Characterization of a recombinant Eimeria acervulina antigen expressed in sporozoite and merozoite developmental stages

Abstract

A cDNA from Eimeria acervulina encoding an immunogenic region of antigens shared between sporozoites and merozoites was cloned and expressed in Escherichia coli. Immunofluorescence staining of sporozoites, merozoites, and coccidial-infected intestinal tissue with monoclonal antibody used to detect the recombinant clone indicated that the epitope was present on internal parasite proteins. Immunostaining of nitrocellulose paper containing protein from both asexual stages revealed numerous sporozoite antigens (18-120 kDa) and only 1 merozoite antigen (150 kDa). Northern blot hybridization assays using this cDNA clone for probing sporozoite and merozoite RNA showed that distinct transcripts were present in both asexual stages. Similar to the immunofluorescence studies, many homologous RNAs were observed in sporozoites (3.7-13 kb) and only 1 prominent hybridizing species was found in merozoites (3.3 kb). The recombinant coccidial antigen, designated MA16, is a 125-kDa beta-galactosidase fusion protein, representing about 10 kDa of parasite protein. In blastogenesis assays, purified recombinant MA16 antigen is capable of activating T lymphocytes obtained from E. acervulina-immune inbred chickens. DNA sequencing of MA16 clone and analysis of the predicted amino acid sequence indicated several putative T cell epitopes that may be responsible for the observed in vitro blastogenic response.