Eimeria tenella: Vitamin Requirements for Development in Primary Cultures of Chicken Kidney Cells

SYNOPSIS. Development of Eimeria tenella was studied in primary cultures of chicken kidney cells maintained in Medium 199 lacking each of the following: vitamin A. biotin, p -aminobenzoic acid, folic acid, nicotinamide, Ca pantothenate, pyridoxine, pyridoxal, riboflavin, thiamin, ascorbic acid, calciferol, α-tocopherol, and menadione. Data obtained concerning numbers of mature schizonts or total numbers of parasites or both indicated that all of the vitamins are needed for 1st- and 2nd-generation schizogony, and all except calciferol and folic acid are needed for gametogony.     

The Effect of Eimeria nieschulzi Infection on Leukocyte Levels in the Rat*

SYNOPSIS. Rats inoculated with 10,000 sporulated oocysts of Eimeria nieschulzi had significantly higher total leukocyte counts on postinoculation days (PI) 1, 5, 6 and 7 when compared to control rats. Relative and absolute neutrophil counts increased concomitantly with a decrease in the relative lymphocyte levels in E. nieschulzi-infected rats on PI day 7. Absolute and relative neutrophil counts in infected rats on PI days 7 and 8 were closely correlated with the host's total oocyst discharge. The E. nieschulzi infection had no significant effect on the relative or absolute levels of monocytes or eosinophils. The described changes in leukocyte levels were not paralleled by a significant change in the erythrocyte count.     

Coccidian Parasites of Intertidal Fishes from Wales: Systematics, Development, and Cytochemistry

SYNOPSIS. Examination of littoral fish Blennius pholis and Cottus bubalis caught at Aberystwyth and Porth Cwyfan, Wales. U.K., revealed 2 species of coccidia.
Eimeria dingleyi sp. n. Oocysts spherical (16.1–19.2) to subspherical (13.9–14.2 × 18.8–20.0) μm, with thin walls; sporulation outside the host to produce ellipsoid sporocysts; endogenous phases in epithelial cells throughout intestine; 26 of 58 B. pholis infected.
Eimeria variabilis (Thélohan) Reichenow. Oocysts spherical (11.9–14.6) to subspherical (9.2–10.9 × 13.9–14.3) μm; sporulation in lining of pyloric ceca and rectum; previously unrecorded schizonts and gametocytes present; 21 of 25 C. bubalis infected.
Electron microscopy revealed that the oocyst wall of E. variabilis consists of a thin membrane whereas the sporocyst wall is thick and 3-layered. Typical oocyst wall-forming bodies were absent from the macrogamete. Cytochemical tests on the endogenous stages of E. dingleyi and E. variabilis indicated that in general they resembled other coccidia in their chemical constitution.     

Scanning Electron Microscopy of Schizogony in Eimeria tenella

SYNOPSIS. Developing 2nd- and 3rd-generation schizonts of Eimeria tenella were found in the ceca of chicks infected orally with sporulated oocysts. Several free 2nd-generation schizonts, which varied in diameter from 11 to 21.6 μm, were found on the epithelial surface of the cecum. Some schizonts appeared to have lost merozoites. Other schizonts were intact, one of which was surrounded by an unbroken membrane that followed the contours of the merozoites. Third-generation schizonts, much smaller than 2nd-generation schizonts and with fewer merozoites, were found only on cut or fractured surfaces of the cecal tissue. Third-generation merozoites appeared shorter and thicker than those of the 2nd-generation and were attached to the schizont residuum. A form with conical protuberances and another with 4 triangular segments were found; they were believed to be developing stages 3rd-generation schizonts.     

