NOTICES 
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to occupational and environmental 
concerns. 
A waiver provision in the section on 
"Prohibitions” will permit NIH sup- 
port and conduct of risk-assessment 
experiments of crucial importance to 
the determination of the safety of this 
work. Recommendations from the sci- 
entific community, the public, and rel- 
evant Federal agencies will be sought 
for their advice on specific projects. 
Waiver decisions will include a careful 
consideration of potential environmen- 
tal impact. 
In summary, a number of safety 
standards and procedural require- 
ments have been included in the PRG- 
NIH to insure minimal environmental 
impact. All experiments exempted 
from the Guidelines are of minimal 
speculative risk and present no signifi- 
cant hazard to health or the environ- 
ment. 
II. Containment 
Analysis of Current Guidelines 
Two approaches to the problem of 
containing potentially hazardous or- 
ganisms form the basis of the safe- 
guards recommended by the guide- 
lines. Each may be viewed as setting 
up barriers between the laboratory 
w'orker and the organisms and be- 
tween the laboratory and the greater 
environment. The first of these ap- 
proaches involves the limitation of the 
actual physical escape of organisms 
and is referred to as "physical contain- 
ment.” The second approach is the use 
of biological barriers, to be described 
later as "biological containment.” 
(The October 1977 EIS on the cur- 
rent guidelines, in response to com- 
ments received on the draft EIS, docu- 
ments in considerable detail the ade- 
quacy of the containment require- 
ments and show’s the bases on which 
judgments in this regard have been 
made.) 
Physical containment 
A major aspect of physical contain- 
ment is the set of standard microbiolo- 
gical practices that have been devel- 
oped over many years and are widely 
used for handling pathogenic organ- 
isms. In the hands of well-trained per- 
sonnel, these procedures have proved 
to be effective both in laboratory and 
clinical settings. A second major 
aspect of physical containment in- 
volves the use of special kinds of 
equipment and facilities (1) for limit- 
ing the spread of aerosols, (2) for de- 
contamination and containing labora- 
tory air and w'astes, and (3) for re- 
stricting access to laboratories. As 
with standard microbiological tech- 
niques, the type of equipment and fa- 
cilities are not new but have been de- 
veloped and used previously for con- 
tainment of known pathogens. 
The guidelines go into some detail 
concerning the practices and facilities 
required for physical containment. 
Four levels are specified: PI, P2, P3, 
and P4, in the order of increasing safe- 
guards. PI consists in the use of the 
standard microbiological practices 
mentioned above. P2 and the next 
higher level, P3, require special proce- 
dures and facilities designed to limit to 
increasing extents any possible acci- 
dental escape of potentially hazardous 
organisms. Finally, P4, the maximum 
level of containment, requires sophisti- 
cated and isolated facilities designed 
for maximum containment. 
Each of the levels from P2 through 
P4 assumes that the standard micro- 
biological practices demanded by PI 
will also be followed. Furthermore, for 
each level, relevant training of person- 
nel is mandatory. The training is to In- 
clude the nature of the potential haz- 
ards, the technical manipulations, and 
instruction in the biology of the rele- 
vant organisms and systems. Specific 
emergency plans, to be used in case of 
accident, are required; and serological 
monitoring is to be provided where ap- 
propriate. 
Biological containment 
Biological containment is defined as 
the use of host cells and vectors with 
limited ability to survive outside of 
very special and fastidious laboratory 
conditions that are unlikely to be en- 
countered by escaped organisms in 
natural environments. This is an inte- 
gral part of the experimental design, 
since the host and vector will need to 
be chosen, in any given experiment, 
with a view to the purpose of the ex- 
periment as well as to containment. 
The guidelines stress that physical 
and biological containment procedures 
are complementary, each serving to 
control any possible failure in the 
order. The use of both in a given ex- 
periment affords much higher levels 
of containment than either alone. 
Therefore, the guidelines always rec- 
ommend both a particular level of 
physical containment and a level of 
biological containment for any given 
experiment. The guidelines explicitly 
recognize that better physical contain- 
ment capabilities are likely to evolve 
as research proceeds and may reduce 
the needs for the standard physical 
containment procedures. Such innova- 
tions are to be considered as part of 
the on-going review of the guidelines 
for appropriate revision. 
The Use of Bacterial Hosts and Vec- 
tors. In recognition of the relation be- 
tween the host-vector system required 
by the experiment and the design of 
suitable biological containment, exper- 
ments using the same host-vector 
system are grouped together. At pres- 
ent, the system of choice for many ex- 
periments is the common laboratory 
bacterium E. coli, strain K-12, and in- 
dependent genetic elements (plasmids 
and bacteriophage) known to reside or 
replicate in this strain. 
There are several factors contribut- 
ing to this (discussed more fully in 
part III). Strain K-12 has been studied 
extensively and can be readily manipu- 
lated for recombinant DNA experi- 
ments. This extensive experience and 
ease of manipulation permit modifica- 
tion of E. coli K-12 and vectors used 
with it by classical genetic techniques 
for the purpose of establishing biologi- 
cal containment. 
The guidelines discuss arguments 
against as well as for the use of E. coli 
K-12. The main aigument against it is 
the intimate association of various 
other strains of E. coli with humans. 
By reason of the prevalence of E. coli 
strains (but not K-12) in mammals, 
the guidelines recommend the cau- 
tious use of E. coli K-12 host-vector 
systems and urge that efforts be ma,de 
to develop alternate hosts and vectois. 
E. coli K-12 appears to be harmless: 
It does not usually establish itself in 
the normal bowel or multiply signifi- 
cantly in the alimentary tract. These 
facts suggest that accidental ingestion 
of a small number of bacteria by a lab- 
oratory worker would not result in 
their extensive spread outside the lab- 
oratory. This property of the organism 
may be altered, however, when the in- 
fected person is taking antibiotics or 
has certain abnormal digestive condi- 
tions, and it is recommended that such 
persons not work for the duration of 
the abnormal circumstances. 
While E. coli K-12 does not establish 
itself as a growing strain in the normal 
bowel, it does remain alive during its 
passage through the intestinal tract. 
Therefore, transfer of plasmid or bac- 
teriophage vectors containing foreign 
DNA from the original E. coli K-12 
host to bacteria resident in the intes- 
tines or encountered after excretion 
must be considered. The guidelines 
take into account the transferability 
of certain vectors in recombinant re- 
search. In brief, host-vector systems 
derived from E. coli K-12 and certain 
plasmids or bacteriophage appear to 
have extremely limited ability to 
spread recombinant DNA molecules. 
Considering, then, the properties of 
E. coli K-12 and the known plasmid 
and bacteriophage vectors, the guide- 
lines conclude that recombinant 
DNA’s manipulated through such 
host-vector systems are unlikely to be 
spread by the ingestion or dissemina- 
tion of the few hundred or thousand 
bacteria that might be involved in a 
laboratory accident, given standard 
microbiological practice. Therefore, 
these existing systems, and analogous 
combinations of E. coli K-12 with 
other vectors and bacteriophage, are 
judged to offer a moderate level of bio- 
FEDERAL REGISTER, VOL 43, NO. 146— FRIDAY, JULY 28, 1978 
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