Fine Structure of the Schizonts and Merozoites of Eimeria acervulina in the Chicken

Fine structural changes observed during development of merozoites were similar in the 1st and later asexual generations of Eimeria acervulina in the chicken. Merozoites were formed at the surface of multinucleated schizonts. The anlage of the merozoite first appeared as a cone-shaped elevation of the surface, in which the inner membrane complex and the organelles of the apical complex developed. The nucleus, Golgi complex, and other organelles were incorporated into the developing merozoite while it was attached by the posterior end to the residual cytoplasm. The inner membrane of the pellicle terminated anteriorly at a polar ring and posteriorly at a thickening suggestive of a posterior polar ring. Twenty-two subpellicular microtubules extended along the merozoite from the anterior polar ring to the posterior end. The nucleus and its Golgi complex were situated in the posterior half of the merozoite and mitochondria, endoplasmic reticulum, refractile bodies, numerous micronemes, and 2 rhoptries in the anterior half. Mitochondria and endoplasmic reticulum were found posterior to the nucleus. The conoid had a spiral structure and 2 anterior preconoidal rings. A micropore just anterior to the nucleus was well developed in the lst-generation merozoites. Polysaccharide granules were found only in the 2ndand 3rd-generation merozoites. Entry of merozoites into villus epithelial cells caused deformity of microvilli. Secondand 3rd-generation schizonts lying close to the brush border deformed microvillar filaments in the superficial web zone. Developing schizonts were within parasitophorous vacuoles in host cells whose endoplasmic reticulum and numbers of mitochondria had increased. Mature schizonts were in cells whose organelles, including the nucleus, had degenerated. Tyzzer (1929) described the life cycle and the morphology of the various stages of Eimeria acervulina in the chicken. E. acervulina has since been frequently used to study the effect of intestinal coccidiosis on the chicken host (Preston-Mafham and Sykes, 1970; Turk and Stephens, 1967; Hein, 1968), but as yet these effects have not been examined at the fine structural level. Lee and Millard (1971a) described the fine structure and development of the macrogamete and oocysts of E. acervulina. The ultrastructure of the schizonts and merozoites and of the changes in host cells parasitized by schizonts are described in this paper. MATERIALS AND METHODS Two-week-old, male White Leghorn chicks, raised coccidia-free, were inoculated with sporulated oocysts of Eimeria acervulina as previously described (Fernando and McCraw, 1973). They were killed at 12, 24, 36, 48, and 56 hr, and 3, 4, and 5 days postinfection, and tissues were processed for electron microscopy. The birds killed up to 56 hr postinfection received 4 million sporulated oocysts and those killed at later periods Received for publication 2 March 1973. * Supported by Ontario Health Research Grant No. PR.297 and by the Ontario Ministry of Agriculture and Food. received 500,000 sporulated oocysts each. A section of the duodenum, about 2 inches proximal to the entry of the bile duct, was cut into pieces, 1 to 2 mm square, and fixed in 3% glutaraldehyde in Millonig's buffer at 4 C. The tissues were fixed from 4 to 48 hr, the best results often being obtained with longer fixation periods. They were washed for 24 hr or longer, first in Millonig's buffer and then in 0.2 mi sucrose solution, buffered at pH 7.4 with 0.1 M phosphate, postfixed in 1% osmium tetroxide in Millonig's buffer for 1 hr, washed in the same buffer, and dehydrated through a graded series of acetone, all at 4 C. The tissues were embedded in Epon and sectioned on a Reichert OM ultramicrotome. Ultrathin sections were mounted on 200-mesh copper grids and stained with lead citrate (Venable and Coggeshall, 1965), followed by a saturated solution of uranyl acetate in 50% ethanol. They were examined and photographed at 60 kv in a Phillips EM 300 electron microscope.