NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
Appendix I.
417
0
s*;
9
m
11
12
13
\¿
10
17
18
Name.
Rhagia .
Gum) r ana
Jaikishen
>> •
Chandra.
Bhaisaki.
Heroli .
Deluli
Jaikishen
Gumyana
ALMORA ASYLUM.
AGE.
S
35
35
35
35
3D
u,
30
36
32
45
••
S
£
A
>>
M
>J
j>
A
„
>>
M
A
1rs
ted.
CU.«
2 2
19
29
29
II
2O
15
12
29
19
Condition of
skin where
vaccinated.
Normal
Sensation di-
minished.
Tubercula-
ted (ear).
Normal
>>
Sensation di-
minished.
Normal
Tubercula-
ted (ear).
Sensation di
minished.
VACCINA-
TION.
Condition of
vesicle.
Normal
j»
•
>>
•
>»
•
tj
•
j>
•
Si
'
Crust from
broken
vesi.de.
Crust from
broken
vesicle.
Date of
vacci-
nation.
24-3-9!
24-3-91
25-3-91
25-3-91
28-3-91
2S-3-91
2S-3-91
28-3-91
25-3-9I
24-3-91
Date of
taking
ymph.
30-3-91
30-3-91
1-4-91
I-4-91
1-4-91
1-4-91
3-4-91
6-4-91
6-4-91
6-4-91
isses.
over-gl£
'S
No.
2
1
2
2
2
2
2
2
2
2
Result of Micros-
copic Examination.
1. No Leprosy Ba-
cilli : epithelial
cells with nuclei.
2. Epithelial de-
bris, lymph cells,
rods, and micro-
cocci. No Le-
prosy Bacilli.
1 Wholly nega-
tive.
1. One or two ba-
cilli takinir fuch-
sin. 2. Debris:
no bacilli.
t. One ort%vo bad 1 -
1}'stained bacilli.
2. Debris : no
bacilli.
1. Few small blue
cocci : no bacilli.
2. Debris : no ba-
cilli.
1. Debris: no ba-
cilli. 2. Debris.:
no bacilli.
, 1. Debt is : no ba-
cilli. 2. Debris:
no bacilli.
1. No Leprosy Ba-
cilli, 'z. No Le-
prosy Bacilli.
i. No bacilli. 2.
Few bacilli stain-
ed blue : no Le.-
prosy BacilH.
1. Numerous cocci
stained blue : no-
Leprosy Bacilli.
2. Numerous cocci
stained blue : no
Leprosy Bacilli.
2 E
4i8 APPENDIX TO REPORT OF THE LEPROSY COMMISSION;
d
19
20
21
22
23
24
25
26
27
2
Name.
Gulavi .
Bhuri .
Bhag-uli .
Ganguli
Dikvi .
Bachali .
Parmati
Naduli .
Gopuli .
Khimuli
ALMORA ASYLUM.
AGE.
SCxJ
0
0
26
50
4°4
0
40
5°3
C
5
S
1
A
99
91
99
99
,,>
99
$9
99
cd -u
v 0
«7
35
10
22
8
5
34
20
10
20
Condition of
skin where
vaccinated.
Ansesthetic
patch
Normal
»»
•
Sensation di-
minished.
Anaesthetic
patch.
j»
•
Sensation di
minished.
9i
•
>>
•
Sensation di
minished.
VACCINA-
TION.
Condition of
vesicle.
Normal
»
•
«
•
»»
•
>?
•
>>
•
Date of
vacci-
nation.
30-3-91
30-3-9'
30-3-91
30-3-91
31-3-91
31-3-9
30-3-91
30-3-9
30-3-9
30-3-9
Date of
:akingf
ymph.
4-4-91
4-4-91
4-4-91
6-4-91
6-4-91
6-4-91
6-4-91
6-4-91
6-4-91
6-4-91
i
a
-a
¿
0
'S
2
2
2
2
2
2
2
2
2
2
Result of Micros-
copic Examination,
1. Unstained ba-
cilli : no Leprosy
Bacilli. 2. Debris:
no Leprosy Ba-
cilli.
1. Blue rods: no
Leprosy Bacilli.
2. Numerouscoc-
ci anddiplococci :
no Leprosy Ba-
cilli.
1. Few blue rods r
no Leprosy Ba-
cilli. 2. Few blue
rods-: no Leprosy
Bacilli.
