A Complete Lipopolysaccharide Inner Core Oligosaccharide Is Required for Resistance of Burkholderia cenocepacia to Antimicrobial Peptides and Bacterial Survival In Vivo

Abstract

Burkholderia cenocepacia is an important opportunistic pathogen of patients with cystic fibrosis. This bacterium is inherently resistant to a wide range of antimicrobial agents, including high concentrations of antimicrobial peptides. We hypothesized that the lipopolysaccharide (LPS) of B. cenocepacia is important for both virulence and resistance to antimicrobial peptides. We identified hldA and hldD genes in B. cenocepacia strain K56-2. These two genes encode enzymes involved in the modification of heptose sugars prior to their incorporation into the LPS core oligosaccharide. We constructed a mutant, SAL1, which was defective in expression of both hldA and hldD, and by performing complementation studies we confirmed that the functions encoded by both of these B. cenocepacia genes were needed for synthesis of a complete LPS core oligosaccharide. The LPS produced by SAL1 consisted of a short lipid A-core oligosaccharide and was devoid of O antigen. SAL1 was sensitive to the antimicrobial peptides polymyxin B, melittin, and human neutrophil peptide 1. In contrast, another B. cenocepacia mutant strain that produced complete lipid A-core oligosaccharide but lacked polymeric O antigen was not sensitive to polymyxin B or melittin. As determined by the rat agar bead model of lung infection, the SAL1 mutant had a survival defect in vivo since it could not be recovered from the lungs of infected rats 14 days postinfection. Together, these data show that the B. cenocepacia LPS inner core oligosaccharide is needed for in vitro resistance to three structurally unrelated antimicrobial peptides and for in vivo survival in a rat model of chronic lung infection.

Phase variation has a role in Burkholderia ambifaria niche adaptation

“…In the Bcc, the complete lipid A-core oligosaccharide is required for resisting polymyxin B and melittin (Loutet et al, 2006). The modification of the LPS profile that we observed between the wild-type and the variant might explain their difference in sensitivity to antimicrobial peptides.…”

“…Bcc isolates are resistant to a wide variety of antimicrobial compounds, including antimicrobial peptides (Mahenthiralingam et al, 2005;Loutet et al, 2006). In the Bcc, the complete lipid A-core oligosaccharide is required for resisting polymyxin B and melittin (Loutet et al, 2006).…”

The suhB gene of Burkholderia cenocepacia is required for protein secretion, biofilm formation, motility and polymyxin B resistance

“…Therefore, we also investigated whether the mutant has a defect in membrane permeability by assessing resistance to the cationic antimicrobial peptide polymyxin B (Loutet et al, 2006(Loutet et al, , 2009. Fig.…”

“…Antibiotic sensitivity assays. Antibiotic sensitivity assays were performed as previously described (Loutet et al, 2006). Cultures were incubated at 37 uC for 24 h in the Bioscreen C system, and the final density was determined by determining the OD 600 .…”

Burkholderia cenocepacia zinc metalloproteases influence resistance to antimicrobial peptides

“…are highly resistant to antimicrobial cationic peptides that typically kill bacteria by membrane disruption. LPS structure has been shown to play an important role in resistance to polymyxins and other peptides in both B. cenocepacia (Loutet et al, 2006) and Burkholderia pseudomallei (Burtnick & Woods, 1999). Other mechanisms of bacterial resistance to antimicrobial peptides have been described, including alteration of surface charges, changes in membrane proteins, efflux pumps/transporters and proteolytic enzymes (Brogden, 2005;Hiemstra et al, 2004;Potempa et al, 2009;Yount & Yeaman, 2005).…”