NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
PLATE
I
[ Ind. J. Vet. Sci. & Anim. Husb., Vol. XI, Part I
EXPLANATION OF PLATE
All figures were drawn with the aid of a camera lucida. Figs. 1-6 were made
from air-dried smears, stained with May Grunwald-Giemsa, and figs. 7-9 were drawn
from sections of spleen which were stained with Heidenhain's iron-haematoxylin and
chromotrop 2 R. In fig. 8 the nucleus of the host cell is omitted. The figures are
2300 times the actual size.
Fig. 1
a
b
c
d
e
Figs. 2 and 3
Fig.4a
b
Fig. 5a
b
Fig.6
Figs.7-9
. . Extracellular form of the parasite seen in smears of spleen and
liver
. . Form showing loose nuclear structure, probably preparing for
division
. . Youngest form with condensed nuclear structure
. . An elongated form with chromatinic material along the periphery
of the nucleus
. . A young form with loose nuclear structure
. . A rounded parasite showing enlarged nucleus and the karyosome
. . Intracellular groups of parasite. Some of them are seen in the
process of binary fission
. . Four parasites in close contact with each other
. . An elongated form with its nucleus already divided into two
. . An extracellular form of parasite showing two stained dots at
either pole and its nucleus preparing for division
. . An extracellular parasite with its nucleus in metaphase
. . An intracellular parasite with two stained dots at either pole.
Its nucleus is in the resting phase
. . Intracellular forms as seen in sections. Fig. 8 shows the nuclei
of some of the parasites in the process of divisions. In figs. 7
and 9 note the central dot-like karyosome in the nucleus
H. N. RAY AND K.
RAGHAVACHARI
31
TABLE I
Temperature, frequency of B. gibsoni and anæmic changes in the spaniel
which on post-mortem examination also revealed the presence of T.
canis in its system
Days
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
Tempera-
ture (°F)
101.6
101.6
102.0
102.2
102.0
101.8
101.6
101.8
102.4
101.8
104.4
105.8
105.4
105.2
104.6
104.2
103.8
104.0
103.8
99.6
98.0
Frequency
of B.
gibsoni
..
..
rare
„
„
„
+
+
rare
„
„
+ + +
+ +
+ + +
+ + +
+ + +
+ + +
+ +
rare
v. rare
„
Anæmic
changes
..
..
..
..
..
..
..
..
..
..
..
..
slight
„
+ +
slight
„
+ +
+ + +
+ + +
+ + +
Remarks
Inoculated intravenously with 5 c.c. of
virulent blood
Feeds well
Do
Do
Do
Do
Do
Do
Do
Do
Do
Do
Do
Do
Do
Do
Do
Feeds fairly
Feeds fairly and dull
Do
Off-feed, died during the night
On the twelfth day the temperature rose to 105. 8°F and its blood was swarming
with B. gibsoni. Anæmic changes in the blood also appeared on this day
and these persisted until the animal died. Three days before death, however,
the parasites were rarely seen in its peripheral circulation and the dog showed
signs of inappetence. It died on the twenty-first night. The organ smears
revealed very few Babesia gibsoni parasites while fairly numerous Toxoplasma
canis were seen in various stages of development. Young extracellular forms
of the parasites measuring 2.4µ in length and 1.5-2.5µ in breadth were
seen in all the organ smears examined (Plate I, fig. 1, b); some of these
extracellular forms showed signs of nuclear division and in this stage they
measured 4.6µ in length and 2-4µ in breadth (Plate I, figs. 1, a, c, d and
4b). Intracellular forms residing in the cytoplasm of the monocytes were
seen in large numbers in the smears made from the spleen (Plate I, figs. 2
and 3). Single intracellular and extracellular individuals showing signs of
nuclear division were met with and, in a few instances, two pink stained
dots were seen, situated at either pole of the dividing nucleus (Plate I,
figs. 5a, b and 6). It is possible that these dots represented the centrosome,
C
32
Toxoplasma canis Infection in a Spaniel
because in one case a strand of fibres could be seen connecting the two dots
while the dividing nucleus remained in the equatorial region. In the mono-
cytes which contained groups of eight or sixteen parasites one could often
see signs of nuclear division in some of the more well-defined individuals
(Plate I, figs. 2, 3 and 8). In none of the preparations could we see a
mass of cytoplasm, with nuclei embedded in it, suggestive of schizogony.
