Muassachusells Gpveyal Hospital

GENERAL HOSPITAL wo / VA MCLEAN HOSPITAL
BAKER MEMORIAL Of 4 f W. FRANKLIN Woop, M.D,

PHILLIPS HOUSE DIRECTOR

 

 

BURNHAM MEMORIAL FOR CHILDREN IN LINCOLN
HALL- MERCER HOSPITAL STORROW HOUSE
HUNTINGTON MEMORIAL HOSPITAL (CONVALESCENTS)
VINCENT MEMORIAL HOSPITAL

 

Dean A. CLARK. M.D.
GENERAL DIRECTOR

October 2, 1956

Dear Dr, Lederberg:

One of the cultures: which I sent you, the Proteus 52
regained from the L form, needs some comment. It has a high
resistance to penicillin and the culture is pleomorphic on
penicillin plates, but most of the colonies are nét‘iof the L-type.
In the same way, it is difficult to decide the nature of the growth
in broth. Such strains may be interesting in some respects but are
not appropriate to study the properties of the L forms and it was a
mistake to send it to you, Usually the resistance to penicillin of
a Proteus recovered from the L form is not increased,

I would like to add a few remarks to the possible connection
of protoplasts, large bodies and L forme. My impression is that
if we regard the protoplasts as the living organisms inside of a
faifly rigid cell wall, the L forms are probably the growth of this
organism without building up the cell wall, However, the large body
is not the protoplast of a single bacterium but it is produced by a
limited multiplication of a single protoplast. This was most clearly
visible with the Bacseroides strain 132 to which I will refer in an
other connection also. The large bodies developed in the broth
cultures of this strain without the addition of penici}lin or any
other growth-inhibiting substances, The sequence: wad this and
almost all bacteria went through the same sequence.at the same time:=

~~ Rares ans gene . weet. as

ers f

Division stopped after 6 ~ 8 hours.
When transfers were made to an agar plate during the development
of large bodies, within a few hours all these various forms were

retransformed to the usual bacilli. The most interesting was the
retransformation of := .--. no

They fall withou€ changing the outliné of the structure into bacilli
indicating that the bacilli were preformed in them:-

I published photographs of this process, It is apparent in this case
that the development of the large bodies was not a physical process
like inkivifion with water, but a growth process involving division
of the living units. The staining of the chromatinic structures
both in strain 132 and in other bacteria indicated the same thing.
IN BOSTON
GENERAL HOSPITAL
BAKER MEMORIAL
PHILLIPS HOUSE

 

Massachusels General Hospital

'o/ hon. Wy MCLEAN HOSPITAL
NOL Th W. FRANKLIN Woop, M.D,

DIRECTOR

 

BURNHAM MEMORIAL FOR CHILDREN —2- IN LINCOLN

HALL-MERCER HOSPITAL
KUNTINGTON MEMORIAL HOSPITAL
VINCENT MEMORIAL HOSPITAL

 

Dean A. CLARK, M.D,

GENERAL DIRECTOR

STORROW HOUSE
(CONVALESCENTS)

for ete fe, - sf

In strain 132 it was very interesting to observe, this process, ©
At about 4 hours most bacilli grew to short filaments about

2-4 bacilli long. The formations of the large body started

by an incomplete division:= ee ‘ a)

The next step was juxtaposition and fusion of the segnents-
producing forms surprisingly similar to those which you
described in B, coli, Usually this fusion was complete

but in some cases a dividing line remained between the structures
developing from the two bacilli:=

Dr, Smith who worked with me wrote a short paper about these forms.
A fusion preceding the formations of large bodies is visible also
sometimes in B, coli and Salmonella in the following way: First a

quadrangular side growth develops in the filaments and later this
fuses into a large body:=

At present, when sexual intermixture of the characteristics and
conjugal paring also have been observed, these few observations
concerning the oregin of large bodies seem significant. Maybe

they will lead to a better understanding of what the large bodies
really are,

The growth of the L forms from the large bodies does not

appear to me a budding similar to that of yeast. A large body,
as is apparent from the way it is formed, does not consist of one
but of many living units and these living mits grow out of it.
In photograph these ehromatintie#ét bodies are stained, They
look about the same in the large bodies as in the bacteria only
that they are arranged in @ sphere, When the large body grows further
(photograph [ ) the chromatinic bodies multiply, ‘The old photograph
(JIL) shows the way of growth and the size of the growing elements
of the cultures, These are smaller than the bacteria but not too
much smaller. The smallest are about 0.3 to 0.5.4, In most cultures
they are larger and may swell to large forms, Their arrangement in the mm
culture indicates that they multiply like bacteria. In electron
Micrographs from Proteus L, there are many granules between 0.15 to
0.3 /« and some less than 0,1“, Thus far the multiplication of
single granules of this size ‘has not been observed, Growth can be
obtained in our media only from large inoc ula, This may explain
why the small gramles do not grow. BECEBEECHSODCRENOSSAOOtRIIIET iin

The growth cycle in large colonies of certain cultures seems to be

different and to consist of growth of the small granules into large

bodies and the subsequent breaking=up of these again into
without multiplication of the small granules in such ¢
orm,

  

granules
IN BOSTON
GENERAL HOSPITAL
BAKER MEMORIAL
PHILLIPS HOUSE

 

 

C7 , ‘I MCLEAN HOSPITAL
DL é V4 W. FRANKLIN Woop, M.D,
on Zinn

DIRECTOR

 

a

BURNHAM MEMORIAL FOR CHILDREN IN LINCOLN

HALL- MERCER HOSPITAL
HUNTINGTON MEMORIAL HOSPITAL
VINCENT MEMORIAL HOSPITAL

 

Dean A. CLARK, M.D.

GENERAL DIRECTOR

STORROW HOUSE
(CONVALESCENTS)

As far aa I can see, there is
nothing in the origin and multiplication of L forms which
contradicts the supposition that they correspond to protoplasts.
They may be protoplasts slightly modbfied or in a special
condition, They are poorly adapted to ouf media and most
cells die and disintegrate.

Would you mind if I made a few experiments with the
two colon bacillus strains in which mating is often observed?
Tf not, could you send me these two strains?

With kind regards,

Sincerely yours,

Kuru i, ok Cia]
Louis Dienes, M.D,

I received your letter of September 27 after this letter

was typed, I would like very much to read the promised

enclosure but it was not in the letter, It is wonderful

that you succeeded to get L forms from K-12, I had some time ago
the idea that the L cultures may be analogous to the haploid yeast
colonies of Winge, It will be very interesting to see whether they
have any genetic function,

It may be of some use if I also make a few experiments with
your colon bacillus strains, I would be very much interested to
know whether what kind of L cultures you get, I leave it entirely
up to you whether it fits into your plans or not,

I am quite excited to learn how your experiments are
developing,