More Is Not Always Better: Co-Occurrence Analysis of Anti-Phage Systems Reveals Prevalent Neutral Interactions in P. aeruginosa

<i>Pseudomonas aeruginosa</i> is a major nosocomial pathogen and a key model for studying phage-bacteria interactions. While a multitude of anti-phage defense systems have been discovered in this species, the functional relationships between them remain poorly characterized. Here, we sequenced 53 clinical <i>P. aeruginosa</i> isolates and uncovered an arsenal of diverse defense systems. Surprisingly, phage infection and adsorption assays revealed that the quantity of defense systems in a given strain did not correlate with its innate phage resistance. Subsequent bioinformatic analysis identified statistically significant positive and negative associations between specific defense systems. Experimental validation of selected pairs demonstrated that certain pairs conferred a spectrum of interaction outcomes, including additive, synergistic, and neutral effects, clearly demonstrating that the outcome cannot be predicted by a simple additive model. Our findings establish that functional relationships between defense systems, rather than their mere abundance, is a critical determinant of the anti-phage defense landscape in clinical <i>P. aeruginosa</i>, with implications for bacterial evolution and phage therapy.