Class II. Mastigophora, Diesing

Sub-Class. FLAGELLATA, Cohn emend. Bütschli.

During the motile part of their life the Flagellata possess one or more flagella which serve for locomotion, and in many cases also for the capture of food. A few groups (Euglenoidinæ, Choanoflagellata) have only one flagellum, others two or several of about equal length (Isomastigoda), or of various lengths (Monadina, Heteromastigoda, Dinoflagellata). The long flagellum is the principal one; the smaller ones on the same organism are accessory flagella. The flagella directed backwards, which occur in the Heteromastigoda and are used for clinging, are termed trailing flagella or tractella. At the base of the flagellum, which is almost always at the anterior end, a Choanoflagellate possesses a cytoplasmic funnel-shaped neck or collar. In the parasitic forms an undulating membrane is often present.

The body of the Flagellata is usually small, generally elongate and of unchangeable form. It is frequently covered by a distinct cuticle, and, in certain groups, by a hard envelope, or it may be more or less loosely enveloped by a gelatinous or membranous covering. An ectoplasmic layer is thin and not always obvious. The granular cytoplasm contains a varying number of vacuoles, one of which may be contractile, and is generally situated near the area from which the flagella arise, that is, at the anterior extremity. The cytoplasm, moreover, contains the nucleus, which is nearly always single; and in many species there are also yellow, brown, or green chromatophores of various shapes, such as occur in plants. Some species feed after the manner of green plants (holophytic), or of plants devoid of chlorophyll (saprophytic); others, again, ingest solid food, and for this purpose usually possess a cytostome; the latter, however, in a few forms is not used for its original function, but is connected with the contractile vacuole. Many parasitic forms feed by endosmosis. A few species possess eye-spots with or without light-refracting bodies.

Variation in the form of the nuclear apparatus occurs. One nucleus only, which may be compact or vesicular, is known in many species. This nucleus is situated either centrally or sometimes near the flagellar end of the body, but its position is subject to variation. The flagella may arise near the nucleus. Other structures, such as an axial filament and a rhizoplast, may be present. Some flagellates are binucleate, the two nuclei—which often differ in size and shape—being separated from each other. One of these nuclei is the principal, vegetative or trophic nucleus; the other is an accessory nucleus, frequently termed the blepharoplast, flagellar or kinetic nucleus. One or more small basal granules are often present at or very near the origin of the flagella.

Multiplication is by fission, usually longitudinal, which may occur in either the free or encysted forms. Division is initiated by that of the nucleus or nuclei (especially the kinetic nucleus). The basal granule divides also. Collars and chromatophores, if present, likewise separate into two. Variation in the method of doubling the original number of flagella occurs. In most organisms, especially uniflagellate forms, the flagellum splits lengthwise, after division of the basal granule, blepharoplast and nucleus. The daughter flagella may be of the same or different lengths and thicknesses. Other flagellates at division are said to produce new flagella in the neighbourhood of the original ones. The daughter organisms in such cases are provided with one or more parental flagella in addition to newly formed ones. It has been stated that in certain cases the parent flagellate retains all its flagella, while new ones arise ab initio in the cytoplasm of the daughter forms.

Multiplication by longitudinal fission may be interrupted sooner or later by the production of gametes, which form zygotes, from which new generations of individuals arise. In many flagellates gamete formation and sporogony are unknown, and asexual reproduction by fission alone prevails.

Incomplete division results in the formation of colonies of individuals. These colonies must not be confused with the aggregation rosettes of flagellates found among the parasitic Mastigophora. The individuals of aggregation rosettes are capable of immediate separation from the rosette at will.

A number of parasitic Flagellata produce non-flagellate stages which are very resistant to external conditions, the assumption of which forms serves to protect the organisms during their transference from one host to another. Such non-flagellate forms possess one or more nuclei, are usually of an oval or rounded contour, and are capable of developing into the full flagellate on the return of more favourable conditions. These forms are often known as the post-flagellate stage of the organism. When ingested by a new host, the post-flagellate coat becomes more flexible, and the phase of the organism which now recommences growth is known as the pre-flagellate stage; it gradually develops into the typical flagellate organism.

Many Flagellata live free in fresh and salt water. They prefer stagnant water, rich in organic products of decomposition, such as puddles, swamps and pools. Those forms developing shells and colonies are, as a rule, adherent. A number of species are parasitic in man and animals, living mostly within the intestine or in the blood.

It is usual to classify the Flagellata in four orders: Euflagellata, Dinoflagellata, Choanoflagellata, and Cystoflagellata, of which only the Euflagellata are of interest to us. This is a group comprising numerous species, for the further classification of which the number and position of the flagella are utilised.

The Euflagellata observed in man belong to the Protomonadina as well as to the Polymastigina. The former possess either only one or two similar flagella, or one principal and one or two accessory flagella. The Polymastigina possess at least three flagella of equal size, or four to eight of unequal size, inserted at different points. An undulating membrane may be present in members of both groups.

It must also be pointed out that unicellular organisms with one or several flagella are not always classified with flagellates, for such forms occur in Rhizopods as well as temporarily in the lower plants. In addition, the examination of the flagellates, especially the parasitic species, is very difficult on account of their diminutive size and great activity; thus it happens that certain forms cannot with certainty be included in the group because their description is insufficient.

Order. Polymastigina, Blochmann.

The Polymastigina contains flagellates with three to eight flagella. Some of the Flagellata parasitic in man belong to the Polymastigina, and to two or three genera that are easily distinguishable.

Genus. Trichomonas, Donné, 1837.

The body is generally pyriform, the anterior part usually rounded, the posterior part pointed. There are at the anterior extremity three (? four) equally long flagella that are sometimes matted together. A blepharoplast (kinetic nucleus) and basal granule are present, together with a supporting structure known as an axial filament or axostyle. In addition there is an undulating membrane, bordered by a trailing flagellum, that commences at the anterior extremity and proceeds obliquely backwards. The nucleus, which is vesicular, is situated near the anterior extremity, and behind it are one or more vacuoles, none of which seems to be contractile. These flagellates are parasitic in vertebrate animals, and live chiefly in the intestine.

Trichomonas vaginalis, Donné.

The form of the body is very variable, and is elongate, fusiform or pear-shaped, also amœboid. The length varies between 15 µ and 25 µ, and the breadth between 7 µ and 12 µ. The posterior extremity is drawn out to a point and is about half the length of the remainder of the body. The cuticle is very thin and the body substance finely granular. At the anterior extremity there are three—some say four—flagella of equal length which are frequently united together, at least at the base, and are easily detached.

There is an undulating membrane (fig. ) which runs spirally across the body, arising from the place of insertion of the flagella, and terminating at the base of the caudal process. A cytostome seldom is recognizable in fresh specimens, but is apparently present. The nucleus is vesicular, elliptical and situated near the anterior extremity.

Multiplication takes place by division (Marchand). Encysted forms are almost unknown.

Trichomonas vaginalis lives in the vaginal mucus of women of various ages, not in normal mucus, but in mucus of acid reaction. It is found in menstruating females as well as in females who have passed the menopause. It occurs in pregnant and non-pregnant women, even in very young girls, provided always that they have a vaginal catarrh with acid reaction of the secretion. Should the acid reaction change, as, for instance, during menstruation, the parasites disappear, as they do likewise on injection of any alkaline fluid into the vagina. A low temperature (below +15° C.) is also fatal to the parasites. These flagellates can pass from the vagina through the urethra into the bladder, and produce severe catarrh, and are not easily removed.

T. vaginalis appeared to be a parasite specific to the female organs and not transmissible to man. However, several observations have since been made that confirm the occurrence of this species in the urethra of the male. The infection apparently takes place through coitus when changes are present in the urethral mucous membrane. At any rate, three cases observed point to this circumstance.

Attempts at experimental transmission to rabbits, guinea-pigs and dogs failed (Blochmann, Dock). So far, the manner in which women become infected is unknown.

Trichomonas intestinalis, R. Leuckart, 1879 = Trichomonas hominis, Davaine, 1854.

Some authors believe that a second trichomonad inhabiting man, Trichomonas intestinalis, R. Lkt., is identical with Trichomonas vaginalis, Donné. Leuckart’s species was based on the discoveries of Marchand (1875) and Zunker (1878), who stated that according to all appearances, and in their opinion, it was the same as Cercomonas intestinalis, Lambl, 1875 (nec 1859), which they found in the fæces of patients suffering from intestinal disorders. The organism is described by them as being pear-shaped and 10 µ to 15 µ in length and 3 µ to 4 µ in breadth. The posterior extremity terminated in a point (fig. 17).

A row of twelve or more cilia was said to commence at the anterior end and extend over the body. Leuckart stated that this parasite, placed by the two authors in the genus Cercomonas, was a Trichomonas, and that they mistook the undulating membrane for cilia, and overlooked the flagella. Notwithstanding its striking similarity with T. vaginalis, it was said to be distinguishable from that species by differences in the undulating membrane. Lambl’s C. intestinalis (of 1875) which corresponds with C. hominis, Davaine (1854), is regarded by Leuckart as a true Cercomonad (characterized by a flagellum and the absence of an undulating membrane, see p. ), and is thus generically distinct from Trichomonas.

The correctness of Leuckart’s judgment in regard to Marchand-Zunker’s flagellate was demonstrated by Grassi’s researches, accounts of which were published soon after. In about 100 cases of bowel complaints in North Italy and Sicily, Grassi found Flagellata in the stools, which he first named Monocercomonas and Cimænomonas, but later termed Trichomonas. However, in opposition to Leuckart, Grassi has also classified Davaine’s C. hominis (= C. intestinalis, Lambl, 1875) as Trichomonas, and most authors have followed his example. Hence arose the use of the name Trichomonas hominis. It was through Janowski (1896) that the former view was again taken up. After a review of the literature, the occurrence of Cercomonads in the intestine of human beings in addition to Trichomonads was considered by the author to have been proved, and he added a description of the Trichomonads. According to this, all morphological distinction between T. vaginalis, Donné, and T. intestinalis, Leuckart, disappeared. On the other hand, it is worthy of note that the smaller size, the more pear-shaped form, and the longer flagella differentiate T. intestinalis (= T. hominis) from T. vaginalis.

The easily deformed pear-shaped body has three free flagella anteriorly, and an undulating membrane with its flagellar border terminating in a short free flagellum posteriorly (figs. 17, 18). The undulating membrane may coil itself spirally round the body. A supporting rod or axostyle projects as a posterior spine. It appears to begin near the nucleus and blepharoplast, which are situated near the more rounded, anterior end of the body. There may be a chromatoid basal supporting line along the body for the undulating membrane. Rows of chromatoid granules are sometimes situated along one side of the axostyle. A cytostome may sometimes be seen. In mice, Wenyon (1907) found these parasites to vary in length from 3 µ to 20 µ. They occur in the cæcum and intestine of mice, where their internal structure seems more obvious than in man. The flagellates divide by longitudinal fission.

T. intestinalis, R. Leuckart, appears to be capable of settling in all parts of the human intestine in which the contents have an alkaline reaction. Trichomonads have been cited as occurring in the oral cavity by Steinberg, Zunker, Rappin and Prowazek; in the œsophagus by Cohnheim, and in the stomach by Strube, Cohnheim, Zabel, Hensen and Rosenfeld. The normal situation seems to be the small intestine. The parasites then appear in the dejecta, especially in various intestinal diseases the course of which is connected with an increased peristalsis. They are also found in healthy persons, from whom they are obtained after the administration of laxatives. They have been regarded by some workers as commensals, which, however, have the power of accelerating the onset of intestinal complaints, or at least of adding to them. They have been found in cases of carcinoma of the stomach, and in other diseases of that organ in which the acid reaction ceased.

Naturally, whether all the reports relate to the same species of Trichomonas must remain undecided. Certain authors (Steinberg, Cohnheim, van Emden) accept several species. Prowazek speaks of a variety of T. intestinalis inhabiting the oral cavity. This was distinguished by a posterior process exceeding the length of the body fourfold, and by a somewhat unusual course of the undulating membrane. The food of this form, which was found in the whitish deposit present, especially in the cavities of carious teeth, consisted almost exclusively of micrococci. Schmidt and St. Artault named the Trichomonads found in pathological products (e.g., gangrene, putrid bronchitis, phthisis) of the lungs of man, as Trichomonas pulmonalis. Trichomonads have also been found by Wieting in lobular pneumonia in the lungs of pigs.

It is still uncertain in what way the infection takes place. Experiments in the transmission of free trichomonads to mammals (per os), in which the same or allied species occur (guinea-pigs, rats, apes), have been without result. Probably encystment is necessary. Such conditions are mentioned by May, Künstler, Roos, Schurmayer, van Emden, Prowazek, Galli-Valerio and Schaudinn. According to Prowazek, intestinal trichomonads of rats become encysted for conjugation. In the cyst an accumulation of reserve food material occurs, causing distension. The nuclei of the conjugants each give off a reduction body and, after fusion, produce the nuclei for the daughter individuals. According to Schaudinn the intestinal trichomonads lose their flagella before conjugation, become amœboid and encyst in twos, the formation of a large agglomeration of reserve substance accompanying this. Galli-Valerio found double-contoured cysts in the fæces of trichomonad-infected guinea-pigs, after the fæces had been kept for a month in a damp chamber. When exposed to heat small flagellates escaped from them. Administration of such material containing cysts resulted in severe infection with trichomonads, and death of the experimental guinea-pigs followed. The cyst wall is clearly a protection against the deleterious acid reaction of the stomach contents. Alexeieff (1911) and Brumpt (1912) think that the trichomonad cysts of man are really fungi, while other workers also doubt encystment among trichomonads. Wenyon (1907) states that T. intestinalis in mice produces spherical contracted forms which escape from the body in the fæces.

Air, water, and under certain circumstances even food may be regarded as vectors for the trichomonads. The occurrence of the organisms in the oral cavity, and still more so in the lungs, is in favour of the air being the transmitting agent. An observation made by Epstein supports the idea of water transmission. Multiplication of the trichomonads, once they have gained access to the body, is effected by longitudinal division commencing at the anterior end (Künstler). “Cercomonads” with several flagella and an undulating membrane, as well as trichomonads, have been observed by Ross in some cases of cutaneous ulcers.

Mello-Leitao (1913) has described flagellate dysentery in children in Rio de Janeiro. He states that it is due to T. intestinalis and Lamblia intestinalis either separately or together. Flagellate dysentery, he thinks, is benign and is the most frequent form of dysentery in infants. The flagellates are pathogenic to infants under three years of age. Escomel (1913) found 152 cases of dysentery in Peru due solely to Trichomonas. Such cases are probably widespread.

Genus. Tetramitus, Perty, 1852.

Tetramitus mesnili, Wenyon, 1910.

Syn.: Macrostoma mesnili, Chilomastix mesnili, Fanapapea intestinalis.

The genus Tetramitus differs from Trichomonas in possessing an undulating membrane inserted in a deep groove or cytostome. There are three anterior flagella. The pear-shaped organism measures 14 µ by 7 µ, but smaller examples occur. T. mesnili occurs in the human intestine, having been described by Wenyon (1910) from a man from the Bahamas in the Seamen’s Hospital, London. Its occurrence is widespread. Alexeieff considers that Macrostoma and Tetramitus are synonymous. The parasite is the same as Fanapapea intestinalis, Prowazek, 1911, from Samoa. Brumpt (1912) found T. mesnili to be the causal agent of colitis in a Frenchwoman. Nattan-Larrier (1912) considers it of little pathological importance.

