THE ROCKEFELLER INSTITUTE
FOR MEDICAL RESEARCH

66TH STREET AMD YORK AVENUE
NEW YORK 21, N. Y.

December ff, 1952

Dear Josh:

This is to answer yours of the 24th and 25th. 1 don't think any
enclosure with the reprints is necessary but thanks for asking anyway.
Re: respreading experimmants. There are few details I can add as illness
and the pressure of other work has kept me from it. ‘he culture used was
SW-354 (Gal+ ST) and the phage PLT-22/78" . I have since done a reconstruction
experiment which essentadally invalidates the conclusions with S‘ as the clones
don't increase in time. “his being due to inhibition of colony formation by
the heavy curds of debri etc rather than differential growth rate. The Gal story
remains intact. The mutant is sufficiently stable that thus far I8ve not had
to correct for spontaneous mutants in time. The experiment is not easily done
in broth as one can't grow populations of sufficient size,in single steps anyway,
which would have sufficient number of transductions per p ting sample. However
in order to avoid the mess with SM and still study the chbacter and also
Aux-prot transductions I shall set up a primitive chemostat and try the experiennt
in broth. So far there has been little difference in the growth rates on EHB and
NA. The inmediate stimulus for this experimant was Hotchkiss! finding of a
similar delay with S* transformations. He believes this to be inhibitional
not segregational which as you say will be difficult to ppove. He now has a
mannitol transformation.
Re: SW-541 and 666 . I merely streaked out the 541 you sent me. The rest was
aK pure speculation, Have separated the + and S components which are stable and
prepared FA from each to do the genetic tests. The full mormex pluses which you mention
but which I've not seen made me hopefu} for another linked transduction,
Re; lytic variants. I have thus far defined a lytic variant as one that
does not induce lysogenicity but does select resistant strains which thus far have
all been roughs. In this sense lytie viruses transduce, Ive not yet tested
whether they kill sensitive cells after adsorption which is probably the best
criterion.
Your experiment with 666 and lysogenicity is most useful in that it
supports FAsphage and also transduction need not result in phage infection.
Why should the a receptor cells be more contaminated with alternate phases
than the donor cells (FA II lacking phase I components)?
Apdarently some time last summer you most flatteringly told Szybalski that
it would take me about three months to explain Schneiders problem. The three months
are now up and I shall report the progress to date.
The problem was this. He had two strains of typhimurium which could be
differentiated by no means( antigenicity ,phagocytic index Btc) other than
one was virulent (single cell will kill) and the other avirulent (10° cells will
kill). If he injected 1000 cells of eack simultaneously all mice die. If he
first injected with avirulent and then after two days with virulent the mice
had a higher chance of survival especially if on a natural diet rather than
a synthetic (40% difference). These experiments have since been repeated with
a marked virulent strain ( marlting the avirulent might have produced some secondary
affectadck we could not have assayed) The mice were sacrificed in time and the fol,
und 4nd plated.With the mixed challenge at zero days the avirulents go way
head establishing a ratio of greater than 100 to 1 and achieve titers of
10° per spleen by five days when the mice start to die. Those mice on the natural diet
have higher titers of virulents and correspondingly die sooner. With the two day
post challenge (avirulents are a few thousand per spl#en at this time) the xvirulents
with the synthetic diet establish themselleves in the spleen in 24 hours but much dep-
ressed when compared to the one day mixed challenge. On the natural diet however
it takes from 48-72 hours before the virulents appear similarly much depressed.
By five days after secondary challenge the virulents on thé weebaa diet outnumber
the avirulents but are rising vdy slowly. At the same time on the natural diet
some mice have equal numbers of the two and others the virulents are increasing
rapidly mirroring the mixed challenge on this diets. All survivors will soon be
sacrificed to see wiiat has happened there. We now know what happens in vivo but
not why it happens. It also give us a smaller mouse assay for this dietary factor ?
previously taking forty mice to test a fractionation now about four so the resolition
of the material won't require tons of extract.

Sincerely,

Norton