NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
PRECIPITATION OF ANTIBODY FROM ANTI-RINDERPEST SERUM
155
and rinderpest virus are inoculated simultaneously into a number of susceptible
hill bulls, while for control a group of healthy bulls is inoculated with virus alone.
From the nature and degree of the reaction shown by the animal, the protective
value of the anti-serum can be approximately estimated.
As our tests were merely to obtain a preliminary indication of the value of
the methods used, in most cases only two bulls were used for each sample. The
period of observation was in all cases 15 days and our results based on the
observations made on the most susceptible animal in each case. These results
are tabulated below :—
Test results of samples 1 and 2.
Sample.
1
1a
1b
2
2a
2b
2c
2d
Controls
Dose of
serum
per 100 lbs.
15 c.c.
„ „
„ „
„ „
„ „
„ „
„ „
„ „
. .
Initial
condition.
Fair.
Rather poor.
Fair.
„
„
„
„
Rather poor.
Fair.
Final condition.
Improved.
Unchanged.
Worse.
Improved.
Unchanged.
„
Worse.
Improved.
Died.
Percentage potency based
on relative loss or
gain in weight.
100
60
22
85
66
64
28
79
. .
The results obtained with samples 1 and 2 furnish instructive indications
on the points raised earlier. In the case of sample 1 the potency of the euglobulin
fraction obtained by water precipitation is markedly superior to that obtained by
precipitation with ammonium sulphate. The difference in the potencies of the
two fractions is as great as to appear definitely beyond any experimental test
error. It is also to be noticed that with the water-insoluble protein the amount of
immune-body separation, although actually somewhat less, is of similar order to
that found by Felton in the case of anti-pneumococcus serum where better methods
of assay are available. The results obtained in the case of sample 2 give similar
indications and it seems, therefore that there exists a definite difference in the
antibody contents of the two euglobulin fractions, one obtained by the water-
precipitation method, and the other obtained by ammonium sulphate. It is of
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INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [III, II
interest to note here that there was no perceptible difference in the potencies of
the two fractions of insoluble proteins obtained at two different pH, namely, 6.8
and 6.4, and it may be mentioned that Felton also did not find much difference
in the potencies of water precipitated fractions of anti-pneumococcus serum when
the pH range was between 6.0 and 7.0.
With regard to the total globulin fraction, the test clearly indicates that more
immune bodies have been precipitated with the total globulin than with the
euglobulin (ammonium sulphate method). This observation is contrary to
Hartley's earlier finding that the whole of the antibodies of anti-rinderpest serum
are associated with the euglobulin fraction; at least this appears to be the case
when we are dealing with the euglobulin fraction which is precipitable by one-third
saturation with ammonium sulphate. It is interesting, however, to find that the
potencies of the total globulin fraction and the water-insoluble protein fraction do
not differ greatly, and this observation seems to indicate that in the so-called
water-insoluble protein (the Felton fraction), we have probably obtained a precipita-
tion of a protein fraction which lies on the border line between the fractions usually
designated as euglobulin and pseudoglobulin, and that the immune bodies of anti-
rinderpest serum are associated not only with the euglobulin fraction, but to a
certain extent also with the pseudoglobulin fraction. In this connection it is of
interest to refer to the work of Chick [1914] and Sörensen [1925], who believe that
it is almost impossible to obtain a complete separation of euglobulin and pseudo-
globulin, which are associated as a labile compound, and to the results of Svedberg
and Sjögren [1928] on the homogeneous nature of native serum globulin. The
recent investigation of Reiner and Reiner [1932] on the physical and chemical
properties of different fractions of serum globulin may also be referred to.
It is considered that the data in this paper are too meagre to permit of further
discussion on this point at present. It is, however, indicated that the total
globulin fraction and to a lesser extent the water-insoluble protein fraction give
better separation of the antibodies of anti-rinderpest serum than the euglobulin
fraction (ammonium sulphate method).