Monocytes containing about thirty or more parasites were also seen and
in these, too, on careful examination, one could see some of the parasites in
the stage of binary fission. From these observations it is considered that
schizogony is absent in Toxoplasma and that the only method of reproduction
so far known is binary fission.
In sections of liver, evidence of marked cellular degeneration was present
and embedded in this mass of degenerating cells were large numbers of extra-
cellular parasites. A proportion of the monocytes in the sections also showed
parasites and in such cases the cytoplasm of infected cells had developed a
vacuole within which the parasites were crowded (Plate I, figs. 7-9). The
nucleus of the parasite exhibited a vesicular appearance with a central karyo¬
some (Plate I, figs. 7 and 9). Sections of the spleen also showed similar
cellular degeneration. In the sections of the lungs and kidney, however,
the parasites were not present to the same extent as in the liver and spleen.
SUMMARY
The occurrence of Toxoplasma canis in a pure-bred spaniel which had
succumbed to experimental Babesia gibsoni infection is reported. Stages
of division encountered in this parasite were suggestive of binary fission and
not of schizogony. A brief resumé of the literature on toxoplasmosis in dogs
is given.
ACKNOWLEDGEMENT
It is our pleasant duty to express our grateful thanks to Dr F. C. Minett
Director, and to Capt. S. C. A. Datta, Officer-In-charge, Veterinary Zoology
Section of the Institute, for kindly going through the manuscript and for
helpful suggestions.
REFERENCES
Blanc, G. (1917). Bull. Soc. Path. Exot. 10, 379
Boez, L. (1921). C. R. Soc. Biol. 85, 479
Carini, A. (1911). Ibid. 4, 518
Carini, A. and Maciel, J. (1913). Ibid. 6, 681
Laveran, A. (1913). Ibid. 6, 294
MacHattie, C. (1938). Trans. Roy. Soc. Trop. Med. & Hyg. 32, 273
Mello, U. (1910). Bull. Soc. Path. Exot. 3, 359
Wenyon, C. M. (1926). Protozoology, Vol. 1
—(1939).
Trans. Roy. Soc. Trop. Med. & Hyg. 33, 10
Yakimoff, W. L. and Kohl-Yakimoff, Nina. (1911). Bull. Soc. Path. Exot. 4, 617
THE DETERGENT EFFICIENCY OF SODA ASH, WOOD
ASH AND MUD ON BRASS AND TINNED STEEL
MILK UTENSILS
BY
H. C. VERMA, D. L. PAUL AND ZAL. R. KOTHAVALLA
Imperial Dairy Institute, Bangalore (S. India)
(Received for publication on 21 October 1940)
T HOROUGH cleanliness of dairy utensils is of great importance. How
to clean these utensils and render them sterile soon after use at the lowest
possible cost forms one of the main problems of the dairyman today. Different
practices are adopted for this purpose in different countries depending on the
conditions, the resources of the men in the trade and several other factors.
The practice with dairies run on modern lines is that, after the preliminary
scrubbing and cleaning is carried out, the utensils are sterilized either with
live steam, boiling water or dry heat. The cleansing agents which are popular-
ly known as ' washing powders ' are of various kinds, namely soap powders,
neutral soda, soda ash, causticized ash, scouring powders and trisodium phos-
phate. A solution of these powders possesses the properties of emulsifying
and saponifying fat and dissolving casein. Some powders also possess abra-
sive properties.
In rural India the use of washing powders is practically unknown because
this cannot be afforded by small milk producers. The practice throughout
the country is to scour the utensils with cheap materials such as wood ash or
fine mud. Scouring is mostly done on the wet utensils. After the utensils
have obtained the necessary degree of polish they are washed with clean
water and exposed to strong sunlight. During the rainy season, instead of
exposure to sunlight, the utensils are inverted for a short time over an open
fire or exposed to smoke. This practice has come down from time im-
memorial and is not without some technical significance. Its effect lies in the
abrasive and chemical action of mud and ash and in the bactericidal action of
sunlight or heat. The apparent primitiveness of the method, however, casts
doubt on its efficiency and it was to determine this efficiency and to compare
the method under conditions such as prevail in India with that followed by
dairies adopting western practices that this investigation was undertaken.