1. Group of vac-
cine cocci stained
blue : no Leprosy
Bacilli. 2. Few
bacilli
stained
blue : no Leprosy
Bacilli.
1. Short thick ba-
cilli taking- pink
stain : no Lepro-
sy Bacilli.
2.
No bacilli.
1. Debris : no Le-
prosy Bacilli. 2.
Debris : no Le-
prosy Bacilli.
1. Debris: no ba-
cilli. 2. Few ba-
cilli taking blue
stain : no Lepro-
sy Bacilli.
1. Numerous cocci
stained blue. 2.
Numerous leu-
cocytes : no Le-
prosy Bacilli.
1. Debris : no ba-
cilli. 2.
Few
cocci; no bacilli.
1. Rods of various
sizes : no Lepro-
sy Bacilli.
2.
Rods of various
sizes; no Leprosy
. Bacilli.
Appendix I.
419
Ô
z
3°3
1
32
33
34
35
36
37
3S
39
40
Name.
Himatna
Jasuli
Dhania .
Karakna.
Khimuli .
Partima.
Motiya .
Kamalua
Budhia .
Telegu .
Sundarsai
Gujia
.
ALMORA ASYLUM.
AGE.
60
25
25
iS
20
40
32
40
45
41
55
60
È
1
A
T
A
33
39
;>
>}
»5TJ
2 1)
<u u
12
19
4
13
13
10
IG
28
I
30
7
17
Condition of
skin where
vaccinated.
Sensation
diminished.
Normal
.
Anaesthetic
patch.
Sensation
diminished.
Normal
3J
•
»»
•
Sensation
diminished.
Anaesthetic
patch.
Normal
VACCINA-
TION.
Condition of
vesicle.
Normal
Purulent
and mixed
with blood.
Normal
Imperfectly
developed.
Purulent .
Normal
3>
•
33
•
Purulent
and mixec
with blood
Immature
vesicle anc
mixed witl
blood.
Normal
.
3»
•
Date of
vacci-
nation.
1-4-91
30-3-91
I-4-91
3-4-91
3-4-91
5-4-91
8-4-91
S-4-91
S-4-91
9-4-91
9-4-91
8-4-91
Date of
:aking
ymph.
7-4-91
7-4-91
7-4-91
10-4-91
10-4-91
11-4-91
13-4-91
13-4-91
13-4-91
13-4-91
13-4-91
13-4-91
jsses.
). ot cover gl;
2
2
2
2
2
2
2
2
2
2
2
2
Result of Micros-
copic Examination.
1. Debris : no Le-
prosy Bacilli. 2.
Debris; no Le-
prosy Bacilli.
1. Debris : no ba-
cilli. 2 Cocci
and diplococci :
no bacilli.
1. Cocci and bacilli
stained blue : no
Leprosy Bacilli. 2.
Cocci and bacilli
stained blue : no
Leprosy Bacilli.
1. Debris: no ba-
cilli. 2. Debris:
no bacilli.
1. Pus cells and de-
veloping- fibrous
tissue : no ba-
cilli. 2. Pus cells :
no bacilli.
1. Bacilli stained
blue and shreds
of tissue. 2. Ba-
cilli stained blue
and shreds of
tissue.
1. N umerous cocci :
no Leprosy Ba-
cilli. 2. No Le-
prosy Bacilli.
1. Debris : no ba-
cilli. 2. Debris :
no bacilli.
1. Debris : no ba-
cilli. 2. Debris :
no bacilli.
1. Cocci and bac-
teria: no Leprosy
Bacilli. 2. Cocci
and bacteria : no
Leprosy Bacilli. J ,
1. Cocci : no ba-
cilli. 2. Cocci :
no bacilli.
1. Debris : no ba-
cilli. 2. Debris :
no bacilli.
2 £ 2
42O
APPENDIX TO REPORT OF THE LEPROSY COMMISSION.-
41
42
43
44
45
Name.
Kalangit
Gumani .
Harna .
Maina .
Dhamia .
ALMORA i
AGE.
S
2S
2S
54
tú
60
g
Ut O
A
»
>!
))
2'S
cö tí
V .S
>m si
3
CO
30
30
12
\SYLUM.
Condition of
Slvlll Willie
vaccinated.
Normal
Anœsthetrc
patch.
