Porin A and α/β-hydrolase are necessary and sufficient for hemolysis induced by Bartonella bacilliformis.

Abstract

Carrion's disease is endemic to the South American Andes and is characterized by fatal hemolytic anemia. This neglected tropical disease is caused by Bartonella bacilliformis, a fastidious and slow-growing pathogen difficult in genetic manipulation. In this study, we determine that porin A and α/β-hydrolase are both necessary and sufficient for hemolysis induced by B. bacilliformis. These genes are identified through a screen of a Tn5 transposon mutant library. Using markerless deletion mutagenesis, porin A and α/β-hydrolase deletion mutants are generated and functionally analyzed by hemolysis assays. In silico analyses predict conserved biological functions and three-dimensional structures of the identified proteins, with the α/β-hydrolase showing structural similarity to known lipases. Site-directed mutagenesis of the α/β-hydrolase active site demonstrates that the catalytic triad (Ser205, Asp267, His310) is essential for its hemolytic function. Screening of a phospholipase inhibitor library comprising 27 bioactive compounds identifies compound 48/80 as a potent inhibitor of hemolysis, with activity in the micromolar range. Unraveling the molecular mechanisms underlying Carrion's disease may facilitate the future development of anti-virulence therapies, a promising strategy particularly in the context of increasing antibiotic resistance of B. bacilliformis.