2601. Identification of Staphylococcus aureus Genetic Factors Associatiated with the Subversion of Macrophage Phagosomal Acidification

Abstract

Background S. aureus is a major medical pathogen contributing to healthcare-associated costs and mortality. Metastatic S. aureus infection is commonly associated with skin and soft-tissue infections. Macrophages as resident tissue phagocytes are essential for bacterial clearance and effectively phagocytose and kill S. aureus at low inocula. Formation of an intracellular reservoir develops when the capacity for macrophages to eradicate S. aureus is overwhelmed at increased inocula. Phagosomal maturation after bacterial ingestion involves sequential fusion with endosomes and lysosomes, reducing luminal pH, facilitating bacterial degradation. In the context of intracellular S. aureus, incomplete phagosomal maturation is demonstrable with impaired acidification and failure of lysosome fusion. Inhibition of phagosomal maturation is dependent upon bacterial factors as it is reversed in heat-killed bacteria. Intracellular survival of S. aureus is associated with reduced antimicrobial effectivity and increased complications. Within the intracellular environment, persistence of S. aureus is associated with adaptation of gene expression which confers greater resistance to antimicrobial effector mechanisms.MethodsAn ordered mutant library of S. aureus provides the opportunity to give a comprehensive evaluation of gene function. The Nebraska transposon (Tn) mutant library contains 1952 sequence-defined Tn insertion mutants derivative of USA300 LAC S. aureus, each with a single non-essential gene deletion. The mutants were labeled with a fluorescent stain activated at pH < 6 and challenged against differentiated human monocyte-derived macrophages for 4 hours. A high-content microscopy screen was developed to identify the bacterial genes associated with impairment of phagosomal acidification.ResultsThe results of the high-throughput screen indicate the global regulators agr and saeR, hemolysin A and catalase are associated with the inhibition of phagosomal acidification.ConclusionThe burden of S. aureus bacteremia and metastatic disease makes the targeting of intracellular S. aureus essential. Identification of bacterial factors associated with impaired phagosomal acidification and maturation offers targets to limit S. aureus infections.Disclosures All authors: No reported disclosures.