Much experimental work has been done by foreign workers on ascertain-
ing the efficiency of the detergent methods followed in western practice using
the reagents mentioned above. The degree of the detergent efficiency of each
reagent has been determined comparatively by the amount of reduction brought
about in the number of bacteria. Incidentally, it may be mentioned that the
term sterilization as is commonly employed in cleaning dairy equipment
does not always imply absolute bacterial sterility. It means sterility to a
degree good enough for practical purposes of handling milk.
Prucha, Weeter and Chambers [1918] found that 170 washed and un¬
steamed milk cans contained bacteria sufficient to contribute 128,000 per ml.,
while 91 washed and steam-sterilized cans would have added only an average
33
C 2
34
Detergent Efficiency of Soda, etc., on Milk Utensils
of 23,000 bacteria per ml. had the cans been filled with milk. In making a
comparative study of the collective influence of 81 steam-sterilized cans with
117 unsterilized cans in which milk was handled, they found that the former
added 6,800 organisms and the latter 285,600 per ml. Ayres, Cook and
Clemmer [1918] found that 60 samples from sterilized pails showed an average
of 6,300 organisms with a maximum of 21,500, while 59 samples from unsteri-
lized pails gave an average of 73,300 per ml. with a maximum of 284,000.
Levine, Peterson and Buchanan [1927] found that sodium hydroxide, sodium
carbonate and trisodium phosphate possess germicidal properties. Mudge and
Lawler [1928] have also found that alkali solutions possess germicidal
property.
Determining the efficiency of cleaning utensils by the traditional method
followed in India by methods used by foreign workers does not appear to have
been done in this country. It was felt that this kind of work would be
instructive. It was, therefore, decided to investigate the detergent efficiency
of the materials used for cleaning dairy utensils in this country, namely soda
ash, wood ash and mud on brass and tinned steel utensils. Since brass utensils
are commonly used in rural areas and tinned utensils in the handling of milk
in dairies, these two materials were selected. The investigation is not claimed
to be exhaustive and complete but it is hoped that the results will illustrate
the values of indigenous cleaning methods.
EXPERIMENTAL
(a) Preparation of utensils.—Four utensils of brass and four of tinned
steel were used. In order to make a fair comparison, the shape, constructional
features and capacity (1 litre) of the utensils were the same. All utensils were
washed and steam-sterilized in a uniform manner. Five hundred ml. of well-
mixed fresh milk were placed in each utensil and incubated at 37°C. to bring
about souring. The utensils were emptied of coagulated milk, rinsed with
500 ml. of water, scrubbed with sterilized coconut coir and water so as to remove
traces of soured milk and finally rinsed out again with 500 ml. of water.
(b) Procedure.—The utensils were then divided for treatment as follows :
Brass No.
1
2
3
4
Tinned steel No.
1A
2A
3A
4A
Material used for cleaning
Control
Wood ash
Mud
Soda ash
The control utensils were scrubbed with sterilized coconut coir only and
rinsed with water ; the other utensils were treated similarly using the respec-
tive cleaning agents. All the utensils were then placed on a drying rack and
exposed to the sun for 30 minutes.
(c) Determining the bacterial counts.—The average bacterial count of the
water used for washing the utensils was 75 per ml. and tests for coliform
organisms, Streptococcus faecalis and Clostridium welchii showed negative
results in both 1 ml. and 10 ml. samples. The bacterial counts and presump-
tive coliform tests of all the utensils were done. For the bacterial counts,
H. C. VERMA, D. L. PAUL AND Z. R. KOTHAVALLA 35
50 ml. of sterile water was used for each control utensil and 75 ml. for
ach treated utensil. Standard nutrient agar was used for the plating and
MacConkey's broth for the presumptive coliform tests. Plating in all cases
was done with 1 ml. direct in triplicate and from this result the bacterial count
of each utensil was calculated. The plates were incubated at 37°C. for 72 hours.
