P. aeruginosa (PA) causes chronic airway infections (CAI) in cystic fibrosis (CF) patients, established from a single ancestral and persistent clone, which typically suffers phenotypic diversification. CAI are characterized by a high prevalence of mutators, mainly driven by a defective mismatch repair system established by a single bacterial lineage which persists throughout the life-time of the patients by using diverse adaptive mechanisms. Hypermutability, due to deficiency in (MRS) is highly selected in CAI and it has been proposed as an adaptive strategy used by PA for persisting, but its role in CAI remains to be completely solved. This work is aimed to study the impact of hypermutability on the PA genome structure through many generations of persistence in CF patients. Cross-sectional and longitudinal analysis were performed by obtaining collections of isolates from 2 CF patients (CFA and CFD) infected each other by a single PA clone. CFA collection was composed by 2 mutator isolates obtained in 2004 and 2007 and a population of 90 isolates obtained from a single sputum in 2010. CFD collection was composed by a nonmutator strain in 1991, 2 mutator strains in 1995 and 2002, and a population of 90 isolates from a single sputum in 2011. Whole genomic sequencing of each earliest isolate belonging from both patients, as well as latest 11 strains from each CFA 2010 and CFD 2011 collection was performed. CFA and CFD collections showed a high selection, prevalence and dominance of mutator strains. A coexistence of different mutator lineages during the CAI with persistence and dominance of only one of them was observed. Mutations in global regulator coding genes were present early during the infection and once occurred they were fixed in the population and persisted throughout the CAI. However a negative selection for most of the generated mutations was observed.
