21730 
Federal Register / Vol. 44. No. 71 / Wednesday. Aprd 11. 1979 / Notices 
DEPARTMENT OF HEALTH, 
EDUCATION, AND WELFARE 
National Institutes of Health 
Recombinant DNA Research; Actions 
Under Guidelines 
agency: National Insitutes of Health, 
PHS. HEW 
action: Notice of actions under NIH 
Guidelines for Research Involving 
Recombinant DNA Molecules. 
summary: This notice sets forth actions 
taken by the Director, NIH. under the 
1978 NIH Guidelines for Research 
Involving Recombinant DNA Molecules 
(43 FR 60108). 
EFFECTIVE DATE: April 11. 1979 
FOR FURTHER INFORMATION CONTACT: 
Additional information can be obtained 
from Dr. William J. Gartland. Office of 
Recombinant DNA Activities. National 
Institutes of Health. Bethesda. Maryland 
20205, (301) 496-6051 
SUPPLEMENTARY INFORMATION: 1 ant 
promulgating today several major 
actions under the NIH Guidelines for 
Research Involving Recombinant DNA 
Molecules These proposed actions were 
published for comment in the Federal 
Register of January 15. 1979 and 
reviewed and recommended for 
approval by the Recombinant DNA 
Advisory Committee (RAC) at its 
meeting on February 15-16, 1979. In 
accordance with Section IV-E-l-b of 
the NIH Guidelines. I find that these 
actions comply with the Guidelines and 
present no significant risk to health or 
the environment. 
Part I of this announcement provides 
background information on the actions 
Part II lists the major actions. 
I. Decisions on Actions Under 
Guidelines 
A. Lower Eukaryotes as Host-Vector 
Systems 
Consideration of the use of lower 
eukaryotes as host-vector systems was 
initiated at a meeting at NIH of a 
Working Group on Lower Eukaryote 
Host-vector Systems on September 16. 
1978 The purpose of the meeting was to 
consider possible alternatives to the EKl 
and EK2 systems. Based on the routes of 
escape, the possibility of mating with 
naturally occurring strains (and. 
therefore, potential transfer of cloned 
material) and the ecological niches 
available to Saccharomyces cerevisiae 
and Neurospora crassa. it was reasoned 
that considerable containment is 
afforded by normai laboratory strains of 
S. cerevisiae and mutationally modified 
strains of N. crassa. In addition, it was 
unanimously agreed that S. cerevisiae 
could meet the criteria of HV2 systems if 
transfer of cloned DNA was prevented 
by mutations that would prevent mating. 
A Working Group report was developed 
from this meeting and presented to the 
Recombinant DNA Advisory Committee 
(RAC) at its October 30-31. 1978 
meeting. The report was modified and 
approved by the RAC. The 
recommendations were summarized and 
published for comment in the Federal 
Register on January 15, 1979. No 
comments were received during the 30- 
day period for comment. The RAC again 
considered the recommendations at its 
meeting on February 15-16. 1979. 
1. N. CRASSA AND S. CEREVISIAE 
AS HVl SYSTEMS. The RAC 
considered the following proposal for 
approval of Neurospora crassa and 
Saccharomyces cerevisiae as HVl 
systems: 
".V crassa and S. cerevisiae as HVl 
systems. Specified strains of ,V. crassa 
which have been modified to prevent 
aerial dispersion and unmodified 
laboratory strains of S. cerevisiae are 
acceptable as HVl systems based on 
their natural containment. The following 
N. crassa strains can be used: 
(1) ini (inositolless) strains 37102. 
37401. 46318. 64001 and 89601. 
(2) csp-1 strain UCLA37 and csp-2 
strains FS 590. UCLA101 (these are 
conidial separation mutants) 
(3) eas strain UCI.A191 (an “easily 
wettable" mutant)." 
At the February 15-16. 1979 RAC 
meeting, the report was summarized and 
some of the advantages of using the 
lower eukaryote host-vector systems 
were described. The organisms are 
structurally and genetically more related 
to the cells of higher eukaryotes and 
there is the possibility of obtaining gene 
expression that may not occur in E. colt. 
particularly due to possibility of 
processing intervening sequences. The 
extensive knowledge of the genetics of 
Saccharomyces and Neurospora will 
enable significant contributions by 
in\estigators who have worked 
extensively with these organisms. 
It was explained that the basis for 
proposing these organisms for HVl 
containment came from considerations 
of the modes of escape, the opportunity 
for establishment in nature, and the 
possibility of transferring cloned 
segments to other species or strains in 
nature. As discussed in the report, there 
is very little opportunity to mate with 
other strains or species in nature and. 
furthermore, there are no natural vectors 
known which might effect interspecific 
transfer or genetic material. 
Saccharomyces cerevisiae and 
Neurospora crassa are nonpathogenic to 
man, animals, and plants, and not 
closely related to known pathogens. 
Therefore, the RAC feels that relevant 
and important experiments can be done 
safely without risk to man. animals, 
plants or the general environment. The 
ecological niche for S. cerevisiae is the 
surface of fruits or the nectar of flowers. 
N. crassa is found abundantly at barren, 
burned-out sites in the semi-tropics. 
Neither organism can colonize the gut of 
man. Large amounts of bread and beer 
are consumed without any pathogenic 
effects of the yeast that are present. 
The RAC voted to accept this section 
of the report by a vote of 19 to 0 with 1 
abstention. 
2. USE OF UNMODIFIED 
LABORATORY STRAINS OF N. 
CRASSA. The RAC considered the 
following proposal for use of unmodified 
laboratory strains of N. crassa: 
"Unmodified laboratory strains of ,V. 
crassa can be used in experiments that 
require the HVl level of containment 
provided that these are carried out at 
physical containment one level higher 
than required for HVl. However, if P3 
physical containment is specified for 
HVl, this level is considered adequate 
for unmodified N. crassa. For P2 
physical containment, special care must 
be exercised to prevent aerial dispersal 
of macroconidia." 
This action is essentially equivalent to 
allowing the use of N. crassa as an 
"HVO" system. The scientific reason for 
proposing this is that a wide variety of 
mutants are available in N. crassa. The 
restriction of experiments to a few 
strains would require considerable 
genetic manipulation and effort which 
could be avoided by use of unmodified 
laboratory strains. 
Several RAC members were 
concerned about the escape of 
Neurospora in aerosols or into the air 
since it forms spores which are freely 
dispersed. Since the intent was to have 
investigators exercise caution in their 
use of wild-type strains, it was proposed 
that the original proposed action be 
made more specific by the addition of 
the statement "including the use of a 
biological safety cabinet" after the last 
word in the paragraph. The RAC voted 
18 to 1 with 1 abstention to accept this 
section as amended. 
However, later in the discussion on 
the equivalence of lower eukaryote HV 
systems with those of E. coli. the RAC 
voted 18 to 0 with 2 abstentions that 
unmodified laboratory strains of N. 
crassa be approved at the present time 
[81] 