In the case of samples 3 and 4 the idea was not only to ascertain the relative
amount of antibodies in the different protein fractions, but also to determine
whether the precipitated fractions can be dissolved in a much less volume of saline
than the volume of the original serum used, and, thus concentrated, if this solution
would show a relatively higher potency without a great increase or indeed without
any increase, in the volume of the actual dose administered. Unfortunately,
owing to difficulty experienced during dialysis, concentration to the degree
originally anticipated was not possible, and where further concentration was
PRECIPITATION OF ANTIBODY FROM ANTI-RINDERPEST SERUM
157
attempted by partial drying under vacuum at a slightly elevated temperature the
results were unsatisfactory owing to a defect in the apparatus at our disposal. The
euglobulin fraction of sample 3 was concentrated four times, i.e., the precipitate
was dissolved in ¼ the volume of the original serum. The globulin fraction of
sample 3 as well as the two fractions from sample 4 were concentrated twice. The
result of the test of these sera and the original sera are given in the accompanying
Table. The dose of serum in the Table is given in terms of the concentrated
fractions.
Test results of samples 3 and 4.
Sample.
3
3 a
3 a
3 b
3 b
4
4 a
4 b
Controls
Dose of serum
per 100 lbs.
15 c.c.
15 c.c.
7.5 c.c.
30 c.c.
15 c.c.
15 c.c.
30 c.c.
30 c.c.
Initial condition.
Fair
Poor
Fair
„
„
Fairly good
Poor
Fair
Final condition.
Worse
„
„
Improved
Worse
Died
Improved
Worse
Same as for samples 1 and 2.
Percentage potency
based on relative
loss or gain in weight.
55
55
37
76
54
0
76
52
The results tabulated above conform in general with the results obtained with
the fractions of samples 1 and 2, the only apparent anomaly being that obtained
with the total globulin fraction (4b) of sample 4. This anomaly is probably
explained by the fact that this sample was one of those in which concentration by
drying in vacuo was attempted. The sample was accidentally overheated and
appeared to be partially charred, but we have included the test result as of interest
in that even in spite of the drastic treatment which the sample received, it still
retained an appreciable protective value.
As far as the main point, viz.—the concentration of antibodies, is concerned
the results are not so striking as in the case of the first point under investigation,
presu mably because the increased dosage used did not compensate for the percent-
age loss in the separation of antibodies. Generally it appears that some success
has been achieved and we are hopeful from the results of the total globulin (3b) of
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INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [III, II
sample 3, and the results of sample 4 and its derived fractions that, with more
experience on a larger scale, standard concentration on a practical basis may yet be
attained.
In conclusion we may remark that this preliminary investigation has covered
roughly the three points for the study of which the present research was started and
the results so far obtained are summarised later on. Much more work will be
necessary before satisfactory conclusions can be formulated but we should like to
mention here that current experiments on a larger scale, which will be published
later, already confirm the indications of the tests reported now. Several difficul-
ties of a practical nature have also been encountered, and in a future communica-
tion these will be discussed.
SUMMARY.
(1) An experimental investigation has been carried out on the fractionation of
the antibody containing proteins of anti-rinderpest serum by several methods and
an attempt has also been made to concentrate the antibodies after fractionation.
The potencies of the different fractions have been estimated by animal inoculation
experiments on " hill bulls " susceptible to rinderpest virus.
(2) The total globulin fraction precipitated by half saturation with ammonium
sulphate appears more potent than either the water insoluble protein fraction, or
the euglobulin fraction obtained by one-third saturation with ammonium sulphate.
(3) Attempts made to concentrate low potency sera by fractionation of the
sntibodies and their subsequent concentration have given encouraging results.
ACKNOWLEDGMENTS.
We desire to express our thanks to Mr. F. Ware. F.R.C.V.S., I.V.S., Director,
Imperial Institute of Veterinary Research, Muktesar-Kumaun, for his encourage-
ment and helpful suggestions throughout the course of this work, and also to
Mr. P. C. Banerji, Veterinary Deputy Superintendent, for his supervision of the
animals while under test.