Presumptive coliform tests were done with 1 ml. and 10 ml. in duplicate at
37°C. for 72 hours.
Each cleansing treatment under discussion was repeated 20 times in both
series. The results obtained are given in Tables I and II.
TABLE I
Results of cleansing treatments
Expt. No.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
Presumptive coli-
form
positive
cases
Brass series
Number
1
(Con-
trol)
6,900
6,750
7,500
17,250
16,500
15,000
19,500
18,450
16,800
13,650
14,400
7,050
13,650
10,800
16,650
9,750
10,500
7,200
9,000
11,700
5
(Wood
ash)
3,750
1,650
2,850
1,875
1,950
1,875
3,150
3,450
1,200
2,775
1,650
1,275
3,225
3,525
1,875
2,475
1,950
2,250
1,800
2,850
1
3
(Mud)
6,000
2,250
3,600
4,875
4,500
5,250
5,525
2,700
2,250
1,725
2,550
2,175
5,700
6,600
3,600
3,000
1,800
2,700
2,175
5,600
4
4
(Soda
ash)
2,775
1,500
1,950
1,725
2,250
1,875
1,350
1,275
1,650
1,050
1,350
1,200
2,050
1,350
1,200
1,875
1,125
975
1,200
1,050
Nil
Tinned steel series
Number
1A
(Con¬
trol)
16,500
9,900
12,000
45,000
60,000
75,300
20,000
75,000
30,450
15,750
62,400
16,500
67,500
50,800
49,500
12,750
13,500
12,300
14,250
16,200
8
2A
(Wood
ash)
7,200
2,850
2,850
3,450
1,200
3,325
1,650
1,125
2,250
2,170
1,800
1,200
2,500
1,800
3,600
1,950
1,425
1,295
2,170
1,875
1
3A
(Mud)
5,400
4,500
2,850
12,750
2,475
4,970
5,700
4,875
1,875
8,250
9,000
6,700
7,600
4,250
4,600
2,525
2,925
3,750
4,200
3,000
5
4A
(Soda
ash)
3,750
2,700
1,720
1,500
975
2,175
575
750
225
1,220
1,200
1,350
1,960
1,550
2,680
900
1,050
600
1,200
1,125
Nil
Table I shows that on comparing with bacterial counts of the control
utensils, a considerable reduction in counts in both series has been brought
about and that the amount of reduction varies with cleaning material used.
As regards both reduction of plate count and destruction of coliform organisms,
soda ash was the most effective and next came wood ash and lastly mud.
36
Detergent Efficiency of Soda, etc., on Milk Utensils
TABLE II
Results of cleansing treatments
(Digest of Table I showing the results more clearly)
Particulars
No. of bacteria :—
Maximum
Minimum .
Average .
No. of cases :—
Below 1000 bac¬
teria
1000 to 3000
3000 to 4000
over 4000 .
Presumptive
coliform posi-
tive
Order of merit
Brass series
Number
1
(Con-
trol)
19,500
6,750
12,450
. .
. .
. .
. .
5
..
2
(Wood
ash)
3,750
1,200
2,370
0
15
5
0
1
2
3
(Mud)
6,600
1,725
3,730
0
10
0
10
4
3
4
(Soda
ash)
2,775
975
1,540
1
19
0
0
0
1
Tinned steel series
Number
1A
(Con¬
trol)
75,300
9,900
33,780
..
..
..
..
8
..
2A
(Wood
ash)
7,200
1,125
2,330
0
17
2
1
1
2
3A
(Mud)
12,750
1,875
5,110
0
6
1
13
5
3
4A
(Soda
ash)
3,750
225
1,460
6
13
1
0
0
1
DISCUSSION AND CONCLUSIONS
The high bacterial counts of the control utensils suggest that proper clean-
ing is necessary in order to keep the bacterial counts of the milk kept in them
as low as possible.
There is a difference between the bacterial counts of the milk in the
control utensils made of brass and tinned steel. Thus, the former contained
12,450 and the latter 33,780 organisms per ml. This is probably due to the
bactericidal action of the copper or the brass on bacteria (objodynamic action).
