Characterization of the Poly-β-1,6- N -Acetylglucosamine Polysaccharide Component of Burkholderia Biofilms

We demonstrated the production of poly-β-1,6-N -acetylglucosamine (PNAG) polysaccharide in the biofilms ofBurkholderia multivorans ,Burkholderia vietnamiensis ,Burkholderia ambifaria ,Burkholderia cepacia , andBurkholderia cenocepacia using an immunoblot assay for PNAG. These results were confirmed by further studies, which showed that the PNAG hydrolase, dispersin B, eliminated immunoreactivity of extracts from the species that were tested (B. cenocepacia andB. multivorans ). Dispersin B also inhibited biofilm formation and dispersed preformed biofilms ofBurkholderia species. These results imply a role for PNAG in the maintenance ofBurkholderia biofilm integrity. While PNAG was present in biofilms of all of the wild-type test organisms, a ΔpgaBC mutant ofB. multivorans (Mu5) produced no detectable PNAG, indicating that these genes are needed forBurkholderia PNAG formation. Furthermore, restoration of PNAG production in PNAG negativeE. coli TRXWMGΔC (ΔpgaC ) by complementation withB. multivorans pgaBCD confirmed the involvement of these genes inBurkholderia PNAG production. While the confocal scanning laser microscopy of untreated wild-typeB. multivorans showed thick, multilayered biofilm, Mu5 and dispersin B-treated wild-type biofilms were thin, poorly developed, and disrupted, confirming the involvement of PNAG inB. multivorans biofilm formation. Thus, PNAG appears to be an important component ofBurkholderia biofilms, potentially contributing to its resistance to multiple antibiotics and persistence during chronic infections, including cystic fibrosis-associated infection.