REFERENCES.
Astros and Rietsch (1900). Compt. Rend. Soc. Biol. 52, 337.
Barr, Glenny and Pope (1931). Brit. J. Exp. Path. 12, 217.
Béclère. Chambon and Menard (1899). Ann. Inst. Past. 13, 81.
Belfanti and Carbone (1898). Zeit. f. Bact. 23, 906.
Brieger and Boer (1896). Zeit. f. Hygiene. 21, 259.
Brieger and Ehrlich (1893). Zeit. f. Hygiene. 13, 336.
Brodie (1897). Jour. Path. and Bact. 4, 460.
Cathcart (1904). Jour. Physiol. 31, 497.
Chick, H. (1914). Biochem. Jour. 8, 404.
PRECIPITATION OF ANTIBODY FROM ANTI-RINDERPEST SERUM
159
Dieudonné (1897). Arbeiten aus dem Kaiserlischen Gesundheitsamte, 13, 293.
Felton, L. D. (1925). J. Inf. Dis. 37, 199, 309.
—
(1928). J, Inf. Dis. 42, 248, 43, 542.
Findlay (1931). Brit. J. Exp. Path. 12, 9.
Frankel, M. (1932). Proc. Roy. Soc. B., 111, 165.
Frankel, M., and Olitzki, L. (1930). Nature, 126, 723.
Gibson (1906). J. Biol. Chem. 1, 161.
Gibson and Banzhaf (1910). J. Exp. Med. 12. 411.
Gibson and Collins (1907). J. Biol. Chem. 3, 233.
Goodner, K. (1930). J. Immunology, 19, 473.
Hartley (1914). Mem. Dept. Agric. India, 1, No. 4, 177—270.
Hartley (1931). A System of Bacteriology, 6, 249. M. R. C. Sers.
Henseval (1919). Compt. Rend. Soc. Biol. 82, 1071.
Hiss and Atkinson (1901). J. Exp. Med. 5, 47, 67.
Laidlaw and Dunkin (1931). J. Comp. Path. Ther. 44, 1.
Landsteiner (1900). Zent. f. Bact. 27, 357.
Ledingham (1907). J. Hygiene, 7, 65.
Ledingham, Morgan and Petrie (1931). Brit. J. Exp. Path. 12, 357.
Mallick and Maitra (1932). Ind. J. Med. Res. 19, 951.
Morgan and Fairbrother (1930). Brit. J.. Exp. Path. 11, 512.
Opic and Barker (1907). J. Exp. Med. 9, 207.
Pick (1901). Beit. Chem. Physiol. u. Path. 1, 351.
Reiner, H. K., and Reiner, L. (1932). J. Biol. Chem. 95, 345.
Seng (1899). Zeit. f. Hygiene, 31, 573
Sörenson (1925). J. Amer. Chem. Soc. 47, 457.
Sternberg (1892). Quoted from Ledingham, Morgan and Petrie, loc. cit.
Svedberg and Sjögren (1928). J. Amer. Chem. Soc. 50, 3318.
Wassermann (1894). Zeit.f. Hygiene, 18, 236.
Weyer, Park and Banzhaf (1929). Amer. J. Path. 5, 517.
A PRELIMINARY REPORT ON THE SUCCESSFUL INFECTION
WITH NASAL SCHISTOSOMIASIS IN EXPERIMENTAL
CALVES
BY
M. ANANT NARAYAN RAO,
Lecturer in Parasitology, Madras Veterinary College.
(Received for publication on 27th March 1933.)
It has been established that, in the nasal lesions and in the nasal discharges
of the so-called nasal granuloma of cattle, schistosome ova of a typical shape are
met with, and from the lesions worms have also been obtained and described. The
experiment recorded here was designed with a view to determine the kind of
cercariæ involved, the intermediary host and also the period of incubation as there
is no recorded evidence of any work having been done.