Gäbel (1914) described an interesting case of seasonal diarrhœa acquired in Tunis, in which a new Tetramitid was the causal agent. The organism was pear-shaped, without an undulating membrane, and measured 6·5 µ to 8 µ by 5 µ to 6 µ. The cytostome was large, and there was no skeletal support. Encystment occurred. Gäbel named the organism Difämus tunensis and considered that it was pathogenic.

Genus. Lamblia, R. Blanchard, 1888.

Syn., Dimorphus, Grassi, 1879, nec Haller, 1878; Megastoma, Grassi, 1881, nec de Blainville.

The body is pear-shaped, with a hollow on the under surface anteriorly. It has four pairs of flagella directed backwards, of which three pairs lie on the borders of the hollow disc, and the fourth arises from the pointed posterior extremity.

Lamblia intestinalis, Lambl, 1859.

Syn.: Cercomonas intestinalis, Lambl, 1859 (nec 1875); Hexamitus duodenalis, Davaine, 1875; Dimorphus muris, Grassi, 1879; Megastoma entericum, Grassi, 1881; Megastoma intestinale, R. Blanch., 1886; Lamblia duodenalis, Stiles, 1902.

The organism is pear-shaped and bilaterally symmetrical. It is from 10 µ to 21 µ long and 5 µ to 12 µ broad and possesses a thin cuticle. Anteriorly an oblique depression is present, which functions as a sucking disc (fig. , s). Its edges are raised above the general surface and are contractile. It corresponds to a peristome and acts as an adhesive organ (fig. , b, c). No true cytostome is present. A double longitudinal ridge, representing axostyles, extends from the sucking disc to the tapering posterior extremity, which is prolonged as two flagella from 9 µ to 14 µ long.

Lamblia intestinalis possesses eight flagella (fig. 19). The first pair of flagella, which cross one another, arise in a groove formed by the anterior edge of the sucking disc. Two pairs of flagella (lateral and median) are inserted on the posterior edge of the disc, while the posterior flagella occur at the tapering posterior extremity of the body. Basal granules are found at the bases of the flagella. The median flagella are most active in movement, the anterior and lateral flagella being less motile, as they are partially united to the body for part of their length.

The nuclear apparatus is situated in the thin, anterior, hollowed part of the body. It is at first dumb-bell shaped, the “handle” of the dumb-bell being formed by a very slight connecting strand, which eventually separates, so that the flagellate becomes binucleate, and thus completes the general bisymmetry of the organism.

There is a karyosome in each nucleus. Other bodies of unknown function, and possibly composed of chromatin, occur on or near the axostyles.

Division has not been observed in the flagellate stages of the Lamblia, but it occurs within the cysts. The resistant cysts (fig. , e) are oval and are surrounded by a fairly thick, hyaline cyst wall. They measure 10 µ to 15 µ by 7 µ to 9 µ, and may be tetranucleate. According to Schaudinn, the cysts arise from the conjugation of two individuals, and nuclear rearrangement occurs.

L. intestinalis occurs in its flagellate stage in the duodenum and jejunum, and rarely as such in the other parts of the intestine. Normally it is found in the large intestine as cysts, which are voided with the fæces. The hosts of Lamblia include Mus musculus, M. rattus, M. decumanus, M. silvestris, Arvicola arvensis and A. amphibius, the dog and cat, rabbit, sheep and man. Cysts voided with the fæces of infected animals reach plants or drinking water, and thence are transferred to man.

The flagellate in these different hosts exhibits some variation in size and in the problematic chromatic bodies. Bensen has suggested the species L. intestinalis from man, L. muris from the mouse and L. cuniculi from the rabbit. It is not certain whether these different species are necessary, as the variation may be due to differences of environment.

Like Trichomonas, Lamblia can multiply under inflammatory conditions of the alimentary tract. Thus they are found in cases of diarrhœa, carcinoma of the stomach, etc. The parasites attach themselves by their sucking discs to the epithelial cells of the gut (fig. 20, c), and though their numbers may be very great, their direct pathological significance is not fully known. Their occurrence in cases of diarrhœa has been explained as being due to the increased peristalsis, which has detached the parasites from the epithelium. Free flagellate forms perish in stools if kept, more especially if the temperature falls below 0° C. or rises above 40° C. Lamblia has often been found in dysenteric diseases, especially in the East, and is said to be the causal agent of certain diarrhœas in India. Mathis (1914) found Lamblia in cases of diarrhœa with dysenteriform stools in Tonkin. He also discovered healthy carriers of Lamblia cysts.

The parasite under discussion was first observed by Lambl (1859) in the mucous evacuations of children. He regarded the parasite as a Cercomonad and termed it Cercomonas intestinalis, which name as a rule is applied to Cercomonas hominis, Davaine, although Stein had already pointed out the difference between the two species. Grassi (1879) observed this species first in mice (calling it Dimorphus muris), and subsequently in human beings in Upper Italy and named it Megastoma entericum. Bütschli and Blanchard then laid stress on the identity of this species with Lambl’s C. intestinalis (1859), and consequently called it Megastoma intestinale. Later, Blanchard drew attention to the circumstance that the generic name Megastoma chosen by Grassi had already been used four times for various kinds of animals, and established the genus Lamblia. Accordingly, L. intestinalis is the valid name, and should be generally adopted.

In Upper Italy the parasite in the encysted condition has also been seen by Perroncito in man. At the same time, Grassi and Schewiakoff began a new investigation of specimens from mice and rats. In Germany, L. intestinalis was found by Moritz and Hölzl, Roos, Schuberg and Salomon. Moritz and Hölzl confirmed the relative frequency of the species. In Königsberg, Prussia, a student found encysted Lamblia in his fæces. One case was reported from Finland by Sievers, another case from Scandinavia by Müller. Frshezjesski and Ucke reported cases from Russia. Jaksch announced the occurrence of the parasite in Austria; Piccardi mentioned their presence again in Italy. They were reported from Egypt by Kruse and Pasquale, and from North America (Baltimore) by Stiles. Noc stated that 50 per cent. of the population of Tonkin harboured Lamblia. Finally, the structure of L. intestinalis has been described by Metzner (1901), and by Wenyon (1907) in mice.

In all these cases L. intestinalis has been observed in the small intestine, or in the evacuations of patients with intestinal diseases. It has also been found in the intestine of healthy subjects. Just as Trichomonas intestinalis may be found inhabiting the stomach in diseases of that organ, in which an alkaline reaction is present (carcinoma), so has L. intestinalis been found to occur under similar circumstances (Cohnheim, Zabel). However, in Schmidt’s case, 1 per cent. hydrochloric acid was certainly stated to be present. Infection takes place by the ingestion of cysts (fig. , e), as was established by Grassi, experimentally on himself. Cereal food-stuffs, contaminated with Lamblia cysts from vermin of the locality, such as rats and mice, serve to convey the infection to man. Such cysts may probably be found in street-dust, etc. Stiles induced infection in guinea-pigs, and Perroncito in mice and rabbits, by means of cysts of Lamblia from human beings. Stiles suspected that flies could transport Lamblia cysts. Mathis (1914) found that L. intestinalis was not amenable to emetine, at any rate in its cystic stage.

Order. Protomonadina, Blochmann.

The smallness of the Protomonadines and their less superficial situation than the Polymastigines, may be the cause that so far as the species occurring in man are concerned, they were formerly less well known. As regards parasitic species, this group may be divided as follows, according to the number of flagella and the presence or absence of an undulating membrane:—

(1) Cercomonadidæ, with one flagellum at the anterior extremity, without an undulating membrane.

(2) Bodonidæ, with two flagella, without an undulating membrane, except in Trypanoplasma.

(3) Trypanosomidæ, with one flagellum, and an undulating membrane along the length of the body in some genera.

Family. Cercomonadidæ, Kent emend. Bütschli.

Small uniflagellate forms, without cytostome.

Genus. Cercomonas, Dujardin emend. Bütschli.

Oval or rounded organisms, with the aflagellar end often drawn out into a tail-like process.

Cercomonas hominis, Davaine, 1854.

Davaine found flagellates in the dejecta of cholera patients. They had pear-shaped bodies, lengthening to a point posteriorly. Their length was from 10 µ to 12 µ, and a flagellum about twice as long as the body projected from one extremity (fig. 21). A nucleus was hardly recognizable. Occasionally a somewhat long structure (cytostome?) appeared at the anterior extremity. The animals moved with remarkable activity. They also attached themselves by means of their posterior extremities and swung about around the point of attachment. Davaine found a smaller variety, only about 8 µ long, in the dejecta of a typhoid patient (fig. 21, b).

The Flagellata observed by Ekeckrantz (1869) in the intestine of man belong to this form—at least to the larger variety—and Tham (1870) reported fresh cases soon after. Lambl’s publication of 1875, which was written in Russian, and became known through Leuckart’s work on parasites, also alludes to apparently typical Cercomonads, which, however, were discovered, not in the intestine, but in an Echinococcus cyst in the liver (fig. 22). The elliptical, fusiform, rarely pear-shaped or cylindrical bodies of the parasites measured 5 µ to 14 µ in length, and were provided with a flagellum at one end, while the other extremity usually terminated in a long point. An oral aperture occurred at the base of the flagellum, and there were one or two vacuoles near the posterior extremity. Longitudinal division was also observed (fig. 22).

As already mentioned, this form, which Lambl termed Cercomonas intestinalis, differs considerably from the form found by the same author in 1859, which received the same designation (cf. Lamblia intestinalis, p. ), but it corresponds with Cercomonas hominis, Davaine. The latter, as well as C. intestinalis, Lambl, 1875, is usually classed with the Trichomonads, but, as has already been remarked (cf. Trichomonas intestinalis, p. ), this cannot be considered correct, as only one flagellum is present.

Cercomonas vaginalis (Castellani and Chalmers, 1909) was found in the vagina of native women in Ceylon.

Other species of Cercomonas have, at various times, been recorded from man. However, the parasitic species of the genus Cercomonas require further investigation.

According to Janowski (1896–7), typical Cercomonads have also been observed in the intestine of man by Escherich, also by Cahen, Massiutin, Fenoglio, Councilman and Lafleur, Dock, Kruse and Pasquale, Zunker, Quincke and Roos, and others. However, it is an open question whether the Flagellata observed by Roos in one of his cases belonged to Davaine’s species, the size showing some deviation (14 µ to 16 µ). In his, as in many other cases, doubts have been raised as to whether the flagellates found in the stools had actually lived in the intestine, or had subsequently appeared in the fæces: for this a surprisingly short time only is necessary. Salomon also appears to have observed Cercomonads (Berl. klin. Wochenschr., 1899, No. 46).

As with T. intestinalis so with C. hominis, it appears that the parasite settles not only in the intestine but also in the air-passages. This is demonstrated by the statements of Kannenberg and Streng of the occurrence of Monads and Cercomonads in the sputum and putrid expectoration in gangrene of the lungs, which no doubt apply to C. hominis (cf. also Artault). Possibly also the Flagellata observed in the pleural exudation by Litten and Roos may be included here; this is the more probable in Roos’s case as the process ensued in the pleura after the breaking through of a vomica.

Perroncito and Piccardi have described encysted stages of Cercomonads.

Monas pyophila, R. Blanch., 1895.

R. Blanchard thus designates a Flagellate that Grimm found in the sputum, as well as in the pus of a pulmonary and hepatic abscess, in the case of a Japanese woman living in Sapporo. The parasites resemble large spermatozoa (fig. 23). The body, 30 µ to 60 µ, has the shape of a heart or a myrtle leaf, and is surrounded by a thick cuticle which is supposed to extend into the interior of the body, dividing it into three parts. A long appendix at the rounded pole is covered for the greater part of its length by the cuticle; the extremity, however, is free and resembles a flagellum. The parasites were very active, frequently changed their shape, and were able to retract the long appendix within the body, which then assumed a round form.

[This organism requires further investigation.]

Family. Bodonidæ, Bütschli.

Protomonadina which are either free-living or parasitic, with two dissimilar flagella, while the possession of an undulating membrane and of a kinetic nucleus or blepharoplast is variable.

There are three genera:—

1. Bodo, Stein, 1878, without a kinetic nucleus and undulating membrane.

2. Prowazekia, Hartmann and Chagas, 1910, with a kinetic
nucleus and without an undulating membrane.

3. Trypanoplasma, Laveran and Mesnil, 1901, with a kinetic
nucleus and undulating membrane.

Of these genera Prowazekia must be discussed. Bodo does not occur in man. Species of Trypanoplasma occur in the blood and in the gut of various fishes, in the seminal receptacle of certain snails, in the gut and genitalia of a flatworm (Dendrocœlum lacteum) and in the vagina of a leech. Closely allied to Trypanoplasma is the genus Trypanophis, parasitic in the cœlenteric cavity of Siphonophores.

Genus. Prowazekia, Hartmann and Chagas, 1910.

The genus was founded for a flagellate parasite, Prowazekia cruzi, discovered in a culture of human fæces in Brazil. Various other species have been referred thereto. The genus is separated from Bodo by the possession of a second nucleus, the so-called kinetonucleus or blepharoplast. It differs from Trypanoplasma in the absence of an undulating membrane. It is heteromastigote, that is, it possesses two dissimilar flagella, one anteriorly directed and the other lateral and trailing.

The principal species are:

Prowazekia urinaria, Hassall, 1859.

Syn.: Bodo urinarius, Hassall, 1859; Trichomonas irregularis, Salisbury, 1868; Cystomonas urinaria, Blanchard, 1885; Plagiomonas urinaria, Braun, 1895.

Hassall in 1859 first found Bodo-like flagellates in human urine. He examined fifty samples of urine from patients suffering from albuminuria and from cholera. The reaction of the urine was alkaline or sometimes only feebly acid. The flagellates were only seen after the urine had been standing for several days. Hassall named the organism Bodo urinarius, and gave a very good description of it with illustrations. The flagellate, which was round or oval, measured 14 µ by 8 µ. The organism had “one, usually two, and sometimes three lashes or cilia.” In 1868 Salisbury described a similar flagellate in the urine under the name Trichomonas irregularis. Künstler in 1883 described the latter parasite under the name B. urinarius. In 1885 Blanchard, considering Künstler’s organism a different parasite from Hassall’s, called it Cystomonas urinaria. Braun, in 1895, gave the name Plagiomonas urinaria. Barrois (1894) considered Künstler’s and Hassall’s organisms to be identical and not to be true parasites of man. Sinton, in 1912, found the flagellate in the deposit, after centrifuging, of a 24-hour old specimen of alkaline urine from a Mexican sailor in the Royal Southern Hospital, Liverpool. Sinton found a kinetic nucleus or blepharoplast in the organism, and therefore placed it in the genus Prowazekia.

The flagellate stage (fig. 24) of the organism is polymorphic, and may be either (a) sausage-shaped, 10 µ to 25 µ in length by 2·5 µ to 6 µ in breadth; (b) round or oval, varying from 4 µ in diameter to oval forms 15 µ by 10 µ; (c) a carrot-shaped form, of varying size up to 25 µ by 4 µ. The kinetic nucleus is large and pear-shaped. Near it are basal granules, closely applied to one another, from which the flagella arise. There is a small cytostome near the roots of the flagella. There is a well-marked karyosome in the nucleus. The movement is jerky. The shorter, anterior flagellum may be used in food-capture. In life, bacteria have been seen to be ingested. Food-vacuoles tend to accumulate at the posterior (aflagellar) end. A contractile vacuole may be present, near the base of the cytostome, and may really be the dilated fundus of the latter. Division occurs by binary fission. The organism can encyst (fig. 25, a), when the flagella are lost, and round or oval cysts are found, 5 µ to 7 µ in diameter. After a time flagella are formed inside the cyst, and the organism emerges therefrom in its typical flagellate form (fig. 25, b-f).

