NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
Medical Officers of the Army of India.
15
Number
of speci-
men.
Number
of corre-
sponding
specimen
of unboil-
ed milk in
Table 1.
Date at
which pro-
cured.
Source.
General
condition when
obtained.
Nature and treatment
of individual
samples.
Subsequent condition
XVIII
XVI July
29th.
Bazaar. Neutral
1. Milk from the bot-
tom ; boiled in a
sterilised tube; in
the incubator at
37°C.
July 30th. Faintly alkaline; quite
fluid.
July 31st. Faintly alkaline ; in
soft loose coagulum with a
little supernatant fluid.
XIX XVII Aug.
3rd.
, ,
,,
1. Nature and treat-
ment as above.
August 4th. Quite fluid and seem-
ingly unaltered.
August 8th. Reaction faintly
acid, almost neutral on drying ;
loosely coagulated and partially
separated into coagulum and
fluid.
XX XVIII Aug.
15th.
, ,
Faintly alka-
line.
1. Nature and treat-
ment as above.
August 16th. Quite fluid ; reac-
tion very faintly alkaline ; non-
coagulant on being boiled anew.
August 17th. Quite unaltered
save that reaction is now neu-
tral.
2. Nature and treat-
ment as above.
August 16th. Quite fluid ; reac-
tion very faintly alkaline.
August 17th. A soft powdery
coagulum and a little superna-
tant fluid ; reaction very faintly
acid.
Table III corresponds to the first table of the series, but shows the changes
occurring in specimens of milk which were boiled when first procured. In order
to facilitate comparison a separate column has been introduced showing the
numbers of the specimens in Table I to which those of the present table corre-
spond. The most striking differences which the boiled, as compared with the
unboiled specimens of milk, showed were in regard to reaction and coagulation.
The results were not, however, of uniform character throughout, for, whilst in
some cases the specimens after a period of 24 hours remained perfectly fluid and
unaltered in reaction, in others, especially if they had only been once boiled,
coagulation had occurred, sometimes only imperfectly, but in others fully
developed and of the normal type, and the reaction, although not so highly
permanently acid as is normal to unboiled specimens, was yet transitorily decid-
edly acid, and in some cases remained feebly so permanently.
The most curious point in regard to this difference in results was that the
occurrence of coagulation and alteration in reaction appeared to be more closely
related to the stratum of the sample from which' the given specimens were ob-
tained than to the fact that they had been exposed to repeated boiling or not.
Medical Officers of the Army of India. 1
7
This difference in the phenomena presented by portions of milk from one
and the same sample according to the level in the fluid from which they have been
obtained is a very striking one. It certainly cannot be ascribed to the presence of
any living ferments in the upper strata and absent in the lower ones, as the results
of cultivations of boiled milk showed that in all cases, and whether the fluid had
been obtained from the upper or lower strata of the sample, sterilisation had been
completely effected in regard to all organisms save Bacillus subtilis, and that
this was present throughout the entire depth. On the other hand, the occurrence
of acid fermentation and coagulation is unequivocally related to the presence and
multiplication of schizomycete organisms in the milk, so that it appears to be
probable that the explanation of the phenomenon must lie either in the actual
ferment developed by the schizomycetes and affecting the milk, having a ten-
dency to accumulate in the upper portion of the fluid, or in peculiarities in the
constitution of the latter as a material to be acted on by it. This, however, is a
point which will be again referred to in connection with an account of the results
of cultivations of specimens of boiled milk.
Ordinary acid fermentation and coagulation thus does not occur in samples
of the lower strata of any common mass of milk as supplied in Calcutta, if these
have been once boiled so as to destroy the normal lacteal schizomycetes present
in them, and if complete sterilisation be effected, the fluid remains permanently
unaltered, save that in some cases a very slight and often fugitive acidity may
gradually manifest itself. If, however, complete sterilisation be not effected, if
the Bacillus subtilis surviving ordinary boiling remain in the fluid, whilst the
normal acid fermentation and coagulation does not occur, coagulative changes
of a different type manifest themselves at a later period. The coagulum in these
cases is not of the firm consistence of that in the case of the ordinary acid
fermentation, nor does it appear in the form of large masses but as a copious
deposit of fine powdery flocculi, easily diffused throughout the fluid, which
thus, after agitation, for a considerable time appears to casual observation as
though it retained its original characters and had not undergone any coagulative
change.
