Bile modulates phage–host interactions in multidrug-resistant Pseudomonas aeruginosa

Biliary tract infections (BTIs) arise within a bile-rich environment that profoundly shapes microbial ecology and pathogen adaptation. Pseudomonas aeruginosa, a major opportunistic pathogen in nosocomial settings, exhibits remarkable physiological plasticity, that enable persistence in such challenging niches. However, the influence of bile on P. aeruginosa's adaptive responses and phage-host interactions remains largely unexplored. Here, we demonstrate that ox-bile imposes concentration-dependent stress on P. aeruginosa strain ZS-PA-35, indicative of host-derived selective pressure. Notably, ox-bile enhances biofilm formation and promotes swarming and twitching motilities while concurrently suppressing swimming motility. Moreover, ox-bile modulates phage susceptibility, likely through altered receptor expression: exposure to ox-bile sensitizes P. aeruginosa to the type IV pili (T4P)-dependent phage phipa2, whereas susceptibility to the lipopolysaccharide (LPS)-targeting phage phipa10 remains unchanged. Genome-wide mutagenesis identified resistance-conferring mutations affecting T4P structures, LPS biosynthesis, and associated regulatory pathways. Among these, phage-resistant mutants ΔpilT and ΔgalU retained high fitness under ox-bile stress, accompanied by enhanced swarming and swimming motilities. Furthermore, in a lysogenic context, ox-bile markedly suppressed prophage accumulation in the T4P-dependent strain ZS-PA-05. These findings reveal that bile acts as a critical environmental cue shaping both adaptive physiology and phage susceptibility in P. aeruginosa, with broad implications for microbiome dynamics and the development of phage-based therapies targeting bile-impacted infections.