A genome-wide association analysis reveals a potential role for recombination in the evolution of antimicrobial resistance in Burkholderia multivorans

Abstract

Cystic fibrosis (CF) lung infections caused by members of the Burkholderia cepacia complex, such as Burkholderia multivorans, are associated with high rates of mortality and morbidity. We performed a population genomics study of 111 B. multivorans sputum isolates from one CF patient through three stages of infection including an early incident isolate, deep sampling of a one-year period of chronic infection occurring weeks before a lung transplant, and deep sampling of a post-transplant infection. We reconstructed the evolutionary history of the population and used a lineage-controlled genome-wide association study (GWAS) approach to identify genetic variants associated with antibiotic resistance. We found the incident isolate was basally related to the rest of the strains and more susceptible to antibiotics from three classes (β-lactams, aminoglycosides, quinolones). The chronic infection isolates diversified into multiple, distinct genetic lineages and showed reduced antimicrobial susceptibility to the same antibiotics. The post-transplant reinfection isolates derived from the same source as the incident isolate and were genetically distinct from the chronic isolates. They also had a level of susceptibility in between that of the incident and chronic isolates. We identified numerous examples of potential parallel pathoadaptation, in which multiple mutations were found in the same locus or even codon. The set of parallel pathoadaptive loci was enriched for functions associated with virulence and resistance. Our GWAS analysis identified statistical associations between a polymorphism in the ampD locus with resistance to β-lactams, and polymorphisms in an araC transcriptional regulator and an outer membrane porin with resistance to both aminoglycosides and quinolones. Additionally, these three loci were independently mutated four, three and two times, respectively, providing further support for parallel pathoadaptation. Finally, we identified a minimum of 14 recombination events, and observed that loci carrying putative parallel pathoadaptations and polymorphisms statistically associated with β-lactam resistance were over-represented in these recombinogenic regions.

Comparative Evolutionary Patterns of Burkholderia cenocepacia and B. multivorans During Chronic Co-infection of a Cystic Fibrosis Patient Lung

“…This publication also reported the complete loss of chromosome III resulting in genome-size reduction as an adaptive trait of B. cenocepacia (Lee et al, 2017), consistent with a previously reported rare in vivo loss of the same mega-plasmid among B. cenocepacia clonal variants emerging during chronic infection (Moreira et al, 2017). Recently, a population genomic study focused on 111 B. multivorans isolates obtained from a Canadian CF patient showed potential parallel pathoadaptation involving genes associated with resistance toward multiple classes of antibiotics (Diaz Caballero et al, 2018).…”

“…Over the last decade, several virulence factors and adaptive traits have been identified in Bcc species (Mahenthiralingam et al, 2005;Drevinek and Mahenthiralingam, 2010;Loutet and Valvano, 2010;Zlosnik and Speert, 2010;Madeira et al, 2011Madeira et al, , 2013Mira et al, 2011;Zlosnik et al, 2011Zlosnik et al, , 2014Sousa et al, 2017;Hassan et al, 2019a). Several studies have used genome-wide methods to identify genetic changes during long term infection by B. dolosa (Lieberman et al, 2011(Lieberman et al, , 2014, B. multivorans (Silva et al, 2016;Diaz Caballero et al, 2018) and B. cenocepacia (Lee et al, 2017;Nunvar et al, 2017) within individual hosts. For example, a retrospective study involving 112 sequential B. dolosa isolates from 14 CF patients, identified several mutated genes involved in genetic variation within individual patients, in particular genes required for expression of surface polysaccharides, lipopolysaccharide O-antigen (OAg) biosynthesis, outer membrane components, iron scavenging, and antibiotic resistance (Lieberman et al, 2011).…”

Epidemiology of E. coli in Cystic Fibrosis Airways Demonstrates the Capacity for Persistent Infection but Not Patient-Patient Transmission

“…The primary limitation of our study -derived from the retrospective cohort analysis of a biobank -is that we only sampled one isolate per morphologically distinct E. coli colony per patient per time point, capturing limited intra-patient diversity. While it has traditionally been assumed that patients are infected with a single strain at a time, recent evidence from studies of other CF pathogens indicates that multi-strain infections can occur (Lee et al, 2017;O'Brien et al, 2017;Clark et al, 2018;Diaz Caballero et al, 2018;Azarian et al, 2019). For this to be true, however, these simultaneously independently infecting strains would have to have identical morphologic appearance on MacConkey agar, an unlikely scenario.…”

“…The identification of genes with multiple independent mutations (i.e. multi-mutated loci), as well as a higher ratio of nonsynonymous to synonymous mutations relative to genes with only single mutations, has recently been used to identify candidate pathoadaptive genes in P. aeruginosa (Caballero et al, 2015) and Burkholderia multivorans (Diaz Caballero et al, 2018) infecting individuals with CF. Thus, we applied a similar approach to identify genes potentially involved in the adaptation of E. coli to the CF lung environment.…”

Rapid evolution drives the rise and fall of carbapenem resistance during an acute Pseudomonas aeruginosa infection

“…Although this link between antibiotic treatment and resistance is straightforward and intuitive, the dynamics of the rise and fall of antibiotic resistance during infections remain poorly understood. Progress in this area has largely come from studies that have used longitudinal sampling of patients to study the emergence of resistance during long-term chronic infections associated with diseases such as cystic fibrosis and tuberculosis [7][8][9][10][11][12][13]. Although these studies have produced important insights into resistance, short-term acute infections, such as hospital-acquired infections by opportunistic and commensal pathogens, represent a very important burden of AMR [14].…”