(1) Typhoid bacilli are naturally resistant to
digestion by proteolytic ferments such as Pepsin
or Trypsin, but when extracted with acetone for
24 hours, they are almost completely dissolved
by 1 2% solutions of Trypsin in 4 - 6 hours.
(2) From such digests of B. Typhosus, by appropriate
methods, bacterial protein derivatives, corresponding
to progressive phases of digestion, can
be isolated.
(3) These derivatives are, alkali and acid metaprotein,
coagulable protein, primary proteose, secondary
proteose and peptone.
(4) Equal amounts of these bacterial products have
been inoculated into rabbits, and the antibody
response compared with that obtained by inoculating
equivalent amounts of dried undigested
Typhoid bacilli.
(5) Metaproteins are definitely antigenic, but compared
with unaltered Typhoid bacilli, they act
only as partial antigens. Agglutinins are not
produced in response to inoculation of Typhoid
metaprotein, and the other antibodies are present
in much less concentration than in the controls.
There is no difference to be made out between
acid and alkali metaprotein. Though these are
the earliest recognisable derivatives of the
unchanged bacterial protein, their combination
with acid or basic radicals brings about a marked
deterioration in their antigenic power.
(6) Coagulated. Protein produced all the recognisable
antibodies against the Typhoid bacillus, but in
lesser concentration than the untreated bacilli.
In spite of the alteration in molecular grouping
associated with coagulation, this antigen proved
more efficient than acid or alkali metaprotein.
(7) Proteoses act as very weak antigens. No agglutinins
or opsonins were formed, and complement
fixation antibodies could not be demonstrated.
Their inoculation caused a slight rise in the
bacteriolytic pourer of the serum, and a positive
precipitin response, demonstrable only in the
lowest dilutions of the serum.
(8) No demonstrable antibodies against the Typhoid
bacillus were formed as a result of inoculations
of bacterial peptone.
(9) Typhoid bacilli digested for 4 hours with 1%
Trypsin were tested as antigens. Inoculation of
these filtered digests produced a rise in the
bacteriolytic power of the serum greater than
that found in the controls receiving undigested
bacilli. Agglutinins, precipitins, opsonins and
complement fixation antibodies were produced, but
to a lesser degree than in the controls.
(10) Typhoid bacilli, extracted with acetone, are not
thereby appreciably weakened in their antigenic
power.
(11) The lipoid material, removed from bacilli by
extraction with acetone, is completely devoid of
antigenic properties.
(12) In experimental animals, the titre of antibodies
in the serum cannot be accepted as an accurate
criterion of the protective power against the
living bacillus.
(13) Evidence is brought forward to show that antibodies
are not all produced together, but in
response to different phases of the digestion of
bacteria. In this respect Douglas' findings
are confirmed.
(14) It is suggested that agglutinins are formed in
response to a very early phase in the cleavage
of the bacterial protein, and that unaltered
native protein is necessary for their production.
(15) There is also evidence that opsonins are produced
in response to an early stage of cleavage
of the protein molecule, though not so early as
is required for the production of agglutinins.
For the production of opsonins, the protein must
not have passed the coagulable stage.
(16) It is further suggested that bacteriolytic antibodies
are produced in association with a later
period of digestion of the bacterial protein
than either agglutinins or opsonins.
(17) The production of precipitins seems to run on a
parallel with that of bacteriolysins, but the
evidence on this point is inconclusive.
(18) It is concluded that agglutinating and precipitating
properties of sera are not very closely
related, but are really distinct phenomena.
(19) Judging from the experiments carried out, it
would appear as if bacteriolysins in a serum are
only capable of being stimulated up to a certain
titre, and this can sometimes be reached after
two or three inoculations. Subsequently in
spite of the inoculation of larger amounts of
antigen, there is a tendency for the bacteriolytic
power to decline. This is especially apt
to occur where undigested bacilli are employed.
(20) The efficacy of so called detoxicated vaccines,
which consist of metaproteins and proteoses, is
discussed. From experience with Typhoid
derivatives, it is concluded that while the
constituents of detoxicated vaccines can act
as antigens and produce antibodies, the immunisation
is only partial. This appears to be due to
the fact that chemically treated antigens undergo
some alteration., as a result of which they
become less effective.
(21) Typhoid bacilli digested for 4 hours with i to
2% Trypsin are much more toxic than undigested
bacilli.
(22) The split products which can be isolated
chemically from digests of Typhoid bacilli are
less toxic than the original organisms.
(23) The primary proteose fraction is the most toxic
of the protein split products derived from the
Typhoid bacillus. The metaproteins are less
toxic than primary proteose, and secondary
proteose is the least toxic of all.
(24) The symptoms and pathological findings in rabbits,
which have received lethal doses of both living
and dead Typhoid bacilli, are described.
(25) The mode of death, and the post -mortem appear-
ances including the congestion of the abdominal
organs, the hyperemia and focal haemorrhages in
the intestine, and the congested and haemorrhagic
condition of the lungs, are very similar to what
is found in sensitised rabbits which have received
a toxic dose of egg globulin and in which
death has occurred comparatively late.
(26) The subacute nature of the illness as compared
with acute anaphylactic death, points to a
solution or partial disintegration of the
bacterial protein being necessary before the
toxic substances are liberated.
(27) The pathological picture in death from inoculation
of Typhoid primary proteose is practically
the same as in death following inoculation of
the whole bacilli or of bacilli which have been
digested with Trypsin.
(28) Primary proteose, derived from egg albumen, has
no power of sensitising animals either towards
itself or towards the native protein from which
it was derived.
(29) The pathological picture following inoculation
of animais with Typhoid bacilli is a non -specific
one, and can be produced not only by colon
bacilli, but by non bacteria]. protein.
(30) The symptoms produced in animals Typhoid
primary proteose are the same as those descrihod
following inoculation with "Peptone ", and
peptone shock seems to be dependent on the
presence of proteoses -- especially primary
proteoses.
(31) From a study of the toxic substances derived
from Typhoid bacilli, it is probable that the
intoxication produced by these bacteria is
dependent upon. -
(a) Preformed toxins of a simple protein nature,
stored up in the bacterial body and liberated
by lysis -- endotoxins.
(b) Protein split products of the bacterial
cell -- e.g. primary proteose.
(32) It is suggested that in man, the native bacterial
proteins may be associated with the intoxication
produced, the system having become sensitised
to these proteins during the incubation period.
