NationalLibraryOfScotland/medical-history-of-british-india huggingface dataset:
3
leucocytic parasites was confined to those described in birds by Danilewsky,
Sacharoff and others, and some doubt existed as to whether the cells in which
these parasites were seen were true leucocytes. Danilewsky named the para-
sites Leucocytozoa, but Laveran considered the cells in which they were
contained were immature red cells and that therefore the parasites were
Hœmamœbœ.
More recently Captain Christophers, I.M.S., 7 has described a characteristic
hæmogregarine in the blood of Gerbillus indicus in Madras. It presents the
following appearances.
Encysted forms, each like a vermicule with a short tail sharply flexed upon
the body, lie in spaces in enlarged and faintly-staining red cells. None could
be identified as a young or developmental form.
Free parasites were present in blood which had been kept moist for some-
time. The nucleus was more elongated and was still situated near the posterior
end. Some of the forms were long and slender.
In fresh preparations, the vermicules, resembling the encysted forms in
appearance, exhibited active movements. They travelled rapidly, gliding along
with the blunt end forward. They were also seen to twist and turn among the
red cells. Some of them were observed emerging from the red cells. Only a
small number kept under suitable conditions exhibited any change.
Many smears from the liver, spleen and bone-marrow were searched for
developmental forms with negative results.
Christophers gives a description of a new species of Hæmatopinus which
was found on the rats and describes the development undergone by the
parasite in the body of this louse. Vermicules were seen on the fourth day in
the midguts of two lice. Others removed on a later date showed large cystic
bodies lying in the body cavity. Each cyst was filled with smaller oval cysts
containing 6 to 8 crescentic bodies. These bodies were placed on a slide in the
intestinal juice of some of the rats and it was found that most remained motion-
less, and some appeared to have undergone dissolution, although in the blood
plasma of the rat these bodies exhibited remarkably active movements.
No further development was observed and no bodies were seen in the
salivary glands or ovaries.
This mammalian hæmogregarine is confined to the red cells and is the first
in which free vermicules have been described in the blood. The discovery made
by Captain Christophers of its developmental cycle in the body of a louse is
of extreme importance.
Still another hæmogregarine parasite has recently been described in a
mammal by Dr. Andrew Balfour 8 in the blood of jerboas (Jaculus jaculus, or
J. gordoni as it has been re-named,) in the Soudan. He has described three
4
forms of this hæmogregarine : encysted forms in the red cells, free vermicules
in the plasma and a third in the liver in the form of cytocysts containing
merozoits and residual protoplasm. Examination of the spleen and bone-marrow
proved negative.
Laveran, 9 having seen Dr. Balfour's specimens, obtained some jerboas from
Tunis belonging to the genus faculus orientalis and found they were infected
with this hæmogregarine. He describes it as follows. The endoglobular
elements 8µ to 8.5µ are elongated reniform bodies with a tail flexed on the
body. One extremity is round and the other pointed. These bodies contain
no pigment and male and female forms could not be distinguished. Free vermi-
cules were seen in the plasma of the peripheral blood. They were more
frequent in liver smears. They resembled the endoglobular parasites.
Spherical bodies measuring from 21µ to 23µ were seen in the liver smears.
These cysts contained sporozoits, which were elongated with one end round
and the other attenuated. The chromatin mass was well defined and large.
Laveran has named this parasite Hæmogregarina balfouri.
In May 1905, while searching for blood parasites in some of the smaller
mammals—hedgehogs, squirrels, flying-foxes and bats, found in and around
Rajkote, Kathiawar—I discovered a parasite in the large mononuclear leucocytes
of the Kathiawar palm squirrel ( Sciurus palmarum or, as it has been re-named,
Funambulus pennantii ).
A specimen was sent to Captain Christophers, who informed me that he
thought the parasite was a hæmogregarine.
In all 52 squirrels were examined and 51 were found to be infected. The
parasites were in large numbers in the majority and remained numerous for many
weeks. It is now four months since the squirrels were first examined and the
infection remains unaltered.
The parasite in fresh preparations.
On examining a drop of blood from an infected squirrel, the parasite is
seen as a clear elongated worm-like body measuring 10µ in length, lying in
the substance of the large mononuclear leucocytes (Fig. 1). The majority
exhibit slow vermicular movements, altering their position in the cells. They
may be seen lying close alongside the nucleus or at right angles to it.
The nucleus becomes compressed, and in some instances almost comes to
encircle the parasite. Many of the nuclei are split into two separate parts with
the parasite lying between. The invaded leucocytes show extremely slight
movements. Each body is oval with one end rounder than the other. The
protoplasm contains a large refractile nucleus placed about the centre and some
dark dots are seen moving in the protoplasm.