TABLE IV.—Results of Plate Cultivations of Boiled Specimens of Milk from
various sources.
Number of
specimen.
Number
of corre-
sponding
specimen
in Table
II.
Date at
which pro-
cured and
boiled.
Source. Reaction.
Nature of
cultivation.
Date at
which set.
RESULT.
I
III Jan. 9th.
Twice boil-
ed ; at 10
A.M. and
1 P.M.
House-
supply.
...
1. Plate inocu-
lation with
0.083 C.C Set
in incubator.
January
10th.
January 11 th Crowded with
colonies of Bacillus subtilis :
a pure cultivation of that
species.
18
Scientific Memoirs by
Number o f
specimen.
Number
of corre-
sponding
specimen
in Table
II.
Date at
which pro-
cured and
boiled.
Source. Reaction.
Nature of
c u l t i v a t i o n .
Date at w h i c h s e t .
RESULT.
Jan. 10th.
Twice boil-
ed at 10 A.M.
and 1 P.M.
2. Plate inocu-
lated with
0.083 C.C. Set
in incubator.
January
11th.
January 13th. A solitary
colony of Bacillus subtilis.
Jan.11th
Treated
as above.
Jan. 11th.
Faintly
acid.
3. Plate inocu-
lated with
0.083 C.C. Set
in incubator.
January
12th.
January 13th, and subse-
quently quite sterile.
II
IV Jan. 16th.
Twice boil-
ed ; at to
A.M. and
1 P.M.
House-
supply.
Jan. 16th.
Neutral.
1. Plate treat-
ed as above.
January
17th.
January 18th. A pure cultiva-
tion of Bacillus subtilis.
Jan. 17th.
1 reated as
above.
Feb. 27th.
Neutral.
2. Plate treat-
ed as above.
Feb.
27th.
February 28th, and subse-
quently remained absolutely
sterile.
III
VI March 8th
Once boil-
ed.
, , March 8th.
Neutral.
1. Plate treat-
ed as above.
March
9t h.
March 10th. A pure cultiva-
tion of multitudes of colonies
of Bacillus subtilis: smell
faint and mouldy.
IV
VII March
10th.
Once
boiled.
, ,
March
10th.
Neutral.
1. Plate set as
above.
March
11th.
March 12th. Surface in greater
part covered by a continuous,
slightly wrinkled stratum of
Bacillus subtilis.
March
11th.
Once
boiled.
March
11th.
Neutral.
2. Plate set as
above.
March
12th.
March 13th. Absolutely
sterile.
V VIII March
12th.
Once
boiled.
, ,
March
12th.
Neutral.
1. Plate set as
above.
March
13th.
March 14th. Containing colo-
nies of Bacillus subtilis in
the proportion of 271,320
per C.C : a pure cultivation.
March
13th.
Once
boiled.
.
March
13th.
Neutral.
2. Plate set as
above.
March
14th.
March 15th. Sterile.
VI
. . .
March
I5th.
Once
boiled.
Bazaar. March
15th.
Neutral.
March
16th.
Acid.
1. Plate inocu-
lated with
0.058 C.C.
and set in
incubator.
March
16th.
March 17th. Surface covered
by a thick, continuous, dry,
tough coating of Bacillus
subtilis.
VII
IX May 9th.
Once boil-
ed.
Zoologi-
cal Gar-
den.
May 9th.
Neutral.
1. Plate set as
above.
May
10th.
May 11th. Absolutely crowded
with colonies of Bacillus
subtilis ; smell smoky.
May 10th,
11th, 12th,
13th, 14th.
Once boil-
ed.
2, 3, 4, 5, 6,
Plates set as
above.