Sinton’s case is interesting. He obtained the flagellate only twice from the same patient, a Mexican then in hospital in Liverpool. The flagellate was not found in the patient’s fæces, nor was it found in the urine on later occasions when taken aseptically.

In cultures Prowazekia urinaria was always found in association with bacteria. The cultures died at a temperature of 37° C., but grew well at 20° C. Various media were useful at the lower temperature, such as urine, salt agar, nutrient agar, serum agar, blood agar, peptone salt solution, and diluted blood serum. The flagellate was, then, considered to be an accidental contamination and not a true parasite of human urine.

Prowazekia asiatica, Castellani and Chalmers, 1910.

The flagellate was found by the discoverers in the stools of patients suffering from ankylostomiasis and diarrhœa in Ceylon. It was referred by them to the genus Bodo, but in 1911 Whitmore further studied it and placed it in the genus Prowazekia. In the stools the flagellate is found either as a long, slender form measuring 10 µ to 16 µ by 5 µ to 8 µ or as a rounded form 8 µ to 10 µ in diameter. Its cytoplasm is alveolar. A rhizoplast connects the basal granules to the kinetic nucleus. There is multiplication and cyst formation as before. The organism is easily cultivated, especially in the condensation water of nutrose agar and maltose agar. The pathogenicity is stated to be nil.

Prowazekia javanensis, Flu, 1912.

Found in agar cultures from the motions of patients at Weltevreden, Dutch East Indies. The flagellates are 12 µ long and 5 µ broad. The lateral flagellum is stated to be attached to the cell body for a short distance. Regarding the karyosome in the nucleus, the author states that the smaller the karyosome the more chromatin is deposited on the nuclear membrane. Flu mentions that the specific name javanensis is a temporary one, as in the course of time it may be shown that there is only one species of Prowazekia.

Prowazekia cruzi, Hartmann and Chagas, 1910.

Found in a culture from human fæces on an agar plate in Brazil, and considered to be a free-living form. The organism is oval or pear-shaped, 8 µ to 12 µ long and 5 µ to 6 µ broad. In human stools at Tsingtau, China, a Prowazekia has been found by Martini which he thinks is the same as Prowazekia cruzi. He considers it to be a cause of human diarrhœa and intestinal catarrh.

Prowazekia weinbergi, Mathis and Léger, 1910.

This species was found in the fæces of men, both healthy and diarrhœic, in Tonkin. It is pear-shaped, 8 µ to 15 µ long by 4 µ to 6·5 µ broad. The flagella occur at the broad end.

The discoverers think that Prowazekia weinbergi is an intestinal inhabitant, but non-pathogenic, since it was found to occur in the fæces even when obtained with aseptic precautions.

Prowazekia parva, Nägler, 1910.

A free-living form found in the slime on the stones at the biological station at Lunz. Another Prowazekia was found in 1914 in tap-water in Calcutta.

Family. Trypanosomidæ, Doflein.

The Trypanosomidæ, broadly considered, are uniflagellate organisms, the flagellum being at the anterior end. The flagellum arises near the blepharoplast (kinetic nucleus), which lies anterior, near or posterior to the nucleus.

The following genera will be considered:—

Trypanosoma—with an undulating membrane along the length of the body.

Crithidia—with a less well-developed undulating membrane anteriorly (see fig. ).

Herpetomonas—including the so-called Leptomonas, with anterior free flagellum only, and no undulating membrane.

Leishmania—non-flagellate forms in mammalian blood, flagellate herpetomonad stages in culture, probably occurring naturally in Arthropods.

Genus. Trypanosoma, Gruby, 1843.

The members of the genus possess a single flagellum, which arises posteriorly, adjacent to a blepharoplast or kinetic nucleus. The flagellum forms a margin to an undulating membrane, and may or may not be continued beyond the body as a free flagellum. Many species are parasitic in vertebrate blood and in the digestive tracts of insects.

Historical.

The history of blood flagellates goes back to the year 1841, in which Valentin discovered in the blood of a brook-trout (Salmo fario L.) minute bodies, from 7 µ to 13 µ in length, with active movements and presenting marked changes in form. Valentin considered the parasite a new species of the old genus Proteus or Amœba, Ehrbg. This announcement led Gluge (1842) to publish a similar discovery he had made in frog’s blood. The latter forms were called by Mayer (1843) Amœba rotatoria, Paramœcium loricatum and P. costatum, while Gruby (1843) called them Trypanosoma sanguinis. Later it was discovered that similar organisms occurred also in the blood of birds (Wedl (1850), Danilewsky) and of mammals. Gros (1845) found them in the mouse and mole, Chaussat (1850) in the house rat, Lewis (1879) in the Indian rat, Wittich (1881) in the hamster. Danilewsky (1886–89) and Chalachnikow (1888) investigated the structure and division of trypanosomes.

In the case of all these forms, there was no discussion as to a pathogenic influence on the host. Opinion, however, as to the action of trypanosomes changed when, in 1880, Evans found flagellates in the blood of horses in India that suffered from a disease endemic there called “surra,” and associated the parasites with the disease. Steel and Evans were successful in transmitting the parasites—first known as Spirochæta evansi, Steel, then as Trichomonas evansi, Crookshank, and finally as Trypanosoma evansi—to dogs, mules and horses. They recognized that the above mentioned flagellates in the blood of the experimental animals were the causal agents of the disease.

From that time there was a considerable increase in the literature, the contents of which have been summarized by Laveran and Blanchard. In 1894 Rouget discovered trypanosomes in the blood of African horses that suffer from “stallion’s disease” (dourine). In 1894 Bruce found similar forms (T. brucei) in the blood of South African mammals suffering from “nagana,” and in consequence attention was drawn to the part which the much dreaded tsetse-fly played in the transmission of “nagana.” In 1901 Elmassian discovered trypanosomes in the blood of horses that were stricken with “mal de caderas,” which is very common in the Argentine. The disease in cattle named “galziekte” (gall-sickness), occurring in the Transvaal, was also at one time attributed to a trypanosome remarkable for its great size, and like some other species, bearing the name of its discoverer (T. theileri).

The study of the species hitherto known has been carried on partly by the above mentioned authors and in part by others, e.g., Rabinowitsch and Kempner, Laveran and Mesnil, Wasiliewski, Senn. It was greatly advanced by the method of double staining (with alkaline methylene blue and eosin) introduced by Romanowsky (1891) and elaborated by Ziemann, Leishman, Giemsa and others. By this means the presence of a terminal flagellum and of an undulating membrane at the side of the flattened and extended body was demonstrated. Laveran and Mesnil (1901) discovered allied flagellates in the blood of the fish, Scardinius erythrophthalmus. These flagellates, now placed in the genus Trypanoplasma, had a second free flagellum in addition to the one bordering the undulating membrane. Trypanoplasms have since been found in both freshwater and marine fishes. The transmission of trypanoplasms of freshwater fishes is effected by leeches. Trypanoplasma varium from Cobitis is transmitted by Hemiclepsis marginata according to Léger, while the Trypanoplasmata of Cyprinus carpio and Abramis brama reach new hosts by the agency of Piscicola according to Keysselitz.

Another ally of the Trypanosomidæ, Trypanophis, lives in the cœlenteric cavity of Siphonophores. It has also an extra terminal flagellum (Poche, Keysselitz). [Trypanoplasma and Trypanophis belong to the Bodonidæ, see p. ].

Finally it was shown that Trypanosomes occurred in human beings. Although Nepveu’s early report of trypanosomes in the blood of malarial patients may be doubtful, subsequent researches by Forde and Dutton demonstrated trypanosomes (fig. ) in the blood of a European, apparently suffering from malaria, living in the Gambia. Dutton (1902) called the human trypanosome, T. gambiense. The expedition despatched by the Liverpool School of Tropical Medicine (1902) to Senegambia found trypanosome infections in six cases among a thousand inhabitants examined.

About the same time attention was devoted to the disease of West African negroes known for a century as “sleeping sickness.” Castellani (1903) was the first to succeed in demonstrating the presence of trypanosomes (at first called T. ugandense) in centrifugalized cerebro-spinal fluid obtained by puncture from cases of sleeping sickness in Uganda. Similar discoveries were made by Bruce, who also found trypanosomes in the blood of those attacked with sleeping sickness. Sambon regarded a species of Glossina as the transmitter. From consideration of the geographical distribution of the disease Christy regarded Glossina palpalis as the transmitter. Brumpt first thought it was G. morsitans, but, later, supported the view of G. palpalis. Bruce, Nabarro and Greig also named the same insect as the transmitter, not only for geographical reasons but also because healthy apes became infected by the bite of certain G. palpalis. The inoculation of cerebro-spinal fluid from subjects of sleeping sickness into the spinal canal of apes (Macacus) had the same result.

Just as the discovery of the malarial parasites called forth a whole flood of research memoirs which were followed by a second series on the relation of the mosquitoes to malaria, so a similar outpouring occurred after the discovery of the pathogenic trypanosomes of mammals and men. In both cases the inquiry was not limited to the stages in man and other vertebrate hosts, but the fate of the parasites in the intermediate (invertebrate) hosts was investigated, and allied species were obtained from many different hosts.

Novy and MacNeal (1903) were the first to cultivate trypanosomes in artificial media (blood-agar).

In 1910 Stephens and Fantham recorded the presence of another human trypanosome, T. rhodesiense, from a case of sleeping sickness in Rhodesia, where G. palpalis was absent. Kinghorn has since demonstrated that T. rhodesiense is transmitted by G. morsitans. Kinghorn and Yorke believe that big game (e.g., antelope) is the reservoir of T. rhodesiense.

The output of literature on trypanosomiasis in men and animals is enormous. To cope with it the Sleeping Sickness Bureau Bulletin was founded in 1908, and it is now (since November 1912) continued as a section of the Tropical Diseases Bulletin, wherein current literature is reviewed.

General.

Trypanosomes occur in the blood of representatives of all the vertebrate classes. Often the trypanosomes occur so scantily in the blood that they are overlooked on examination. A useful aid in detecting the flagellates in such cases consists in the use of cultures of the blood of the host on artificial media. Stimulated by the medium multiplication occurs, and hence the parasites are more easily detected. [For the composition of such culture media see Appendix.]

There is a periodicity in the appearance of the trypanosomes in the peripheral blood of the host, due to alternating phases of multiplication and of rest on the part of the parasites. Such periodicity has been established both by biological and enumerative methods. Again, a seasonal variation has been observed in the occurrence of certain trypanosomes in the peripheral circulation of the hosts; for example, some trypanosomes (e.g., T. noctuæ in birds) are found only in the summer in the blood, while in the winter they occur in the internal organs.

Recent cultural researches have established that trypanosomes, e.g., T. americanum, may be present in very small numbers in hosts, such as cattle, which are quite unharmed by them, and in which the presence of these flagellates formerly was never suspected (“cryptic trypanosomiasis.”) However, the majority of the trypanosomes occurring in domestic animals are usually deleterious or even lethal to their hosts. Many wild animals, such as various species of antelope, harbour trypanosomes without being injured thereby. In such cases it is probable that the vertebrate hosts have been so long parasitized in the past, that they have become tolerant and immune to the effects of the flagellates. Should such trypanosomes of wild animals be transmitted to domesticated stock or man, they may re-acquire their initial virulence and become pathogenic to the new host. As a general statement, the newer a parasite is to its host the greater is its virulence. For example, T. gambiense, T. rhodesiense and T. brucei are innocuous to big game in Africa, but are pathogenic to man and domestic animals respectively. Pathogenic trypanosomes appear to have a wider range of hosts, that is, to be less limited to one specific host than non-pathogenic forms. Thus, T. rhodesiense is pathogenic to man and all laboratory animals, while it is non-pathogenic to antelopes and their kind.

Morphology.

The general structure of the various trypanosomes shows much uniformity, though variations in size and shape occur. Typically the body is elongate and sinuous. The flagellar end tapers gradually to a point, the aflagellar extremity usually being rounded or more blunt. In some trypanosomes there is much diversity in size, the organisms varying from long, slender forms to short, stumpy ones; in other species relative constancy of size is maintained. The former are known as polymorphic trypanosomes, the latter as monomorphic forms.

Two nuclei are present. The main or principal nucleus, sometimes termed the trophic nucleus, is often situated towards the centre of the body; it is frequently of the vesicular type, containing a karyosome. The blepharoplast or kinetic nucleus is posterior to the nucleus, and usually is rod-like. The flagellum arises close to the blepharoplast, and forms an edge to the undulating membrane. It may or may not extend beyond the limits of the undulating membrane. If it does so, the unattached part is known as the free flagellum. Sometimes a small granule is found at the origin of the flagellum. This is the basal granule, and is considered by some to function as the centriole of the kinetic nucleus.

The undulating membrane is a lateral extension of the ectoplasm or periplast, and is the main agent in locomotion. It is edged by the flagellum, which forms a deeply stainable border to it. Within the membrane substance, often arranged parallel with its edge, are a number of fine contractile elements, the myonemes. These contractile elements may also occur on the body of the trypanosome. They are easily seen in some large trypanosomes, but are difficult of demonstration in others, owing to their great fineness.

Multiplication of trypanosomes in the blood is brought about by binary longitudinal fission (fig. ). Division is initiated by that of the blepharoplast and nucleus. The division may be equal or subequal, whereby differences in size of individuals partly arise. Multiple division by repeated binary fission, without complete separation of the daughter forms, is known in some trypanosomes (e.g., T. lewisi), and rosettes of parasites thereby are produced (fig. 27).

The classification of trypanosomes is very difficult. Laveran (1911) has suggested the examination of the relative length of the flagellum as a diagnostic character, and so arranged these flagellates in mammals in three groups. The first group included those trypanosomes always having part of the flagellum free (e.g., T. evansi, T. vivax); the second group comprised forms without a part of the flagellum free (e.g., T. congolense), while the third group included forms some members of which have free flagella, while others have not (e.g., T. gambiense). Bruce (1914) and Yorke and Blacklock (1914) have also devised classifications.

Resting stages of some trypanosomes have been found in the internal organs of their vertebrate hosts. The formation of these oval, Leishmania-like bodies will be noted in individual cases later. Similar small oval bodies form an important phase in the life-history of T. cruzi, which multiplies normally by multiple fission or schizogony into these oval, daughter elements, and not by binary longitudinal fission in the circulating blood.

Polymorphism in trypanosomes (e.g., T. gambiense, T. rhodesiense) is now interpreted as a phenomenon resulting from growth and division. Long, thin forms are those about to divide. Fully mature forms are shorter and broader. Various intermediate types occur and represent growth forms. Formerly, polymorphism was interpreted in terms of sex, thin forms being regarded as males, broad forms as females, while the intermediate types were termed indifferent. Conjugation was not observed, and there is no evidence in support of the sexual interpretation.

The transmission of trypanosomes from one vertebrate host to another is usually accomplished by the intermediation of some biting arthropod in the case of terrestrial animals, while leeches are usually considered to act as transmitters in the case of the trypanosomes occurring in aquatic animals. Developmental phases of the life-histories of trypanosomes occur in the invertebrate transmitters, and will be considered in individual cases.

Trypanosoma gambiense, Dutton, 1902.