5
The narrower end shows a distinct bend upwards simulating a tail. There
is no clear space between the edge of the parasite and the substance in which
it lies, and no pigment granules are seen in the protoplasm.
In fresh blood films containing a large number of parasites, some of these
bodies are seen escaping from the cells. After watching one, which appears
particularly active, the more rounded end will be seen to cause the cellular
envelope to bulge. It ruptures and the parasite is extruded. No space is left
behind, to show where the parasite was lying.
Besides the encysted forms many are seen lying free in the plasma.
They have the same appearance as those in the cells (Fig. 2). The tail is
most distinct when the parasite is on its side. It is quite short and ends
abruptly. Most of the granules are seen between the nucleus and the tail.
These forms are usually lying so that the tail is below, and is seen as a band
across the protoplasm. The majority exhibit slow vermicular movements,
never moving out of the field.
In films from three squirrels with swarming parasites free vermicules were
seen (Fig. 3). Each is an elongated spindle-shaped body measuring 13µ to
14µ in length and from 3µ to 4µ in breadth. They were very active, perform-
ing rapid serpentine-like movements, twisting and curling about. They would
approximate both ends and suddenly separate them. They attached themselves
to a red cell by one end, and whirled round so fast that it was impossible to
follow their movements. They very quickly glided out of sight. The nucleus
was situated about the centre and consisted of a single mass. Many refractile
dots were seen towards one end.
The parasite in stained films.
The parasite stains readily with Romanowsky's stain and Leishman's modi
-fication. It appears lying in the substance of the leucocyte. No distinct capsule
can be made out by staining in the usual way, but by prolonged staining with
Romanowsky's stain and also with Giemsa's, a faint pink outline is seen around
the parasite. This under higher magnification consists of minute chromatic
particles.
Each body is oval in shape with one extremity larger and rounder than the
other (Fig. 4). It measures 10 µ in length and 5µ across the broadest part.
The protoplasm stains light-blue, somewhat darker at the two ends.
The nucleus is a large irregularly quadrilateral shaped mass, which lies about
the centre. It occupies the whole of this part of the parasite encroaching on the
sides. It stains deeply but not uniformly and contains three to eight vacuoles.
The substance is at parts much darker than at others. Six to eleven large chro
-matic dots are seen in the protoplasm, more in one extremity than in the other.
The parasite is most frequently seen lying close alongside the nucleus which
6
adapts itself to the shape of the parasite (Fig. 4). In most of the cells no
protoplasm is seen between the nucleus and the parasite. They may be also
commonly seen lying between two parts of the nucleus (Fig. 5). These may be
quite detached or united by a thin strand of nuclear tissue.
On three occasions two parasites were seen in one leucocyte (Fig. 6). In
one cell they are lying close to each other, between the two detached parts of
the nucleus. In the other cell, which was much distended, the two parasites are
lying on either side of the drawn-out nucleus.
In more than one film from a heavily-infected animal, the nuclei of the
parasites were seen to consist of two separate parts (Fig. 7), situated towards the
extremities of the parasites. They may be quite circular or elongated. These
parasites appeared in all other respects exactly similar to those described above.
The protoplasm of all the invaded leucocytes stains feebly as compared
with the normal cell (Fig. 8). Even with the most intense staining with Giemsa's
stain the protoplasm is indistinct and the parasite appears lying close to a
nucleus.
Films were made from the blood of a Madras squirrel, when the protoplasm
of the large mononuclear leucocytes was stained an intense blue (Fig. 8).
The presence of the parasite in the cell seems to cause a very definite change
in the protoplasm as shown by its be haviour to stains. In one case where out of
300 large mononuclear leucocytes, 180 contained parasites, almost every one of
the remaining mononuclear leucocytes consisted of a nucleus with mere traces of
protoplasm.
One leucocyte showed two vacuoles lying on each side of the partially
divided nucleus (Fig. 8). This would suggest that at one time these cells had
harboured parasites, but that previous to the making of the film, they had left
the cells. No free forms were seen close to these cells, which might have
proved that the parasites were only extruded during the preparation of the film.
The cells are certainly friable and break up when the film is made.
Figure 9 shows a parasite in a stained film in the act of passing out of the
cell. The rounded end has already left the cell, while the more attenuated end is
still in the body of the cell.
On staining the free forms with Giemsa's stain the minute structure of the
parasite is clearly seen (Fig. 10). One end is globular, while the other is narrow,
protoplasm consisting of a loose network, which stains blue. Scattered through-
out it are minute faintly-staining pink dots, which in some places are collected
together, giving a uniform pink appearance, while in others they are scattered
and give the appearance of stippling.