Eimeria dispersa and Eimeria gallopavonis: Infectivity, Survival, and Development in Primary Chicken and Turkey Kidney Cell Cultures

SYNOPSIS. Eimeria dispersa (turkey strain) and Eimeria gallopavonis sporozoites were inoculated into primary cultures of chicken kidney (CK) and turkey kidney (TK) cells. Eimeria dispersa sporozoites were more infective in either cell type than those of E. gallopavonis : at 4 hr, the percentage of infection was 67-98 for E. dispersa but only 23-56 for E. gallopavonis . E. dispersa also survived better in culture: at 2 days, losses of E. dispersa in both cell types were only 4-19%, whereas losses of E. gallopavonis were 35-47% in TK cells and 60–95% in CK cells. However, E. gallopavonis developed further than E. dispersa . Location and increase in numbers of intracellular stages at 4 days indicated that E. dispersa proceeded through 2 schizogonic generations before development stopped.     

The Life Cycle of Eimeria dispersa Tyzzer, 1929 in Turkeys

SYNOPSIS. The life cycle of a turkey strain of Eimeria dispersa Tyzzer was studied in Beltsville Small White turkeys. There were 4 asexual generations. Mature schizonts of the first generation were present 30 h postinoculation (PI); those of the 2nd, 3rd, and 4th generations were present 48, 72, and 96 h PI, respectively. Average size of schizonts and number and size of merozoites for each generation were as follows: first , 14.3 × 13.0 μm with 19.2 merozoites, each 4.5 × 1.2 μm; second , 8.0 × 7.2 μm with 13.5 merozoites, each 4.5 × 1.1 μm; third , 8.9 × 8.9 μm with 15.1 merozoites, each 5.6 × 2.1 μm; fourth , 11.6 × 10.5 μm with 6.7 merozoites, each 8.2 × 2.0 μm. Sporozoites and developmental stages of the first generation were in close association with an epithelial cell nucleus and located between the brush border and the "row" of epithelial cell nuclei; developmental stages of the other 3 generations were not associated with a nucleus and were located just under the brush border. Early macrogametes and microgametocytes were present 96 h PI. Development was confined to the epithelial cells of the villus and extended from the tip of the villus to ∼ 1/2 the distance down the sides in all areas of the intestine except the cecum. The prepatent period was between 114 and 120 h. Percentage of sporulation was 15, 57, and 90, at 24, 36, and 48 h, respectively. Sporulated oocysts averaged 24.5 × 20.2 μm.     

Thymidylate Synthetase as a Chemotherapeutic Target in the Treatment of Avian Coccidiosis*

SYNOPSIS. Thymidylate synthetase (E.C.2.1.1.45) has been demonstrated in unsporulated oocysts of Eimeria tenella. The properties of this enzyme have also been investigated in Tetrahymena pyriformis, as a protozoan model, and 7-day-old chick embryo, as a host model. The enzymes from E. tenella and chick embryo were inhibited by all concentrations of MnCl₂ and MgCl₂ tested. Tetrahymena pyriformis thymidylate synthetase was stimulated by low concentrations of both these cations but was inhibited by high concentrations. Subsequent data refer to chick embryo, E. tenella and T. pyriformis respectively: the apparent K_m was 5.89 μM, 5.94 μM, and 0.53 M for the substrate dUMP: and 5.13 μM, 1.10 μM and 4.65 μM, respectively for the cofactor N⁵N¹⁰-methylenetetrahydrofolate. The pH optimum for the enzyme from both chick embryo and T. pyriformis was 8.0, with Tris-HCl buffer; activity of E. tenella thymidylate synthetase was still increasing at pH 8.2. The E. tenella enzyme was found to have a molecular weight of 4.6–4.9 × 10⁵ daltons. The effects of nucleotides, inhibitors, and the omission of assay components on each enzyme are presented. Thymidylate synthetase from E. tenella is not greatly different from that of chick embryo, but does not resemble the enzyme from T. pyriformis. A case for using thymidylate synthetase as a chemotherapeutic target in the treatment of Eimeria infections remains. Indeed Eimeria may be considered as a model for infections caused by other protozoan parasites, such as Toxoplasma and Plasmodium, provided that suitable inhibitors can be found that are not toxic to the host.     

