CONICET | Buscador de Institutos y Recursos Humanos

Chlamydia trachomatis (CT) is an obligate intracellular pathogenand the leading cause of sexually transmitted bacterial infectionsglobally. CT resides inside a vacuole or ?inclusion? and undergoes alife cycle involving the infectious elementary body and the replicatingreticulate body. Polymorphic membrane proteins (PMPs) are afamily of Chlamydia-specific autotransporter proteins with proposedroles in adhesion and antigenic variation. However, due to technicallimitations in genetic manipulation of this bacterium, the role of PMPsin CT biology remains poorly elucidated. By chemical mutagenesiscoupled with whole genome sequence analysis, we obtained 3 mutantswith nonsense mutations in pmpA/B/C. Due to the presenceof extra mutations/genome (DpmpA: 6; DpmpB: 17; DpmpC: 3) webackcrossed these mutants with a Wt strain to obtain recombinantscarrying only the mutations in pmp genes. We obtained cleaned recombinantsfor pmpA/C. For pmpB mutant we were able to clean14 of 17 mutations. We carried out IFU (Infectious Forming Units)assays in HeLa cells and observed that the generation of infectiousprogeny in the recombinants carrying nonsense mutations for pmpA/B/C was significantly impaired compared to Wt. These data pointout that PMPs are important for efficient propagation of CT in epithelialcells. Additionally, by confocal microscopy and electron microscopy,DpmpC strain showed an aggregation phenotype that couldnot be trans-complemented with the Wt strain in coinfection assays,suggesting a role for PmpC in preventing aggregation of individualbacteria inside the chlamydial inclusion. We are currently using arecently developed technology for gene deletion in CT by insertionof a group II intron in order to obtain pmpA, pmpB and pmpC nullmutants to evaluate the role of PMPs in CT pathogenesis in a micemodel of infection.