The Chlamydia trachomatis insulinase/protease ctl0175 is required for efficient recovery upon IFNgamma-induced stress

PANZETTA, MARÍA E.; BASTIDAS, RJ; VALDIVIA, RH; SAKA HA

Buenos Aires

Congreso; SAIB - Reunión de Sociedades Conjuntas de Biociencias; 2017

SAIB

Chlamydia trachomatis (CT) is the most common sexually transmitted bacterial pathogen in humans. CT frequently causes asymptomatic, persistent infections leading to serious complications, particularly in young women. CT is an obligate intracellular bacterium and alternates between two developmental forms: the infectious elementary body (EB) and the replicative reticulate body (RB). In presence of stressors such as interferon-γ (IFNγ) or penicillin, CT enters a viable but non-cultivable, ?persistent? state, allowing immune evasion for long periods of time. Once the stressors are removed, CT resumes replication and continues to propagate. CT has been considered a genetically intractable pathogen until very recently, thus the bacterial factors regulating chlamydial persistence remain poorly elucidated. Aiming to identify the genetic determinants involved in chlamydial persistence, we carried out a high throughput screening using a collection of ~800 fully sequenced CT mutants obtained by chemical mutagenesis and identified 8 mutants with defects in recovery upon IFNg- and/or penicillin-induced stress. We focused our study on a mutant strain (M275) carrying 4 point mutations. M275 shows a significant decrease in progeny generation after IFNγ-induced stress compared to the Wt. We then used lateral gene transfer to map the mutation/s responsible for the persistence defect. Recombinant strains that have cleaned the mutations in bioF and ptr genes show Wt recovery linking the defect to either ptr or bioF mutations. In parallel, we disrupted ptr in CT L2 by insertional gene inactivation with an intron carrying a spectinomycin resistance cassette (aadA). The ptr::aadA strain reproduced the defect in progeny-generation after IFNg-induced stress, linking ptr to chlamydial persistence. Our results strongly suggest that CTL0175, a non-characterized protease encoded by ptr, plays a role in chlamydial persistence upon IFN-induced stress.