Syn.: Trypanosoma hominis, Manson, 1903. Trypanosoma nepveui, Sambon, 1903. Trypanosoma castellanii, Kruse, 1903. Trypanosoma ugandense, Castellani, 1903. Trypanosoma fordii, Maxwell Adams.

In vertebrate blood Trypanosoma gambiense is polymorphic, for long, thin forms may be seen in contrast with short, stumpy forms, as well as intermediate forms (fig. , a—c). This polymorphism has been interpreted in terms of sex, especially by German investigators, following Schaudinn (see above). However, there is no evidence of conjugation, and the polymorphic forms are more easily interpreted in terms of growth and division, for the long thin forms are potential dividing organisms, and the stumpy or short parasites, with little or no free flagellum, are the adult individuals.

Morphology of T. gambiense in the Circulating Blood.

T. gambiense varies from 13 µ to 36 µ in length, its average length being 24·8 µ, as was determined in 1913 by exact biometrical methods by Stephens and Fantham. Three forms of parasite occur. According to Miss Robertson, the relatively short forms from 13 µ to 21 µ long may be regarded as the mature or “adult” type of parasite in the blood. They carry on the cycle in the vertebrate. From them intermediate forms, which are longer than the “adult” but at first have the same breadth, arise by growth. They possess a free flagellum. The intermediate forms grow into long individuals, which are those about to divide. The products of division give rise, directly or indirectly, to the adult forms.

The organism has an elongate body with an anterior or flagellar end and a blunter posterior or non-flagellar end. The protoplasm is finely granular, large inclusions being rare. The central nucleus is oval and large, often containing most of its chromatin concentrated as a karyosome, with small granules only scattered near or on the fine nuclear membrane. The blepharoplast is either rounded or rod-shaped. The undulating membrane is thrown into folds and is bordered by the flagellum. A small basal granule may be present near, or at the actual origin of the flagellum.

Multiplication in the vertebrate is brought about by longitudinal division. According to the recent account of division by Miss Robertson, the blepharoplast doubles, then the flagellum splits for the greater part of its length, and the daughter flagella separate, one being shorter than the parent flagellum. The nucleus often shows two well marked dark granules on the membrane at opposite poles, and these appear to act as centrosomes. Nuclear constriction occurs and the halves gradually separate. Finally the two daughter organisms become free, the aflagellar end splitting last. The products of division may be equal or unequal. Repeated division goes on in the general circulation until the blood swarms with parasites. Then the trypanosomes gradually disappear, and a period occurs when it is practically impossible to demonstrate the parasite in the blood. At such a period, trypanosomes can be obtained by puncture of the enlarged lymphatic glands or of the spinal canal, or can be found in the internal organs, more particularly in the spleen, lungs, liver and bone-marrow. In the latter organs, latent bodies are produced (fig. 29, d—f) which are capable of again becoming flagellates and entering the general circulation. Their formation was described by Fantham (1911). The parasite contracts, the blepharoplast migrates towards the nucleus, a very thin coat differentiates around the two nuclei and a certain amount of cytoplasm, and the parts exterior to the coat disintegrate, leaving a small, oval body behind. Fuller details are given in connection with T. rhodesiense. Laveran (1911) considers that latent bodies are “involution” forms, but acknowledges that they can flagellate and become infective in fresh blood.

No multiplication of the trypanosomes within the cells of the lung, liver or spleen of infected monkeys was found by Miss Robertson in her recent researches.

There appear to be negative periods in infected monkeys, since, although trypanosomes may occur in their blood at such times, they are not infective to Glossina.

Development in Glossina palpalis.—The principal accounts are those by Sir D. Bruce and his colleagues (1911), and by Miss Robertson (1912), whose results will be followed. According to the latter investigator T. gambiense never enters the body cells of the fly (G. palpalis), nor does it penetrate the gut wall into the body cavity. Practically no crithidial stage occurs in the fly’s main gut, but a trypanosome facies is retained therein.

After the trypanosomes are ingested by the fly during a meal of infected blood, sooner or later multiplication occurs. This development usually begins in the middle or posterior part of the mid gut, and trypanosomes of varying sizes are produced. After the tenth or twelfth day, many long, slender trypanosomes (fig. 30, a) are found, which gradually move forwards into the proventriculus. Such long, slender forms represent the limit of development in the lumen of the main gut. The proventricular type, developed about the eighth to the eighteenth or twentieth day, is not infective; it may occur in the crop, but is not to be found permanently there. Between the tenth and the fifteenth days multinucleate forms of trypanosomes are found, and may be styled multiple forms (fig. 30, b). Some of these latter may be degenerative.

Invasion of the Salivary Glands of the Fly.—Long, slender trypanosomes from the proventriculus pass forward into the hypopharynx. They then pass back along the salivary ducts, about sixteen to thirty days after the fly’s feed. The trypanosomes reach the salivary glands as long, slender forms. In the glands they become shorter and broader, attach themselves to the surrounding structures, and assume the crithidial facies (fig. 30, c, d). As crithidial forms they remain attached to the wall and multiply in the glands. These crithidial stages differentiate into the short, broad trypanosome forms, capable of swimming freely (fig. 30, e).

Miss Robertson considers the development in the main gut to be indifferent multiplication, and that salivary fluid seems necessary to stimulate trypanosomes to the apparently essential reversion to the crithidial type. The second development in the salivary gland is the essential feature. The short, stumpy forms of trypanosomes (fig. 30, e) finally produced in the salivary glands are alone infective. No conjugation of trypanosomes occurs in the fly. Only about 5 per cent. of captive tsetse flies fed on trypanosome-infected blood become infective, but they probably remain infective for the rest of their lives.

J. G. Thomson and Sinton (1912) have obtained in cultures the various trypanosome forms of T. gambiense seen in the fly’s main gut.

Duke (1912) found T. gambiense in a species of antelope, the situtunga (Tragelaphus spekei), on Damba Island in Victoria Nyanza. Wild G. palpalis could be infected therefrom. The antelope may then act as a sleeping sickness reservoir in that district, but men are apparently the chief reservoir.

Trypanosoma nigeriense, Macfie, 1913.

Macfie has recently (August, 1913) described a human trypanosome from the Eket district of Southern Nigeria. It is common in young people. The disease produced does not seem to be of a virulent type in Nigeria, and does not occur in epidemic form. In the early stages the glands of the neck are enlarged. In the later stages—cases of which are rarer—lethargy appears. The parasite is a polymorphic trypanosome, morphologically almost indistinguishable from T. gambiense, though it may be slightly shorter. Macfie recorded the occurrence in his preparations of a few trypanosomes appearing to have a flagellum free during their whole length. Some of the parasites, as seen in a sub-inoculated guinea-pig, are very small (8 µ long). Other trypanosomes have their nuclei displaced somewhat anteriorly. This parasite may only be a variety of T. gambiense. The parasite is perhaps spread by Glossina tachinoides.

Trypanosoma rhodesiense, Stephens and Fantham, 1910.

The parasite was found in the blood of a young Englishman who had contracted sleeping sickness in the Luangwa Valley, North-eastern Rhodesia, in the autumn of 1909. The patient had never been in an area infested with Glossina palpalis.

(1) Morphology.—The morphology of the parasite in man and sub-inoculated rats was studied by Stephens and Fantham in 1910. They pointed out a morphological peculiarity in the presence of certain trypanosomes with posterior nuclei in sub-inoculated animals, that is, parasites in which the nucleus (trophonucleus) was situated towards the posterior or aflagellar end, close up to or even beyond the blepharoplast or kinetic nucleus (fig. 31, 4, 5). When the nucleus was beside the blepharoplast, the former was seen to be kidney-shaped (fig. 31, 4). The posterior nuclear forms were of the stout and stumpy variety, and about 6 per cent. of the stumpy forms were found to have their nuclei displaced from the centre. The anterior or flagellar end of these trypanosomes often contained chromatoid granules. T. rhodesiense varies in length from 12 µ to 39 µ; short stumpy forms vary from 13 µ to 21 µ, intermediate forms from 21 µ to 24 µ, and long, slender forms from 25 µ onwards. The average length is 24·1 µ.

Certain regular periods occur in the course of the trypanosomiasis when few or no flagellate trypanosomes are found in the peripheral blood of the patient or of the sub-inoculated animal. These periods can be explained in terms of morphology, for the trypanosomes are capable of assuming a non-flagellate form in the internal organs of the host, particularly in the lungs and in the spleen. Such forms are known as “latent” or “resting” forms. The term “latent body” was first used by Moore and Breinl in 1907 in connection with T. gambiense. Fantham (1911) has described the process of formation of latent from motile forms and the reconversion of the latent bodies into active flagellates. Fresh preparations of splenic blood or lung blood containing trypanosomes were made. A trypanosome gradually withdrew or cast off its flagellum, concentrated its cytoplasm, and became more or less elongate oval. Nucleus and blepharoplast approached one another and came to lie more or less side by side. Then an opaque line often made its appearance around the nuclear area and differentiated as a slight envelope or covering, the cytoplasm external to this merely degenerating. The small, oval, refractile body (fig. , d—f) thus formed was a non-flagellate latent body, 2 µ to 4 µ in diameter, like Leishmania or the non-flagellate, multiplicative forms of T. cruzi (fig. ), and remains temporarily inactive in the internal organs of the host. After this period of inactivity, the non-flagellate body, recuperated by its rest, begins to elongate again. The nuclei separate. From a small vacuole-like portion the flagellum differentiates and forces out the ectoplasm, which assumes the form of the undulating membrane with its flagellar border. Subsequent growth results in the production of the typical trypanosome form, which re-enters the circulating blood and multiplies by longitudinal binary fission. Division of the parasite prior to the formation of a latent body may occur and division of the latent forms themselves is known, though less common. Consequently latent bodies, like the flagellate forms themselves, show diversity in size. The blepharoplast of the latent bodies is sometimes less well marked than in Leishmania (see fig. , d-f). Laveran’s views on these bodies have already been given on p. .

(2) Animal Reactions.—The posterior nuclear trypanosomes were found in all sub-inoculated animals, such as rats, guinea-pigs, dogs, mice, Macacus, rabbits and horses, but were not seen in the human patient, as few trypanosomes occurred in his peripheral blood. R. Ross and D. Thomson found a periodic, cyclical variation in the number of the parasites in the patient’s blood from day to day, the cyclical period being about a week (fig. ). Fantham and J. G. Thomson (1911) found a similar periodic, cyclical variation in the trypanosomes in the blood of sub-inoculated rats, guinea-pigs and rabbits. On counting the parasites in the blood of similar animals inoculated with T. gambiense, they established, by enumerative methods, that T. rhodesiense was more virulent than T. gambiense, while Yorke also showed this marked virulence of T. rhodesiense in practically all laboratory animals. In other words the duration of infection in the case of T. rhodesiense was shorter. It was also found that T. rhodesiense was resistant to atoxyl. The patient, from whom the original strain was obtained, died about nine months after the probable date of infection. Some patients infected with T. rhodesiense have died in an even shorter period, such as four or five months.

In sheep and goats T. rhodesiense causes an acute disease, marked by high fever, œdema of the face, and keratitis, as shown by Bevan and others, death resulting after a relatively short period. T. gambiense gives rise, in these animals, to no symptoms except fever, which may be overlooked. T. rhodesiense produces keratitis in dogs.

Stannus and Yorke (1911) observed T. rhodesiense in animals inoculated from a case of sleeping sickness in Nyasaland. Sir D. Bruce and his colleagues have shown (1912) that T. rhodesiense is the parasite usually found in man and in animals sub-inoculated from cases of sleeping sickness in Nyasaland. It has since been found in German East Africa and Portuguese East Africa, while Ellacombe has described a case from North-western Rhodesia.

(3) Serum Reactions.—Interesting experiments on this subject were performed during 1911 and 1912 by various French investigators.

(a) Action of Immune Serum (Mesnil and Ringenbach): (1) A goat was infected with T. rhodesiense. Twenty-two days later its serum mixed with T. rhodesiense was injected into a mouse. Result: Protection. (2) The serum mixed with T. gambiense was injected into a mouse. Result: Infection.

(b) Action of Baboon Serum.—Contrary to T. gambiense, T. rhodesiense is very susceptible to human and baboon sera. Mesnil and Ringenbach showed that a dose of 1 c.c. of baboon (Papio anubis) serum cured mice infected with T. rhodesiense. In the same dose it acted very feebly on T. gambiense.

(c) Action of Human Serum.—1 c.c. of human serum cured T. rhodesiense mice in three out of four cases; on T. gambiense mice there was no appreciable effect.

Laveran and Nattan-Larrier have shown the same, namely, that human sera act on T. rhodesiense, but are quite without action on T. gambiense.

(d) Trypanolytic Reactions.—Mesnil and Ringenbach have also shown that the sera of animals (man, monkey and guinea-pig) infected with T. gambiense are trypanolytic for the homologous trypanosome, that is, T. gambiense, but have no action on the heterologous trypanosome, that is, T. rhodesiense.

(4) Cross Immunity Experiments.—(a) Mesnil and Ringenbach immunized a monkey (Macacus rhesus) against T. gambiense. It was inoculated with T. rhodesiense on June 7, 1911; on June 27 trypanosomes appeared, the infection being slight; on July 4 it died. A control died in ten and a half days.

(b) Laveran immunized a goat and mice against T. gambiense. When they had acquired a solid immunity, they were inoculated with T. rhodesiense. They became infected like the controls.

(c) Laveran and Nattan-Larrier immunized a ram against T. brucei, it subsequently became infected with T. rhodesiense.

(d) Laveran immunized a ram and a sheep against different strains of T. brucei. Inoculated with T. rhodesiense they both acquired acute infections and died. Conclusion: T. rhodesiense is not T. brucei.

When the converse set of experiments is tried, namely, immunizing an animal against T. rhodesiense, and then inoculating with T. gambiense, the difficulty immediately arises that it is impossible to immunize an animal against T. rhodesiense, owing to its virulence. But a partial and transitory immunity to T. rhodesiense can be obtained by treating the infected animal with drugs, such as arsenophenylglycin. The results, so far as they go, seem to show that an animal immunized against T. rhodesiense is immune not only to T. rhodesiense, but also to T. gambiense, a fact which, according to Mesnil and Léger, does not invalidate the specificity of T. rhodesiense, but tends to show that the two trypanosomes are closely related.

(5) Mode of Transmission and Reservoir.—Kinghorn has shown that T. rhodesiense is transmitted by Glossina morsitans in which it undergoes development. Kinghorn and Yorke found that about 16 per cent. of the wild game examined in Northern Rhodesia was naturally infected with T. rhodesiense. The wild game examined included waterbuck, hartebeest, mpala, bushbuck and warthogs. One native dog near the Nyasaland border was found infected, but not domestic stock. Taute doubts whether T. rhodesiense really occurs in wild game. Approximately 3·5 per cent. of the tsetse flies fed on infected animals may become permanently infected with T. rhodesiense, and capable of infecting clean animals. Furthermore, a tsetse fly when once infective probably remains infective for the rest of its life.

Kinghorn and Yorke, however, have shown that climatic conditions, namely, those of temperature, also affect the infectivity of the tsetse fly, as the ratio of flies capable of transmitting T. rhodesiense to those incapable of transmitting the virus is 1 : 534 in hot valley districts (e.g., Nawalia, Luangwa Valley, temperature 75° to 85° F.), while on elevated plateaux (e.g., Ngoa, on the Congo-Zambesi watershed, temperature 60° to 70° F.) the ratio falls to 1 : 1312.

Mechanical transmission by the tsetse fly does not occur, if a period of twenty-four hours has elapsed since the infecting meal.