» »
VACCINA-
TION.
Condition of
vesicle.
Normal
Purulent .
Normal
Crust of im-
perfect ve-
sicle.
Date of
vacci-
nation .
8-4-91
10-4-91
S-4-9»
9-4-91
9-4-91
Date of
taking-
1 Y m p h •
13-4-91
13-4-91
13-4-91
14-4-91
14-4-91
isses.
to
>
0
0
No,
2
2
2
2
2
Result of Micros-
copic Examination.
1. Bacteria: no Le-
prosy Bacilli. 2.
Bacteria : no Le-
prosy Bacilli.
1. Pus cells: no Le-
prosy Bacilli. 2.
Pus cells : no Le-
prosy Bacilli.
1. Debris : no ba-
cilli. 2. Debris :
no bacilli.
I. Debris : no ba-
cilli. 2. Debris :
no bacilli.
1. Large rods tak-
ing- blue stain :
no Leprosy Ba-
cilli. 2. Debris:
no bacilli.
V.—CULTIVATION
EXPERIMENTS. 29
Hansen, in his original paper, described both rod-formed bodies
(Bacillus Leprse) in tubercles, and jointed threads obtained in speci-
mens of blood which, when kept in a moist chamber, developed into
a mycelial growth, as probable specific microscopical features of true
leprosy. He also figured these granular mycelial masses, and spoke
of them as a culture, but it is almost certain that this must be looked
upon as a contamination of the specimen, and since the Bacillus
Leprae never occurs in blood, this growth could not have been a deve-
lopment of these. It is clear also that Hansen himself did not regard
the mycelium in this light.
The cultivation experiments of Neisser so are the earliest, and the
details may be given in his own words.
1. An unbroken tubercle was thoroughly cleansed with alcohol,
then removed with every possible precaution against foreign infection,
and finely divided up with needles made sterile in a flame. Particles
of this broken up material were employed in the following three
culture methods :—
{a) With blood serum in hollow-ground microscope slides. The
hollow spaces were very capacious and contained much air.
( 2!) ) The references available in India for this section are etccelingly few.
(*') Virchow's Archiv. 188!, 1SS6.
Appendix I.
421
(b) In test tubes, carefully cleansed by heat and acids, which
were partly filled with blood serum or with alkaline steri-
lized flesh-extract-solution.
(c) Under cover-glasses with blood serurn, in a space where
evaporation was checked. All these preparations were
kept permanently in a warm chamber at 35 o —39°C.
1. Cultivations from pus and juice of leprous nodule which were
protected in capillary glass tubes from accidental fouling. The tubes
Contained much air and were sealed.
Neisser describes three bacilli : first, movable rods which are
perfectly free from distortion of any kind ; second, motile rods with
a minute ball-like swelling at one or both extremities or in the
middle ; these rods are somewhat longer than the first variety and
frequently sharply bent ; third, bacilli which are characterised by
being distinctly longer and broader than either the first or second
kind ; each of these rods contains one to four clear unstained spaces
(vacuolation). It is possible, according to this observer, that the
breaking up of the rod into granules (kokkothrix) which lie in a row
adhering together because of the hull which invests the bacillus, is
a retrogressive metamorphosis, and that the bacilli of the second and
third kinds exhibit veritable spore-formation. Lastly, both in cultures
and in microscopical preparations of leprous tubercles which, after
experimental introduction, had been encapsuled in the peritoneal
cavity, the bacilli grew into a thread occasionally four times the
normal length, but in these no sign of anything which could be called
a spore-formation was evident.
From a careful examination of Neisser's figures, especially I and
Io, it appears not unlikely that, as he claims, a veritable culture was
obtained. Further, the three kinds of bacilli described by him are to
be seen in juice from a leper tubercle. Of the actual existence of
two—the first and third kinds—there can be no question, but the
relation of these to each other is not so clear ; both stain excellently
with the Koch-Ehrlich method and resist decolorization with acids ;
and also Gram's method clearly defines both of them, especially the
latter. Neisser's first kind is the normal Bacillus Lepræ; the second
we believe to be possibly a distortion, for a swelling upon a rod can
easily be caused to appear and disappear, as the bacillus slowly turns
under the cover-glass; and the appearance seen is figured on Plate
III, Fig. VIII ; the third form, as far as we can judge from his Figs.
