloning of Genes Coding for the

Biosynthesis of Molecules Toxic for Vertebrates), 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-A is amended to read:

Appendix C-II-A. Exceptions