May
11th,
12th,
13th,
14th,
15th,
May 12th, 13th, 14th, 15th,
16th. Abundant crops of
Bacillus subtilis.
Medical Officers of the Army of India.
19
Number
of speci-
men.
Number
corre-
spondin g s p e c i m e n
in
Table II.
Date at
w h i c h p r o -
cured and
boiled.
Source. Reaction. Nature of culti-
vation.
Date
at
which
set.
RESULT.
May 15th.
Twice boil-
ed ; 9 A . M.
and 12-30
P.M.
7. Plate inocu-
lated with
0 058 C. C.
and set in in-
cubator.
May
16th.
May 17th, and subsequently
absolutely sterile.
VIII
. . . May 9th.
Once boil-
ed.
Zoologi-
cal Gar-
den.
. . . 1. Plate set as
above.
May
10th.
May 11th. Covered by a con-
tinuous, dense, dry, wrink-
led, thick coating of Bacillus
subtilis ; smell musty.
IX
X May 21st.
Once boil-
ed.
House-
supply.
May 21st.
Faintly
acid ; neut-
ral on dry-
ing.
Ditto
May
22nd.
May 23rd. Full of colonies
of Bacillus subtilis.
X
XI May 23rd.
Once boil-
ed.
„
May 23rd.
Neutral,
becoming
faintly
alkaline.
Ditto
May
24th.
May 25th. Crowded with co-
lonies of Bacillus subtilis,
but not a pure cultivation, a
certain number of colonies
of a large Micrococcus being
also present.
XI XIII May 27th.
Once boil-
ed.
Bazaar May 27th.
As above.
May 28th
Acid, be-
coming
neutral.
Ditto
May
28th.
May 29th. Absolutely crowd-
ed with colonies of Bacil-
lus subtilis.
XII
XIV May 29th.
Once boil-
ed.
„
. . . Ditto
May
30th.
May 31st. Full of colonies
of Bacillus subtilis.
XIII
XV May 31 st.
Once boil-
ed.
„
. . . Ditto
June 1st June 2nd. An abundant
growth of Bacillus subtilis.
June 1st,
2nd, 3rd,
4th, 5th.
Once boil-
ed.
2, 3, 4, 5, 6.
Plates set as
above.
June
2nd, 3rd,
4th, 5th,
6th.
Yielded abundant crops of
Bacillus subtilis.
XIV
. . . June 7th.
Boiled ; 10
A.M. and
6 P.M.
„
Neutral 1. Plate set as
above.
June
8th.
June 9th. Crowded with
small colonies of Bacillus
subtilis.
2. Plate set as
above.
June
9th. June 10th. Ditto ditto.
June 8th
and 9th.
3. Plate set as
above.
June
11th.
June 12th. Perfectly sterile.
Treated as
above.
June 10th,
11th.
4. Plate set as
above.
June
12th.
„ 13th.
,,
,,
Kept for
some hours
in a boiling
water-bath.
5. Plate set as
above.
June
13th.
, , 1 4 t h .
, ,
, ,
XV
...
July 20th.
Once boil-
ed.
,,
,,. 1. Plate set: in-
oculated 0.03
C. C. and set
in incubator.
July
21st.
July 22nd. Covered by an
almost universally confluent
stratum of colonies of Bacil-
lus subtilis.
D 2
20
Scientific Memoirs by
Table IV bears the same relation to Table II that the previous Table bears
to Table I. It shows the results of a series of cultivations of boiled specimens
of milk. These are very uniform and show clearly that a single and brief boil-
ing is sufficient to clear normal samples of the fluid from all living organisms
save Bacillus subtilis, which then multiplies with excessive rapidity. The num-
bers of colonies of the latter organism occurring in cultivations of boiled milk are
illustrated in the four photographs on the right side of the plate. In connection
with Bacillus subtilis there are two phenomena in these experiments specially
noteworthy—first, the evidence which is afforded that the other schizomycete
organisms normally occurring in milk in Calcutta exert a repressive action on the
development of Bacillus subtilis, and, second, the variations occurring in regard to
the persistence of Bacillus subtilis in different specimens of milk. The repres-
sive influence of the other normal schizomycetes inhabiting the fluid on the
development of Bacillus subtilis was very clearly demonstrated throughout the
entire series, the amount of the latter species developed in any cultivation of
unboiled milk being extremely limited, and its appearance almost entirely con-
fined to cultivations set immediately after the samples of milk were obtained,
and, therefore, at a stage when they contained relatively small numbers of the
other common schizomycetes as compared with those present at later periods.
