Exob.
8-1

STATUS OF EXOBICLOGY PROGRAM

Some elements of the strategy of exobiologicai investigation have been
discussed in previous reports (cf. Westex material as summarized by Space Science
Board; Lederberg, J., 1960 "Exobiology - Experimental Approaches to Life Beyond
the Earth," Science 132: 393-400, August 12). The first investigations of a
new planetary habitat should certainly include a general survey at various magni-
fications for visual! evidence of life: until the results of these are in, it is hard
to judge what further steps should be taken in investigating the biology of possible
larger organisms. Needless to say, if the first look does give evidence of intelligent
culture or of larger organisms, considerable effort would be devoted to characterizing
these in later experiments. Meanwhile, a great deal can be planned for and done in
microbiological investigation starting from first principles. There are many reasons
to justify a strong emphasis on microbiology in planetary Investigations just as
there are for laboratory studies here. These include:

1) The greater likelihood of a successful result. In some habitats only
micro-organisms might be present; on the other hand, it is difficult to conceive
of 4 habitat which contained larger forms but did not contain micro-organisms.

2) Ubiquity. Even on a richly populated planet like ours, micro-organisms
can be found in almost any small sample of the atmosphere, the surface dust, or
bodies of water. This is much less true of larger organisms.

; 3) Metabolic diversity. Micro-organisms should afford a wider variety of
metabolic capabilities from which we might draw inferences about the comparative
biochemistry of the planet.

4) Ease of handling. With any luck at all, at least some species of
micro-organisms would be very much easier to cultivate and to contain for detailed
experimental investigation.

5) importance for terrestrial ecology. Both small and large organisms of
external origin constitute potential hazards if inadvertent!y brought to the earth;
One might anticipate much greater difficulties in removing micro-organisms and in
controlling them if they should escape. Therefore, we should obtain as much
information as possible about them in their natural habitat before programming
experiments that involve the actual return of pianetary samples to the earth.

How then might we procede to investigate the distribution of micro-organisms?
We should be prepared to meet the expectations: that: ,

1) A scarcity of moisture may result in a very sparse level of life.

2) The nutrition and biochemistry of the planetary organisms may differ
quite markedly from terrestrial ones.

We must, therefore, stress the most efficient methods of detecting micro-
organisms and also those methods which do not require a too specific identification
of earthly biochemical components. However, it is impossible to do a general
experiment in the abstract; and concrete experiments necessarily involve
some measured compromises on these principles. Further considerations in experi-
mental design are the necessary limitations on the size of the pay load, the

J. Lederberg
Stanford Universit

9/4/60 - |
2.

necessity for its automatic functioning, and the narrowness of the effective communi-
cation channel for returning the date to the earth. We should also keep in mind

that apart from the existence of life on another planet, we are most deeply interested
in a comparative study of its basic biochemical systems, and in particular, whether
other types of compounds can take the place of nucleic acids and proteins of
terrestrial organisms.

Proposed methods for exobiological study involve a combination of techniques
and a systematic presentation is likely to be rather artificial. The following
approaches may be kept in mind.

1) Chemical composition and metabolic effect of the micro-organisms. This
approach should be assiduously studied; the chief discouragement is the possible
scarcity of material to analyze. However, in combination with, for example, con-
trolled cultivation of the organisms, this approach will probably give us the most
detailed information on the intrinsic biochemistry of the exo-organisms. Studies
will be made to determine whether such characteristic substrates as ATP (adenosine-
triphosphate) are sufficiently rapidly metabolized by small numbers of organisms to
warrant further developments along this line.

2) Cultivation of the organisms on prepared media. This is the habitua}
technique of the microblologist and will doubtless have very high priority when
we can make firmer decisions on exo-microbial nutrition. Where water is a limiting
factor in a planetary biology, this may prove to be the most important nutrient.
If the communication channel is so narrow that only a few bits are available, an
efficient black box design, incorporating a test of microbial growth, and giving
a simple yes/no answer might be desired. However, in view of our uncertainties as
to the optimal nutrients, it would be wise to test a variety of alternative media.
Such a system might be miniaturized by culturing the organisms on moistened spots
on a moving tape rather than in bulk culture. The likelihood that the communication
channel, though still restricted, might be more ample suggests a more detailed
study of the samples. The two most promising procedures appear to be microscopy
and spectrophotometry, or some combination of the two.