There were two ways open to the writer of elucidating the points in question.
One way was by infecting different kinds of freshwater mollusca, bred in the
laboratory, with the miracidia obtained from the ova in the nasal lesions and
utilising the cercariæ thus obtained for infection in bovines. The other, the
easier alternative, was to obtain different kinds of mollusca from the freshwater
ponds or tanks, used by cattle in villages, where nasal schistosomiasis is endemic,
and to utilise for infection the apharyngeal cercariæ discharged by the snails.
The latter method was adopted. To effect this the writer, in June 1932, visited
Nammiandal, a village in North Arcot District, where nasal schistosomiasis is
enzootic and collected snails mostly Planorbis exustus (Deshayes) from the tank
there. These snails discharged two species of apharyngeal cercariæ, viz., one
resembling cercariæ of Schistosoma spindalis described by Soparkar in 1921 and
another resembling Cercariæ Indicœ XXX Sewell, 1921. Only 1.2 per cent. of the
snails discharged the former kind of cercariæ and almost 3 per cent. discharged
the latter variety.
In September 1932 the writer again collected various kinds of freshwater
snails from Nammiandal and Bandarlapalli, another enzootic area in Chittoor
District, and the two kinds of cercariæ were seen in Planorbis exustus. It was also
noted that some Limnea leuteola (Lamarck), collected in the latter area, were
infected with Cercariæ Indicœ XXX. The noticeable feature again was that the
percentage rate of infection in the mollusca with cercariæ resembling Cercariæ
( 160 )
SUCCESSFUL INFECTION WITH NASAL SCHISTOSOMIASIS IN CALVES
161
Indicæ XXX was much more than with the cercariæ resembling those of S. spin¬
dalis. In order to compare the difference in incidence in areas where this disease
is less prevalent, snails were obtained in September 1932 from Conjeevaram and it
was found that the rate of infection in the mollusca with Ceroariæ Indicæ XXX
was almost negligible when compared with the rate of infection with cercariæ of
S. spindalis. In the same month a large number of snails were also collected from
the Spur Tank, Madras, in which buffaloes wallow practically all the year round
and where no cases of Nasal Schistosomiasis have been noticed. It was found that
from the snails collected here only a small percentage of Planorbis exustus dis-
charged cercariæ resembling those of S. spindalis, and none resembling the other
was present.
From the above it was clear that Cercariæ Indicæ XXX predominated over
those of S. spindalis in the villages Nammiandal and Bandarlapalli where Nasal
Schistosomiasis is endemic; hence it was thought possible that the former may
develop into the adult Schistosomes found in Nasal Schistosomiasis. With this
idea in view one calf (No. 409) was infected with the few available cercariæ from
two or three snails that remained out of the collection obtained from Conjeevaram
at the end of September.
In December 1932, soon after the rains, the writer collected on 3rd December
1932 and on 28th December 1932 two batches of snails amounting, in all, to 2,975
Planorbis exustus and 2,050 Limnea leuteola from the pond in Bandarlapalli. The
rate of infection in these was found to be 1.27 per cent. and 1.71 per cent. respec-
tively. Two more calves (Nos. 410 and 411) were obtained, and these two along
with the previous one were infected by three different methods. Calf No. 410 was
drenched with water containing approximately 4,000 cercariæ per dose; calf
No. 411 was infected by the nose by applying to the mucous membrane of the
nostrils with a pipette, about 4,000 cercariæ per dose; and calf No. 409 was
drenched with water containing approximately 2,000 cercariæ per dose and at the
same time about 1,000 cercariæ were applied to the mucous membrane of the nose.
