Investigating how different bacterial species interact with one another remains a critical question for understanding clinical pathologies, which often present as co-infections.  Enterococcus faecalis is an opportunistic pathogen responsible for a range of clinical manifestations including urinary tract infections (UTI’s), endocarditis, and bacteremia, and accounts for 65-80% of nosocomial infections caused by Enterococcus species.  Pseudomonas aeruginosa is often co-isolated with E. faecalis in clinical infections and studies have shown that the presence of these pathogens together can affect their growth, pathogenicity, and likely clinical outcomes.  Understanding these dynamic multispecies interactions is therefore critical to developing therapeutics, which is especially relevant given the rising antibiotic resistance in both of these bacterial species.  We are using multiple approaches to characterize these interactions, guided by co-infection in vivo studies using an E. faecalis transposon insertional mutagenesis library (Tn-Seq).