Chemical analysis of cellular and extracellular carbohydrates of a biofilm-forming strain Pseudomonas aeruginosa PA14

Abstract

Background: Pseudomonas aeruginosa is a Gram-negative bacterium and an opportunistic pathogen, which causes persisting life-threatening infections in cystic fibrosis (CF) patients. Biofilm mode of growth facilitates its survival in a variety of environments. Most P. aeruginosa isolates, including the non-mucoid laboratory strain PA14, are able to form a thick pellicle, which results in a surface-associated biofilm at the air-liquid (A-L) interface in standing liquid cultures. Exopolysaccharides (EPS) are considered as key components in the formation of this biofilm pellicle. In the non-mucoid P. aeruginosa strain PA14, the "scaffolding" polysaccharides of the biofilm matrix, and the molecules responsible for the structural integrity of rigid A-L biofilm have not been identified. Moreover, the role of LPS in this process is unclear, and the chemical structure of the LPS O-antigen of PA14 has not yet been elucidated. Principal Findings: In the present work we carried out a systematic analysis of cellular and extracellular (EC) carbohydrates of P. aeruginosa PA14. We also elucidated the chemical structure of the LPS O-antigen by chemical methods and 2-D NMR spectroscopy. Our results showed that it is composed of linear trisaccharide repeating units, identical to those described for P. aeruginosa Lanýi type O:2a,c (Lanýi-Bergman O-serogroup 10a, 10c; IATS serotype 19) and having the following structure: -4)-α-L-GalNAcA-(1-3)-α-D-QuiNAc-(1-3)- α-L-Rha-(1-. Furthermore, an EC O-antigen polysaccharide (EC O-PS) and the glycerol-phosphorylated cyclic β-(1,3)-glucans were identified in the culture supernatant of PA14, grown statically in minimal medium. Finally, the extracellular matrix of the thick biofilm formed at the A-L interface contained, in addition to eDNA, important quantities (at least ~20% of dry weight) of LPS-like material. Conclusions: We characterized the chemical structure of the LPS O-antigen and showed that the O-antigen polysaccharide is an abundant extracellular carbohydrate of PA14. We present evidence that LPS-like material is found as a component of a biofilm matrix of P. aeruginosa. © 2010 Coulon et al.