Developmental Cycle in the Fly.—The period which elapses between the infecting feed of the flies and the date on which they become infective varies from eleven to twenty-five days in the Luangwa Valley, according to Kinghorn and Yorke. Attempts carried out at laboratory temperature on the Congo-Zambesi plateau, during the cold season, to transmit T. rhodesiense by means of G. morsitans were always unsuccessful. The developmental cycle of the trypanosome in the fly is influenced by the temperature to which the flies are subjected (as stated above). The first portion of the developmental cycle proceeds at the lower temperatures (60° to 70° F.), but higher temperatures are necessary for the completion of the development of the trypanosome. Kinghorn and Yorke found that the trypanosomes may persist in the fly, at an incomplete stage of their development, for at least sixty days when the climatic conditions were unfavourable.

The first portion of the developmental cycle of the trypanosome takes place in the gut of the fly. Invasion of the salivary glands of the tsetse is secondary to that of the intestine, but is necessary for the infectivity of the fly. A relatively high mean temperature, 75° to 85° F., is essential for the passage of the trypanosomes into the salivary glands and the completion of their development therein.

Kinghorn and Yorke state that the predominant type of trypanosome in the intestine of infected G. morsitans was a large broad form, quite different from that which is most common in the salivary glands. The trypanosome in the glands resembles the short form seen in the blood of the vertebrate host. The authors quoted state that both the intestinal and salivary gland forms of infective G. morsitans are virulent when inoculated into healthy animals.

Bruce and colleagues have quite recently (June, 1914) published an account of their investigations of T. rhodesiense in G. morsitans in Nyasaland. (Incidentally it may be remarked that Bruce considers T. rhodesiense to be identical with a polymorphic strain of T. brucei—see pp. , ). The development of T. rhodesiense takes place in the alimentary canal and salivary glands, not in the proboscis, of the tsetse fly. In feeding experiments with laboratory bred flies, as well as with a few wild flies, fed on infected dogs or monkeys, only 8 per cent. of the flies were found to be infected on dissection. Of such infected flies, however, only some allow of the complete development of the trypanosomes within them, in other words only about 1 per cent of the flies become infective. The length of time which elapses before a fly becomes infective varies from fourteen to thirty-one days, averaging twenty-three days, when kept at 84° F. (29° C.). The dominant intestinal type of flagellate in the fly is that seen in the proventriculus, which contains many long, slender trypanosomes. These proventricular forms find their way to the salivary glands, wherein crithidial and encysted forms are seen. They change into “blood forms,” which are short, stumpy trypanosomes and are infective. “The infective type of trypanosome in the salivary glands—corresponding to the final stage of the cycle of development—is similar to the short and stumpy form found in the blood of the vertebrate host.” The cycle is thus very similar to that of T. gambiense in G. palpalis (fig. ).

Culture.—J. G. Thomson (1912), and subsequently Thomson and Sinton, succeeded in cultivating T. rhodesiense in a modified Novy-MacNeal medium. The development obtained resembled that of the trypanosome in the intestine of Glossina.

General Note on Trypanosomes with Posterior Nuclei.

Posteriorly placed nuclei have been found to occur not only in T. rhodesiense by Stephens and Fantham (1910), but also in T. pecaudi by Wenyon (1912), in T. brucei by Blacklock (1912), and in T. equiperdum by Yorke and Blacklock (1912).

Recently Stephens and Blacklock (1913) have shown that two trypanosomes, different morphologically, have been confused under the name T. brucei. One of these is polymorphic (i.e., it exhibits long and slender as well as short and stumpy forms) and came from Uganda, while the other is monomorphic and is the original Zululand strain described by Bruce from cattle suffering from “nagana.” Bruce (1914) considers that morphological change has occurred in T. brucei in its passage through laboratory animals, and thus explains the diversity of views. The posterior nuclear forms described by Blacklock occurred in the Uganda strain of T. brucei. (See p. .) Similarly, a posterior nuclear form, T. equi, has been separated from T. equiperdum. (See p. .)

Again, Bruce and his colleagues on the Royal Society Commission investigating sleeping sickness in Nyasaland, have stated (April, 1913) that “evidence is accumulating that T. rhodesiense and T. brucei (Plimmer and Bradford) are identical.” The exact identity of trypanosomes showing posterior nuclei is, then, far from settled, although Laveran by cross immunity tests has declared that T. brucei is distinct from T. rhodesiense. No one has yet seen posterior nuclei in T. gambiense.

Trypanosoma cruzi, Chagas, 1909.

Syn.: Schizotrypanum cruzi, Chagas, 1909.

The trypanosome was discovered by Chagas in the intestine of the bug, Triatoma (Conorhinus) megista, in Brazil, and then in the blood of a small monkey bitten by the bug. A little later it was found in the blood of a child, aged two years, suffering from irregular fever, extreme anæmia and enlarged glands in the State of Minas Geraes, Brazil. Chagas found that he was able to infect many of the usual laboratory animals with the trypanosome, by allowing the bug to bite them. He was also able to culture the parasite on blood agar.

Chagas found the Reduviid bug, Triatoma megista, in the houses of the poorer inhabitants of the Brazilian mining State, and that it attacked the people, more especially the children, at night, biting the face. On this account the insect is called “barbeiro” by the inhabitants. The bite is somewhat painful. The disease has since been found in other parts of Brazil, e.g., Matta de São João in Bahia province, Goyaz, Matto Grosso and São Paulo provinces, as well as in Minas Geraes.

Morphology.—The trypanosome has a large blepharoplast or kinetic nucleus. It is stated to occur both free and in the red blood corpuscles in the peripheral blood. It is about 20 µ long, on an average.

Two forms of the parasite (fig. 33, 6, 7) are described in the human blood. In one free form there is a large egg-shaped blepharoplast and the posterior (aflagellar) end of the parasite is drawn out. The blepharoplast (kinetic nucleus) may have a chromatin appendage. The nucleus is oval or band-like, containing a karyosome. The flagellum, starting close to the blepharoplast or its appendage, has a free portion of variable length. The other free form in the blood has a more or less round, terminal blepharoplast, smaller than in the first form, without a chromatin appendage as a rule. The body of this second form is decidedly broader than that of the first mentioned.

The dimorphism has been interpreted sexually, the first mentioned forms being termed males, the second ones females. The correctness of this interpretation is very doubtful.

No sign of longitudinal division was ever seen in the peripheral blood or in the internal organs. The “endocorpuscular” forms may be completely or partially enclosed in the red cell or only attached thereto (fig. 33, 1-5). At the beginning of infection the endocorpuscular forms are the more numerous. Some authorities, however, doubt these stages.

Life-history in the Vertebrate Host.—Chagas found fluctuations in the number of the parasites in the peripheral blood. He believes the increase of the parasites to be periodic.

The investigations of Chagas and of Hartmann have revealed two types of multiplication which take place in the internal organs of the vertebrate host.

(a) The first type—which possibly belongs to another organism, Pneumocystis carinii, see p. —occurs in the capillaries of the lungs. The flagellate parasite entering the lung capillaries loses its flagellum and undulating membrane. Its body becomes curved, and the two ends fuse, and so an oval mass is formed (fig. 33, 8-11). In some cases the blepharoplast disappears, in other cases it blends or fuses with the nucleus. The nucleus of the rounded parasite then divides into eight by successive divisions (fig. 33, 12-15). Next the body, which is surrounded by its own periplast, also divides, giving rise to eight tiny daughter individuals or merozoites (fig. 33, 15). The merozoites lie inside the periplast, which acts as a sort of “cyst wall.” The merozoites are said to exhibit dimorphism, and Chagas has interpreted the dimorphism in terms of sex. The daughter forms, produced by the parent trypanosomes which kept their blepharoplasts, themselves have blepharoplasts as well as nuclei, and have been termed “males” or “microgametes.” The merozoites, arising from parent trypanosomes which lost their blepharoplasts, have themselves only nuclei, and have been called “females” or “macrogametes.” In the case of the so-called “female” forms the single nucleus divides into two unequal parts, of which the smaller becomes the blepharoplast, and a flagellum is formed later. The so-called “males” possess early a rudiment of a flagellum. Both kinds of merozoites escape from the parent periplast wall, and enter red blood corpuscles. They grow into flagellates within the corpuscles, and then become free as adult trypanosomes in the blood-stream.

(b) The second mode of multiplication is one of asexual reproduction (schizogony or agamogony). It was first described by Hartmann from hypertrophied endothelial cells of the lungs. It has since been found in the cardiac muscle, in the neuroglia of the central nervous system, and in striped muscle (fig. 34). In laboratory animals it has also been found in the testicle and suprarenal capsules. In these tissues the parasite is intracellular, appearing as a small rounded body with nucleus and blepharoplast, without flagellum or undulating membrane. In other words the parasite is Leishmania-like in the body tissues, and recalls the organism of kala-azar.

Chagas considers this second mode of multiplication to be strictly asexual. By this means the number of parasites in the vertebrate host is increased, and symptoms are produced. On the other hand the first mode of multiplication, seen in the lung capillaries, is considered by Chagas to be a process of gametogony, in which sexual forms are differentiated. He finds that (1) the adult trypanosomes exhibit a dimorphism in human blood rarely seen in artificially infected guinea-pigs. In these guinea-pigs (infected from guinea-pigs) the so-called gametogony in the lungs is seldom seen. (2) The intermediate host, Triatoma (Conorhinus), becomes infective if fed directly on infected human blood, but very rarely so if fed on guinea-pigs. Chagas is led to believe that the occurrence of sexual forms constantly in the blood of man implies a greater resistance to infection on the part of man than on the part of guinea-pigs or other animals, assuming the general hypothesis that the formation of gametes represents a reaction of the Protozoön to unfavourable conditions. In human infection the number of parasites is always less than in laboratory animals, and their presence in the blood is transitory, lasting from fifteen to thirty days in acute cases. In many cases examination of the tissues at death has shown the presence of parasites in patients who did not exhibit them in the general circulation.

Life History in the Invertebrate Host.—About six hours after the ingestion of infected blood by the bug (Triatoma megista), the kinetic nucleus of the trypanosome moves towards the nucleus, and the flagellum is usually lost (fig. 35, 1-5). The parasite becomes rounded and Leishmania-like (fig. 35, 3-5), and multiplies rapidly by division. After a time, multiplication having ceased, the rounded forms become pear-shaped and develop a flagellum at the more pointed end. Crithidial forms (fig. 35, 7) are thus produced and pass into the intestine, where they multiply and may be seen in about twenty-five hours after the ingestion of blood. The crithidial forms may also be found in the rectum and fæces. The last stage in the invertebrate is a small, trypanosome-like type, long and thin with a band-like nucleus and conspicuous kinetic nucleus. These parasites are found in the hind gut and in the body cavity. They find their way into the salivary glands, and are the forms (fig. 36) which are transmissible to a new vertebrate host. The development in the bug takes about eight days altogether, after which time the bugs are infective.

There are thus three principal phases in the development of T. cruzi in Triatoma megista: (1) A multiplicative phase (Leishmania-like) in the stomach of the bug, (2) a crithidial phase, which is also multiplicative, in the hind-gut, and (3) a trypanosome phase, which is “propagative,” and apparently passes through the wall of the alimentary canal into the body cavity and so into the salivary glands.

Brumpt found that T. cruzi could live in Cimex lectularius, C. boueti, and Ornithodorus moubata. The Cimex fæces may be infective. Blacklock found multiplication of the parasite in C. lectularius.

Culture.—The trypanosome can be cultivated on Novy-MacNeal’s blood agar, and the cultural forms resemble those described in the bug.

Possible Reservoir.—Chagas thinks that probably the armadillo or “tatu” (Dasypus novemcinctus) may be the reservoir of T. cruzi. He also thinks that Triatoma geniculata is a transmitter; it lives in the burrows of the armadillo. Other carriers may be Triatoma infestans and T. sordida.

Clinical Features.—The trypanosomiasis of Brazil, produced by T. cruzi and spread by Triatoma spp. has received various names, such as oppilação, canguary, parasitic thyroiditis, and coreotrypanosis. It is also known as the human trypanosomiasis of Brazil, South American trypanosomiasis, and Chagas’ disease.

Chagas reports two principal forms—acute and chronic. The acute infection is rare, and is characterized by increase in the volume of the thyroid gland, pyrexia, a sensation of crackling in the skin, enlarged lymphatic glands in the neck, axilla, etc., while the liver and spleen are increased in volume. Sclerosis of the thyroid gland is found at autopsy and fatty degeneration of the liver. During an attack of fever, trypanosomes are found in the blood. The acute form was only observed in children.

In the chronic form Chagas reports several varieties: (a) A pseudo-myxœdematous form, occurring in most cases, especially up to the age of 15. There is hypertrophy of the thyroid gland or at least signs of hypothyroidism, general hypertrophy of glands, disturbance of heart rhythm, and nervous symptoms. (b) The myxœdematous form is characterized by similar symptoms, especially by considerable swelling of the thyroid body, and myxœdema of the subcutaneous cellular tissue; sometimes there is a true pachydermic cachexia. (c) In the nervous form there are motor disturbances, aphasia, disturbances of intelligence or signs of infantilism, athetosis of the extremities and idiocy. There are also paralytic symptoms of bulbar origin, disturbances of mastication, phonation and deglutition, and in some cases convulsive attacks. (d) The cardiac form, characterized by disturbance of the heart rhythm. In all these forms the parasite is found at autopsy in the nervous substance, brain, bulb and heart.

Vianna (1911) has studied the histopathology of the disease. Some of the chief points are: in the heart muscle destruction of the sarcoplasm, followed by interstitial myocarditis; in the central nervous system invasion of the neuroglia cells and inflammatory reaction; in the suprarenal capsule invasion of medulla or cortex; inflammatory reaction can also be seen in the kidneys, the hypophysis and thyroid gland.

Recently Chagas states that “schizotrypanosomiasis” has been found in a child 15 to 20 days old, and that Trypanosoma cruzi has also been found in a fœtus—the mother being infected with the trypanosome. The trypanosomiasis can, then, be transmitted hereditarily.

Trypanosoma lewisi, Kent, 1881.

The trypanosome has a nucleus somewhat displaced anteriorly, about one-third of the way from the anterior (flagellar) end of the body, a relatively straight edge to the undulating membrane, and a rod-shaped blepharoplast (fig. 37, A). It averages about 25 µ long and 1·5 µ broad.

Much attention has been devoted in recent years to the elucidation of the life history of the rat parasite, Trypanosoma lewisi. It is usually non-pathogenic to its host. It has been shown that the trypanosome can be transmitted from rat to rat by the rat-flea, Ceratophyllus fasciatus, and by Ctenocephalus canis (the so-called dog-flea). (See also p. ). The flagellate may also persist, but doubtfully develop, in the rat-louse, Hæmatopinus spinulosus. These researches may now be summarized.

Life Cycle in the Vertebrate Host.—After infection of a rat, the trypanosomes usually appear in the animal’s blood in five to seven days. This incubation period applies either to a natural or an artificial infection. The trypanosomes first observed in the rat’s blood are diverse in form (fig. 37), being small, medium and large in size. This diversity is explained by the rapid multiplication taking place. A trypanosome may divide by equal longitudinal fission (fig. 37, C, D), but more commonly multiple fission occurs (fig. 37, G, H), and is unequal. Rosette forms are produced, in which the parent form can be recognized by its long flagellum (fig. 37, H) and attached to it are daughter individuals, smaller in size, from which flagella are growing. Minchin and J. D. Thomson (1912) find that the daughter forms may be set free sometimes with a crithidia-like facies (fig. 37, I), the blepharoplast being anterior but near to the nucleus. The daughter forms, when set free, may themselves divide by binary or multiple fission, in the latter case forming rosettes (fig. 37, K). Rosette forms were described by Moore, Breinl and Hindle in 1908.