7 and 13, we have frequently seen and have also, we believe, cultiva-
ted. Whether it is a bacillus possessing pathogenic power it is
impossible to say. This form, which it is very probable is the same
422
APPEND IX TO REPORT OF THE LEPROSY COMMISSION:
as that described by Neisser, we shall speak of as the vacuolated
bacillus In a later paper Neisser, 31 as the result of extended obser-
vations, inclines to the view that the vacuoles, or spaces in the
bacillus, are veritable spores, though in no instance has he seen an
isolated spore. The ball-like swelling in the second kind he does not
regard as a spore-formation. He also confirms the presence of the
hull of the bacillus, stating that it can easily be seen after staining a
preparation with watery dye. The presence of this hull is botanically
of interest ; it occurs also in the Bacillus Tuberculosis, 82 Pneumococ-
cus, 33 and several other forms.
Further culture experiments by Neisser consisted in using solid
blood serum and hard-boiled fowl or duck eggs as a nutrient medium,
and minute pieces of excised tubercle, or blood from a tubercle, as
material for inoculation. The culture was maintained at a tempera-
ture of 37°—38°C. The growth was exceedingly slow. Cultivation
in generations was not successful. Specimens from detached bacillary
islets on the medium yielded what the author held to be a genuine
cultivation of the Bacillus Lepræ.
Hansen 34 in 1882 described a culture on serum of bacilli which
were motile. He used methylene-blue as a stain, but according to
Neisser this will not even tinge the genuine Bacillus Lepræ. This is
not a strong argument against Hansen's culture, for in our hands
methyl-blue stains the Bacillus Lepræ, though great differences exist
in the samples of this dye, so that the discordant results of various
observers are conceivable.
It is impossible to be quite certain that a veritable cultivation of
the Bacillus Lepræ was obtained by Neisser. On Plate XII, 34 Figs.
1, 5, 6, and especially 10, have the appearance of true cultures, but
when it is considered that this observer wholly failed to cultivate the
bacillus in generations, an actual multiplication in fluids which pro-
bably did occur is all that can be conceded.
Manson, 35 some years before Hansen had discovered the Bacillus
Lepræ, attempted by incubation of leper juice to discover the then
unknown germ. Sealed capillary tubes were inserted into eggs which
were placed under a hen. Beyond the development of streptococci,
both free and in the zoogloea state, these experiments gave wholly
negative results.
( 31 ) Virchow's Archiv. 1886.
( 32 ) Lutz and Unna. Dermatol. Studien Heft I. 1886.
(33) Friedländer. Fortschr. d. Med. Bd. III, 1885.
( 34 ) Virchow's Archiv. 1881.
(35) Journal of the Leprosy Investigation Committee, No. 1, p. 40.
Appendix I.
423
Arning 36 states "that there is no difficulty whatever in growing
the Leprosy Bacillus, by simply allowing pieces of leprous tissue to
macerate in ordinary water, at ordinary temperatures, giving but
sufficient time ; but the resulting growths are not pure cultures in a
strict sense."
The introduction by Koch of solid nutrient media has, to a large
extent, caused nearly all observers who have attempted to cultivate
the Bacillus Lepræ to abandon cultivation in fluid media which in
the hands of Pasteur had led to many discoveries. In the history of
the attempted cultivation of the specific bacillus of leprosy, the great
majority of experiments have consisted in the removal of a piece of
leprous tubercle and the direct implantation of this upon a solid
nutrient surface, such as agar, glycerin agar, or solidified serum. 37
The successful growth of Bacillus Tuberculosis by this method is by
no means easy, and uniformly negative results with the Bacillus Lepræ
have been obtained by the vast majority of observers. 38 Quite recently
Sheridan Delépine and Slater 39 record numerous attempts to cultivate
the bacillus, which were made independently at two different periods
during the life of the patient On both occasions the results were
negative. The latter observer two years previously had attempted
identical cultivation experiments.