The results stand out in favour of soda ash treatment which is recognized
as suitable for cleaning purposes. Wood ash treatment did not give results
as good as soda ash but closely approached it. Wood ash contains calcium
oxide, hydroxide, carbonate and phosphate and also much potassium and
sodium carbonate and other alkali salts. The alkali carbonates and the
H. C. VERMA, D. L. PAUL AND Z. R. KOTHAVALLA 37
scouring properties of the other constituents of wood ash contribute to this
detergent action. Wood ash is thus a valuable cleaning agent and its cost
is negligible. The mud treatment takes third place but still shows a con-
siderable effect on lowering the bacterial count.
SUMMARY
An investigation of the cleaning powers of soda ash, wood ash and mud
on brass and tinned steel milk utensils has shown that all possess detergent
properties descending in efficiency in the above order.
ACKNOWLEDGEMENT
The authors are thankful to Dr W. L. Davies, D.Sc., Ph.D., F.I.C., N.D.A.,
Director of Dairy Research, for his valuable suggestions.
REFERENCES
Ayres, S. H.; Cook, L. B. and Clemmer, P. W. (1918). U. S. Dept. Agric. Bull. 642
Levine, M. ; Peterson, E. G. and Buchanan, J. H. (1927). Ind. and Eng. Chem. 19, 1338
Mudge, C. S. and Lawler, B. M. (1928). Ind. and Eng. Chem. 20, 378
Prucha, M. J.; Weeter, H. M. and Chambers, W. H. (1918). Ill. Agric. Expt. Sta.
Bull. 204
A NOTE ON ENCEPHALITOZOON CUNICULI INFECTION
IN A RABBIT*
BY
H. N. RAY, M.Sc., PH.D. (LOND.)
Systematic Protozoologist
AND
K. RAGHAVACHARI, G.M.V.C.
Veterinary Inspector
Imperial Veterinary Research Institute, Mukteswar
(Received for publication on 23 September 1940)
(With Plate II)
E NCEPHALITOZOON CUNICULI is known to produce spontaneous
encephalitis [Levaditi, Nicolau and Schoen, 1924] and nephritis [Smith
and Florence, 1925] in the rabbit. Levaditi and his co-workers found the
organism in the brain and kidney of the rabbit and were not only able to infect
rabbits but also rats, mice and dogs. Wright and Craighead [1922] demons-
trated this parasite also in the spleen, liver and myocardium. Its presence
in the urine suggested to them the possible mode of infection, and later Levaditi
and his collaborators were actually able to demonstrate the infectivity of the
urine. Wenyon [1926], in reviewing the literature on the subject, expressed
uncertainty regarding the systematic position amongst protozoa of this
parasite, although Levaditi and his co-workers believed it to be a micro¬
sporidian. They demonstrated certain structures in the parasite which, they
thought, were pansporoblasts, which ultimately gave rise to spores. They,
however, failed to demonstrate a polar capsule and a filament in the spore.
According to Wenyon [1926] their work required confirmation. It may be
mentioned that microsporidia have not so far been recorded in any species
of warm-blooded animal. In this article, the occurrence of E. cuniculi is
recorded for the first time in India and a description is given of the various
stages in the development of the parasite, as met with in kidney smears from
an infected rabbit. Certain of these stages, when considered in their proper
sequence, are suggestive of a much closer affinity of this organism with the
microsporidia than with any other group of protozoa.
It should, however, be mentioned that the minute size of this organism
makes elucidation of its different stages a matter of considerable difficulty.
We do not claim to have seen the polar filament, but anterior to the uninucleate
sporoplasm we have observed a clear area which closely resembles the polar
capsule of many known microsporidia. The spore would appear to have only
a single capsule, each pansporoblast giving rise to a single spore. We, how-
ever, refrain from placing it under any known family or genus of microsporidia
until a more detailed study of this parasite has been carried out.
*Paper presented to the 27th annual meeting of the Indian Science Congress, 1940.
38
LATE
II
[ Ind. J. Vet. Sci. & Anim. Husb., Vol. XI, Part I
EXPLANATION OF PLATE
All figures, except fig. 9, were drawn from smears with the aid of a camera lucida.