Lingard, some years ago, described as a distinct species, T. longocaudense, certain forms with markedly elongate posterior ends (fig. 37, E). According to Minchin, “these forms appear to arise by binary fission” (fig. 37, D). These long drawn-out forms “are of constant occurrence and very numerous at a certain stage of the multiplication period.” It is about the eighth or tenth day after infection that the multiplication of T. lewisi is at its maximum in the rat’s blood. About the twelfth or thirteenth day the trypanosomes seen in the blood appear uniform. According to Minchin (1912) the rat “gets rid of its infection entirely sooner or later, without having suffered, apparently, any marked inconvenience from it, and is then immune against a fresh infection with this species of trypanosome.” There is, then, a cycle of development in the vertebrate host. Minchin notes that the records of the pathogenicity of T. lewisi in rats, causing their death, need further investigation.

T. lewisi inoculated into dormice (Myoxus nitela) and jerboas may become pathogenic thereto.

Carini found cysts in the lungs of rats infected with T. lewisi. He thought the cysts were schizogonic stages of the trypanosome, comparable with those found in the lungs of animals sub-inoculated with T. cruzi. Delanoë (1912) has found, however, that such cysts, containing eight vermicules, occurred in rats uninfected with T. lewisi. Delanoë concludes that the pneumocysts are independent of T. lewisi, and represent a new parasite, Pneumocystis carinii. The pneumocysts may be allied to the Coccidia, and must be considered when investigating the life-cycle of a trypanosome in a vertebrate host. Some of the stages of T. cruzi may possibly be of this nature.

Life-cycle in the Invertebrate Host.—This occurs in fleas, and has been investigated in considerable detail by Minchin and Thomson in Ceratophyllus fasciatus, and by Nöller in Ctenocephalus canis and Ctenopsylla musculi.

When infected rat’s blood is taken up by the flea, the parasites pass with the ingested blood direct to the mid-gut of the Siphonapteran. In the flea’s stomach they multiply in a somewhat remarkable manner, namely, by penetration of the cells of the lining epithelium, and division inside the epithelial cells. Inside these lining cells the trypanosomes first grow to a large size and then form large spherical bodies, within which nuclear multiplication occurs (fig. 38, A-F). Any one of these large spherical bodies contains at first a number of nuclei, blepharoplasts and developing flagella, the original flagellum still remaining attached for a time. The cytoplasm then divides into daughter trypanosomes which are contained within an envelope, formed by the periplast of the parent parasite. Inside the periplast envelope are a number of daughter trypanosomes “wriggling very actively; the envelope becomes more and more tense, and finally bursts with explosive suddenness, setting free the flagellates, usually about eight in number, within the host-cell” (fig. 38, F). The daughter forms escaping from the host cell into the stomach lumen of the flea are fully formed, long trypanosomes.

The trypanosomes (fig. 38, G) pass into the flea’s rectum. The next phase is a crithidial one. The parasites become pear-shaped, in which the blepharoplast (kinetic nucleus) has travelled anteriorly past the nucleus towards the flagellum (fig. 39). The crithidial forms attach themselves to the wall of the rectum, and multiply by binary fission (fig. 39, D). A stock of parasites is thus formed which, according to Minchin and Thomson, “persist for a long time in the flea—probably under favourable conditions, for the whole life of the insect” (fig. 39, A–I).

From the crithidial forms of the rectum, according to Minchin, small infective trypanosomes arise by modification morphologically (fig. 39, J–M). The flagellum grows longer and draws out more the anterior part of the body, the blepharoplast migrates posteriorly, behind the nucleus, and carries with it the flagellar origin. These trypanosomes are small, but broad and stumpy (fig. 39, N), and can infect a rat. Minchin and Thomson formerly considered that the small, stumpy, infective trypanosomes pass forwards from the rectum into the stomach, and “appear to be regurgitated into the rat’s blood when the flea feeds.” However, the small infective trypanosomes were previously described by Swellengrebel and Strickland. They may be found in the flea’s fæces. Nöller (1912) has found that the development of T. lewisi proceeds quite well in the dog flea (Ctenocephalus canis) in Germany. Wenyon confirms this, and states that the human flea, Pulex irritans, and the Indian rat-flea, Xenopsylla cheopis, are also able to serve as true hosts for T. lewisi.

Nöller stated that rats were not infected with T. lewisi by infective fleas biting them, but by the rats licking up the fæces passed by the fleas while feeding. This is not in agreement with Minchin and Thomson’s earlier views of regurgitation, which, apparently, they have now abandoned. Wenyon (1912) confirms Nöller’s experiments. He took a dog flea, containing infective trypanosomes in its fæces, and allowed it to feed on a clean rat. The fæces of the flea, passed while feeding, were carefully “collected on a cover glass and taken up in culture fluid with a fine glass pipette.” The contents of the pipette were discharged into the mouth of a second clean rat. Injury to the rat’s mouth was carefully avoided. The first rat, on which the infective flea was fed, did not become infected, while the second rat, in whose mouth infective flea fæces were placed, became infected in six days.

When infective forms of T. lewisi have been developed within the gut of a rat flea, they may enter and infect the vertebrate host by (a) being crushed and eaten by the rodent; (b) the rat may lick its fur on which an infected flea has just passed infective excrement; or (c) the rat may lick, and infect with flea excrement, the wound produced by the bite of the flea.

The time taken for the full development of T. lewisi in the flea is about six days. The intracellular phase is at its height about the end of the first day; the crithidial phase, in the flea’s rectum, begins during the second day; the stumpy, infective trypanosomes are developed in the rectum about the end of the fifth day.

Wenyon writes that, “the fleas, when once infected with T. lewisi, remain infected for long periods, for though many small infective trypanosomes are washed out of the gut at each feed, those that remain behind multiply to re-establish the infection of the hind gut. Further, the infection is still maintained even if the flea is nourished on a human being, so that fresh human blood does not appear to be destructive to the infective forms in the flea.”

The best method of controlling fleas during experiments is that due to Nöller. He adopted the method of showmen who exhibit performing fleas, and secure them on very fine silver wire.

Of fleas fed on an infected rat only about 20 per cent. become infective. About 80 per cent. are immune. If fleas are examined twenty-four hours after feeding, trypanosomes will be found in all, so that many of the parasites are destined to degenerate.

It may be of interest to note that Gonder (1911) has shown that a strain of T. lewisi resistant to arsenophenylglycin loses its resistance after passage through the rat-louse, Hæmatopinus spinulosus. These experiments suggest that physiological “acquired characters” may be lost by passage through an invertebrate host.

Trypanosoma brucei, Plimmer and Bradford, 1899.

Trypanosoma brucei was discovered by Sir D. Bruce in 1894 in cattle in Zululand and was named T. brucei by Plimmer and Bradford in 1899 in honour of its discoverer. This trypanosome is of considerable economic importance, as it is responsible for the fatal tsetse fly disease, or “nagana,” in cattle, horses and dogs. The disease is widely distributed in Africa and is transmitted from host to host by the tsetse, Glossina morsitans, and other species of Glossina. The virus is maintained in nature in certain big game, such as wildebeest, bushbuck and koodoo, which thus act as living reservoirs of disease from which the tsetse may become infected. These reservoir hosts are not injured, apparently, by the presence of the parasites.

T. brucei is rapidly fatal to the small laboratory animals, such as rats and mice. Horses, asses and dogs practically always succumb to its attacks, while a very small number of cattle recover from “nagana.” The disease is characterized by fever, destruction of red blood corpuscles, severe emaciation and by an infiltration of coagulated lymph in the subcutaneous tissue of the neck, abdomen and extremities giving a swollen appearance thereto. The natural reservoirs in which T. brucei has been long acclimatized are unaffected by the trypanosomes, while the newer hosts, such as imported cattle in Africa, are rapidly destroyed by their action.

The general morphology and life history in the vertebrate host is that of a typical trypanosome (fig. 40). Its length is from 12 µ to 35 µ, its breadth from 1·5 µ to 4 µ. Multiplication by longitudinal division proceeds in the peripheral blood (fig. ), while latent, leishmaniform bodies are produced in the internal organs.

Bruce and colleagues have quite recently (June, 1914) described the development of a Zululand strain of T. brucei in G. morsitans. The tsetse flies were bred out in Nyasaland. In vertebrate blood the brucei strain was polymorphic. The development was like that found for T. gambiense in G. palpalis (fig. ), and by Bruce and colleagues for T. rhodesiense in G. morsitans in Nyasaland. Long trypanosomes were found in the proventriculus of the tsetse. Crithidial, rounded or encysted, and immature “blood forms” occurred in the salivary glands; and finally infective, stumpy, “blood forms” were differentiated in the salivary glands. The period of development of T. brucei in G. morsitans takes about three weeks, and then the fly becomes infective. Bruce believes that T. rhodesiense of Nyasaland and T. brucei of Zululand are the same, their cycles of development in G. morsitans being “marvellously alike.” (But see Laveran, p. 80.)

T. brucei has been cultivated with difficulty by Novy and MacNeal, using blood agar. The best treatment for nagana is arsenic in some form.

It is probable that more than one trypanosome has been confused under the name T. brucei, more especially as the occurrence of many species of trypanosomes in various animals in Africa was not suspected until comparatively recent times. It has been shown by Stephens and Blacklock (1913) that the original Zululand strain of T. brucei was monomorphic, while the organism sent from Uganda, and at the time believed by Bruce to be the same as the Zululand trypanosome, has been found to be polymorphic, with morphological resemblances to T. rhodesiense. Stephens and Blacklock have suggested the name T. ugandæ for the polymorphic trypanosome, which, however, has marked resemblances with Trypanosoma pecaudi, and they are, perhaps, identical. T. pecaudi was the name given by Laveran in 1907 to the causal agent of “baleri” in equines and sheep in the French Sudan. T. pecaudi, which is dimorphic, is widely distributed in Africa. An extremely small number of both T. pecaudi and T. ugandæ have been shown to possess posterior nuclei. T. pecaudi is transmitted by various species of Glossina, and is said to develop in the gut and proboscis of the fly.

On the other hand, Bruce and colleagues (1914), examining a strain sent from Zululand in 1913, state that T. brucei is polymorphic. Bruce (1914) suggests that passage through laboratory hosts has influenced and altered the morphology of the parasite.

Trypanosoma evansi, Steel, 1885.

Syn.: Spirochæta evansi, Steel, 1885; Hæmatomonas evansi, Crookshank, 1886; Trichomonas evansi, Crookshank, 1886.

Trypanosoma evansi, first found by Evans in 1880, in India, is the causal agent of the disease known as “surra.” The malady affects more particularly horses, mules, camels and cattle in India and neighbouring countries, such as Burma and Indo-China. It occurs also in Java, the Philippines, Mauritius and North Africa. Elephants may be affected. A serious outbreak among cattle in Mauritius occurred in 1902, the disease being imported into the island. The symptoms are fever, emaciation, œdema, great muscular weakness and paralysis culminating in death.

T. evansi varies from 18 µ to 34 µ in length and 1·5 µ to 2 µ in breadth. It has a pointed posterior extremity, and, anteriorly, there is a free portion to the flagellum (fig. 41). It is possibly monomorphic, but a few broad forms occur. The trypanosome multiplies by longitudinal fission in the blood. Rounded leishmaniform stages occur in the spleen of the vertebrate host, which stages Walker (1912) considers to be phases of schizogony.

The parasite is transmitted in nature by various species of Tabanus and Stomoxys, though at present little is known of the life-history within these invertebrate hosts.

Dogs are said to contract the disease by feeding on animals dead of surra.

A variety of T. evansi is the cause of “mbori” in dromedaries in Africa (Sahara and Sudan). Another possible variety, or closely allied form, is T. soudanense, the causal agent of “el debab” in camels and horses in North Africa, especially Algeria and Egypt.

An extraordinary example of the possible infection of a human being with an animal trypanosome is recorded in the case of Professor Lanfranchi, of the Veterinary School, Parma. The Professor became infected with trypanosomes, although only nagana and surra were maintained in his laboratory, and he himself had never visited the tropics. He suffered from irregular attacks of fever and was œdematous, but his mind remained clear. The identification of the trypanosome from Lanfranchi’s blood has been a matter of great difficulty. Apparently Mesnil and Blanchard (1914) consider the strain found in the patient is almost indistinguishable in its reactions from T. gambiense, though the parasite is monomorphic. Lanfranchi considers that he was infected with T. evansi.

Trypanosoma equinum, Voges, 1901.

Syn.: Trypanosoma elmassiani, Lignières.

Trypanosoma equinum was found by Elmassian to be the cause of the fatal disease, “mal de caderas,” of horses and dogs, in South America (Paraguay, Argentine, Bolivia). The name refers to the fact that in the disease, as in other trypanosomiases, the hind quarters become paralysed. Cattle are refractory to inoculation.

T. equinum is about 22 µ to 24 µ long and about 1·5 µ broad (fig. 42). Although this trypanosome is very active, yet it is characterized by the blepharoplast (kinetic nucleus) being very minute or even absent, as the granule sometimes seen may be the basal granule of the flagellum.

The mode of transmission of T. equinum is not known with absolute certainty. Migone has shown that the parasite causes a fatal disease in the large South American rodent, the capybara (Hydrochœrus capybara). This animal appears to be a reservoir of the parasite. Dogs may become infected by eating diseased capybaras, and it is suggested that the infection is spread from the dogs to horses by the agency of fleas. Some authorities consider that T. equinum may be spread by various Tabanidæ and by Stomoxys. Neiva (1913) doubts all these modes of transmission in Brazil, and suggests Chrysops or Triatoma as vectors.

Trypanosoma equiperdum, Doflein, 1901.

Syn.: Trypanosoma rougeti, Laveran and Mesnil.

The malady of horses known as “dourine” or “mal du coït” is due to a trypanosome, T. equiperdum, discovered by Rouget in 1894. “Dourine”—also known as “stallion disease” or “covering disease”—is found among horses and asses in Europe, India, North Africa and North America. The trypanosome is transmitted by coitus, and so far as is known not by insect agency.

The progress of the disease may be considered under three periods. The period of œdema, when signs of œdema of the genitalia are seen. The œdema is generally painless and non-inflammatory. This period lasts about a month. It is succeeded by the period of eruption, which sets in about two months after infection. Circular œdematous areas (“plaques”), often about the size of a two-shilling piece, appear under the skin of the sides and hind quarters, and also, at times, under the skin of the neck, thighs and shoulders. The eruption is variable, but usually lasts about a week and leaves the animal in an enfeebled condition. Gland enlargement and swelling of the joints and synovia also may occur. The third period of the disease is described as that of anæmia and paralysis. The animal becomes very anæmic, emaciation is marked, superficial non-healing abscesses often form, and conjunctivitis and ulcerative keratitis can occur. Paralysis ensues, and in from two to eighteen months the animal dies. In the acute form of the disease the animal may die after the first period from acute paralysis.

It is difficult to find the trypanosomes in naturally infected animals, and they are best obtained from the plaques of the eruption. Apparently the parasite occurs more in the lymph than in the blood.

Ruminants are said to be refractory to this trypanosome.

