Genus Sarcocystis

In this genus the cysts are found in the striated and heart muscles, and are usually divided into compartments by internal septa. Synonyms of this name are Miescheria Blanchard, 1885 and Balbiania Blanchard, 1885.

Sarcocystis is common in many species of animals. It is found in the great majority of sheep, cattle, horses and swine, and is often seen in wild ducks. It is extremely rare, however, in carnivores such as the dog and cat, and the reports of its presence in these animals require verification. Dogs cannot be infected experimentally (Pfeiffer, 1891).

More than 50 species of Sarcocystis have been named, but it is not at all certain that they are all valid. They are differentiated on the basis of the host in which they occur, the structure of the cyst wall and the size of the trophozoites. However, Sarcocystis is not very host-specific. The rat and guinea pig can be infected with the form from the mouse, the mouse, guinea pig, chicken and duck with that from the sheep, and the rat and mouse with that from the pig. In addition, the same species does not look the same in all hosts. For example, in the guinea pig the trophozoites of the form from the mouse are only half their former size and the cysts do not have alveoli. Finally, the structure of the cyst wall may vary with its age even in the same host. The specific names below are therefore used more as a matter of convenience and custom than from any conviction that they are all necessarily valid.

Morphology

The cysts are known as Miescher's tubules and are easily visible to the naked eye. They are usually cylindroid or spindle-shaped, running lengthwise in the muscles, but they may also be ellipsoidal or rather irregular. They vary in size depending in part on the host. The ellipsoidal cysts in the sheep may reach 1 cm in diameter, but considerably smaller ones are the rule. Those in the duck are 1 or 2 mm in diameter and 1 cm or more long.

The cyst wall varies in appearance with the species. There are 3 types. In one, e.g., Sarcocystis muris of the mouse, it is smooth. In another, e.g., S. platydactyli of the gecko, it has an outer layer of radial spines, villi or fibrils called cytophaneres. In a third, e.g., S. tenella of the sheep, the wall is smooth in the young cysts, acquires a layer of cytophaneres as the cysts develop, and then loses it when they become old.

The cyst wall of S. tenella is composed of 2 thick layers (Ludvik, 1958). The inner one is homogeneous and contains nuclei. Extensions from it form septa between the compartments in the cyst. The outer layer contains no nuclei and appears spongy in electron micrographs. It forms the cytophaneres. The inner layer contains RNA and the outer a polysaccharide. The cyst wall is essentially negative to the periodic acid-Schiff test, altho the cytophaneres stain slightly according to Frenkel (1956a).

The cyst wall of S. miescheriana differs from that of S. tenella in being composed of only a single layer with a complicated surface structure (Ludvik, 1960). The cytoplasm of the wall is granulated, and fine septa project from its inner surface to divide the interior of the cyst into small compartments. The outer surface of the cyst wall is spongy, with a fine honeycomb structure. It sends numerous parallel, hollow, finger-like projections or villi into the surrounding muscle tissue. These villi may be as much as 8 to 10 u long, and are circular or ellipsoidal in cross section and about 0.7 to 0.8 u in diameter. They contain slender, long double fibrils 100 A thick.

There is a difference of opinion as to whether the cyst wall is formed by the parasite or the host. A few authors, such as Wang (1950), think that the host forms both layers, but this view is probably not correct. Some, such as Chatton and Avel (1923) and Barretto (1940), think that the parasite forms both layers. Others, such as Scott (1943), think that the parasite forms the inner layer and the host the outer; still others, such as Babudieri (1932), think that the whole cyst wall is formed by the parasite in S. muris and similar species, and that one layer is formed by the parasite and the other by the host in S. tenella and similar species. According to Ludvik (1960), the singlelayered cyst wall of S. miescheriana is quite certainly formed by the parasite, and the villi which project into the muscle tissue take up nutritive material from the host.

The trophozoites are banana-shaped when mature, with the anterior end slightly pointed and the posterior end rounded. They are 6 to 15 u long and 2 to 4 u wide, varying in size with the species. They move by gliding or body flexion, twisting, turning, or following a spiral path.

Throphozoite of Sarcocystis tenella

Ludvik (1958, 1960) described their structure in S. tenella and S. miescheriana on the basis of electron microscope, cytochemical and light microscope studies. At the anterior end within the pellicle is a polar ring 0.4 to 0.5 u in diameter, and within it is a hollow, truncate cone 0.3 to 0.4 u long known as a conoid. From the polar ring 22 to 26 fine fibrils run backwards in the pellicle the full length of the body. In some individuals short, club-shaped structures similar to the toxonemes of Toxoplasma can be seen in the cytoplasm beneath the pellicle.

The cell body is divisible into 3 zones. The anterior third of the body, the socalled fibrillar zone, is filled with a large number (about 300 to 350) of parallel, equidistant fibrils or perhaps channels about 50 mu in diameter, the sarconemes. They probably arise from the conoid, and they end abruptly. Just under the pellicle on the dorsal (convex) side about the middle of the fibrillar zone is a disc-shaped granule which stains with Bodian silver.