Fig. 9 was drawn from a section of kidney, which was fixed in Duboscq and Brasil's
modification of Bouin's fluid and stained with Heidenhain's iron-haematoxylin and
chromotrop 2 R. The smears were either air-dried or fixed in osmic vapour and stained
with May Grunwald-Giemsa
Fig. 1a ..
b, c, cl ..
d, e, f ..
g, h, i ..
Fig. 2 ..
Fig. 3 ..
Fig. 4 ..
Figs. 5, 6a and 6b
Figs. 7 and 8 ..
Fig. 9 ..
Fig. 10 ..
Fig. 11 ..
Fig. 12 ..
Fig. 13 ..
Fig. 14 ..
Planont—found free in a smear
Planont—showing binary fission
Planont—showing nuclear components for four, three and five
individuals respectively
Planont—showing meronts in chains (× 2300)
Urinary tubule cells showing planonts (×2300)
A tubule cell showing schizonts (×1700)
Tubule cell showing planonts and meronts (×1700)
Groups of meronts (×1700)
Tubule cell showing early pansporoblasts (fig. 7, ×1700 ; fig. 8,
×2300)
Section showing the lumen of the tubule obliterated by the
presence of planonts and meronts (×1166)
Formation of spores in the cytoplasm of a tubule cell (× 2300)
Group of tubule cells showing fully-formed spores (×875)
A portion of the same, highly enlarged (×2300)
Tubule cell showing large number of pansporoblasts, each showing
five nuclei (×1700)
A fully-formed spore. (Note the vacuolated area at one pole)
(×2300)
40
Encephalitozoon cuniculi Infection in a Rabbit
meronts resulting from schizogony underwent further schizogony or divided
by binary fission was, however, very difficult to ascertain.
The next stage encountered in our preparation may be interpreted as
pansporoblasts. The cytoplasm of a pansporoblast became loose in structure
and the nucleus was often observed to have broken up into five very small
nuclei (Plate II, figs.7, 8 and 13). Owing to the minute size of the nuclei,
it was extremely difficult to ascertain the means by which they were utilized
in the process of forming a spore. In some of the preparations in which the
spore wall was not fully formed, we could frequently see one or two nuclei
situated at one of its poles, and it is possible that these two nuclei formed
the spore wall (Plate II, fig.10). In other preparations we could often
see a nucleus situated at the margin of the vacuole which was placed anterior
to the sporoplasm. Possibly this nucleus is used in the formation of the polar
capsule (Plate II, fig.10). The sporoplasm at an early stage often showed
two nuclei but in the mature spore we could always find a single nucleus.
The sporoplasm was seen to occupy one end of the spore, which at the other
end exhibited a vacuolated area very much resembling the polar capsule
(Plate II, figs. 11, 12 and 14). It may, however, be mentioned that no
structure resembling a polar filament was encountered inside this so-called polar
capsule. It was also observed that a single pansporoblast gave rise to a single
spore. The host cell was often seen to be packed with numerous spores and
in such cases cytoplasmic hypertrophy was noted, the nucleus remaining
unaltered. A fully formed spore is ovoid in shape and measures 2-2.5µ in
length and 1-1.5µ in breadth.
SYSTEMATIC POSITION
Regarding the systematic position of this parasite, it is not possible to
express a definite opinion at this stage of our observations but certain charac-
teristics of the parasite, as enumerated below, would appear to show that,
as already mentioned, it has a closer relationship with the microsporidia than
with any other member of the protozoa :—
(1) the organism has an early amoeboid stage;
(2) multiplication takes place either by binary fission or schizogony ;
(3) the resisting phase or spore is formed from a pansporoblast, which
initially is a single nucleated individual but in the process of
formation of the spore breaks up into several nuclei;
(4) at one end of the spore there is a vacuolated area which has a very
close resemblance with the polar capsule of other microsporidia.
As, however, no polar filament was encountered inside this polar capsule
it is not possible to assign this parasite to any particular group of microsporidia.
TRANSMISSION EXPERIMENT
Two rabbits were inoculated, one subcutaneously and one intraperi¬
toneally, with a suspension of infected kidney tissue. These animals were
kept under observation for over two months, but they failed to become
infected.