Time Series Genomics of Pseudomonas aeruginosa Reveals the Emergence of a Hypermutator Phenotype and Within-Host Evolution in Clinical Inpatients.

Pseudomonas aeruginosa, a common opportunistic pathogen, is one of the leading etiological agents of nosocomial infections. Many previous studies have reported the nosocomial transmission and epidemiology of P. aeruginosa infections. However, longitudinal studies regarding the dynamics of P. aeruginosa colonization and infection in health care settings are limited. We obtained longitudinal samples from aged patients with prolonged intensive care unit (ICU) stays (~4 to 19 months). P. aeruginosa was isolated from 71 samples obtained from seven patients and characterized by whole-genome sequencing. The P. aeruginosa isolates were assigned to 10 clonal complexes, and turnover of main clones was observed in sequential sputum samples from two patients. By comparing intraclonal genomic diversities, we identified two clones that had significantly higher numbers of single nucleotide polymorphisms and variations in homopolymeric sequences than the other clones, indicating a hypermutator phenotype. These hypermutator clones were associated with mutations T147I/G521S and P27L in the MutL protein, and their mutation rates were estimated to be 3.20 × 10 and 6.59 × 10 per year per nucleotide, respectively. We also identified 24 recurrently mutated genes that exhibited intraclonal diversity in two or more clones. Notably, one recurrent mutation, S698F in FptA, was observed in four clones. These findings suggest that convergent microevolution and adaption of P. aeruginosa occur in long-term ICU patients. Pseudomonas aeruginosa is a predominant opportunistic pathogen that causes nosocomial infections. Inappropriate empirical therapy can lead to prolonged hospital stays and increased mortality. In our study of sequential P. aeruginosa isolates from inpatients, high intrahost diversity was observed, including switching of clones and the emergence of a hypermutator phenotype. Recurrently mutated genes also suggested that convergent microevolution and adaption of P. aeruginosa occur in inpatients, and genomic diversity is associated with differences in multiple-drug-resistance profiles. Taken together, our findings highlight the importance of longitudinal surveillance of nosocomial P. aeruginosa clones.