The results obtained in the case of the eighth cultivation recorded in the pre-
sent Table are specially instructive in this respect as compared with those
recorded in regard to corresponding cultivations of a specimen of the same
Number
of speci-
men.
Number
of corre-
sponding
specimen
in
Table II.
Date at
which pro-
cured and
boiled.
Source. Reaction. Nature of culti-
vation.
Date
at
which
set.
RESULT.
XVI XVIII July 25th.
Once boil-
ed.
Bazaar Neutral 1. Plate inocu-
lated with one
common needle
dipped in a ste-
rilised, neutral
solution of salt
into which one
looped needle
dipped in the
milk had been
introduced.
July
31st.
August 1st. Covered by a
continuous stratum of Bacil-
lus subtilis.
XVII
...
August
3rd. Once
boiled.
„
August
3rd.
Neutral.
August 8th
Faintly
acid.
1. Plate set as
above.
August
8th.
August 9th. Covered by a
continuous stratum of Bacil-
lus subtilis.
XVIII
XX August
15th.
Once
boiled.
,,
Neutral
Ditto
August
16th.
August 17th. An abundant
pure cultivation of Bacillus
subtilis.
Medical Officers of the Army of India.
21
sample of milk which had not been boiled. The latter are entered under No. IX
of Table II. It will be seen that while the two cultivations of the unboiled speci-
men failed to show any evidence of the presence of Bacillus subtilis, that of the
boiled specimen within 24 hours became covered by an enormous growth of the
species in the form of a continuous, thick, dry, wrinkled stratum covering the
entire surface of the plate. Judging by the very feeble development manifesting
itself in any of the cultivations of unboiled samples of milk, it would not appear
probable that the proportion of rods and spores of Bacillus subtilis originally
present in the fluid is considerable, as, were it so, the species would probably
show more evidence of success in the struggle for existence with its compan-
ions. Direct evidence of this was obtained by means of a series of successive
cultivations of one sample of boiled milk, the results of which are recorded in
Table V—
TABLE V.—Results of Successive Cultivations of a Specimen of Boiled Milk.
Nature of
specimen.
Source, and
Date when
obtained
and boiled.
No. of
plate
cultiva-
tion.
Time at which set.
RESULTS.
Milk once boil-
ed in a steri-
lised tube;
reaction neu-
tral.
Bazaar-
September
6th.
...
. . . . . . . September 7th. Quite fluid ; reaction neutral.
September 8th. Quite fluid ; reaction faintly acid.
September 9th. Separated into fluid and large, loose
coagulum of powdery flocculi ; reaction neutral,
very faintly alkaline on drying.
September l0th. Three distinct strata present ;
1st at the surface, a thick layer of cream ; 2nd, in-
termediate, a large amount of yellowish fluid ; 3rd
inferiorly, an abundant deposit of powdery, white,
flocculi ; easily diffused by shaking, when the milk
for a considerable time resumed a normal appear-
ance; reaction very faintly acid.
1 Immediately
after boiling;
0.03 C. C. in-
oculated.
September 7th. Quite sterile.
2 September 7th,
24 hours after
boiling ; 0.03
C. C. inocula-
ted.
September 8th. A pure and very abundant growth
of Bacillus subtilis in the form of innumerable,
discrete colonies, and a generally diffused, tough,
dry film extending over the whole surface.
3 September 9th,
72 hours after
boiling ; inocu-
lated by a
needle dipped
in sterilised
salt solution
inoculated by
one looped
needle from the
milk.
September 10th. A pure and crowded growth of
Bacillus subtilis.
