Diagnostically significant antigens of Treponema pallidum subsp. pallidum: identification, serological efficacy, and characterisation of the major antigenic determinants.

Grace, Christopher (2000). Diagnostically significant antigens of Treponema pallidum subsp. pallidum: identification, serological efficacy, and characterisation of the major antigenic determinants. PhD thesis The Open University.

DOI: https://doi.org/10.21954/ou.ro.0000ff8b

Abstract

The antibody response to native antigens derived from Treponema pallidum (Nichols strain) was investigated by western blotting. In early primary disease the responses were most frequently to the major immunodominant antigen, TpN47, and the flagellar proteins. Strong responses to several other lipoproteins, especially TpN44.5, TpN17 and TpN15, were observed in all other disease stages. Generally, this confirmed the importance of TpN47, TpN44.5, TpN17, and TpN15 as diagnostlcally significant antigen.

Western blot analysis using recombinant proteins showed the four major lipoproteins, TpN47, TpN44.5, TpN17, and TpN15, to be important disease specific antigens, with TpN47 particularly reactive in early primary infections. TpN24-28 was found to be disease specific but only reactive with relatively few strongly positive sera, and so of relatively little diagnostic significance. Further analysis of the other recombinant proteins by enzyme immunoassay confirmed them as important diagnostic antigens, but no single recombinant protein was capable of detecting all cases of serologically confirmed syphilis. However, an enzyme immunoassay using a solid phase coated with a combination of all four major lipoproteins proved highly specific and showed improved sensitivity over a commercially available enzyme immunoassay based on native antigens.

Synthetic peptide epitope mapping studies of the four major lipoproteins showed that the antigenic sequences within these proteins were not simple linear epitopes. The complicated picture of several reactive regions across the proteins, each containing multiple epitopes, was typical of the type of responses seen when a substantial part of the antibody response is directed towards conformational epitopes.

Further analysis of the epitopes present within the four major lipoproteins using phage display isolated several long protein sequences which represented high affinity antibody binding motifs. The size of the sequences suggests that the retention by the solid phase is due either to conformational epitopes, or to a mixture of epitope types producing high affinity binding through multipoint attachment. The sequences identified have potential as antigens for use in diagnostic tests.