Excystation of Eimeria tenella Sporozoites Impaired by 1 Antibody Recognizing Gametocyte / Oocyst Antigens 2 GAM 22 and GAM 56 * 3 4

18 The nucleotide sequence data reported in this paper appear in the DDBJ, EMBL and 19 Genbank® nucleotide sequence database with accession number CS000361. Abstract 1 Eimeria tenella is the causative agent of coccidiosis in poultry. Infection of chicken 2 intestine begins with ingestion of sporulated oocysts releasing sporocysts which in turn 3 release invasive sporozoites. The monoclonal antibody E2E5 recognizes wall-forming 4 bodies type II (WFBII) in gametocytes and the WFBII-derived inner wall of oocysts. 5 Here, we describe that this antibody also binds to the stieda body of sporocysts and 6 significantly impairs in vitro excystation of sporozoites. Using affinity chromatography 7 and protein sequence analysis, E2E5 is shown to recognize EtGAM56, the E. tenella 8 ortholog of the Eimeria maxima gametocyte-specific GAM56 protein. In addition, this 9 antibody was used to screen a genomic phage display library presenting E. tenella 10 antigens as fusion proteins with gene VIII product on the surface of phagemid particles 11 and identified the novel 22 kDa histidine-and proline-rich protein EtGAM22. The 12 Etgam22 mRNA is predominantly expressed at the gametocyte stage as detected by 13 Northern blotting. Southern blot analysis in combination with data from the E. tenella 14 genome project revealed that Etgam22 is an intronless multi-copy gene with 15 approximately 12-22 copies in head to tail arrangement. Conspicuously, Etgam56 is also 16 intronless and is localized adjacent to another gam56-like gene, Etgam59. Our data 17 suggest that amplification is common for genes encoding oocyst wall proteins.