Within-host evolution of Burkholderia pseudomallei during chronic infection

Abstract

Burkholderia pseudomallei, the causative agent of melioidosis, is an opportunistic environmental bacterium found in tropical areas. Melioidosis has a wide spectrum of clinical presentations but most common is pneumonia. People with certain risk factors such as cystic fibrosis (CF) are more likely to contract the disease upon exposure. B. pseudomallei generally causes acute disease but in some instances give rise to chronic infection. People with CF are more prone to develop chronic infection and the molecular mechanisms behind this chronic presentation are not well understood. This thesis aim was to investigate B. pseudomallei evolution within CF patients during chronic infection using genomics and transcriptomics. First, sequential isolates from one of the only two known patients with chronic infection without CF were whole genome sequenced (WGS). Results from this analysis showed no within-host evolution of the isolates over time but did identify a rare polyclonal infection of two B. pseudomallei isolates of the same sequence type. Next, paired isolates from seven chronically infected CF patients were WGS and we found that mutations accumulated over time in these isolates. We identified traits common among chronic CF pathogens such as multi-antibiotic resistance, reductive evolution, metabolic adaptation, and virulence factor attenuation. Also, the first B. pseudomallei isolate with a hypermutator phenotype was identified. Additionally, two new potential resistance mechanisms towards two of the five clinically relevant antibiotics for treatment of melioidosis were identified. Transcriptomic analysis of five of the seven isolate pairs grown in artificial sputum media to mimic the C Flung conditions showed differential expression of genes in four of the isolate pairs, and confirmed the role of key genetic mutations in altering gene expression of certain pathways. Combined these studies have created a first glimpse into the pathogenesis of chronic B. pseudomallei infections in CF, and my results will provide potential new avenues for targeted therapeutic management for CF patients infected with this recalcitrant pathogen.