Developmental gene expression in Eimeria bovis

By differential screening of stage-specific cDNA libraries of Eimeria bovis, we have identified and isolated a large set of genes that are regulated during development of the sporozoites and merozoites. Duplicate lifts of cDNA libraries constructed from partially sporulated oocysts and merozoites were probed with radioactively labeled first-strand cDNA prepared from partially sporulated oocyst and merozoite mRNA. Out of 60000 plaques screened in each case, over 250 plaques from the partially sporulated oocyst library preferentially hybridized with the oocyst cDNA probe and 67 plaques from the merozoite library preferentially hybridized with the merozoite cDNA probe. Three of the oocyst phage and 7 of the merozoite phage were selected for further characterization. Northern analysis revealed a common pattern of mRNA expression for the oocyst cDNA clones. Consistent with the results of the differential screen, no hybridization to merozoite RNA was detected with any of these 3 oocyst cDNA clones. The expression of the merozoite cDNA clones was more complex, with 3 different classes of merozoite genes being identified based on their pattern of developmental regulation. Although each of the merozoite clones was expressed to some extent during sporulation, in all cases, expression was higher in merozoites than in partially sporulated oocysts, consistent with the restriction of expression defined by the differential screen. Sequence analysis revealed that 2 of the merozoite cDNA clones encode elongation factor 1 alpha and the ubiquitin/ribosomal protein fusion, and 1 of the sporozoite cDNAs displays a significant identity to insulin-degrading enzyme. The developmental expression of E. bovis genes involved in protein synthesis and degradation provides additional evidence for the importance of regulation of protein metabolism during parasite development.