The calves got a daily dose of cercariæ from 6th December 1932 to 14th
December 1932 and again from 31st December 1932 to 5th January 1933. The
idea in giving repeated doses for a few days was to simulate the supposed natural
infection. Perhaps the animals in the villages get daily doses of infection while
drinking water from a pond and this is continued as long as they drink from the
same infected pond. The pond water at Bandarlapalli was subjected to an
examination regarding the apharyngeal furcocercarial content and it was found
that on an average there were three furcocercariæ per cubic centimeter of water.
Assuming that an average-sized bovine drank about 6 gallons of water per day,
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INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [ III, II
the total dose of cercariæ swallowed would be about 15,630, in addition to what
might burrow through the skin. Though in the natural mode of infection these
animals might get comparatively larger doses than it is possible to give under
experimental conditions, yet it would appear that Nasal Schistosomiasis can be
produced in calves with comparatively smaller doses of cercariæ.
On the 3rd day of infection in the nose, the muffle of calves Nos. 409 and 411
showed dryness, the edges showed scaling of the epithelium and the mucous
membrane too was reddened with small red dots on it. These symptoms disappeared
in about a week after the first course of application of cercariæ. Similar symptoms
were again noticed during the second course of application of cercariæ to the nose.
The nasal washings and fæces of the three calves were periodically examined from
the last week of January 1933. From about the 15th February calves Nos. 409 and
411 showed a thin discharge from the nose and occasional sneezing was observed in the
latter. On 28th February 1933 a few small beads of pus were seen in the nostrils
of calf No. 411 and when the discharge was examined, it showed ova resembling
those found in nasal schistosomiasis and all of them contained live miracidia. On
3rd March 1933 calf No. 409 also showed suppurative foci on the mucous membrane
of the nose, and on 4th March 1933 ova were seen in the nasal discharge as in the
other. This experiment shows that the minimum period of incubation in the two
calves is roughly 3 months. Calf No. 410 has been showing a discharge from the
nose but no ova have been found in it up to the time of writing this paper. So far
none of the calves showed ova of S. spindalis in their fæces.
It is my pleasant duty to acknowledge my indebtedness to Mr. T. J. Hurley,
M.R.C.V.S., D.V.S.M., Principal, Madras Veterinary College, for the valuable
suggestion of infecting one calf in the nose only and the calf infected in this way
has shown the disease first. I am also grateful to Mr. P. T. Saunders, O.B.E.,
M.R.C.V.S., Director of Veterinary Services, Madras, for obtaining sanction to
conduct these experiments, and to Messrs. A. Ramaswamy Ayyar, K. P. Dorai¬
swamy, L. S. Parameswara Ayyar and S. Vaidynatha Mudaliar for all the help
rendered by them in this work. My thanks are also due to Mr. R. Narasinga Rao,
District Veterinary Officer, Madras, for providing me with snails from Conjeevaram
on two occasions.
FIG. 1. Lateral view of the anterior end ×830.
FIG. 2. Dorsal view of the anterior end × 830.
FIG. 3. Posterior extremity of female × 830.
Setaria buxi.
ON A FEW NEMATODES PARASITIC IN GOATS AT MUKTESAR
BY
G. D. BHALERAO, M.Sc.,
Helminthologist, Imperial Institute of Veterinary Research, Muktesar.
(Received for publication on 3rd August 1932.)
(With three text-figures.)
Setaria buxi n. sp.
Only one female specimen was obtained from the peritoneal cavity of a hill goat
at Muktesar by Mr. Gurubux Singh who very kindly placed it at my disposal for
which I wish to express my indebtedness to him.
The worm is of medium size, measuring 45.2 * in length and the maximum
thickness which is attained at some distance behind the œsophagus is 0.47. It is
whitish in colour and tapers towards both the extremities ; the posterior end being
somewhat spirally coiled. The cuticle is marked with fine longitudinal striations.
The nerve ring encircles the œsophagus at a distance of 0.145 from the anterior
extremity. The cervical papillæ are situated 0.12 posterior to the nerve ring. The
head is provided with two lateral and. four pairs of submedian papillæ, the latter
of which are rather inconspicuous. The cephalic portion appears dome-shaped.
