# Transactions of the Royal Society of Tropical Medicine and Hygiene/Volume 2/The Parasite of Kala-Azar and Allied Organisms

TRANSACTIONS
of
The Society of Tropical Medicine and Hygiene.
THE PARASITE OF KALA-AZAR AND ALLIED ORGANISMS.
By Captain W. SCOTT PATTON, I.M.S.
Of the King Institute of Preventive Medicine, Madras.
(Friday, January 15, 1909.)
Shortly after my return to England I was asked to read a paper on "Kala-Azar" before this Society, and on considering what particular aspect of the disease I was most competent to deal with, I decided to limit myself to the parasite, so to-night I propose to give you some account of my observations on this organism and its allies. I have no doubt you are all well acquainted with the parasite, the discovery of which has opened up an entirely new field of research of great practical importance. Although we now know a great deal about it and its two allies, the parasites of infantile splenomegaly and Oriental sore, there is considerable doubt as to their exact systematic position. Laveran and Mesnil, I believe, still regard the parasite of kala-azar as belonging to the genus Piroplasma, while you will remember Ross placed it as an entirely new genus, Leishmania. The view ​that this parasite is a hsemocytozoon can no longer be entertained, and Christophers' recent studies on the extracorporeal life-cycle of Piroplasma canis in the tick have shown that it does not pass through a flagellate stage similar to that of the parasite of kala-azar. I am aware that certain observers claim to have discovered flagellate stages in some Piroplasmata; their observations, however, are far from convincing. I have recently seen some of these so-called piroplasma flagellates, and have no hesitation in saying that they do not bear any resemblance to the characteristic flagellates of the parasite of kala-azar. It should also be noted that these flagellate stages of Piroplasmata have so far only been seen in test-tubes or in the blood of the vertebrate host, whereas they ought to occur naturally in ticks or other invertebrates.
There is, however, no doubt that the parasite of kala-azar and its two allies, as seen in man, are but stages in the life-histories of flagellates, and the important question to answer is, do they belong to any known group of the parasitic flagellata? As our knowledge of the life-cycles of these organisms is exceedingly limited, I think it somewhat premature to say they belong to an entirely new genus, and I hope to show you that they do belong to a group which has long been recognized as occurring in the alimentary tracts of insects. From a study of the flagellates of the parasite of kala-azar in the test-tube, as well as in the bed-bug Cimex rotundatus, I have no hesitation in saying it is not a trypanosome; for, like all other observers who have studied this stage, I have never seen anything similar to the well-known vertebrate trypanosome.
The most important suggestion as to the nature of the parasite of kala-azar was undoubtedly made by ​Rogers, who, on comparing it with Herpetomonas muscae domesticae, as figured by Prowazek, recognized the close similarity in the build of the two flagellates. Unfortunately, Prowazek erroneously described the adult flagellate of H. muscae domesticae as possessing two flagella closely united together, and, in addition, its life-cycle according to his account is exceedingly complicated. For more than a year I have made an exhaustive study of this flagellate, the type species of the genus Herpetomonas. In Madras about 100 per cent, of flies, caught in the bazaar meat shops, are infected with it, and as a result of my observations on the living parasite as well as in stained preparations, I am unable to accept Prowazek's view of its flagellar apparatus, for I find the adult flagellate has a single flagellum. I can only assume that Prowazek happened to examine flies in which the majority of the adult flagellates were undergoing the early stages of simple longitudinal division; I have frequently seen this in Madras. I also find that the life-cycle of the flagellate of the house-fly is very simple, and is similar to that of the parasite of kala-azar so far as we know it at present.
You will remember Prowazek began his description of H. muscae domesticae at its adult flagellate stage, and entirely omitted the stage preceding it, thus giving anyone who read his paper an entirely erroneous idea of its life-cycle. As you will see later, the earlier stages in the development of the parasite are of great importance, as it is then when it is so like the parasite of kala-azar.
