High affinity iron proteins involved in catecholamine utilization contribute to B. pertussis intracellular survival

YANINA LAMBERTI; JUAN HILARIO CAFIERO; HUGO VALDEZ; JANA HOLUBOVA; VECEREK BRANISLAV; PETER SEBO; MARÍA EUGENIA RODRIGUEZ

CABA

Congreso; 11th International Bordetella Symposium; 2016

Several studies indicate that B. pertussis (Bp) may have an intracellular phase during infection which might contribute to its persistence. Recently we have shown that upon entry into macrophages, Bp survives in compartments in which it has access to transferrin and undergoes an adaptive response characterized by changes in the levels of proteins involved in stress response, cellular metabolism, and virulence. Only two proteins probably involved in iron acquisition, BfrD and BfrE, were found increased during the intracellular phase. Both receptors are implicated in iron uptake from transferrin via catecholamines in B. bronchiseptica, however, their functionality in Bp has not been demonstrated. Objective: Evaluate the relevance of BfrD and BfrE in Bp survival inside macrophages. Material and methods: The genes encoding BfrD and BfrE were deleted by homologous recombination. The adhesion, phagocytosis and intracellular survival capacity of the double mutant strain (BpΔbfrDE), a complemented strain (Bp∆bfrDEc.) and the wild type strain (Wt) inside THP-1 cells were evaluated by immunofluorescence microscopy, polymyxin B protection assay, CFU counts and FISH staining of intracellular viable bacteria at 24 and 48 hours post-infection (p.i.). Results: No differences in the adhesion or the phagocytosis between the different strains were observed. However, Bp∆bfrDE showed a significant decrease in intracellular survival already after 24 hours p.i. (p