Medical Officers of the Army of India.
25
canal will naturally regard it as the result of imperfect diagnosis; those, on the
other hand, who regard the presence of numerous Comma-Bacilli in the intestinal
contents of choleraic patients as a result, not a cause, of the morbid state, will
see in it a confirmation of their opinion. In any case, it is a phenomenon
calculated to excite a certain reserve in the acceptation of a belief in the absolute
value of the presence of cultivable Comma-Bacilli as specially diagnostic of the
nature of any case as a true case of cholera. If we place implicit faith in the
diagnostic value of the presence of cultivable Comma-Bacilli in the alvine dis-
charges, we must believe that the diagnosis of cholera in Calcutta is very imper-
fect, and that the registration of the prevalence of the disease is liable to be
correspondingly greatly in excess of its actual prevalence, but if we do not,
we can only see additional ground for scepticism in regard to the essential
causal relation existent between the presence of the Bacillus within the intes-
tinal tract and the development of choleraic phenomena in the organism.
Returning from this digression, it remains to consider the phenomena
manifesting themselves in cases where Comma-Bacilli have gained access to
samples of milk. Three distinct series of experiments specially relating to these
were carried out. In the first the results following the access of Comma-Bacilli
to samples of milk of ordinary quality, as represented by the milk commonly
procurable in the Calcutta bazars, were the special subject of investigation; in
the second the enquiry was directed to the phenomena presenting themselves in
cases where the common normal lacteal schizomycetes had been partially re-
moved by means of boiling, so that the intrusive Comma-Bacilli had no longer
to contend with any living elements in the milk save Bacillus subtilis; in the
third, the effects manifesting themselves in cases where the milk, in place of
having its schizomycete population merely reduced, as in the previous series of
experiments, has had it absolutely abolished ere inoculation of Comma-Bacilli
—in other words the behaviour of sterilised milk as a medium for Comma-Bacilli
— w a s p a r t i c u l a r l y e n q u i r e d i n t o . T h e r e s u l t o f t h e s e e x p e r i m e n t s a r e s u m m a r i s e d
in the three following tables.
The first of these, Table VI, deals with the phenomena attendant on
the introduction of large masses of Comma-Bacilli into common samples of milk,
which had not been exposed to any special preliminary treatment.
E
26
Scientific Memoirs by
TABLE VI.—Results of inoculation of specimens of unboiled milk with
Comma-Bacilli.
No. of
speci- men.
Nos. of corre-
sponding spe-
cimens in
Tables I & II.
Nature and
treatment of
specimen.
Date
of
inocu-
lation.
Nature of
cultivations.
Date
at
which
set.
RESULTS.
I.
II.
I
XIV XVII Milk in a steri-
lised tube ;
inoculated
with 0.059 of
sterilised salt
solution full
of Comma-
Bacilli from
a tube of 48
hours' growth
15th
July.
......
. . . 16th July. Coagulated, but not fully
separated into coagulum and fluid ;
reaction permanently acid ; smell
caseous.
17th July. Fully coagulated and
separated ; reaction strongly and
permanently acid.
19th July. Reaction violently acid
smell very faint.
1. Plate ; inocu-
lated by a loop-
ed needle.
16th
July.
17th July. Smell mouldy, non-
choleraic ; crowded with colonies
of the common, lacteal, oval and
straight Bacilli ; no colonies of
commas to be found.
2. Plate ; inocu-
lated by a com-
mon needle.
17th
July.
18th July. Smell mouldy ; almost
a pure cultivation of the common,
small, straight, lacteal Bacilli ;
Bacillus subtilis present in very
small amount and apparently in
the form of a thin diffused surface
growth.
3. Plate ; inocu-
lated by a com-
mon needle.
19th
July.
21st July. Only two colonies, and
these both of large, lacteal Micrococ-
cus.
I I . . .
... Milk in a steri-
lised tube;
inoculated as
above with
0.03 solution
containing
Comma-Ba-
cilli of 48
hours' growth.
20th
July.
......