In order, therefore, to clear up this difficulty I will describe in some detail the life-history of H. muscae domesticae, the complete account of which I shall reserve for another occasion. After having studied a large number of these parasites I have found it convenient ​to divide their life-cycles into three stages, preflagellate, flagellate and postflagellate, and it should be clearly
Plate I.
understood there is no hard and fast line dividing one stage from the other; they merge into each other. You will, however, see that by studying these flagellates ​along these lines it is easy to work out their complete life-histories, and, above all, to find out how they pass from one host to another. In the case of H. muscae domesticae in Madras the flagellate and postflagellate stages are readily found, but owing to the fact that a large number of flies have to be examined before the preflagellate stages can be studied, this important portion of the life-cycle of the parasite is very apt to be missed.
The preflagellate (Plate I., figs. 1 to 3a) stage is usually found in the midgut, the parasites lying in masses within the peritricheal membrane; they are round or slightly oval bodies, measuring on an average about 5.5 μ in breadth, and contain a nucleus, a blepharoplast, and a number of granules. They multiply by simple longitudinal fission or by multiple segmentation (figs. 2, 2a, 3, 3a), so that very soon a large number of these bodies is formed. The flagellate stage is characterized by the formation of the flagellum, a single stout filament which projects freely from a point in close proximity to the blepharoplast (fig. 4); the resulting flagellates elongate and later divide by simple longitudinal division (figs. 5, 6 and 7). The nucleus of the adult flagellate lies about the centre, and the blepharoplast, a large rod-shaped body, is nearly always situated a short distance from the anterior end; the flagellum is a single filament which projects freely almost at once from the rounded anterior end. The early stage of the process of longitudinal fission begins by a splitting of the root of the flagellum, and this, as well as further stages of the same change, led Prowazek to believe H. muscae doinesticae has a double flagellum. Had he, however, studied the method of formation of the flagellum he would not have fallen into this error. It is quite a common thing to find nearly all the adult flagellates in a fly exhibiting ​this appearance of a double flagellum. If the mid-guts of a number of flies are next examined in the fresh condition, in some, the flagellates will be seen collecting together towards the rectum, they attach themselves in rows by their flagellar ends to the gut-wall, and now further changes are seen to occur. The most external parasites begin to shorten, and while this change is taking place the flagellum degenerates and is eventually shed (figs. 8 and 9). A palisade of parasites is now formed, the most internal showing the changes noted above, while the most external are seen as round bodies devoid of flagella, and many of these latter forms are seen dividing again (fig. 10). Eventually, all the bodies round up and are connected together by a sticky substance, and these masses of cells constitute the cysts (fig. 11); this I regard as the postflagellate stage. These cysts are passed out in the faeces of the fly en masse and are well adapted to resist desiccation. It is these bodies which are again sucked up accidentally by the flies as they feed. Such is a short account of the life-cycle of H. muscae domesticae; it is exceedingly simple, for I can find no evidence of the complicated sexual changes described by Prowazek.
Turning now to another herpetomonad, H. lygaei, which I have recently described, you will see that it has a very similar life-cycle, only in this case the parasite is almost identical with that of kala-azar. In its preflagellate stage it is a small round or pear-shaped body (Plate II., fig. 1), measuring from 3.5 μ to 4 μ in length; it has a nucleus and blepharoplast and is found lying free in the mid-gut of Lygaeus militaris. The parasite divides, by simple fission into two, and each half again divides, so that four oval bodies are formed (figs. 2, 3 and 4). These bodies now pass on to flagellation somewhat ​irregularly (figs. 5, 6 and 7), so that one may have developed into an adult flagellate while the remainder are dividing further and remain attached to its anterior end (fig. 7). The adult flagellates divide by simple longitudinal division (figs. 8 and 9), and eventually collect m the hind-gut of the bug, where they attach themselves in rows; they now shorten (fig. 10), the flagella degenerate (fig. 11) and are shed, and they divide (fig. 12) at least twice (fig. 13), resulting in small pear-shaped bodies, the cysts, which are passed out in the fæces of the bug (fig. 14).
