H-035. Investigating the Role of ndvB in Gene Expression in Pseudomonas aeruginosa Biofilms

T. W. Beaudoin, T-F. C. Mah;
Univ. of Ottawa, Ottawa, ON, CANADA.

Bacterial cells are often found as surface attached communities called biofilms that portray a vastly different phenotype when compared to their planktonic counterparts. The formation of these sessile communities confer several advantages to these bacteria, paramount amongst these is an increase in antibiotic resistance. The common opportunistic pathogen Pseudomonas aeruginosa has been well studied for its ability to form biofilms and is clinically relevant in patients with Cystic Fibrosis leading to persistent, recurring infections that are difficult to treat. A previous genetic screen has led to the identification of several mutants that show typical biofilm architecture and planktonic antibiotic resistance, but have a drastically reduced biofilm resistance profile. A mutant with a strong biofilm-specific antibiotic-sensitive phenotype had a transposon-insertion in the ndvB gene. ndvB is important for the expression of periplasmic and extracellular glucans. We hypothesize that these extracellular glucans act as diffusible signals that can modulate gene expression within the biofilm. The goal of the current study is to identify genes that are controlled by ndvB expression via an Affymetrix microarray analysis of RNA extracted from wild type and ndvB- biofilms of P. aeruginosa PA14 strains. The antibiotic sensitivity phenotypes of the wild type and ndvB mutant strains was confirmed by the use of minimal bactericidal concentration assays, where biofilms are grown for 24 hours, then treated with antibiotics and assessed for viability. RNA for microarray analysis was obtained from 48 hour biofilms grown in 6-well microtitre plates and extracted using an on-column method. The identification of differentially expressed genes and confirmation via quantitative real-time PCR is in progress to investigate the role that ndvB plays in the multi-factorial mechanism of biofilm-specific antibiotic resistance.