T. equiperdum is about 25 µ to 28 µ in length on an average, but varies from 16 µ to 35 µ. Its cytoplasm is relatively clear, and does not show chromatic granules (fig. 43). It is stated to be monomorphic.

It has been shown recently by Blacklock and Yorke (1913) that there is another trypanosome giving rise to dourine in horses. This trypanosome is dimorphic (resembling T. pecaudi and T. ugandæ), and is named T. equi. Previously T. equiperdum and T. equi had been confused.

Uhlenhuth, Hübner and Worthe have demonstrated the presence of endotoxins in T. equiperdum. These endotoxins may be set free by trypanolysis.

Trypanosoma theileri, Bruce, 1902.

This parasite, 60 µ to 70 µ long, and 4 µ to 5 µ broad, is distinguished for its large size, though it is not so large as T. ingens from Uganda oxen, whose length may be 72 µ to 122 µ, and breadth 7 µ to 10 µ. The posterior end of T. theileri is drawn out. Small forms of the flagellate are known, 25 µ to 53 µ in length. Probably other forms of the parasite have the nucleus posterior, and these flagellates were formerly separated as T. transvaaliense (Laveran, 1902). Myoneme fibrils may be seen on its body. The pathogenicity of this organism is doubtful, it was formerly thought to be the causal agent of “gall-sickness” in cattle in South Africa. T. theileri also occurs in Togoland, German East Africa, and Transcaucasia. Allied or identical parasites occur in cattle in India.

Trypanosoma theileri, specific to cattle, is perhaps transmitted by the fly Hippobosca rufipes in South Africa.

Trypanosoma hippicum, Darling, 1910.

Trypanosoma hippicum causes the disease of mules known as “murrina.” It was found in mules imported to Panama from the United States. It can live in other equines. The parasite varies from 18 µ to 28 µ in length, and is from 1·5 µ to 3 µ broad. Its undulating membrane is little folded. The trypanosome has a noticeable blepharoplast. It can penetrate mucous membranes, and it is thought that the trypanosome may be transmitted during coitus. It may also be spread mechanically by species of Musca, Sarcophaga and Compsomyia, sucking the wounds of infected animals and carrying over the trypanosomes to wounds on healthy ones.

Endotrypanum schaudinni, Mesnil and Brimont, 1908.

This organism was discovered in the blood of a sloth (Cholœpus didactylus), in South America (French Guiana). It possesses special interest, in that the best known form of the organism is endoglobular, inhabiting the erythrocytes of the sloth. A free trypanosome in the same animal was considered to be different from the endoglobular form, which was somewhat like a peg-top, and possessed a short flagellum. Darling (November, 1914) has seen the organism in Panama. He describes free crithidial forms in shed blood, but not in the blood-stream of the sloth.

Trypanosoma boylei, Lafont, 1912.

This is a parasite of the Reduviid bug, Conorhinus rubrofasciatus. The insect attacks man in Mauritius, Réunion and other places. Lafont infected rats and mice by intraperitoneal injection with the gut-contents of infected bugs. Trypanosomes appeared in the mice. Other flagellate types were assumed by the parasites in the bug.

Monomorphic Trypanosomes.

A number of trypanosomes, characterized by relative uniformity in size and structure, may be considered under this heading. They occur in cattle, sheep, goats and horses in Africa, especially West Africa. Morphologically, they are characterized by the posterior (aflagellar) part of the body being swollen, while the anterior part narrows. The nucleus is central and situated at the commencement of the narrowing of the body. The blepharoplast is almost terminal, the undulating membrane is narrow and not markedly folded, so that the flagellar border lies close to or along the body. The flagellum may or may not possess a free portion.

Some recent workers have considered that T. brucei (Zululand strain) and T. evansi are also monomorphic, but they do not exhibit the general characteristics outlined above. T. brucei and T. evansi have already been considered separately.

The monomorphic trypanosomes, as defined above, include:—

Trypanosoma vivax, Ziemann, 1905.

This trypanosome occurs in cattle, sheep and goats, and was first found in the Cameroons. It is fatal to cattle. Equines are also affected. Antelopes are the possible reservoirs of the trypanosome. It is probably transmitted by Glossina palpalis and other tsetse flies. Its movement is very active. It possesses a free flagellum (fig. 45) and it averages 23 µ to 24 µ in length. T. cazalboui (Laveran, 1906)—the causal agent of “souma” in bovines and equines in the French Sudan—is probably synonymous with T. vivax.

Trypanosoma capræ (Kleine, 1910) is allied, but is somewhat broader and more massive. It was found in goats in Tanganyika.

Trypanosoma congolense, Broden, 1904.

Probable synonyms.—Trypanosoma dimorphon, Laveran and Mesnil, 1904; Trypanosoma nanum, Laveran, 1905; Trypanosoma pecorum, Bruce, 1910; Trypanosoma confusum, Montgomery, 1909.

This trypanosome causes disease among horses (e.g., Gambia horse sickness), cattle, sheep, goats, pigs, and dogs. It is widely distributed in Central Africa (e.g., Gambia, Congo, Uganda, Nyasaland), the strain probably being maintained naturally in big game. It is transmitted by various Glossinæ, and perhaps by Tabanus and Stomoxys. It is said to develop in the gut and proboscis of Glossina palpalis and G. morsitans. The trypanosome averages 13 µ to 14 µ in length and has no free flagellum (fig. 46). It is about 2 µ broad. Formerly T. nanum and T. pecorum were said to differ in their pathogenicity, the former being said not to infect the smaller laboratory animals. Yorke and Blacklock (1913), however, consider that the virulence varies and that these trypanosomes are probably the same.

The T. dimorphon originally obtained by Dutton and Todd (1903) in Gambian horse sickness has been shown to be a mixture of T. vivax and T. congolense.

Trypanosoma simiae (T. ignotum) is like T. congolense. It averages 17·5 µ long. It is virulent to monkeys and pigs.

Trypanosoma uniforme, Bruce, 1910.

This trypanosome was found in oxen in Uganda. It can be inoculated to oxen, goats and sheep, but is refractory to dogs, rats and guinea-pigs. It has been found in antelopes. It resembles T. vivax, but is smaller (fig. 47), averaging 16 µ in length. A free flagellum is present. It is transmitted by Glossinæ.

Many other trypanosomes occur in mammals, while birds, reptiles, amphibia (fig. 48) and fish also harbour them. The discussion of these forms does not come within the scope of the present work. They are dealt with in Laveran and Mesnil’s “Trypanosomes et Trypanosomiases,” 2nd edit., 1912.

General Note on Development of Trypanosomes in Glossina.

Before concluding the account of trypanosomes, it may be of interest to remark that several African trypanosomes develop in various species of Glossina, and are found in different parts of the alimentary tract and in the proboscis. Thus (a) T. vivax, T. uniforme and T. capræ develop in the fly’s proboscis (labial cavity and hypopharynx) only; (b) T. congolense, T. simiæ and T. pecaudi develop first in the gut of the fly and then pass forward to its proboscis; and (c) T. gambiense and T. rhodesiense develop first in the gut and later invade the salivary glands of the tsetse. The proboscis or the salivary glands in such cases are termed by Duke the anterior station of the trypanosome, wherein it completes its development.

Adaptation of Trypanosomes.

These flagellates may exhibit power of adaptation to changes of environment, such as those due to the administration of drugs, change of host, etc. A few examples of such mutations may be briefly considered:—

(1) Blepharoplastless Trypanosomes.—T. brucei may become resistant to pyronin and oxazine. Accompanying this drug resistance is a change in morphology, namely, the loss of the blepharoplast (Werbitzki). A race or strain of blepharoplastless trypanosomes may be thus produced which retains its characteristic feature after as many as 130 passages (Laveran). Oxazine is the more powerful drug, and it acts directly on the blepharoplast. (Compare the natural blepharoplastless character of T. equinum.)

(2) Reference has been made on p. to the experiments of Gonder, who showed that a strain of T. lewisi rendered resistant to arsenophenylglycin lost its resistance after passage through the rat louse. This is in marked contrast with the retention of drug resistance during passage by inoculation from rat to rat.

(3) T. lewisi from the blood of a rat when transferred to a snake seems largely to disappear, as very few flagellates are seen. When blood from the snake is inoculated into a clean rat, then trypanosomes reappear in the rat, but they are not all like those originally inoculated. It seems certain that, in such a case, changes in form and virulence of the trypanosome have occurred. Similar experiments were made with T. brucei from rats to adders and other animals and back to rats. Changes in the form and virulence of T. brucei occurred.

These interesting experiments were performed by Wendelstadt and Fellmer.

Genus. Herpetomonas, Saville Kent, 1881.

Herpetomonas is a generic name for certain flagellates possessing a vermiform or snake-like body, a nucleus placed approximately centrally, and a blepharoplast (kinetic nucleus) near the flagellar end. There is no undulating membrane (fig. , a). The organisms included in this genus certainly possess one flagellum, while according to Prowazek (1904) Herpetomonas muscæ-domesticæ, the type species, possesses two flagella united by a membrane. Patton, Porter and others affirm, however, that the biflagellate character of H. muscæ-domesticæ (from the gut of the house-fly) is merely due to precocious division. The matter is further complicated by the generic name Leptomonas, given by Kent in 1881, to an uniflagellate organism found by Bütschli in the intestine of the Nematode worm, Trilobus gracilis. This parasite, Leptomonas bütschlii, has not yet been completely studied. Until these controversial points relating to the identity or separation of Herpetomonas and Leptomonas have been satisfactorily settled, we may retain the better known name Herpetomonas for such uniflagellate, vermiform organisms. However, the name Leptomonas, having been used by Kent two pages earlier in his book (“Manual of the Infusoria”) than Herpetomonas, would have priority if the two generic names were ultimately shown to be synonymous.

A full discussion of these interesting and important flagellates hardly comes within the purview of the present work; brief mention can only be given here to certain species.

The Herpetomonads occur principally in the digestive tracts of insects, such as Diptera and Hemiptera. They are also known in the guts of fleas and lice, but are not confined to blood-sucking insects. One example, H. ctenocephali (Fantham, 1912) occurs in the digestive tracts of dog fleas, Ctenocephalus canis, in England, France, Germany, Italy, India, Tunis, etc. It is a natural flagellate of the flea, and might easily be confused with stages of blood parasites in the gut of the dog flea. Dog fleas are stated by Basile to transmit canine kala-azar, which is believed to be the same as human infantile kala-azar. Confusion is further likely to arise since herpetomonads pass through pre-flagellate, flagellate and post-flagellate or encysted stages; pre- and post-flagellate stages being oval or rounded and Leishmania-like. The post-flagellate stages are shed in the fæces, and are the cross-infective stages by means of which new hosts are infected by the mouth. The possible presence of such natural flagellates must always be considered when experimenting with fleas, lice, mosquitoes, etc., as possible vectors of pathogenic flagellates like Leishmania and Trypanosoma. H. pediculi (Fantham, 1912) occurs in human body lice. See further remarks on pp. , .

Laveran and Franchini (1913–14) have recently succeeded in inoculating Herpetomonas ctenocephali, from the gut of the dog flea, intraperitoneally into white mice, and producing an experimental leishmaniasis in the mice. A dog was also infected. They have also succeeded in infecting mice with H. pattoni—a natural flagellate of the rat flea—by mixing infected rat fleas with the food of the mice, and by causing them to ingest infected fæces of rat fleas. Further, they have shown that infection with the herpetomonas occurs naturally by this method, that is, by the rodents eating the fleas and not by the insects inoculating the flagellates into the vertebrates when sucking blood. These experiments shed an interesting light on the probable origin of Leishmania and its cultural herpetomonad stage, which were very probably once parasitic flagellates in the gut of an insect.

Fantham and Porter (1914–15) have shown that young mice may be inoculated or fed with Herpetomonas jaculum, from the gut of the Hemipteran, Nepa cinerea (the so-called “water-scorpion”), with fatal results. The pathogenic effects are like those of kala-azar. They also showed that the post-flagellate stages of the herpetomonads seemed most capable of developing in the vertebrate.

A herpetomonad, H. davidi, has been found in the latex of species of the plant-genus Euphorbia in Mauritius, India, Portugal, etc. It is apparently transmitted to the plants by Hemiptera. The plants sometimes suffer from “flagellosis.”

Franchini (1913) has described a new parasite, Hæmocystozoon brasiliense, from the blood of a man who had lived in Brazil for many years. It possesses flagellate and rounded stages, and is closely allied to the herpetomonads.

Genus. Crithidia, Léger, 1902, emend. Patton, 1908.

Crithidia is the generic name of vermiform flagellates with a central nucleus, a blepharoplast or kinetic nucleus in the neighbourhood of the principal nucleus, and a rudimentary undulating membrane bordered by a flagellum arising from a basal granule, which is the centrosome of the kinetic nucleus (fig. b). The anterior or flagellar end of the body is attenuated and fades off as the undulating membrane.

Crithidia fasciculata, the type species, was found by Léger in the alimentary canal of Anopheles maculipennis. Crithidia occur in bugs, flies, fleas, and ticks. Some of them are found in the body-fluid of the invertebrate host as well as in the gut. Others may be restricted to the body cavity or intestine respectively. C. melophagia from the sheep-ked, Melophagus ovinus, and C. hyalommæ from the hæmocœlic fluid of the tick, Hyalomma ægyptium, pass into the ovaries and eggs of their hosts, and the young keds or ticks are born infected.

C. fasciculata has been shown by Laveran and Franchini to be inoculable into white mice, producing a sort of experimental leishmaniasis therein. In one case cutaneous lesions were produced like those of Oriental sore.

Crithidia are natural flagellates of Arthropoda, with their own pre-flagellate, flagellate and post-flagellate stages, and must not be confused with transitory crithidial stages of trypanosomes.

Genus. Leishmania, Ross, 1903.

With an oval body containing nucleus and blepharoplast (kinetic nucleus) but no flagellum. An intracellular parasite in the vertebrate host.

Included in the genus Leishmania are three species, namely:—

(1) Leishmania donovani, Laveran and Mesnil, 1903, the parasite of Indian kala-azar, a generalized systemic disease, usually fatal, occurring in subjects of all ages.

(2) Leishmania tropica, Wright, 1903, the parasite of Delhi boil, Oriental sore, Aleppo button—a localized, cutaneous disease, usually benign.

(3) Leishmania infantum, Nicolle, 1908, the parasite of infantile kala-azar, occurring in children (and a few adults) around the shores of the Mediterranean. The disease is perhaps a form of Indian kala-azar, and the parasite is probably identical with L. donovani.

These diseases may be termed collectively leishmaniases. The morphology of the various species is practically identical.

Leishmania donovani, Laveran and Mesnil, 1903.

Syn.: Piroplasma donovani, Laveran and Mesnil.

The parasite of Indian kala-azar was demonstrated in 1900 by Leishman from a post-mortem examination of a case of “Dum-Dum fever,” but details were not published till May, 1903. In July, 1903, Donovan found similar bodies from cases in Madras. Rogers succeeded in cultivating the parasite in July, 1904. The original centre of the disease was probably Assam; it occurs also in Madras, Ceylon, Burma, Indo-China, China and Syria. A variety of this leishmaniasis is found in the Sudan. The patient becomes emaciated, with a greatly enlarged spleen. There is anæmia and leucopenia.

