Serpin family protease inhibitors (SERPINs) in protozoans are key regulators of multiple biological functions, including the entry of parasites into host cells. Despite their importance, the molecular mechanisms by which SERPINs facilitate host invasion remain largely unresolved. Here, we demonstrate that Eimeria tenella SERPIN1 (EtSERPIN1), present on the sporozoite surface, plays a functional role in cell adhesion and invasion. To identify the host membrane component mediating EtSERPIN1-driven invasion, we applied GST pull-down and yeast two-hybrid approaches, revealing a specific interaction with annexin A2 (ANXA2). Recombinant GgANXA2 bound to sporozoites effectively, and pre-treatment of host cells with either anti-GgANXA2 antibody or recombinant GgANXA2 protein significantly inhibited EtSERPIN1 attachment. Furthermore, recombinant GgANXA2 reduced sporozoite infection in DF-1 cells and in chickens. These findings indicate that the EtSERPIN1–GgANXA2 interaction is critical for E. tenella sporozoite adhesion and invasion. Finally, functional assays with recombinant proteins showed that GgANXA2 significantly decreased sporozoite infectivity, as shown by lower parasite loads in the chicken cecum. Immunization with recombinant EtSERPIN1 provided strong protective effects, including improved body weight gain, reduced cecal lesions and oocyst shedding, and elevated mucosal antibodies. This study highlights EtSERPIN1 as a promising candidate for therapeutic targeting via the GgANXA2 pathway.
