Date of Award:

5-1969

Document Type:

Dissertation

Degree Name:

Doctor of Philosophy (PhD)

Department:

Animal, Dairy, and Veterinary Sciences

Department name when degree awarded

Zoology

Committee Chair(s)

Datus M. Hammond

Committee

Datus M. Hammond

Abstract

In vitro development of Eimeria auburnensis was studied in cell cultures. The cell types used were established cell lines of adult bovine kidney (Madin-Darby) and embryonic human intestine cells (Int 407); also cell line cultures of embryonic bovine trachea and embryonic bovine spleen cells were used. Monolayer cell cultures were serially propagated in disposable prescription bottles and transferred to Leighton tubes for use in experiments. Coverslips from Leighton tubes cultures inoculated with sporozoites were examined at regular intervals with phase-contrast microscopy for as long as 20 days after inoculation. After examination, coverslips were fixed in Schaudinn's fluid or Stieve's fixative and stained with iron hematoxylin or PAS-AO. Multiple infections were seen in all cell types; a maximum of 14 sporozoites were observed in a single kidney cell. Development proceeded only to the trophozoite stage in the human intestine cells. Mature first-generation schizonts were found in all other cell types. Generally, the most favorable results were obtained with kidney cells, although the largest schizonts occurred in spleen and trachea cells. Development to mature schizonts occurred in 3 of 6 experiments with trachea cells, but no development beyond intracellular sporozoites was observed in the others. Few intermediate or mature schizonts were seen in BES cells. Some mature schizonts occurred in each of 3 experiments with these cells. Intracellular sporozoites were observed to undergo a change in morphology of the nucleus, and in size of the nucleus and nucleolus. Sporozoites became enlarged in size and underwent nuclear division to form sporozoite-shaped schizonts. These retained the appearance of sporozoites, but had 2 to 5 nuclei. The enlarged sporozoites or sporozoite-shaped schizonts developed cytoplasmic outpocketing and transformed directly into trophozoites or spheroidal schizonts; trophozoites were rarely seen. Crescent bodies of different shapes and sizes were present as early as 3 days after inoculation; they were rarely seen in large intermediate or mature schizonts. Refractile bodies became smaller as schizonts increased in size. Numerous intermediate schizonts developed in the kidney cells, a smaller number in the trachea cells, and only a few in spleen cells. In each of these cell types, few mature schizonts were observed. Enlargement of the host cell nucleus and nucleolus was first observed in cells harboring the enlarged sporozoites or sporozoite-shaped schizonts; relatively little further enlargement occurred in later stages. Many of the host cells in each cell type had 2 nuclei and occasionally 3 or 4 nuclei occurred in a single host cell. Degenerative changes in the parasite and the host cell usually were noticed about 2 days after the occurrence of the first mature schizont. Degenerative changes of the host cell included increased vacuolization as well as granulation and darkening of the cytoplasm. In schizonts where vacuoles appeared, cell outlines became irregular, nuclei became indistinct, and particulate matter accumulated in the parasitophorous vacuole. Degenerative later occurred in non-infected cells; in some experiments little degeneration occurred. Schizonts in cell cultures were smaller and more variable in size than those which developed in calves.

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