Genome evolution drives transcriptomic and phenotypic adaptation in during 20 years of infection.

The opportunistic pathogen chronically infects the lungs of patients with cystic fibrosis (CF). During infection the bacteria evolve and adapt to the lung environment. Here we use genomic, transcriptomic and phenotypic approaches to compare multiple isolates of collected more than 20 years apart during a chronic infection in a CF patient. Complete genome sequencing of the isolates, using short- and long-read technologies, showed that a genetic bottleneck occurred during infection and was followed by diversification of the bacteria. A 125 kb deletion, an 0.9 Mb inversion and hundreds of smaller mutations occurred during evolution of the bacteria in the lung, with an average rate of 17 mutations per year. Many of the mutated genes are associated with infection or antibiotic resistance. RNA sequencing was used to compare the transcriptomes of an earlier and a later isolate. Substantial reprogramming of the transcriptional network had occurred, affecting multiple genes that contribute to continuing infection. Changes included greatly reduced expression of flagellar machinery and increased expression of genes for nutrient acquisition and biofilm formation, as well as altered expression of a large number of genes of unknown function. Phenotypic studies showed that most later isolates had increased cell adherence and antibiotic resistance, reduced motility, and reduced production of pyoverdine (an iron-scavenging siderophore), consistent with genomic and transcriptomic data. The approach of integrating genomic, transcriptomic and phenotypic analyses reveals, and helps to explain, the plethora of changes that undergoes to enable it to adapt to the environment of the CF lung during a chronic infection.