In 1887 40 Bordoni-Uffreduzzi described at length his culture-
experiments with the Bacillus Lepræ. He was as unsucessful as
previous observers in the direct implantation of a leprous nodule on
to a solid nutrient medium. The chief result of such a proceeding
was a growth of Streptococcus Pyogenes. He considered that the
Leprosy Bacilli which abound in leprous nodules are enclosed in
cells, and probably are not alive, though, if the view of Unna and
Kühne be accepted, the bacillary heaps lie outside the cells. In the
marrow of bones of a dead leper, the Italian observer found free
bacilli, and from this material he raised cultivations of the Bacillus
Lepræ on peptone-salt, glyceriue-serum ; it grows also upon glycerine-
agar. However, the bone-marrow has been examined by Kaurin 41
in many cases, and he never succeeded in finding the Bacillus Lepræ.
it does not appear that many observations on bone-marrow have
been made, but at any rate in the Haversian canals and spaces of the
( 36) Journal of the Leprosy Investigation Committee, No. 2, p. 117.
( 37 ) Fraenkel and Franke repeatedly failed to cultivate the Xerosis Bacillus in this manner
by direct implantation on solid media. Schreiber, however, succeeded in this direction.
(38) Among others cf. Trinidad Leper Asylum Reports, 1888.
( 39 ) British Medical Journal, May 23rd, 1891, page 1127.
(40) Zeitschrift für Hygiene, Vol, III, page 178. This important paper was unfortunately
not available in India,
( 4l ) Journal of the Leprosy Investigation Committee, Voi. 2, p. 63.
424 APPENDIX TO REPORT OF THE LEPROSY COMMISSION:
bones of the little finger, which presented typical mutilation, masses
of bacilli, both discrete, and lodged within undoubted lepra cells, have
been found by Delépine and Slater. 42 Sawtschenko 43 has also worked
out the histology of bone-leprosis.
Bordoni-Uffreduzzi describes the cultivated bacillus on peptone-
glycerine serum at a temperature of 35 o —37°C. as a light-yellow
taenia-form stripe with irregular edges along the needle-track. The
serum is never liquefied, and no growth ever occurs in the condensa-
tion water. On glycerine-agar a similar culture develops if a great
quantity of bacillary material be streaked upon the nutrient surface.
If a smaller amount be streaked, isolated lens formed colonies with
jagged edges and of varying size develop. On glycerine-agar plates,
both on the surface and deeply in the medium, colonies may be seen
with a power of 100 diameters, which are grey, net-like growths with
irregular edges. It is evident that this growth cannot be confounded
with cultures of Bacillus Tuberculosis. The isolated bacilli in form
and measurement resemble those of the tissues, but a striking pecu-
liarity is the frequent occurrence, at one or both ends of a bacillus, of
a swelling in which an arthrospore may be seen. This appearance
may also be recognised in the tissues, but it is an open question
whether this does not represent an involution-form. Isolated spores
or the development of a rod from such an arthrospore, have not been
observed. The bacillus is quite immotile. The tinctorial behaviour
of the cultivated Bacillus Lepræ is characterised, firstly, by remaining
absolutely unstained by alkaline methyl-blue, 44 in which it resembles
the bacillus of the tissues, and secondly, by staining with much
greater difficulty in Ehrlich's fuchsin, indeed, with about the same
facility as the Bacillus Tuberculosis does. The Bacillus Leprœ of the
tissues, however, stains much more readily than the latter. In a great
number of experiments on rabbits and other animals, Bordoni-Uffre-
duzzi wholly failed to establish the pathogenic nature of his cultiva-
tion.
Roux, Nocard, Chantemesse and Cornil 45 have failed to confirm
the above cultivation experiments, but Gianturco 46 has succeeded in
obtaining a culture which Bordoni-Uffreduzzi admits is absolutely
(42) LOC. cit.
(43) Sawtschenko über Osteomyelitis Leprosa. Centralblatt. f. Bact. und Parasitenkunde
Bd. V., 1889. According to Sawtschenko the vacuoles in lepra cells contain a liquid which
is secreted by the bacilli. The absorption of bones proceeds from lepra clusters in the
bone-marrow and in the dilated Haversian. canals.
( 44 ) Koch and Wesener have stained Bacillus Lepræ, and this can be confirmed with
great ease
\ill\
( 46 ) Abstract in Baumgarten's Jahresbericht, Vol. 11, ,8Sg.
Appendix I.
425
identical with his own. At a temperature of 37 o C, after seven days
a colony is developed of feebly motile bacilli, which are more slender
when grown on glycerine-blood-serum than when on glycerine-agar.