The large chromatic dots are arranged irregularly (Fig. 11). There seems
to be no definite order in their position, which might suggest any differentiation
into types. The nucleus is large and stains deeply.
7
Some of the free forms are more attenuated than others (Fig. 13). The
protoplasm stains dark-blue, and the pink stippling is marked. The nucleus,
which is elongated and stains deeply, shows no vacuoles.
Figure 14 represents a vermicule from the peripheral blood stained with
Romanowsky's stain. It measured 17.5µ in length and 3.5µ. in breadth. The
ends are attenuated, one more so than the other. The nucleus is circular and
consists of a single mass without vacuoles.
Leucocyte counts of uninfected and infected squirrels.
Almost the first thing that strikes the eye on examining a blood film taken
from a heavily infected animal, is the presence of a large number of mononuclear
leucocytes. After making a series of counts it was thought that there was a
marked increase in the number of these cells. To settle this point the blood of
Madras squirrels in which the parasite was not found was examined. It is
necessary to mention, that these squirrels were found to harbour the trypanosome
discovered by Major Donovan, I.M.S., and described by Laveran and Mesnil.9
Blood counts in four cases gave the following figures. Five hundred cells
were counted in each film :—
Squirrel I.
Polymorphonuclear leucocytes
•
•
•
. . 45.8
Large mononuclear leucocytes
.
.
.
. 25.
Lymphocytes
.
.
.
.
.
.
.
. 20.8
Myelocytes
.
.
.
.
.
.
.
. 5.8
Basophiles
.
.
.
.
.
.
.
.
2.4
Eosinophiles
.
. .
.
.
.
.
.2
Trypanosomes few.
Squirrel 2.
Polymorphonuclear leucocytes
•
•
•
•
• 43.2
Large mononuclear leucocytes
.
. .
.
. 32.
Lymphocytes
.
.
.
.
.
.
. 18.
Myelocytes
•
•
•
.
.
•
•
.
4.8
Basophiles
.
.
.
.
. .
.
.
2.
Eosinophiles
•
•
•
.
. .
.
.
•
No trypanosome found.
Squirrel 3.
Polymorphonuclear leucocytes
.
.
.
.
. 32.2
Large mononuclear leucocyte
.
.
.
. 28.2
Lymphocytes
. . .
.
.
.
.
. 31.6
Myelocytes
.
.
.
.
.
.
. 5.6
Basophiles
.
. .
.
.
.
.
. 1.6
Eosinophiles
.
. .
.
.
.
. . .8
Trypanosomes abundant.
8
Squirrel 4.
Polymorphonuclear leucocytes
.
.
.
.
. 42.6
Large mononuclear leucocytes
.
.
.
.
. 23.6
Lymphocytes
.
.
.
.
.
.
.
. 27.6
Myelocytes
.
.
.
.
.
.
.
.
4.8
Basophiles
.
.
.
.
.
.
.
. 1.2
Eosinophiles
. . .
.
.
.
.
.
.2
Trypanosomes few.
Squirrels with different degrees of infection were then examined and the
figures were as follow :—
Squirrel I, with intense infection.
Polymorphonuclear leucocytes .
.
.
.
9.4
Large mononuclear leucocytes
. .
.
.
. 80.8
Lymphocytes
. .
.
.
.
.
9.4
Basophiles
.
.
.
.
.
.
.4
Myelocytes
.
.
.
.
.
.
. . . . . .
Eosinophiles
.
.
.
.
.
.
. . . . . .
Total number of parasites seen
.
.
. 326
Free parasites seen
.
.
.
.
6
Squirrel 2, with severe infection.
Polymorphonuclear leucocytes
. .
.
.
. 17.8
Large mononuclear leucocytes
.
.
.
.
.
64.8
Lymphocytes . . .
.
.
. 13.8
Myelocytes
. . . .
.
.
.
1.6
Basophiles
. . .
.
. 1. 8
Eosinophiles
.
.
.
.
.
.
.
.2
Total number of parasites seen .
.
.
1 0 0
Free parasites seen .
.
. .4
Squirrel 3, with a mild infection.
Polymorphonuclear leucocytes
. .
.
.
. 33.8
Large mononuclear leucocytes
.
.
..
.
. 41.4
Lymphocytes
.
. . .
.
.
. 20.2
Myelocytes
. . . .
.
.
. 2.6
Basophiles
. .
.
.
.
.
.
.
.
.6
Eosinophiles . . . . .
.
.
.
1 . 4
Total number of parasites seen .
.
. 50
Free parasites seen .
.
.
