Department of Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, Australia
David R Thomas
Department of Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, Australia; Infection and Immunity Program, Department of Microbiology and Biomedicine Discovery Institute, Monash University, Clayton, Australia
Department of Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, Australia; Infection and Immunity Program, Department of Microbiology and Biomedicine Discovery Institute, Monash University, Clayton, Australia
Sacha Pidot
Department of Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, Australia
Ruth C Massey
School of Microbiology, University College Cork, Cork, Ireland; School of Medicine, University College Cork, Cork, Ireland; APC Microbiome Ireland, University College Cork, Cork, Ireland; School of Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom
Department of Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, Australia; Microbiological Diagnostic Unit Public Health Laboratory, Department of Microbiology and Immunology, Doherty Institute, University of Melbourne, Melbourne, Australia
Staphylococcus aureus infections are associated with high mortality rates. Often considered an extracellular pathogen, S. aureus can persist and replicate within host cells, evading immune responses, and causing host cell death. Classical methods for assessing S. aureus cytotoxicity are limited by testing culture supernatants and endpoint measurements that do not capture the phenotypic diversity of intracellular bacteria. Using a well-established epithelial cell line model, we have developed a platform called InToxSa (intracellular toxicity of S. aureus) to quantify intracellular cytotoxic S. aureus phenotypes. Studying a panel of 387 S. aureus bacteraemia isolates, and combined with comparative, statistical, and functional genomics, our platform identified mutations in S. aureus clinical isolates that reduced bacterial cytotoxicity and promoted intracellular persistence. In addition to numerous convergent mutations in the Agr quorum sensing system, our approach detected mutations in other loci that also impacted cytotoxicity and intracellular persistence. We discovered that clinical mutations in ausA, encoding the aureusimine non-ribosomal peptide synthetase, reduced S. aureus cytotoxicity, and increased intracellular persistence. InToxSa is a versatile, high-throughput cell-based phenomics platform and we showcase its utility by identifying clinically relevant S. aureus pathoadaptive mutations that promote intracellular residency.
