Response of Staphylococcus aureus physiology and Agr quorum sensing to low shear modelled microgravity

Staphylococcus aureus is commonly isolated from astronauts returning from spaceflight missions. Previous analysis of BRIC-23 omics data generated from S. aureus low Earth orbit cultures indicated significantly increased expression of the Agr quorum sensing system and its downstream targets in spaceflight samples compared to ground controls. In this current study, the rotary cell culture system (RCCS) was used to investigate the effect of low shear modelled microgravity (LSMMG) on S. aureus physiology and Agr activity. When cultured in BRIC-23 medium and temperature conditions, S. aureus LSMMG cultures exhibited decreased agr expression and altered growth compared to normal gravity control cultures, which are typically oriented with gas permeable membrane on bottom of the high aspect rotating vessel (HARV). However, when grown in an inverted gravity control orientation (membrane on top of the HARV), reduced Agr activity was observed relative to both traditional control and LSMMG cultures, signifying that oxygen availability may affect the observed differences in agr expression. Metabolite assays revealed increased lactate and decreased acetate excretion in both LSMMG and inverted control cultures. Secretomics analysis of LSMMG, control, and inverted control HARV culture supernatants corroborated these results, with inverted and LSMMG cultures exhibiting decreased abundance of Agr-regulated virulence factors and increased abundance of low-oxygen metabolic proteins. Collectively, these studies suggest that orientation of the HARV gas permeable membrane can affect S. aureus physiology and Agr quorum sensing in the RCCS, a variable that should be considered when interpreting data using this ground-based microgravity model.