3) Direct optical examination of the organisms (microscopy and spectro-
photometry). These methods involve the measurement of light intensity as a function
of position (i.e., the image of a particle) or of the spectral wave length (i.e.,
its absorption spectrum). If the technical problems can be surmounted, the combina-
tion of form and spectral data would give a most powerful method of biological
analysis.

a) Type of microscope. Most living celis, including bacteria, are
essentially transparent to the customarily visible wave lengths and must be processed
in particular ways in order to achieve useful contrast. The traditional way of
doing this in the laboratory is by staining, a procedure which can be used for a
considerable degree of chemical specification, but which adds to the mechanical
complications. Two principal possibilities are left for the observation of
unstained objects: microscopy in the ultra-violet at 2600-2800 & and phase
contrast microscopy. The advantage of ultra-violet microscopy would be the
relative chemical selectivity at different wave lengths, for example, the very
high absorption by nucleic acids at around 2600 A. However, to take full advantage
of this selectivity, additional information on the absorption at one or more other
wave lengths would be desirable. Likewise, with phase contrast microscopy, the
3.

greatest information would be obtained from a system which also discriminated
between particles which were opaque vs. transparent in ordinary transmission micro-
scopy. Therefore, the efficient use of either UV or phase contrast microscopy
Suggests the development of some method of color discrimination. This does not
necessitate the transmission of full color data, but rather the interposition of a
"filter logic’ to pass only those signals that fulfill stated criteria (i.e.,
transparent in visible and opaque in UV or phase contrast). Such finesse should
be more appropriate for electronic than photographic recording of the signals, and
our preliminary efforts have been directed to evaluating a UV microscope-vidicon
chain. Recent work on image-intensifying UV camera tubes (The Westinghouse

Ebicon being constructed for the orbital UV telescope) may be extremely useful;
other arguments favor the adoption of Flying-spot technique into the spectrophoto-
metric-microscope system.

b) Preliminary data reduction in situ. The limited communication
channel suggests the most strenuous efforts to eliminate irrelevancy and unwanted
redundancy from the transmissions. These arise from at least two sources within
the object being looked at:

1) A probable abundance of biologically uninteresting material,
especially mineral particles, among which it may be difficult to discriminate
cells and

2) High point-to-point correlations tn the object being looked
et - for example, under phase contrast, bacteria often resemble homogeneous elipsoids.

The waste resulting from the second factor might be minimized by differential
and edge-contrast discrimination techniques; the discrimination of color, and
perhaps also of form, might be used to monitor a decision as to which elements
were worth transmitting. {Needless to Say, at least a few unfiltered pictures
would be desirable also.) Waste from the first source could be minimized by
methods for the concentration of living material and the rejection of other
extraneous particles. The cultivation of the organisms, if this succeeds, would
be an admirable way to do this. However, in the expectation of possible
failure, thought should be given to other methods for the separation of cells
from various kinds of debris.

The problems of data reduction summarized here have some analogy to the
problems involved in the effective use of satellite reconnaissance (Samos )
(cf., Katz's review in Astronautics, July, 1960. ).

4) Collection of sumples. So far the least attention has been given this
problem, perhaps in the anticipation of productive developments in other fields.
For example, the auger and corer that have been proposed for the lunar round-trip
should be quite applicable to the Present purposes except that the specimens
would be processed within the payload rather than being returned. In addition,
the choice of procedures for collection would depend on the types of examination
that were adopted. At the present time, three alternatives are evident.

a) The collection of atmospheric dust for direct microscopic examination on
adhesive tapes - the flypaper proposal. b) The collection and concentration of
larger amounts of atmospheric dust by impaction or filtration - methods now in
Standard use in aerial microbiology. c) The soil sampler.
The first method might be applicable for direct Inspection and also culti-
vation on the tapes themselves. However, if the payload weight permits, it would
be safer to collect a larger sample and enrich it by the flotation of cells in
a dense medium as described further.

5) Present judgments. At the present time, 1 would suggest a program
with the following objectives. a) The collection of. from 10-100 grams of dust
or soil. b) The fractionation of the semple by flotation and c) direct
examination of the organic fraction by 1) phase contrast/transmission dual
microscopy and 2) microspectrophotometry between 2400 and 8000 R, either on
Single particles or on cleared samples of the organic fraction. c) Data
reduction and programming to transmit only the more likely observations. Pert
of the sample should also be cultivated on a nutrient tape and then also
examined in like fashion.

it will be difficult to make more refined choices and decisions until the
hardware has been designed and constructed to the point where it can be tried
out extensively on a larger variety of terrestrial samples. One of our necessary
and most important functions long before space flight will be the calibration of
these procedures end the search for the kinds of artifacts that must be avoided.

6) Further refinements and later experiments. With appropriate support
and cooperation of industrial organizations a workable system incorporating
some or all of the above elements should be ready in ample time for Saturn
flights to Mars circa 1965. If time still permits, additional refinements can
be added for the first missions and must be planned for succeeding ones. These
might include a) More complete cytochemical tests of observed particles, in
particular for DNA and for proteins. tt should be possible to adjoin these
tests to the microscopic observation. b) Command vs. zutomatic control of the
instrumentation to allow for unexpected developments at the target. c) Larger
scale enrichments for Martian organisms. These might depend on good guesses
or acquired information on the chemistry of the Martian surface and on the
physiology of the Martian organisms, and could then expand this knowledge.