<i>Pseudomonas aeruginosa</i> differentially influences antibiotic-resistant <i>Staphylococcus aureus</i> emergence and expansion in hyperglycemic environments

Individuals with poorly controlled diabetes mellitus often develop multispecies skin and soft tissue infections, with <i>Staphylococcus aureus</i> and <i>Pseudomonas aeruginosa</i> among the most prevalent bacteria isolated from infection sites worldwide. Diabetic infections are recalcitrant to conventional antibiotic regimens and may be a reservoir for emergent antibiotic-resistant bacterial strains. Supporting this, we have previously shown that rifampicin treatment elicits the emergence and expansion of rifampicin-resistant (Rif-r) <i>S. aureus</i> only in diabetic mice, potentially due to greater bacterial outgrowth increasing the frequency of resistance-conferring mutations. However, whether <i>S. aureus</i> exhibits altered resistance outcomes during multispecies diabetic infections is unclear. During co-infection with <i>P. aeruginosa</i> under normoglycemic conditions, <i>S. aureus</i> exhibits reduced growth and altered susceptibility to several antibiotics. In contrast, we previously observed that glucose availability allows <i>S. aureus</i> to largely overcome <i>P. aeruginosa</i>-mediated growth inhibition. Here, we explored <i>S. aureus</i> resistance outcomes under hyperglycemic conditions in the context of co-infection with <i>P. aeruginosa</i> during antibiotic challenge. We found that <i>P. aeruginosa</i> exoproducts regulated by the <i>Pseudomonas</i> quinolone signal quorum sensing system inhibit the emergence but not the expansion of Rif-r <i>S. aureus in vitro</i> under glucose-replete conditions. In contrast, we recovered equivalent Rif-r <i>S. aureus</i> burdens from diabetic mice during mono- and co-infection with <i>P. aeruginosa</i>. These results demonstrate that the diabetic infection microenvironment is conducive to emergent Rif-r <i>S. aureus</i> despite external pressures elicited by <i>P. aeruginosa</i>.IMPORTANCEPoorly controlled diabetes mellitus confers an increased susceptibility to bacterial infections, with <i>Staphylococcus aureus</i> and <i>Pseudomonas aeruginosa</i> frequently isolated from diabetic skin wounds. <i>S. aureus</i> readily develops antibiotic resistance during diabetic mono-infection under antibiotic pressure, but whether this occurs during diabetic co-infection is unclear. Under normoglycemic conditions, secreted <i>P. aeruginosa</i> factors alter <i>S. aureus</i> tolerance to several antibiotics. Here, we show that these <i>P. aeruginosa</i> exoproducts further inhibit the emergence of antibiotic-resistant <i>S. aureus</i> regardless of glucose availability <i>in vitro</i>, but this does not occur during subcutaneous co-infection in diabetic mice. These results provide initial insights regarding conditions that may inhibit <i>S. aureus</i> resistance development in hyperglycemic environments but underscore the influence of the host infection microenvironment in shaping resistance outcomes.