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要旨:
Introduction: Acinetobacter baumannii is a Gram-negative pathogen that causes a multitude of nosocomial infections. The Acinetobacter trimeric autotransporter adhesin (Ata) belongs to the superfamily of trimeric autotransporter adhesins which are important virulence factors in many Gram-negative species.
Objective: We evaluated the impact of Ata in host-pathogen interaction by analysing adhesion, invasion, host-cell-modulation and apoptosis in human endothelial and epithelial cells.
Materials and Methods: Adhesion of A. baumannii to human host cells was investigated using primary endothelial and epithelial cells under static and dynamic flow conditions. Furthermore, an ex vivo organ infection model employing human umbilical cord veins was employed. Host cell invasion was analysed by a gentamicin protection assay. Induction of apoptosis was determined using propidium iodide/annexin V staining with subsequent FACS analysis and the Caspase-Glo assay (Promega). Chemokine secretion upon infection was analysed using Multi-Analyte ELISArray Kits (Qiagen). Pathogenicity in vivo was evaluated using the Galleria mellonella infection model.
Result: Phylogenetic profiling using 3,052 Acinetobacter spp. genomes revealed that ata is present in 78% of all A. baumannii isolates but only in 2% of the closely related but less pathogenic species A. calcoaceticus and A. pittii. Employing a markerless ata deletion mutant of A. baumannii ATCC 19606 and complemented controls, we could show that adhesion to and invasion into human endothelial and epithelial cells under static conditions both clearly depend on Ata. This adhesion defect was even more evident when dynamic infection conditions were applied or when human umbilical cord veins were infected ex vivo with A. baumannii. Infection of primary human umbilical cord vein endothelial cells (HUVECs) with A. baumannii led to the secretion of the pro-inflammatory cytokines interleukin (IL)-6 and IL-8 in a time- and Ata-dependent manner. Ata induced apoptosis, but not necrosis, of HUVECs after 16 h post infection by activation of caspase-3 and caspase-7. Ata deletion mutants were furthermore attenuated in their ability to kill larvae of G. mellonella and to survive in larvae when injected at sublethal doses.
Conclusion: These results strongly suggest that Ata is an important multifunctional virulence factor of A. baumannii that triggers multiple important steps for the initiation of successful infections in different host cells.
