Identification and heterologous expression of a Sarcoptes scabiei cDNA encoding a structural antigen with immunodiagnostic potential.
The mite Sarcoptes scabiei causes sarcoptic mange (or scabies), a disease of considerable human and veterinary significance. An S. scabiei cDNA clone of about 2 kb was isolated from a S. scabiei var. hominis expression library by immunological screening using blood serum from a naturally infected chamois (Rupicapra rupicapra). The nucleotide sequence of the identified cDNA contains an open reading frame of 1930 bp that encodes a 642 amino acid polypeptide. This polypeptide shows tandem repeats of a glycine-serine rich 20 residue sequence followed by a unique C-terminal glutamate rich 54 residue sequence. The cDNA or the deduced polypeptide did not show significant similarities to any of the sequences in the databases. A carboxyl-terminal fragment of this polypeptide (residues 380 to 642) was efficiently expressed in Escherichia coli as a fusion with Glutathione S-transferase and then was used to produce a specific antiserum. The antigen encoded by the cDNA was located at the integument of the mite's epidermis and the cavities surrounding its vital organs. Western blot analysis of mite extracts using the specific antiserum against the recombinant protein identified antigens larger that 60 kDa indicating that the isolated cDNA did not contain the full ORF. Moreover, we designed a diagnostic assay based on the carboxyl-terminal fragment of the antigen for the identification of infected animals.
Casais
Rosa
R
Servicio Regional de Investigación y Desarrollo Agroalimentario, Laboratorio de Sanidad Animal, Jove, Gijón, Asturias, Spain.
Prieto
Miguel
M
Balseiro
Ana
A
Solano
Paloma
P
Parra
Francisco
F
Martín Alonso
José M
JM
D000595:Amino Acid Sequence; D000818:Animals; D000941:Antigens / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D015153:Blotting, Western / Q000662:veterinary; D018076:DNA, Complementary / Q000032:analysis* / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay / Q000662:veterinary; D004926:Escherichia coli; D005260:Female; D005786:Gene Expression Regulation; D015723:Gene Library; D015511:Goat Diseases / Q000175:diagnosis* / Q000469:parasitology; D006041:Goats; D008297:Male; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000276:immunology; D048608:Rupicapra; D012520:Sarcoptes scabiei / Q000235:genetics* / Q000276:immunology; D012532:Scabies / Q000175:diagnosis / Q000469:parasitology / Q000662:veterinary*; D020080:Tandem Repeat Sequences
D000941:Antigens; D018076:DNA, Complementary; D011994:Recombinant Proteins