Persistence and Microevolution of Pseudomonas aeruginosa in the Cystic Fibrosis Lung: A Single-Patient Longitudinal Genomic Study

Abstract

Background: During its persistence in cystic fibrosis (CF) airways, P. aeruginosa develops a series of phenotypic changes by the accumulation of pathoadaptive mutations. A better understanding of the role of these mutations in the adaptive process, with particular reference to the development of multidrug resistance (MDR), is essential for future development of novel therapeutic approaches, including the identification of new drug targets and the implementation of more efficient antibiotic therapy. Although several whole-genome sequencing studies on P. aeruginosa CF lineages have been published, the evolutionary trajectories in relation to the development of antimicrobial resistance remain mostly unexplored to date. In this study, we monitored the adaptive changes of P. aeruginosa during its microevolution in the CF airways to provide an innovative, genome-wide picture of mutations and persistent phenotypes and to point out potential novel mechanisms allowing survival in CF patients under antibiotic therapy.Results: We obtained whole genome sequences of 40 P. aeruginosa clinical CF strains isolated at Trentino Regional Support CF Centre (Rovereto, Italy) from a single CF patient over an 8-year period (2007–2014). Genotypic analysis of the P. aeruginosa isolates revealed a clonal population dominated by the Sequence Type 390 and three closely related variants, indicating that all members of the population likely belong to the same clonal lineage and evolved from a common ancestor. While the majority of early isolates were susceptible to most antibiotics tested, over time resistant phenotypes increased in the persistent population. Genomic analyses of the population indicated a correlation between the evolution of antibiotic resistance profiles and phylogenetic relationships, and a number of putative pathoadaptive variations were identified.Conclusion: This study provides valuable insights into the within-host adaptation and microevolution of P. aeruginosa in the CF lung and revealed the emergence of an MDR phenotype over time, which could not be comprehensively explained by the variations found in known resistance genes. Further investigations on uncharacterized variations disclosed in this study should help to increase our understanding of the development of MDR phenotype and the poor outcome of antibiotic therapies in many CF patients.

Genome-wide association study of Pseudomonas aeruginosa adaptations to the cystic fibrosis lung

“…There have been several longitudinal studies examining changes in the bacterium over the course of a patient's lifespan and infection [17,18,19,182]. Many of these changes during chronic infection include resistance to antibiotics, changes in expression of virulence factors, increases in mutation rate, formation of biofilms and changes in metabolism [14,15].…”

“…PA has been widely studied in the context of CF. There are longitudinal studies that will follow one or a few patients and their infections over time [16,17,18,19], studies of epidemic strains [20,21] and samples from CF clinics [11,22,23]. This has helped characterize the evolution and colonization of PA in a CF lung environment.…”

“…Strains often become mucoid during the early stages of a chronic infection, but may lose this mucoidy in late infections [7,17,24]. They will form biofilms, develop resistance to antibiotics, adapt metabolic pathways, change in virulence, and have an increased mutation rate [6].…”

“…The bacteria become resistant over time, especially in the case of chronic infections. Longitudinal studies have repeatedly observed the evolution of resistance by sampling isolates from a single patient over the course of several years [16,17,19]. There are a variety of mechanisms that confer antimicrobial resistance (AMR), and these mutations within CF PA isolates are related to the treatment an individual has received over the course of their infection.…”

“…The leaves highlighted in green are from a study of the Liverpool epidemic strain. The other highlighted leaves, in orange and pink, were from a longitudinal studies of PA samples from single patients, one in Toronto (orange) [16], the other in Italy (pink) [17]. This phylogeny was subsequently used to ensure that positive associations were not due to shared…”