title: A recombinant Eimeria protein inducing interferon-gamma production: comparison of different gene expression systems and immunization strategies for vaccination against coccidiosis.
abstract: A rabbit antiserum against an 18- to 27-kD native protein fraction (F3) from Eimeria acervulina merozoites identified a cDNA (3-1E) containing a 1086-base pair insertion with an open reading frame of 170 amino acids (predicted molecular weight, 18,523). The recombinant 3-1E cDNA expressed in Escherichia coli produced a 60-kD fusion protein and a 23-kD protein after factor Xa treatment of the fusion protein. Both proteins were reactive with the F3 antiserum by western blot analysis. A rabbit antiserum against a synthetic peptide deduced from the amino acid sequence of the 3-1E cDNA reacted with a 27-kD recombinant 3-1E protein expressed in Sf9 insect cells and a 20-kD native protein expressed by E. acervulina sporozoites and Eimeria tenella sporozoites and merozoites. By immunofluorescence staining, a monoclonal antibody produced against the recombinant 3-1E protein reacted with sporozoites and merozoites of E. acervulina, E. tenella, and Eimeria maxima. Spleen lymphocytes from E. acervulina-immune chickens showed antigen-specific proliferation and interferon (IFN)-gamma production upon stimulation with the recombinant 3-1E protein, indicating that the protein activates cell-mediated immunity during coccidiosis. Immunization of chickens with either the E. coli- or Sf9-expressed recombinant 3-1E protein with adjuvant, or direct injection of the 3-1E cDNA, induced protective immunity against live E. acervulina. Simultaneous injection of the recombinant 3-1E protein, or the 3-1E cDNA, with cDNAs encoding chicken IFN-gamma or interleukin (IL)-2/15 further enhanced protective immunity. These results indicate that the recombinant E. acervulina 3-1E cDNA or its polypeptide product may prove useful as vaccines against avian coccidiosis.
authors.affiliation: Immunology and Disease Resistance Laboratory, Livestock and Poultry Sciences Institute, BARC-East, U.S. Department of Agriculture, Beltsville, MD 20705, USA.
authors.forename: H S
authors.initials: HS
authors.lastname: Lillehoj
authors.forename: K D
authors.initials: KD
authors.lastname: Choi
authors.forename: M C
authors.initials: MC
authors.lastname: Jenkins
authors.forename: V N
authors.initials: VN
authors.lastname: Vakharia
authors.forename: K D
authors.initials: KD
authors.lastname: Song
authors.forename: J Y
authors.initials: JY
authors.lastname: Han
authors.forename: E P
authors.initials: EP
authors.lastname: Lillehoj
mesh_terms: D000818:Animals; D015153:Blotting, Western / Q000662:veterinary; D003001:Cloning, Molecular; D003048:Coccidiosis / Q000276:immunology / Q000517:prevention & control / Q000662:veterinary*; D004539:Eimeria / Q000235:genetics* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary; D005786:Gene Expression Regulation; D007106:Immune Sera; D007371:Interferon-gamma / Q000096:biosynthesis*; D008969:Molecular Sequence Data; D011201:Poultry Diseases / Q000276:immunology / Q000469:parasitology / Q000517:prevention & control*; D015800:Protozoan Proteins / Q000276:immunology / Q000494:pharmacology*; D011817:Rabbits; D011994:Recombinant Proteins / Q000276:immunology / Q000494:pharmacology; D017422:Sequence Analysis, DNA / Q000662:veterinary; D014611:Vaccination / Q000662:veterinary*; D019444:Vaccines, DNA / Q000276:immunology
chemical_list: D007106:Immune Sera; D015800:Protozoan Proteins; D011994:Recombinant Proteins; D019444:Vaccines, DNA; D007371:Interferon-gamma