.
. 3
9
Squirrel 4, with mild infection.
Polymorphonuclear leucocytes
.
.
. .
. 27.5
Large mononuclear leucocytes
.
.
.
.
. 59.5
Lymphocytes
.
.
.
.
.
. 10.5
Basophiles
.
.
.
.
.
.
.5
Myelocytes
.
.
.
.
.
.
......
Eosinophiles
.
.
.
.
.
.
. 2.5
Total number of parasites seen .
. . 27
No free forms.
It will be seen in the case of the first of these that the large mononuclears
formed the bulk of the white cells, and, on comparing this figure with any of those
from the Madras squirrels' counts, it will be at once apparent that there is a
great increase in the numbers of this leucocyte. The Madras squirrel is taken
as an example of an uninfected animal.
It was of some importance to have settled this point and, to bring it
out more clearly, total counts by the field method were made from the heart-
blood of the two species. The following figures were obtained :—
Madras squirrel.
1st count.
2nd count.
Polymorphonuclear leucocyte
.
.
10,600
10,900
Large mononuclear leucocytes
.
.
4,200
4,000
Kathiawar squirrel.
1st count.
2nd count.
Polymorphonuclear leucocyte
.
.
8,600
7,900
Large mononuclear leucocyte
.
.
10,500
11,200
This shows that we are dealing with a distinct mononuclear leucocytosis and
that it varies in proportion to the degree of infection. The fact that a mam-
malian protozoan, which has the property of invading a special leucocyte,
causes a change in the structure and an increase in the numbers of these
in the peripheral blood may have some significance in relation to the origin of
leucocythæmias.
In making the above counts, films were taken from the animals as near as
' possible about the same time of day.
The distribution of the parasite in the body.
Smears were made from the spleen, liver, kidneys, pancreas, testicles, intes-
tinal mucosa and muscles.
The parasites were always present in considerable numbers in the spleen.
They were unusually large and almost equally round at both ends (Fig. 12).
The protoplasm was blue and contained the usual chromatic dots. The nucleus
10
in these forms was voluminous, staining less intensely and in some was worm-like
in appearance. No vermicules and no young or developmental forms were seen.
Special attention was directed to the liver in view of the developmental
forms found in H. balfouri. Fourteen squirrels were shot and smears were
made from their livers. In all the parasite was found both free and in the mono-
nuclear leucocytes, but no developmental forms. Further a heavily infected
animal was killed. Thirty-four thick smears were made from the liver, six from
the spleen and six from the kidneys. These were stained with Romanowsky's
stain and were systematically examined, first with a No. 3 Leitz, then with a 1/12
oil immersion. No cyst or other form suggestive of development were seen.
The parasites were numerous, both free and encysted, and had the appearances
of those in the spleen (Fig. 12). The parasites were scarce in the other tissues.
The majority of the Kathiawar squirrels are infected with a worm in the
abdomen which is seen free and encysted in the abdominal muscles and
diaphragm. No filarial embryos were seen in the blood.
The parasite in the body of the squirrel louse.
This louse has been identified for me by Professor Neumann and Dr. Newstead
as an undescribed species of Hæmatopinus.
It is found in large numbers among the long hairs of the squirrel. It clings
tenaciously to the hairs and is comparatively slow in its movements. The
ova measure .6 m.m. and are laid on the hairs of the legs and tail.
Male.—When full-grown is .9 m.m. in length, the head is triangular in shape
with two antennæ springing from the upper angles. Each consists of five seg-
ments. The third is most attenuated and the basal most globular. A large
spine springs from the upper surface of this segment. The eyes are small and
are situated behind the last antennal joint.
One short and one longer spine spring from the dorsum of the head.
The junction of the head and thorax is marked by a constriction. The
thorax consists of a large dorsal chitinous plate. It has one spine opposite
the first leg and two opposite the second. The legs, three on each side,
consist of four segments. The third leg is the most developed. The basal
segment is large and has a spine on the under-surface. The third segment is
smaller and not quite so globular. It has a few spines on it. The second is larger
and has two or three marked spines. The first segment is globular and has the
claw attached to it by a triangular joint. Below the claw there is a large pad.
The size of the various segments of the legs increases from before backwards :
this is most marked in the case of the claws.
11
The abdomen consists of nine segments. On the dorsum of each segment
there is a fringe of hairs and on each side there is a group of four or five.
The last segment carries one very long hair on each side. This segment ends in
a conical-shaped process from which the penis, a bifid structure consisting
of two long chitinous rods, protrudes.
Female.—The head of the female is smaller than that of the male ; the third
antennal segment is not attenuated as in the male, and there is a marked spine
on the basal segment.
