Supplementary Figures and Tables from The rapid <i>in vivo</i> evolution of <i>Pseudomonas aeruginosa</i> in ventilator-associated pneumonia patients leads to attenuated virulence

<i>Pseudomonas aeruginosa</i> is an opportunistic pathogen that causes severe airway infections in humans. These infections are usually difficult to treat and associated with high mortality rate. While colonizing the human airways, <i>P. aeruginosa</i> could accumulate genetic mutations that often lead to its better adaptability to the host environment. Understanding these evolutionary traits may provide important clues for the development of effective therapies to treat <i>P. aeruginosa</i> infections. In this study, 25 <i>P. aeruginosa</i> isolates were longitudinally sampled from the airways of four ventilator-associated pneumonia (VAP) patients. Pacbio and Illumina sequencing were used to analyse the <i>in vivo</i> evolutionary trajectories of these isolates. Our analysis showed that positive selection dominantly shaped <i>P. aeruginosa</i> genomes during VAP infections and led to three convergent evolution events, including loss-of-function mutations of <i>lasR</i> and <i>mpl</i>, and a pyoverdine-deficient phenotype. Specifically, <i>lasR</i> encodes one of the major transcriptional regulators in quorum sensing, whereas <i>mpl</i> encodes an enzyme responsible for recycling cell wall peptidoglycan. We also found that <i>P. aeruginosa</i> isolated at late stages of VAP infections produce less elastase and are less virulent <i>in vivo</i> than their earlier isolated counterparts, suggesting the short-term <i>in vivo</i> evolution of <i>P. aeruginosa</i> leads to attenuated virulence.