Molecular cloning and expression of an immunodominant 53-kDa excretory-secretory antigen from Trichinella spiralis muscle larvae.
A Trichinella spiralis cDNA expression library was constructed in lambda gt11 from muscle larvae mRNA and immunologically screened to identify genes encoding previously described immunodiagnostic excretory-secretory (ES) antigens. Screening the library with T. spiralis infection serum from swine or rabbit antiserum to T. spiralis ES antigen identified one clone, designated TsA-12, that contains a cDNA transcript 539 bp in length and codes for an apparent 123-kDa beta-galactosidase fusion protein that does not cross-react with Trichuris suis or Ascaris suum infection serum. Western blots of T. spiralis extracts and immunoperoxidase staining of tissue sections from muscle larvae using antibodies to purified TsA-12 demonstrate homology between TsA-12 and the 53 kDa diagnostic antigen from ES products (designated Ts.53) and localize the homologous native antigen to the stichocyte cells of the parasite. ELISA tests using TsA-12 as antigen, detected antibodies to T. spiralis in experimentally-infected mice as early as 14 days post-inoculation with maximum antibody titers being reached at 28 days post-inoculation. The TsA-12 dscDNA hybridizes to mRNA sequences expressed in both the muscle larvae and adult stages; however, concomitant expression of the native antigen is not observed within adult ES products. Southern blots of homologous and heterologous genomic DNAs probed with 32P-labeled TsA-12 dscDNA fragments verify TsA-12 as a T. spiralis specific sequence that is present in multiple copies within the parasite genome.
Zarlenga
D S
DS
Biosystematic Parasitology Laboratory, ARS-USDA, Beltsville, MD 20705.
Gamble
H R
HR
D000595:Amino Acid Sequence; D000818:Animals; D000947:Antigens, Helminth / Q000235:genetics* / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular; D004274:DNA, Recombinant / Q000737:chemistry; D005260:Female; D015698:Genomic Library; D015801:Helminth Proteins / Q000235:genetics* / Q000276:immunology; D007814:Larva / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009132:Muscles / Q000469:parasitology*; D012333:RNA, Messenger / Q000737:chemistry; D051381:Rats; D011919:Rats, Inbred Strains; D011993:Recombinant Fusion Proteins / Q000276:immunology; D014234:Trichinella / Q000235:genetics* / Q000276:immunology
D000947:Antigens, Helminth; D004274:DNA, Recombinant; D015801:Helminth Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C097419:excretory-secretory antigen, Trichinella