Expression of diphtheria toxin in Streptococcus mutans and induction of toxin-neutralizing antisera.
The nontoxic full-length diphtheria toxin (DTX), fragment A (DTA), and fragment B (DTB) were each genetically fused to the major surface protein antigen P1 (SpaP) of Streptococcus mutans. Repeated attempts to express the recombinant DTX and DTB in the live oral vaccine candidate Streptococcus gordonii were unsuccessful, whereas DTA could be readily expressed in this bacterium. However, the recombinant DTX, DTB, and DTA could be expressed in the related oral bacterium S. mutans. Western blotting and enzyme-linked immunosorbant assay (ELISA) using anti-DTX and anti-P1 antibodies demonstrated the expression of the three fusion proteins in S. mutans. Mouse antisera raised against the recombinant S. mutans recognized the native DTX in Western immunoblotting. The antibodies raised against S. mutans expressing the recombinant DTX and DTA neutralized the cytotoxicity of the native toxin in a Vero cell assay, but the neutralization titers were relatively low. The potential of using S. gordonii as a live vaccine against diphtheria faces major challenges in the expression of DTX in this organism and in the induction of high-titer toxin-neutralizing antibodies.
Lee
C W
CW
Department of Applied Oral Sciences, Faculty of Dentistry, Dalhousie University, Halifax, NS, Canada.
Halperin
S A
SA
Morris
A
A
Lee
S F
SF
D000818:Animals; D000907:Antibodies, Bacterial / Q000097:blood* / Q000276:immunology*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002522:Chlorocebus aethiops; D004167:Diphtheria Toxin / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009500:Neutralization Tests; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D013295:Streptococcus mutans / Q000235:genetics / Q000254:growth & development / Q000276:immunology / Q000378:metabolism*; D014709:Vero Cells
D000907:Antibodies, Bacterial; D001426:Bacterial Proteins; D004167:Diphtheria Toxin; D008562:Membrane Glycoproteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C055764:S-layer proteins; C029795:diphtheria toxin fragment A; C039171:diphtheria toxin fragment B