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Tubulins are heterodimeric molecules responsible for the polymerization of microtubules in apicomplexan parasites. The alpha-tubulin, a subcellular structural protein of Eimeria acervulina, was cloned and expressed in Escherichia coli as an alpha-tubulin-GST fusion protein. Immunogenicity of the recombinant protein was studied in chickens by subcutaneous injection of 50, 100, or 150 microg of the protein with or without Freund's adjuvant. Immunization with 150 microg alpha-tubulin-GST protein in combination with Freund's adjuvant conferred partial protection against E. acervulina oocyst challenge, as shown by a 36% reduction in oocyst shedding, a marked decrease in intestinal lesion score and a significant increase in body weight gain in comparison with the nonimmunized controls. The results suggest that alpha-tubulin protein may be used as an effective vaccine antigen for the control of Eimeria infection.

Live attenuated coccidiosis vaccines could be used as powerful carriers, expressing exogenous viral and bacterial antigens, to induce protective immunity against pathogenic organisms. We investigated the ability of Eimeria tenella to express an exogenous gene in vitro. Eimeria tenella sporozoites were transfected with the plasmid pH4-2EYFP-Actin3 containing the yellow fluorescent protein gene (yfp) and inoculated into primary chicken kidney cells (PCKCs), followed by incubation at 41 C in a 5% CO2 chamber. Fluorescent sporozoites were observed as early as 15-20 hr post-inoculation (PI). Fluorescence displayed by the expressed YFP protein was visible throughout the schizogony and gametogony stages of the tranfected E. tenella. Fluorescent oocysts were found between 200-327 hr PI. Higher fluorescence intensity was observed in the nucleus than in other compartments of the transfectants, while little or no fluorescence was seen in the refractile globule. The diversity of schizonts, particularly of the first generation, was presented by fluorescent nuclei arranged in different patterns. Our results demonstrated the ability of E. tenella to express an exogenous gene throughout the endogenous development in vitro. Completion of the endogenous development of transfected E. tenella in cell cultures will facilitate the study of foreign antigen expression in Eimeria spp., paving the way for the development of an Eimeria spp. vector vaccine that also carries and delivers other vaccines by oral administration.

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