March 30, 1955

Dear Bruce:

I am afraid that despite having focussed all my energy on trails
for the last three months (not to mention what I had spent last year)
that I have not been ahle to com up with anything more than negative
results, that is findings which do no mre, to my mind, than make it
still impossible to choose anong any of the interpretations. There has
been a by product of some technical improvements, however, which may
eventually help bring a mere definite solution. These would include the
almost trivial expedient of concentrating the treated bacteria, which
makes the rapid isolation of numerous actile initials very easy, and
pour—platings of nusbera of initials, or of single clones.

The principal negative finding is that, for a sample of a given col-
lection of initialea, the incidence of trails depends directly on the
consentration of the motility agar. This seeas to me to lead directly
to the notion of an accidental determination of trailse/no trails. I
would agree that it is likely that a many-chained clone has a greater
likelihood of producing a trail: if you look at a dispersion of #itt/
Fla* bacteria in motility agar, you will see that a large proportion
of bacteria are enmeshed, while others swim more freely in the appabent
interstices. If an initial produces several motile offspring, there will
obviously be a higher chance that one of them will be able to swim free.
I have not noticed any consistent differences in motility that would
help distinguish many-chained from single-chahhed cells, but I haw not
had mich difficulty of the sort you mntion, of immobilization on the
beach. (When a cell gets st&ck, I have found it easy to dislodge at by
prodding the cell not with the capillary tip, but with a droplet of ofl
pushed in and out against the cell). I have to do more experiments on
the quantitative variation, but the foblowing table will illustrate the
effect of diluting the motility agar (standard .4% agar, 8% gelatih) with
mitrient broth. (The plates were incubated about 8 hours, then brought
to room teaperature to limit the extension of swirms).The input in each
case was 0.1 ml of a suspension of 500 motile initials (##f(probably closer
to 550, since not every collected cell is necessarily counted in a mpid
harvest) in 1 ml broth.

Medium siagle cols. clusters trails swarms total
(2-5 or 10)

MGa standard 53 2 2 2 a9

70% + 30% broth 39 18 11 3 71

60% + 40% broth 19 i 19 3 52

In other experiments, I have had anywhere from 50 - 95% trails per total
viable cells, while standard MGA has generall given from 5-15%. In the
more dilute agar, the trails are not only more numerous, but individually
more prolific, becoming more inflorsscent than linear. I am very doubtful
that one can establish a reasonable standard for a "trail-former" when the
incidence 1s so depemient on the details of the medium. Platings of
single clones have given a similar pictured, with several instances of
2-7 well developed trails per clone (in dilute agar), which is only a
corollary of the above.
Evidently, the pedigree analysis will have to be done the hard way. I am concerned
that the expressivity of motility is so variable, especially whn the clones resume
the lag-log phase, during which they tend to become more sluggish as they enlarge.
I have in Wind to look for modifications of the medium which will facilitate the
diagnosis of motility. For example, in staled broth, TM2 has a very erratic be-
havior, with frequent stops and reversals, while in fresh medium the same cells
move in a more persdstent dignified manner, changing course usually only after

a collision. It is noB a pH effect; I don't know what it is. I only mention this
to indicate what may be possible; the particular observation is not of mich use,
though I think a larger fraction of cells do score ag motile in the staled mediun.

At any rate, I mist comfess to some uneasiness in working under the tension of

your own discomfiture at being held up. I do fully sympathize with you in wanting

to get an account off our respective chests, but my own conclusion is that a
definitive publication would be ill-advised right now. Rather than continue in
haste to push for a definite answer, I revert to a previous proposal to write a
shorter preliminary account which covers what we would agree on together. It will
then be for either or both of us to stick out our singular necks. I am preparing a
new draft, ani will send it (and some illustrative photographs) at the earliest
cpportunity. (Unfortunately we ran out of gelatin about 2 weeks ago, and have not
yet been able to replenish. Difco remains the only staisfa tory brand; I don't know
why, though for many purposes the medium without gelatin is satisfactory). I still
think that the Proc Nat Acad Sei would be the best vehicle for this kind of paper;
there will be ho difficulty in getting it in, and since the work was firmly founded
on your phenomenally productive, if brief, stay here, why all the more reason! I am
afraid that the JG reaches very few geneticists, and Genetics , J. Genetios, Heredity
all have too stramg an atmosphere of definitive archives. Nature and Seleace, on the
other hand, should be for even more discursive accounts. J. Bact. would not be

out of the question, but PNAS seems to m just about right. Would Proc Roy Soc be
entirely out of the qhestion, or is it difficult and time consuming to publish there?
Anyhow the substance is more important than the vehiele, so now, my own notions
having been better fixed, I will get to my own part.

In re terninology s I have so far had the most enthusiastic response to "chains" and
"catenate" (as I hope I mentioned, these are not my own invention, but J. Crow's).
Several muremgke have suggested sticking to the English chain in preference to

Latin catena, whichi is all right with me, if expressions like mgx chain (or chainly)
inheritance, and single, few-, many-chain(ed) cells will go. The English adjectives
in ly. or ed tend to have a narrow reference than the Latin in ate, etc., but there
is no real reason for this. The mst sensible adjectival form is the same as the
noun,chain, but there are too many pedants who object to that much flexibility.

 

Since I burdened you with the news of Mr. Wright in the first place, I should give
you the better picture we have now. After remaining comatose for two weeks (sic),

he then made a sudden ani dramatic recovery. He walked out of the hospital yesterday,
is improving considerablg in compensating for residuals (principally left arm and
hand) amd ought to be back at work this summer. He may still have a difficult adjust—
ment, depending how far his (now) monoplegia recedes. In all, he has come out so far
ineredibly better than we had had any reasonable basis to hope.

With Esther's bak too,
Yours sincerely,

PéS. You did promise to send me a more detailed argument on the transishasdon of
B in the pedigrees