CHARACTERIZATION OF AN IMMUNODOMINANT VARIABLE SURFACE-ANTIGEN FROM PATHOGENIC AND NONPATHOGENIC ENTAMOEBA-HISTOLYTICA

A 125-kD surface antigen of Entamoeba histolytica is recognized by 73% of immune sera from patients with amoebic liver abscesses. Using pooled human immune sera a cDNA clone (λcM17) encoding this antigen (M17) has been isolated from a λgtll expression library of the virulent stain E. histolytica HM1:IMSS. Monospecific antibodies, purified by binding to phage lysate of λcM17, and mAb FA7 reacted exclusively with the 125-kD antigen by Western blot analysis. Surface binding and cap formation are observed with patient sera, purified monospecific antiserum, and mAb FA7. Corresponding genomic clones (pBSgM17-1/2/3) were isolated by hybridization with the cDNA clone. These contained an open-reading frame of 3345 bp, which is in good agreement with the mRNA size of ~3.0 kb as revealed by Northern hybridization with λcM17. The inferred amino acid sequence predicts a 125,513 dalton protein that contains 17 potential Winked glycosylation sites and is unusually rich in tyrosine and asparagine residues. A distinctly hydrophobic NH2-terminal region may serve as membrane anchor or signal sequence. In contrast to conservation of an immunodominant epitope recognized in pathogenic and nonpathogenic strains by monoclonal FA7 and human immune sera, amplification and sequence analysis of a 1,400-bp fragment of this gene from a fresh nonpathogenic isolate by use ofthe PCR demonstrate regions of significant sequence divergence in this antigen. A 1% sequence variability among different isolates ofthe pathogenic strain HM1:IMSS and a 12-13% variability between pathogenic and nonpathogenic strains are revealed by comparison to published partial amino acid sequences (Tannich, E., R.D. Horstmann, J. Knobloch, and H.H. Arnold. 1989. Proc. Nad. Acad. Sci. USA. 86:5118.). Some restriction enzymes were found that allowed PCR diagnosis ofnonpathogenic and pathogenic isolates with the exclusion ofE. histolytica-like Laredo, suggesting that a detailed study of nonpathogenic and pathogenic isolates in relation to the M17 antigen sequence will provide a basis of differentiating isolates. © 1990, Rockefeller University Press., All rights reserved.