Eimeria tenella ROP kinase EtROP1 induces G0/G1 cell cycle arrest and inhibits host cell apoptosis

Coccidia are obligate intracellular protozoan parasites responsible for human and veterinary diseases. Eimeria tenella, the aetiologic agent of caecal coccidiosis, is a major pathogen of chickens. In Toxoplasma gondii, some kinases from the rhoptry compartment (ROP) are key virulence factors. ROP kinases hijack and modulate many cellular functions and pathways, allowing T. gondii survival and development. E. tenella's kinome comprises 28 putative members of the ROP kinase family; most of them are predicted, as pseudokinases and their functions have never been characterised. One of the predicted kinase, EtROP1, was identified in the rhoptry proteome of E. tenella sporozoites. Here, we demonstrated that EtROP1 is active, and the N‐terminal extension is necessary for its catalytic kinase activity. Ectopic expression of EtROP1 followed by co‐immunoprecipitation identified cellular p53 as EtROP1 partner. Further characterisation confirmed the interaction and the phosphorylation of p53 by EtROP1. E. tenella infection or overexpression of EtROP1 resulted both in inhibition of host cell apoptosis and G0/G1 cell cycle arrest. This work functionally described the first ROP kinase from E. tenella and its noncanonical structure. Our study provides the first mechanistic insight into host cell apoptosis inhibition by E. tenella. EtROP1 appears as a new candidate for coccidiosis control.     

La Genèse des Mérozoïtes chez la Coccidie Eimeria necatrix. Etude Ultrastructurale*

RESUME. Les schizontes de 2 ème génération d'Eimeria necatrix ont étéétudiés au microscope électronique. La différenciation des mérozoïtes est associée à la dernière mitose, qui ne semble pas différer essentiellement des précédentes. Les mérozoïtes se développent à la périphérie du schizonte. Le conoide et 22 microtubules sous pelliculaires, probablement induits par les centrioles, et le complexe membranaire interne ainsi que les précurseurs des rhoptries, qui semblent issus de l'appareil de Golgi, apparaissent auprès de chaque pôle nucléaire, sous la membrane du schizonte. Ces organites sont les premiers inclus dans les ébauches de mérozoïtes. Puis, le noyau, le dictyosome et les vésicules multimembranaires pénètrent dans les futurs mérozoïtes. Les micronèmes, probablement formés par l'appareil de Golgi, et les grains d'amylopectine sont produits plus tard, quand les mérozoïtes se séparent du reliquat cytoplasmique. Le mode de genèse de ces divers organites et les relations entre le dernière mitose et la différenciation sont discutés. SYNOPSIS. Second generation schizonts of Eimeria necatrix were studied with the aid of the electron microscope. Differentiation of daughter merozoites is associated with the last mitosis, which is not significantly different from the earlier ones. The merozoites develop at the periphery of the schizont. The conoid and 22 subpellicular microtubules, probably induced by centrioles, and the inner membranes complex and the rhoptry anlagen which seem to be produced by the Golgi apparatus, appear close to each nuclear pole, just near the schizont membrane. These organelles are the first to appear in the merozoite anlagen. Then, nucleus, dictyosome and multimembranous vesicles enter the budding merozoites. Micronemes, probably originating from Golgi apparatus, and amylopectin granules are produced later, when daughter merozoites separate from the residuum. The genesis of these various organelles and the relation between the last mitosis and differentiation are discussed.     

柔嫩艾美耳球虫(Eimeria tenella)早熟株与亲本株致病性和繁殖力的研究

通过对经15代选育的柔嫩艾美耳球虫(E. tenella)山西株的早熟株与其亲本株的繁殖力和致病性进行比较研究,证实早熟株的潜隐期比亲本株缩短21 h,繁殖力下降40%左右;对致病性的研究显示,早熟株感染后对鸡只增重、AC I的影响较小,对11日龄雏鸡的半数感染量和半数致死量较亲本株增大,肠道病变记分较亲本株下降。由此认为,该早熟株符合球虫早熟株的特性,可用于鸡球虫病早熟苗的制作。