Long-Term Evolution ofBurkholderia multivoransduring a Chronic Cystic Fibrosis Infection Reveals Shifting Forces of Selection.
Burkholderia multivoransis an opportunistic pathogen capable of causing severe disease in patients with cystic fibrosis (CF). Patients may be chronically infected for years, during which the bacterial population evolves in response to unknown forces. Here we analyze the genomic and functional evolution of aB. multivoransinfection that was sequentially sampled from a CF patient over 20 years. The population diversified into at least four primary, coexisting clades with distinct evolutionary dynamics. The average substitution rate was only 2.4 mutations/year, but notably, some lineages evolved more slowly, whereas one diversified more rapidly by mostly nonsynonymous mutations. Ten loci, mostly involved in gene expression regulation and lipid metabolism, acquired three or more independent mutations and define likely targets of selection. Further, a broad range of phenotypes changed in association with the evolved mutations; they included antimicrobial resistance, biofilm regulation, and the presentation of lipopolysaccharide O-antigen repeats, which was directly caused by evolved mutations. Additionally, early isolates acquired mutations in genes involved in cyclic di-GMP (c-di-GMP) metabolism that associated with increased c-di-GMP intracellular levels. Accordingly, these isolates showed lower motility and increased biofilm formation and adhesion to CFBE41o-epithelial cells than the initial isolate, and each of these phenotypes is an important trait for bacterial persistence. The timing of the emergence of this clade of more adherent genotypes correlated with the period of greatest decline in the patient's lung function. All together, our observations suggest that selection onB. multivoranspopulations during long-term colonization of CF patient lungs either directly or indirectly targets adherence, metabolism, and changes in the cell envelope related to adaptation to the biofilm lifestyle.IMPORTANCEBacteria may become genetically and phenotypically diverse during long-term colonization of cystic fibrosis (CF) patient lungs, yet our understanding of within-host evolutionary processes during these infections is lacking. Here we combined current genome sequencing technologies and detailed phenotypic profiling of the opportunistic pathogenBurkholderia multivoransusing sequential isolates sampled from a CF patient over 20 years. The evolutionary history of these isolates highlighted bacterial genes and pathways that were likely subject to strong selection within the host and were associated with altered phenotypes, such as biofilm production, motility, and antimicrobial resistance. Importantly, multiple lineages coexisted for years or even decades within the infection, and the period of diversification within the dominant lineage was associated with deterioration of the patient's lung function. Identifying traits under strong selection during chronic infection not only sheds new light ontoBurkholderiaevolution but also sets the stage for tailored therapeutics targeting the prevailing lineages associated with disease progression.
Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Lisbon, Portugal.
Inês N
IN
Silva
Centre of Molecular and Environmental Biology, Department of Biology, Universidade do Minho, Braga, Portugal.
Pedro M
0000-0003-3234-4265
PM
Santos
Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Lisbon, Portugal.
Mário R
MR
Santos
Department of Pediatrics, Faculty of Medicine, Centre for Understanding and Preventing Infection in Children, University of British Columbia, Vancouver, Canada.
James E A
JE
Zlosnik
Department of Pediatrics, Faculty of Medicine, Centre for Understanding and Preventing Infection in Children, University of British Columbia, Vancouver, Canada.
David P
DP
Speert
Department of Molecular, Cellular and Biomedical Sciences, University of New Hampshire, Durham, New Hampshire, USA.
Sean W
SW
Buskirk
Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan, USA.
Eric L
EL
Bruger
Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan, USA.
Christopher M
CM
Waters
Department of Microbiology and Molecular Genetics, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
Vaughn S
0000-0001-7726-0765
VS
Cooper
Institute for Bioengineering and Biosciences, Instituto Superior Técnico, Universidade de Lisboa, Lisbon, Portugal; Department of Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Lisbon, Portugal.
Leonilde M
0000-0002-6838-4245
LM
Moreira
Burkholderia multivorans; biofilm; c-di-GMP signaling; chronic infection; cystic fibrosis; evolution; lung function decline; respiratory infection; within-host adaptation