A primate model system was used to identify antigens uniquely recognized in the context of infection. Serum antibody titres were measured in cynomolgus monkeys challenged urethrally with serovar L2 elementary bodies (EBs). High-titre sera from these primates were used, in parallel with antisera against killed EBs, to differentially screen an expression library of serovar L2 DNA. Four clones were recognized only by antisera from infected monkeys. Sequence analysis revealed that three of these immunoreactive clones overlap a common ORF, designated ORF D242 (encoding p242), in the genome database. The fourth clone contains two complete ORFs, each encoding 32 kDa proteins that share identity with TroA and TroB (ORFs D067 and D068 in the database, respectively). lmmunoblot analysis of lysates expressing TroA, TroB and p242 fusion proteins showed that p242 and TroA, but not TroB, were detected by the sera collected from infected primates. Antibodies directed at TroA and p242 were also detected in sera from several -infected patients, demonstrating that these proteins are also recognized by humans following infection. Immunoblot analysis with antibody against TroA and p242 also demonstrated that both antigens are present in higher abundance in infected ChoK1 cells relative to purified EBs. Immunofluorescence microscopy shows that TroA and p242 are both localized to intracellular developmental forms at the margins of growing inclusions. Collectively, these studies identify two proteins that are under-represented in EBs and are recognized uniquely in the context of infection.


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