The middle third of the body contains a large number of spherical granules 0.4 to 0.5 u in diameter, the so-called central granules. They impregnate with osmium and stain intensely with Heidenhain's hematoxylin but not with Giemsa. In the same region are many minute granules, some of which contain volutin and others RNA. There are also 1 or 2 large vacuoles which stain with neutral red.

The posterior third of the body contains the nucleus. It is an ellipsoidal vesicle almost as wide as the body, and contains a relatively small number of chromatin granules and an endosome which stains with Bodian silver. The nucleus is surrounded by a large number of small vacuoles and granules, many of which contain glycogen, and these extend to the posterior end of the body. Among them lie 1 to 3 serpentine mitochondria 0.15 to 0.2 u in diameter and 2 u or more long.

In addition to the above structures, a network of fibrils forming a characteristic rectangular pattern can be seen on the surface following silver impregnation by the Klein or Chatton technics.

Life Cycle

Several differing accounts have been given of the life cycle of Sarcocystis. Pitfalls in its study have been discussed by Scott (1943). There is now general agreement that the life cycle is simple, without sexual stages, and that no intermediate host is involved.

Animals become infected by ingesting trophozoites, either in unbroken cysts in the muscles or free in the feces of other animals. Smith (1901, 1905) was the first to show that infection took place by the oral route, and was able to maintain the infection with S. muris in mice for 7 years by feeding infected mouse muscle.

The trophozoites presumably pass thru the intestinal wall, enter the blood stream and are carried to the striated muscles, where they enter the muscle cells. They are found in the striated and heart muscles. They are especially common in the wall of the esophagus, but are also found in the tongue, masseter muscle, diaphragm, throat, neck, body and limb muscles, and even in the eye muscles and Purkinje fibers of the heart among other places. In ducks they are most commonly found in the breast muscles.

There is a latent period of a month to 6 weeks or more during which almost nothing is known of what happens. The first stage in the muscle cell is a one-celled, irregularly rounded ("amoeboid") naked parasite. This divides by repeated binary fissions (Scott, 1943) into a number of rounded cells 4 to 8 u in diameter which are enclosed in a cyst wall. Betegh and Dorcich (1912), Erdmann (1914) and Arai (1925) thought that schizogony takes place at this stage, but Scott (1943) did not agree, and Frenkel (1956a) considered its existence doubtful.

The rounded cells have been called sporoblasts, pansporoblasts or prosporoblasts, but these names all carry the connotation that the trophozoites are spores, and the cells are better called cytomeres (Grasse, 1953) or trophoblasts. They continue to reproduce by binary fission, and become pressed together and polygonal. Later they change into ellipsoidal and then into banana-shaped trophozoites.

As multiplication proceeds, the cyst grows and is divided into chambers or compartments by septa arising from the inner layer of the cyst wall. The process continues, new trophoblasts are formed at the periphery of the cysts, produce new trophozoites, and new septa are laid down and new compartments formed.

The trophozoites themselves also reproduce by binary fission. This process was described by Ludvik (1958). The nucleus first begins to enlarge and the dispersed chromatin forms large granules and variously curved structures. The nucleus is indented in the middle of its anterior edge and becomes horseshoeshaped. The cell loses its banana shape and becomes broadly spindle-shaped, with a rounded posterior end. The central granules become dispersed thru the whole cell and diminish in size. A medial sac-like structure begins to be separated off from the posterior part of the horseshoeshaped nucleus, and the central granules disappear. The sac-like structure becomes detached from the nucleus and gradually divides into 2 halves which later, after the true nuclear division has been completed, disappear. The horseshoeshaped nucleus divides into 2 longitudinal segments. The conoid and cytoplasm in the anterior third of the cell also divide into 2 longitudinal halves with a clear streak between them. The newly formed nuclei become shorter and their chromatin gradually disperses. Cell division now begins, starting from the conoid at the anterior end and proceeding posteriorly. The nuclei round up, their nucleoplasm becomes thicker, and they move toward the posterior part of the newly forming cells. New central granules appear in the cytoplasm in front of the nuclei. The daughter cells remain attached at their posterior ends for a time and then separate entirely.

Finally, as the cyst itself becomes older, the trophozoites in the central compartments degenerate and disappear. After the cyst becomes mature, its wall breaks down and the trophozoites are released. They enter the blood stream, reach the digestive tract, and pass out in the feces. They have also been found in the nasal secretions of sheep (Scott, 1943).

Quite a different account has been given by Spindler and his associates, who believe Sarcocystis to be a fungus rather than a protozoon. Spindler and Zimmerman (1945) reported that they had isolated an Aspergillus-like fungus from sarcocysts from pig muscles, and that 25 out of 50 pigs injected with or fed material from the cultures had sarcocysts in their muscles 4 to 6 months later, while the control pigs were negative. They also said that pigs, rats and mice fed the cysts passed yeast-like bodies in their urine or feces which produced a similar fungus upon culture, and they found these bodies in the kidneys of infected mice and in clumps attached to the walls of the ileum and cecum of infected rats and mice.