The mouth is surrounded by an oval chitinous peri-buccal ring which has greater
diameter dorso-ventrally. It is more elevated from the head than that in the
species S. labiulo-papillosa and S. equina, the specimens of which were at my
disposal for comparison. It is deeply notched laterally and projecting from, its
walls on each side there appears, in the lateral view, a tooth-like process. Arising
from the peri-buccal ring are a pair of simple dorso-ventral lips which are not
notched as in S. labiato-papillosa and a pair of lateral lips which are crescentic.
The œsophagus is divided into an anterior muscular portion measuring 0.28 in
length and the posterior glandular portion which measures 2.84. The ratio that
the entire œsophagus bears to the whole body is 1 : 14.5. The tail measures 0.355
in length and the width of the body at the anus is 0.09. At the end of the tail
there is an irregular knob which bears on one side a well-developed spine. At
a distance of 0.014 from the tip of the tail there is a pair of small papillæ and
situated at 0.035 from it is a pair of much larger papillæ. The vulva is situated
at a distance of 0.28 from the anterior extremity. The eggs measure approximately
0.026—0.034 ×0.009—0.013.
* All measurements in millimeters.
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INDIAN JOURNAL OF VETERINARY SCIENCE AND ANIMAL HUSBANDRY [ III, II
Yorke and Maplestone [1926] listed seventeen species under the genus Setaria,
out of which eight species : S. bicoronata, S. cælum, S. cornula, S. scalprum,
S. transversata, S. effilata, S. bidentata and S spelaea have been insufficiently
described and Thwaite [1927] is somewhat sceptical in regard to their inclusion in
this genus. Out of the remaining nine species Purvis [1931] has definitely shown
that S. digitata is a synonym of S. labiato-papillosa, since he found that the outline
of the lateral lips and the nature of the terminal knob of the tail of the female, the
characters which were supposed to distinguish the two species from one another,
vary from specimen to specimen. Boulenger [1921] and Thwaite [1927], however,
regard these as two distinct species. Five additional species of the genus have
been described by Thwaite [1927], viz., S. poultoni, S. southwelli. S. boulengeri,
S. pillersi and S. Yorkei. Subsequently to this Issaitschikow and Rajewsky [1928]
added S. tundra, Rajewsky [1929] added the species S. altaica and the species
S. loveridgei was described by Sandground [1928]. The account of the last one was
not accessible to me in India. Out of these species the present specimen comes
closer to the species S. labiato-papillosa, S. poultoni and S. equina; but from all
these it can be distinguished by the following features :—
(1) The nature of the peribuccal ring.
(2) The form of the dorso-ventral lips.
(3) The nature of the tail.
For these reasons it is considered to be a new species which is the first one of
the genus hitherto recorded from the goat. Baylis [1929] lists only five species of
the genus from the domestic animals, the present one forming, therefore, the sixth
of the series. It is proposed to designate the new species as Setaria buxi after
Mr. Gurubux Singh.
Specific diagnosis.—Setaria: Male.—Unknown. Female.—Length 45.2, maxi-
mum thickness 0.47. Peribuccal ring more elevated and its walls bearing tooth-like
processes projecting into the lateral notches. Lateral lips crescentic, dorso-ventral
lips without a notch. The ratio of the œsophagus to the body being approximately
1:14.5. Glandular portion of œsophagus being approximately 11 times longer than
the anterior muscular portion. Nerve ring, cervical papillae, and vulva 0.145, 0.265
and 0.28 respectively from the anterior extremity. Tail 0.355 long, with two pairs
of papillæ : one of which being much longer than the other, which is situated closer
to the tip ; the terminal knob irregular with a large lateral spine. Eggs measure
0.026—0.034 ×0.009—0.013 approximately.
Host.—Hill goat (Capra sibirica).
Location.—Peritoneal cavity.
Locality.—Muktesar (U. P., India).