The involvement of the low-oxygen-activated locus of Burkholderia cenocepacia in adaptation during cystic fibrosis infection

Abstract

Chronic infection with opportunistic pathogens including Burkholderia cepacia complex (Bcc) is a hallmark of cystic fibrosis (CF). We investigated the adaptive mechanisms facilitating chronic lung infection in sequential Bcc isolates from two siblings with CF (P1 and P2), one of whom also experienced intermittent blood-stream infections (P2). We previously showed increased lung cell attachment with colonisation time in both P1 and P2. WGS analysis confirmed that the isolates are closely related. Twelve genes showed three or more mutations, suggesting these were genes under selection. Single nucleotide polymorphisms (SNVs) in 45 regulatory genes were also observed. Proteomic analysis showed that the abundance of 149 proteins increased over 61-months in sputum isolates, and both time- and source-related alterations in protein abundance between the second patient’s isolates. A consistent time-dependent increase in abundance of 19 proteins encoded by a low-oxygen-activated (lxa) locus was observed in both sets of isolates. Attachment was dramatically reduced in a B. cenocepacia K56-2Δlxa-locus deletion mutant, further indicating that it encodes protein(s) involved in host-cell attachment. Time-related changes in virulence in Galleria mellonella or motility were not observed. We conclude that the lxa-locus, associated with anoxic persistence in vitro, plays a role in host-cell attachment and adaptation to chronic colonization in the hypoxic niche of the CF lung.

A Universal Stress Protein upregulated by hypoxia may contribute to chronic lung colonisation and intramacrophage survival in cystic fibrosis

“…These late chronic infection isolates also showed increased attachment to CF lung epithelial cells [12]. Importantly, all six USPs encoded on the lxa locus showed increased abundance with time of colonisation [11]. Among these lxaencoded usp genes, BCAM0276, encodes a UspA family stress protein which consistently showed increased abundance in the later isolates from two chronically colonised patients and was associated with increased gene expression [11].…”

“…Having characterised USP76, we wanted to examine whether other USPs expressed within the locus share the same functions. We chose a uspA gene downstream of BCAM0276 as it was upregulated by 201-fold in response to low oxygen and showed greatest increase in protein abundance (up to 16 fold) in late stage isolates [10,11]. A targeted single deletion mutant was generated for the BCAM0292 gene; however, despite many attempts we were unable to complement the strain with the BCAM0292 gene.…”

“…Sass et al (2013) identified a 50 gene locus which was dramatically upregulated under low oxygen conditions and designated the low oxygen activated (lxa) locus [10]. We subsequently showed that 19 proteins encoded on the lxa locus showed increased abundance in late infection isolates from chronically colonised CF patients [11]. These late chronic infection isolates also showed increased attachment to CF lung epithelial cells [12].…”

“…Importantly, all six USPs encoded on the lxa locus showed increased abundance with time of colonisation [11]. Among these lxaencoded usp genes, BCAM0276, encodes a UspA family stress protein which consistently showed increased abundance in the later isolates from two chronically colonised patients and was associated with increased gene expression [11]. Previously this USP (USP76) was reported to be upregulated almost 60-fold in B. cenocepacia strain J2315 response to low oxygen [10] and up to 40-fold in a comparative transcriptomic study of B. contaminans isolates from CF patients [13].…”

Parasite Survival and Disease Persistence in Cystic Fibrosis, Schistosomiasis and Pathogenic Bacterial Diseases: A Role for Universal Stress Proteins?

“…Viability loss is also observed in oxygen-free environments, indicating the importance of the gene in B. cenocepacia during oxygen depletion. A study conducted by Cullen and co-workers found that among the 19 proteins that play a potential role in electron transfer, metabolism and regulation encoded by the lxa gene elevated during chronic infection, six are USPs and a heat shock protein [ 57 ]. All the proteins exhibited an increase (of up to 40-fold) in production during chronic lung infection.…”