... 21st July. Coagulated and partly
separated ; reaction acid.
23rd July. Reaction strongly acid.
1. Plate ; inocu-
lated by a
needle dipped
in sterilised
salt solution,
inoculated by
a looped needle
from the milk.
21st
July.
22nd July. An abundance of
colonies present ; 3 distinct species ;
1st bluish and flattened, consisting
of the common, small, straight,
lacteal Bacilli ; 2nd, prominent and
yellowish, consisting of common, lac-
teal, oval Bacilli ; 3rd, two colonies
in the form of greenish yellow su-
perficial patches, consisting of
minute, straight Bacilli.
2. Plate ; inocu-
lated as above.
23rd
July.
24th July. Numerous colonies of
common, lacteal, oval Bacilli and
Micrococcus.
III XV XVIII Milk treated
as above.
25th
July.
......
... 26th July. Exactly in the same con-
dition as a corresponding non-in-
oculated specimen of the same
milk ; in very soft coagulum but
not yet separated ; reaction per-
manently acid.
No. of
speci-
men.
Nos. of corre-
sponding spe-
cimens in
Tables I & II.
Nature and
treatment of
specimens.
Date
of
inocu-
lation.
Nature of
cultivations.
Date
at
which
set.
RESULTS.
I.
II.
27th July. Fully coaguated and
separated ; reaction strongly acid.
28th July. Reaction violently acid.
1. Plate; inocu-
lated by a
needle dipped
in sterilised salt
solution inocu-
lated by a loop-
ed needle from
the milk.
26th
July.
27th July. Smell musty, non-
choleraic ; crowded with colonies of
the common lacteal Schizomycetes ;
not a trace of Comma-Bacilli.
2. Plate ; inocu-
lated as above.
27th
July.
28th July. Numerous colonies of
common lacteal schizomycetes ; not
a trace of Comma-Bacilli.
IV IXVI XIX Milk in a steri-
lised tube ;
i n o culated
with 0.03
solution, con-
taining Com-
ma-Bacilli o
f 4 8 h o u r s '
growth.
29th
July.
. . . . . .
. . . . . . 30th July. Exactly like its corn-
panion, non-inoculated tube ; re-
action acid ; slightly frothed ; at
first in soft loose coagulum, after-
wards undergoing separation.
1 . P l a t e ; i n o c u -
lated as above.
30th
July.
31st July. Exactly like its com-
panion plate of non-inoculated
milk ; smell mouldy ; numerous
colonies of common lacteal schizo-
mycetes ; no Comma-Bacilli.
V XVII . . .
" 3r
Aug.
. . . . . .
. . . 5th July. Strongly acid ; fully coa-
gulated and separated.
1. Plate ; inocu-
lated as above.
4th
July.
5th July. Smell mouldy ; numerous
colonies of common lacteal schizo-
mycetes ; none of Comma-Bacilli.
2. Plate ; inocu-
lated as above.
5th
July.
6th July. An abundant crop of
colonies of common lacteal schizo-
mycetes ; no Comma-Bacilli.
VI XVIII XX Milk treated
as above,
save that the
Comma-Ba-
cilli were de-
rived from a
cultivation of
only 24 hours'
standing.
15th
Aug.
. . . . . .
. . . . . . 16th August. Exactly in the same
condition as its companion speci-
men of non-inoculated milk ; strong-
ly acid ; coagulated, and partly
separated ; somewhat frothed at the
top.
1. Plate ; inocu-
lated as above.
16th
Aug.
17th August. Exactly like a plate-
cultivation of the same milk set on
the 15th previous to inoculation,
save that the number of colonies of
common lacteal schizomycetes is
considerably greater ; not a trace of
colonies of Comma-Bacilli.
The amount
of the solution
containing
Comma-Ba-
cilli was equal
to about
0.0029 of the
bulk of milk.
Medical Officers of the Army of India.
27
The six experiments recorded in this Table were practically identical in
nature and results. The actual quantity of fluid containing Comma-Bacilli intro-
duced into the milk varied slightly in the different experiments, and so did the
E2