The abdomen is made up of nine segments and is about twice as long as
in the male. The dorsal surface has two chitinous bands for each segment.
The last segment ends in a flat surface and has a fringe of six hairs placed
laterally. On the ventral aspect the hairs are arranged as in the male, except
that the first segment has four, the seventh has two and the eighth three. The
legs are the same as in the male.
Each female lays on an average five eggs. The young hatch out into fully
developed lice after three to four days.
This louse is closely related to H. antennatus, Osborn, and H. spinniger,
Denny.
On one occasion a small acarid was found parasitic on a louse, clinging
to the abdomen.
Lice were the only octoparasites that were seen on the squirrels and at
Captain Christophers' suggestion I decided to try to ascertain whether develop-
mental changes are undergone by the parasite in this louse.
A young male squirrel with swarming parasites was selected and 42 lice were
placed on it from other squirrels. On the second day two lice were removed and
examined. The midguts were dissected out and searched in the fresh condition.
Active vermicules were seen exactly similar to those in the peripheral blood. On
staining one of these, it was found to measure 15.5 µ was spindle-shaped with
one extremity slender and more pointed (Fig. 15). The protoplasm stained blue
throughout and had a stippled appearance due to minute chromatic dots. The
nucleus was situated about the centre, and consisted of a circular mass of proto-
plasm which stained uniformly dark. Between it and the thin end were six
large chromatic dots. No other form was seen in the midgut.
On the fourth day two more lice were examined, but besides the vermicules
nothing else was seen which might have suggested further development.
On the fifth day an adult female louse was removed. The midgut was
dissected out without being ruptured. It contained active vermicules and
encysted parasites as the blood was fresh. The remainder of the louse was
carefully examined and one vermicule was seen slowly moving about in the body
cavity. On staining it was found to measure 18 µ with one end attenuated and
12
slightly bent, the other was broader (Fig. 16). The protoplasm stained blue,
darker towards the pointed end. Two very distinct vacuoles were seen, one
at the, extreme end and the other deeper and to the side of the broader end.
Seven large chromatic dots were seen near the vacuoles. The nucleus consisted
of a collection of chromatic rods arranged irregularly.
Lice were examined regularly for ten days after being placed on the
squirrel, but, with the exception of vermicules, nothing suggesting any further
development was seen either in the gut or in the body cavity. No parasitic
bodies were seen in the salivary glands or ovaries.
Twenty-four lice were placed on a Madras squirrel from which all the lice
that could be found were removed. This squirrel was kept separate but up to
the twelfth day the peripheral blood remained free from infection.
Conclusions.
It will be seen that the description of this parasite, agrees in every detail
with that of a hæmogregarine. The parasite is highly specialised, in that it
selects the large mononuclear leucocyte for its host. Though closely related to
Leucocytozoön canis (James), it differs in not having a cytocyst and in possessing
a tail. I therefore propose provisionally naming it Leucocytozoön funambuli.
As in the case of Hæmogregarina gerbilli (Christophers), no developmental
forms were found in the organs and the infection remains unaltered for long
periods. The squirrels, though harbouring two parasitic worms, were to all
appearances as active as the Madras species. It is particularly interesting to
note, that trypanosomes were never found in the blood of the Kathiawar
squirrel, which seems to point to a localised distribution of the blood parasites in
very closely related mammals.
13
References to literature.
1. Wroughton.—Journal of the Bombay Natural History Society, Vol. XVI. No. 3.
2. Laveran.— Contribution à l' Étude des grandes Hœmogrégarines des Grenouilles:
Société de Biologie LIX. p. 172.
3. Stephens.—A new Hæmogregarine in an African toad. The Thompson Yates and
Johnstone Laboratories Report, Vol. VI (New Series), part I. Jan.
1905.
4. James.—On a parasite found in the white corpuscles of the blood of dogs. Scientific
Memoirs by Officers of the Medical and Sanitary Departments of the
Government of India (New Series), No. 14.
5. Bentley.—Preliminary note upon a leucocytzoön of the dog. British Medical
Journal, May 6th, 1905, p. 988.
6. Laveran.—Académie des Sciences T CXLI, p. 295 (Séance du 31 juillet 1905).
7. Christophers.— Hæmogregarina gerbilli. Scientific Memoirs by Officers of the Medical
and Sanitary Departments of the Government of India (New Series),
No. 18.
8. Balfour.—Third stage in the sexual cycle of the hæmogregarine of Jerboas. British
Medical Journal, June 17th, 1905, p. 1330.
9. Laveran and Mesnil.—Trypanosomes et Trypanosomiases. p. 103, 1904.