Chlamydia trachomatis is a leading cause of sexually transmitted diseases and a number of strategies have been developed to produce vaccines to prevent its transmission. The purpose of this study was to map the neutralizing epitopes of C. trachomatis major outer-membrane protein (MOMP) serovar K by using anti-MOMP antibodies and synthetic peptides. Seven anti-MOMP monoclonal antibodies and three polyclonal antisera were produced and characterized. Their fine specificity was defined by direct binding assay on 15 peptides of 10 amino acid residues, overlapping by five residues, corresponding to the four variable domains (VDI-VDIV : residues 64-85, 139-160, 224-237 and 287-319) of MOMP serovar K. Our data confirmed that a neutralizing epitope is found in VDIV, defined by peptides K12 and K13. This epitope is 296rTLNPTIAG304, which has never been reported as a neutralizing epitope of serovar K. Another neutralizing epitope, defined by peptide K2, has been identified in VDI. This epitope is in the same position as 71VAGLEK76, a peptide with neutralizing activity found in serovar A, but they are not identical because antibodies against peptide K2 do not bind to this epitope. No neutralizing epitope was found in the two other variable domains (VDII and 111). In summary, two neutralizing sites, one in variable domain I and one in variable domain IV, were identified in serovar K.

The major outer membrane protein (MOMP) is a primary target antigen for the development of chlamydial vaccine. This protein is composed of four variable domains (I to IV) flanked by constant regions. Some of the variable domains contain antigenic determinants that elicit a neutralizing antibody response. Murine monoclonal antibodies (MAbs) against three nonoverlapping epitopes of MOMP were developed. One of these, called DP1O, bound to all serovars, as shown by immunoblot analysis, and neutralized chlamydial infectivity for hamster kidney (HaK) cells in a complement-independent in vitro assay. Furthermore, analysis of the fine specificity of this MAb showed that it recognized a synthetic peptide contained within variable domain IV of the MOMP. Anti-idiotypic antibodies (aId) directed against this anti-MOMP MAb were produced in rabbits. These aId specifically bound to the relevant idiotype (DP10) and inhibited the binding of anti-MOMP MAb (DP10) to MOMP preparations in a dose-dependent fashion. The specificity of our aId for the binding site of anti-MOMP MAb is further suggested by the binding inhibition of affinity-purified aId to DP1O by the synthetic peptide defined by the idiotype. In addition, these aId also reacted with anti-MOMP antisera from rats and mice, suggesting an idiotypic cross-reactivity between these species. Finally, immunization of naive mice with aId induced an antibody response directed against the peptide defined by our anti-MOMP MAb and with neutralizing activity. Taken together, these data suggest that aId mimic a neutralization site on MOMP and could serve as a surrogate antigen to induce protective immunity against Chlamydia trachomatis.

The major outer membrane protein of Chiamydia trachomatis has been extensively studied and is still considered one of the most promising candidates for development of a synthetic vaccine. Neutralizing epitopes in variable domains I, II, and IV have already been reported. In variable domain I, residues 69 to 78 have been identified as a neutralizing epitope for some of the C-and C-related complex serovars (A, C, I, J, L3, and K). It is not known whether epitopes located at the same position in B-complex serovars are neutralizing. To clarify this point, rabbit polyclonal antibodies directed against the peptide 6TGNAVAPS78 from the B serovar were produced. Rabbit antisera were further rendered peptide specific by purification on a peptide-bovine serum albumin-Sepharose affinity column. Peptide-specific rabbit immunoglobulin reacted with five of the B-complex serovars (B, Ba, E, Li, and L2) by immunoblot and by direct-binding enzyme-linked immunosorbent assay. Furthermore, this peptide-specific rabbit immunoglobulin neutralized the chlamydial infectivity of both serovars B and E for HaK cells in a complement-independent in vitro assay. The importance of these results stems from the fact that peptide 69T1'TGNAVAPS78 was able to induce an antibody response directed against Band B-related complex serovars, including serovar E, which is responsible for a high proportion of genital infections. This peptide could therefore be considered for the construction of a multivalent synthetic vaccine. Chlamydia trachomatis is the leading cause of sexually transmitted diseases in industrialized countries and is also a major cause of preventable blindness in underdeveloped countries (13). On the basis of pathogenicity, antigenicity, and nucleic acid composition, human-pathogenic strains of C. trachomatis have been subdivided into 15 serovars (Li, L2, L3, A through K, and Ba). By their antigenic properties, these serovars were grouped into three complexes: the B complex (B, Ba, D, E, Li, and L2), the intermediate complex (F, G, K, and L3), and the C complex (A, C, H, I, and J) (7, 16). Intermediate serovars can also be subdivided into B-related (F and G) and C-related (K and L3) complex serovars. All 15 serovars of C. trachomatis bear a major outer membrane protein (MOMP), which constitutes 60%, by weight, of the chlamydial outer membrane proteins (4). The MOMP genes encode a highly conserved protein structure that contains four evenly spaced domains, with sequences varying among the different serovars, called the four variable domains