Spindler, Zimmerman and Jaquette (1944) were unable to infect pigs directly with sarcocysts in pig muscles, but they observed that the pigs became infected if they ate their own feces. They fed pork containing sarcocysts to pigs, dogs, cats, rats, mice and chickens. These subsequently passed a stage in their feces and/or urine which was infective for swine. Their observation, incidentally, may perhaps explain the remark of Scott (1943) that feeding experiments in sheep indicate that the trophozoites of S. tenella must undergo some change before they can infect other sheep.

Spindler (1947) described a network of jointed, hypha-like structures in cysts from a sheep and a duck, and said that the trophozoites appeared to be exogenous growths on these structures. However, Grasse (1953) commented that his illustrations were not convincing, and that the structures he described appeared to be the result of marked alterations in the true ones. Frenkel (1956a), too, disagreed with Spindler. He found no fungal characteristics in morphological studies of organisms from man, the sheep, mouse, rabbit, squirrels and the duck. Unlike fungi, the trophozoites and cyst walls did not give a positive reaction with the periodic acid-Schiff stain. Sarcocystis from cottontail rabbits and house mice failed to grow on the media customarily used for fungi. Frenkel concluded that these organisms neither look nor behave like fungi.

Scott (1943), too, and others cited by him were unable to cultivate organisms from the cysts. Only Ciesla (1950) has reported positive results. He observed "sporozoites" in cultures from cysts from cattle, and said that these eventually turned into round corpuscles with a quick, convulsive type of movement which budded into branched chains of mycelia.

The weight of the evidence thus indicates that Sarcocystis is a protozoon and not a fungus.

Pathogenesis

Sarcocystis is not generally considered very pathogenic. However, Scott (1943a) believed that it is of greater economic importance than is usually supposed.

Light or moderate infections produce no noticeable signs, but in very heavy infections there may be lameness, weakness, emaciation, paralysis and even death.

The sarcocyst destroys that part of the muscle fiber which it occupies, and as it grows it may cause pressure atrophy of adjacent cells. Calcification may also occur. There is ordinarily little if any cellular reaction around the cysts. Focal myocarditis and myositis develop when the cysts break down. Destombes (1957) described a marked inflammatory reaction around the cysts followed by necrosis and calcification in swine, but saw no such reaction in cattle. Spindler, Zimmerman and Jaquette (1946) found that pigs with 40 or more cysts per gram of diaphragm were unthrifty and showed signs of muscular stiffness.

Gastrointestinal signs and lesions may occur after ingestion of the cysts. Scott (1943) reported extensive destruction of the epithelium together with a bloody serous exudate in the ileum of young rats fed sarcocysts from sheep, and the animals appeared ill and disinclined to move about. Spindler, Zimmerman and Jaquette (1946) observed vomiting, diarrhea, inappetence and temporary posterior paralysis in pigs fed infected muscles, urine or feces.

The cysts contain a powerful endotoxin known as sarcocystin, which is highly toxic for rabbits, mice, and sparrows, but probably less toxic for rats, sheep and some other animals. Sarcocystin acts on the central nervous system and also affects the heart, adrenal glands, liver and intestinal wall. It is filtrable, and is destroyed by heat. Small amounts cause a febrile reaction in the rabbit, while large amounts produce collapse, severe diarrhea and death. According to Sato (1926), the intravenous minimum lethal dose for the rabbit of the extract from S. fusiformis from the ox is 0.05 mg per kilogram body weight.

Immunity

Animals can be immunized against sarcocystin by repeated injections of untreated or formalin-treated toxin. The serum of immunized animals will protect other animals against the toxin.

The close relationship between Sarcocystis and Toxoplasma is attested by the fact that both react with cytoplasm-modifying antibody in the Sabin-Feldman dye test (described below under Toxoplasma). As a matter of fact, cross reactions between the two are not uncommon. Muhlpfordt (1951) and Awad and Lainson (1954) found that the sera of laboratory animals fed S. tenella from sheep reacted positively to the dye test with Toxoplasma trophozoites. The sera of sheep naturally infected with S. tenella also gave positive reactions. Awad (1954) went a step further, and developed a modified dye test for Toxoplasma, using S. tenella trophozoites. These trophozoites gave positive results with the sera of animals infected with either Toxoplasma or Sarcocystis.

Epidemiology

Seasonal infection during the late spring, summer and early fall has been reported in sheep, swine and horses in the temperate zone (Scott, 1943). Repeated infections of sheep in successive seasons were reported by Scott (1943). He had the impression that the older the animals, the more heavily they were parasitized.

Diagnosis

Because of the absence of recognizable signs, Sarcocystis infections are almost always diagnosed after death. The larger cysts are easily seen with the naked eye, and the small ones can be found by histologic examination.

Cultivation

Sarcocystis has not been cultivated, unless the claims of Spindler and Zimmerman (1945) and Ciesla (1950) are accepted.

Treatment

None known.

Prevention and Control

Since Sarcocystis infections are acquired thru fecal contamination of food or drink, infections can be prevented by measures designed to prevent such contamination. Sanitation and good management should be effective.