I have already described in great detail the development of the parasite of kala-azar in twelve bed-bugs (Ciniex rotundatus), and if you refer to my illustrations you will find its life-cycle, so far as it is known, is very similar to that of the two herpetomonads I have just described. All three parasites are very similar in their preflagellate stages, except that H. muscæ domesticæ is much larger; they develop and multiply in the same way, and in their adult flagellate stage they have the same general morphological build. They all undergo their developmental stages in the intestinal tracts of insects, but the fact that one of them passes its preflagellate stage in man has led most observers to regard it as something quite distinct from a Herpetomonas. The parasite of kala-azar is certainly not a sporozoon; it remains throughout its life-cycle, as far as we know it, a typical flagellate. With a complete knowledge of the life-cycle of such a closely allied form as H. lygæi it is possible to conjecture that the parasite of kala-azar, in order to get back to man, must pass back again to a non-flagellate stage, postflagellate, and that this change is most likely to take place somewhere in the neighbourhood of the biting parts of the bed-bug. Had I known the ​complete life-cycle of H. lygæi when I had my heavily-infected bugs I might have been able to solve this problem. This emphasizes the importance of studying these seemingly unimportant parasites, and I am sorry to say there are some people who still think it is a waste of time to study such forms, but there is no doubt that a knowledge of their life-cycles is of the utmost importance in helping to explain those of more complex pathogenic protozoa.
I have now shown you that the parasite of kala-azar is a Herpetomonad, and that in its preflagellate stage it may be indistinguishable from a harmless species; this truth is of importance, because in spite of other data the two closely allied human forms—the parasites of infantile splenomegaly and Oriental sore—are still regarded as identical with the parasite of kala-azar. I know at least three closely allied Herpetomonads which are very similar in their preflagellate stages to H. muscæ domesticæ so that this similarity in one stage is of little importance in distinguishing between these species. The parasite of kala-azar should, therefore, be known as H. donovani (Laveran and Mesnil), the parasite of infantile splenomegaly as H. infantum (Nicolle), and the parasite of Oriental sore as H. tropica (Wright), and I would suggest the diseases produced by these parasites should be collectively known as Herpetomoniases.
It is most unfortunate that all attempts to transmit H. donovani to animals have entirely failed; quite recently I inoculated dogs with the parasite, with negative results. It seems to me, therefore, that it will be well-nigh impossible to prove finally that C. rotundatus is the carrier of this dangerous parasite. It is to be hoped, therefore, that exhaustive feeding experiments, similar to those I carried out in Madras, will be conducted with ​H. infantum and H. tropica. However, I feel bound to say here that there is too great a tendency at present to cultivate these parasites in test-tubes; these experiments are undoubtedly highly interesting, yet they will not give us the clue as to how the parasites are transmitted from man to man. Feeding experiments with blood-sucking insects in the case of these Herpetomonads are very difficult and tedious, owing to the fact that unless a very good case is selected it is almost impossible to follow one or two developing parasites in the guts of blood-sucking insects. Further, such experiments must be carried out with great precision, and the observer must possess a good working knowledge of the many harmless gut flagellates which occur in nearly every insect. So far as I am aware, no attempts have been made to discover the transmitting agent of H. infantum or H. tropica.
Discussion.
Dr. Sambon said that he wished to express his very high appreciation of Captain Patton's work on kala-azar. Captain Patton had pointed out that the cell-enclosed "Leishman-Donovan bodies" became numerous in the peripheral blood of kala-azar patients in the later stages of the disease, and he had shown by feeding experiments that in the mid-gut of the rounded bed-bug (Cimex rotundatus) they assumed the flagellate form obtained by Major Leonard Rogers in artificial culture media. Furthermore, he had endeavoured to work out the entire life-history of the parasite, not only by following its development in the bed-bug, but also by studying allied organisms in other insects.
The slow rate of the bed-bug's dispersion and the