The parasite, commonly known as the Leishman-Donovan body, is intracellular (fig. , 2, 3). It is found in the endothelial cells of the capillaries of the liver, spleen, bone-marrow, lymphatic glands and intestinal mucosa, and in the macrophages of the spleen and bone-marrow. Some host cells may contain many parasites. It is rather rare in the circulating blood, but may be found in the blood from the femoral, portal and hepatic veins. It does not occur in the red blood corpuscles as was formerly thought. The parasites liberated from the endothelial cells are taken up by the mononuclear and polymorphonuclear leucocytes. The Leishman-Donovan body is the resting stage of a flagellate. As found in man it is a small, oval organism, about 2·5 µ to 3·5 µ in length by 2 µ in breadth, and containing two chromatinic bodies, corresponding to the nucleus and kinetic nucleus (blepharoplast) of a flagellate. The latter element is the smaller and more deeply staining, and is usually placed at the periphery, transversely to the longer axis of the oval organism. There is sometimes a very short, slightly curved filament to be seen, which may be a rhizoplast. Multiplication takes place by binary or multiple fission. The presence of the parasite used to be demonstrated by splenic or hepatic puncture; nowadays it can be demonstrated in peripheral blood, e.g., of the finger, or by culture of infected blood.

L. donovani can be cultivated in citrated splenic blood, under aerobic conditions, at 22° to 25° C. This was first accomplished by Rogers (1904). It is not so easily culturable as L. infantum on the Novy-MacNeal-Nicolle medium. L. donovani is inoculable with some difficulty into experimental animals—in India, white rats, white mice, dogs and monkeys (Macacus spp.), have been inoculated. The Sudan variety, somewhat less virulent, is inoculable to monkeys. Row also produced a local lesion in Macacus sinicus by subcutaneous inoculation of L. donovani. Parasites taken from such a local lesion were found to be capable of producing a generalised infection in Macacus sinicus and white mice.

In cultures the various species of Leishmania all grow into herpetomonad, uniflagellate organisms (fig. 50, 10), about 12 µ to 20 µ in body length. On this account Rogers and Patton place the Leishman-Donovan body within the genus Herpetomonas. The method of culture may be used in diagnosing leishmaniases.

Kala-azar is very probably an insect-borne disease. Patton suspects the bed-bug to be the transmitter and finds (fig. 50, 4-6) that the Leishman-Donovan body can develop into the flagellate stage in the digestive tract of the bed-bug. Feeding experiments are unsatisfactory, since there are very few cases in which the parasites occur in sufficient numbers in the peripheral blood to make the infection of the insect possible, or at any rate easy. In examining the alimentary tracts of insects for possible flagellate stages of Leishmania, it must be remembered that in many insects natural flagellate parasites, belonging to the genus Herpetomonas, may occur therein; such natural insect flagellates may be harmless, and have no connection with the life-cycle of L. donovani. Natural herpetomonads are known to occur in the alimentary tracts of flies, mosquitoes, sand-flies, fleas and lice, but not in bed-bugs. Further, if such flagellates are able to be inoculated into and live within vertebrate hosts, producing symptoms like those of leishmaniasis, the origin of kala-azar is indicated (see pp. , ).

Leishmania tropica, Wright, 1903.

Syn.: Helcosoma tropicum, Wright, 1903; L. wrighti, Nicolle, 1908; Ovoplasma orientale, Marzinowsky and Bogrow.

It is believed by some that the parasite was first described by Cunningham in 1885, and studied by Firth in 1891, being called by him Sporozoon furunculosum. If these earlier studies were of the parasite, then its correct name is L. furunculosa, Firth, 1891.

The benign disease produced by this parasite has received many names, among the best known being Oriental sore, Tropical sore, Delhi boil and Aleppo button. These names, however, are not happy ones, as cutaneous leishmaniasis (e.g., on the ear) is now known to occur in the New World, for example in Mexico, Venezuela, Brazil and neighbouring States. However, it may be necessary to subdivide cutaneous leishmaniases later.

In the Old World the disease occurs in India, Persia, Arabia and Transcaucasia. It is also known in Algeria, Northern Nigeria, Egypt, Sudan, Crete, Calabria, Sicily and Greece.

The boils often occur on the face, and before ulceration the parasites may be found in the cells at the margin and floor of the “button.” In searching for parasites the scab should be removed and scrapings made from the floor and edges. Where lesions occur atrophy of the epidermis takes place, and infiltration of mononuclear cells (e.g., plasma cells, lymphoid and endothelial cells) follows. The parasites are intracellular, being found inside mononuclear cells. In non-ulcerating sores, Cardamitis found some free parasites. Non-ulcerating forms are said to occur in the Sudan. In the Old World the sores are often limited to exposed surfaces of the body. Infection of mucous membranes (such as the lip, palate, buccal and nasal membranes) may occur, especially in South America, and are often known there as “Espundia.” Christopherson (1914) has recorded a case in Khartoum.

Leishmania tropica is equally well cultivated on Novy-MacNeal-Nicolle medium or on citrated blood. The usual temperature for cultivation is 22° to 28° C., though Marzinowski claims to have cultivated the parasite at 37° C. L. tropica can be inoculated into monkeys and dogs, with the production of local lesions. Material from a human sore or flagellates from a culture may be thus successfully inoculated. Also infected material may be rubbed directly into a scarified surface. The incubation period is long, extending over several months. The duration of the disease may be from twelve to eighteen months. Recovery from one attack of tropical sore confers immunity, and the Jews in Bagdad inoculate their children with the disease on a part of the body which will be covered, and so secure immunity in adult life.

The mode of transmission of L. tropica is unknown. Wenyon (1911) has found that the parasite develops into the flagellate stage in the digestive tract of Stegomyia fasciata in Bagdad. Patton (1912) has found similar development in the bed-bug in Cambay. The house-fly, Phlebotomus and Simulium have been suspected as transmitters in different parts of the world.

An interesting announcement has been made recently (May, 1913), that Neligan has found that L. tropica occurs in dogs in Teheran, Persia, producing ulcers on the dogs’ faces (cf. natural occurrence of L. infantum in dogs—see p. ). Yakimoff and Schokhor (1914), have found the disease in dogs in Tashkent.

Gonder (1913) has performed some interesting experiments showing the relation of infantile kala-azar to Oriental sore. Gonder infected mice with L. infantum and with L. tropica. He used culture material and injected intraperitoneally or intravenously. In each a general infection resulted, with enlargement of the liver and spleen. Later, however, mice injected with Oriental sore (North African variety) developed peripheral lesions on the feet, tail and head, and the lesions contained Leishmania. No such peripheral lesions developed in the case of the mice infected with the kala-azar virus. Gonder suggested that Oriental sore, like kala-azar, is really a general infection overlooked in its earlier stages, and that it is in the later stages that peripheral lesions on the skin are developed. Row (1914) also obtained a general infection in a mouse by the injection of cultures of L. tropica from Oriental sore of Cambay.

Leishmania infantum, Nicolle, 1908.

Infantile splenic anæmia has been long known in Italy. It also occurs in Algeria, Tunis, Tripoli, Syria, Greece, Turkey, Crete, Sicily, Malta, Spain and Portugal. This leishmaniasis is, then, distributed along the Mediterranean littoral; also in Russia. Cathoire (1904) in Tunis and Pianese (1905) in Italy were among the first to see the parasite. Nicolle then found the parasite in patients in Tunis, and further found spontaneous infection in dogs. The patients are usually children between the ages of 2 and 5 years. There are a few cases known in which the infantile type of leishmaniasis occurred in youths and adults of the ages of 17 to 19, while one patient in Calabria was 38 years old. The symptoms are like those of Indian kala-azar. Several Italian investigators and others consider that L. infantum is the same as L. donovani, and that the latter name should be used for the parasite of Mediterranean leishmaniasis. This view, as to the identity of L. donovani and L. infantum, seems coming into general favour.

There are, however, differences between the Indian and infantile kala-azars, in addition to the ages of the patients affected, thus: (a) As regards cultures, it is found that L. infantum is readily grown on the Novy-MacNeal-Nicolle (“N.N.N.”) medium (saline blood-agar), and that sub-cultures are easily obtained; in citrated blood L. infantum grows with difficulty. The reverse is the case with regard to culture media for L. donovani, which grows with difficulty on the N.N.N. medium, but relatively easily in citrated splenic blood. (b) Considering inoculability into experimental animals, it is found that L. donovani is inoculated generally with some difficulty into white rats, white mice and monkeys, and with greater difficulty into dogs, while L. infantum can be inoculated into several experimental animals, especially into dogs and monkeys, with ease. (c) At present L. donovani is not known to occur spontaneously in animals, but L. infantum is found naturally in dogs in the Mediterranean region, and the disease in dogs is often referred to as canine kala-azar. Kittens have occasionally been found infected. However, these differences must not be emphasized too much.

The material for cultivation is obtained from punctures of spleen, liver or bone-marrow of cases infected with L. infantum. It is not always easy, however, to infect from cultures, as the cultural flagellates inoculated into the body are often phagocytosed.

Similarly, the material for animal inoculation is obtained from emulsions of infected spleen, liver or bone-marrow. Dogs and monkeys are easily inoculated with such material; Nicolle inoculates into the liver or the peritoneal cavity. Mice, white rats, guinea-pigs and rabbits only show slight infections after such inoculations.

Dogs infected experimentally with infantile leishmaniasis may show either acute or chronic symptoms. The acute course occurs more often in young dogs, and is usually fatal in three to five months. The chronic course is found more commonly in older dogs, and may last seventeen to eighteen months. In acute forms there is irregular fever, progressive wasting, diarrhœa occasionally, motor disturbances involving the hind quarters, and the animal dies in a comatose condition. In the chronic form the animal may appear well, except for loss of weight. The parasites may be found in the internal organs of these experimental dogs, but are not numerous in the peripheral blood except at times of high fever. Experimental monkeys live about three months.

It may be interesting to record the number of dogs found to be infected naturally with leishmaniasis in various countries. In Tunis, Nicolle and Yakimoff found about 2 per cent. infected out of about 500 dogs examined. Sergent in Algiers found 9 infected out of 125 dogs examined. In Italy and Sicily, Basile found about 40 per cent. of the dogs to be infected out of 93 examined at Rome and Bordonaro. Cardamitis found 15 infected out of 184 examined in Athens. In Malta, Critien found 3 infected out of 30 dogs examined. Alvares found 1 infected dog out of 19 examined in Lisbon. Pringault has recently (December, 1913) found an infected dog in Marseilles. Yakimoff and Schokhor found 24 per cent. infected out of 647 dogs examined in Turkestan.

The distribution of the parasites in the body of the human patient is much the same as in the case of Indian kala-azar. Critien records the finding of parasites in the mucous flakes of the stools of a three-year-old Maltese child. Intestinal lesions rarely occur in infantile leishmaniasis.

Ætiology.—Infantile leishmaniasis is stated to be transmitted by fleas, especially dog fleas, Ctenocephalus canis (= Pulex serraticeps), and by Pulex irritans. Children living in contact with infected dogs may be bitten by infected dog fleas, and so contract the disease. Basile (1910–11) and Sangiorgi (1910) state that they found L. infantum parasites in the digestive tract of the dog flea. After searching they found infected dog fleas on the beds, mattresses, and pillows used by children suffering from the disease. Franchini (1912) thinks that Anopheles maculipennis may be concerned in the transmission.

Basile tried a number of experiments to show that infantile leishmaniasis is transmitted by fleas, thus:—

(1) Fleas were taken from a healthy dog. They were placed in vessels containing infected spleen-pulp and allowed to feed thereon. The fleas were then killed and dissected, and portions of the gut-contents examined for parasites. The remainder of the gut was emulsified and injected into a young puppy, whose bone-marrow had been shown previously to be uninfected. Basile states that the puppy became infected. The parasites are said to increase in number in the flea’s gut.

(2) Two healthy pups, each a month old, and born in the laboratory, were placed in a disinfected, flea-proof cage. A few days after, an infected dog was placed in the cage, so that fleas from the infected dog could pass on to the puppies. A month later the two pups became infected, parasites being found in them after liver puncture. A number of control puppies from the same litter remained uninfected and in good health.

(3) Basile next used other laboratory-born puppies, a month old. Four of the litter were placed in a disinfected, flea-proof gauze cage in Rome. The cage was isolated from other dogs. Fleas obtained from an infected area in Sicily were placed in the cage. The puppies were examined by hepatic puncture, but were found to be negative for two months. Then two of the puppies showed infection, and six days later the remaining two puppies were found to be infected, and all four died. They showed irregular temperatures, and were getting thin. Control puppies remained healthy.

From these experiments Basile concludes that fleas transmit leishmaniasis. However, Basile did not exclude the possible occurrence of natural herpetomonads in the gut of the fleas. Herpetomonas ctenocephali is known to occur in the gut of Ctenocephalus canis. A natural Herpetomonas is also known in the gut of Pulex irritans, as well as a Crithidia (C. pulicis, Porter). These natural flagellates of the fleas pass through non-flagellate stages, like the Leishman-Donovan body. In consequence Wenyon and Patton, among others, have criticized Basile’s results. Further, other investigators, such as Wenyon and Da Silva (1913), have repeated Basile’s flea experiments and been unable to confirm them.

In feeding and inoculation experiments the incubation period of the parasite may be long, and so it is necessary to wait a long time to see whether the parasite will develop.

Immunity.—Nicolle has tried some experiments with L. infantum and L. tropica. He finds that in animals recovery from an attack of the former confers immunity against infection by the latter and vice-versâ.

Laveran records that a monkey having an immunity against L. infantum was also immune to L. donovani.

As mentioned on p. , Laveran and Franchini (1913), working in Paris, have succeeded in inoculating Herpetomonas ctenocephali, a natural flagellate in the gut of the flea, Ctenocephalus canis, into white mice. Leishmaniform stages of the flea flagellate were recovered from the peritoneal exudate, blood and organs of the mice some weeks after inoculation. The parasites may also be conveyed by way of the digestive tract of the vertebrate. Similar experiments have succeeded with H. pattoni. These experiments go to show, together with those of Fantham and Porter with H. jaculum (see p. ), that, in the words of the latter authors, “it may be expected that the various leishmaniases, occurring in different parts of the world, will prove to be insect-borne herpetomoniases.”

Genus. Histoplasma, Darling, 1906.

Under the name Histoplasma capsulatum, Darling described small round or oval parasites, enclosed in a refractile capsule, and each containing a single nucleus. The bodies were found in cases of splenomegaly in Panama. They occurred in the endothelial cells of the small blood-vessels of the liver, spleen, lungs, intestine and lymphatic glands, and also within the leucocytes. A few flagellates were stated to occur in the lungs. The parasite has usually been placed near Leishmania, but recently Rocha-Lima has stated that Histoplasma is a yeast.

Genus. Toxoplasma, Nicolle and Manceaux, 1908.

The genus was created for crescentic, oval or reniform parasites, 2·5 µ to 6 µ by 2 µ to 3 µ, possessing a single nucleus and multiplying by binary fission. They occur in mononuclear and polymorphonuclear cells in the blood, spleen, liver, peritoneum etc. (fig. 51). The parasites have been found in the gondi, dog, rabbit, mole, mouse, pigeon and other birds. Although various species names have been given to the parasites in these hosts, it seems probable, from cross infection experiments, that there is but one species with several physiological races. Splendore (1913) has described a flagellate stage.

in spleen. [1 is about the size of a red blood corpuscle, as drawn in the figures]. Magnification not stated. (After Castellani.)]

Castellani (1913–14) has described similar parasites from a case of splenomegaly, with fever of long standing, in a Sinhalese boy. The bodies were found in the spleen and more rarely in the blood (fig. 52). Castellani has named them Toxoplasma pyrogenes. Further researches are needed.