The bacilli also possess terminal swellings (arthrospores). Both
Campana 47 and Katz 48 using every precaution, have failed to culti-
vate the Bacillus Lepræ. The former observer, in the course of two
years, made about five hundred experiments and employed various
nutrient media and material from leprous tubercle. In consequence
of the total failure of these experiments he has thrown doubt on
the genuineness of the culture obtained by Bordoni-Uffreduzzi.
Extensive experiments which were made in Trinidad, with total
want of success, and consisting in the direct implantation of the
centre of a non-ulcerated leprous tubercle, taken with every possible
precaution, upon a nutrient solid medium, were repeated in India
with similar result. No culture was formed, but a staphylococcus
growth spread very slowly from the piece of tissue, and by the end
of ten weeks the implanted piece was obscured by this growth.
The following methods were adopted in our experiments :—
1. It had been ascertained that in blisters raised over unbroken
leprous tubercles, specific bacilli were frequently found.
It is also
well known that the pigment particles in tattoo-marks do not pass
into such a fluid. It was therefore hoped that by this means a separ-
ation could be effected between the living and dead bacilli within
the tubercle. Serum from blisters over the normal skin of the back
of a leper was received into sterile vaccine capillary tubes, and a
small quantity of the fluid of a blister raised over a tubercle was
added to this. The tube, which had an air-space for about one-third
of its length, was then sealed and maintained at the temperature of
the body.
2. Pieces of the centre of a leprous tubercle, removed with all pos-
sible precautions, were introduced into bouillon, and maintained at
the temperature of the room, which during the clay averaged 27°C.
3. An unbroken tubercle was thoroughly cleansed with carbolic
soap and perchloride solution. A deep puncture with a sterile needle
was made into the tubercle, and perfectly clear leper juice was ex-
pressed. This was received into sterile pieces of capillary tube, about
\ill\
long, and the tube and contents at once dropped into
bouillon. This was maintained at the temperature of the room.
In all the above instances microscopical examination showed that
( 47 ) Abstract in Baumgarten' s Jahresbericht, Vol. III, 1889.
( 48 ) Abstract in Baumgarten's Jahresbericht., Vol. III.
426
APPENDIX TO REPORT OF THE LEPROSY COMMISSION:
the material used as the source of cultures contained Leprosy Bacilli.
The results of these three methods will now be given in detail.
From what has been previously mentioned, it would appear that
the direct implantation of leprous material upon solid nutrient media
gives absolutely negative results. Among the causes of this it may
be suggested that the Bacillius Lepræ, like the bacillus of malignant
oedema, 49 is exquisitely anäerobic, or may be incapable of becoming
a facultative saprophyte, or that in the preparation of the piece of
leprous tissue, the hot knives used to make the section cause the
formation of a layer of coagulated material, through which the bacilli
cannot make their way, or, lastly, because the tissue contains for the
most part, only dead bacilli. The last conjecture appears most
probable, since the others admit of direct experiment.
If a section of skin be made through a leprous tubercle the bacilli
are found in quantities, and fluid expressed from the neoplasm abounds
in bacilli. The epidermis from the corneous to the pigmentary layer
in unbroken tubercles is wholly free from bacilli, while the cutis and
subcutaneous tissue is crowded with them. A blister raised over a
tubercle contains a few leucocytes and at times bacilli. In some cases
these latter are absent. We believed that the majority of these bacilli
were alive and capable of being cultivated. '
Cultivation Experiments with Blister Fluid.
Blisters were raised over tubercles on the ears, face and wrists
and at the same time over normal skin between the shoulder blades.
After avoiding all sources of fouling, sterilised vaccine capillary tubes
were charged with a minute quantity of fluid from a blister over a
tubercle, and also with seven or eight times the quantity of fluid from
a blister over the normal skin of the same leper. They were then
sealed. Each tube also contained air for about one-third of its length.
The dates of these experiments ranged from March 31st to May
13th. In all cases samples of the blisters, both over the tubercles
and on the back, were examined for the bacillus. Being at this time
without an incubator, the tubes were packed with a small thermometer
in a cylindrical box and carried in the axilla, a proceeding adopted by
Spallanzani in the last century when studying the action of the diges-
tive juices on food. A month later these tubes were examined and
resealed, and in several cultures were seen. A tube containing a
culture was now sterilised as far as was possible without injury to the
( 49 ) Discovered b\ Koch and first cultivated in bouillon with absence of oxygen by Pasteur.