B-190. Pseudomonas aeruginosa AlgZ Utilizes a Biofilm Specific Mechanism to Regulate Hydrogen Cyanide

W. L. Cody, Jr.1, C. L. Prichett1, A. Frisk2, M. Wolfgang3, M. J. Schurr1;
1Univ. of Colorado, Aurora, CO, 2Tulane Univ. Hlth. Sci. Ctr., New Orleans, LA, 3Univ. of North Carolina, Chapel Hill, NC.

Pseudomonas aeruginosa is an opportunistic pathogen that causes chronic infection in patients suffering from the genetic disorder cystic fibrosis. The two-component transcriptional regulator AlgR regulates a variety of virulence factors, including alginate and rhamnolipid production, twitching motility, biofilm formation and hydrogen cyanide production. In this study, the role of the putative sensor AlgZ on the regulation of cyanide production was examined. Measurements of cyanide production and hcnA::lacZ transcriptional fusion analyses in algZ and algR mutants revealed decreased cyanide production and hcnA expression in non-mucoid strains and increased cyanide production and hcnA expression in mucoid backgrounds. Cyanide production was also significantly elevated in cells grown on agar plates as compared to the same strains grown in broth. A conserved aspartate at position 54 of AlgR, which is required for twitching motility, but not alginate production, affected cyanide production. However, modification of this conserved aspartate did not alter the ability of AlgR to switch from an activator in mucoid cells to a repressor in non-mucoid strains on HCN production. Nuclease protection mapping of the hcnA promoter identified two previously unidentified start sites required for cyanide production. Strains that produced an unexpectedly low amount of cyanide were found to contain deletions of the newly identified hcnA start sites. Taken together, these observations provide data showing that the putative sensor/regulator pair, AlgZ and AlgR, co-regulate HCN production through a biofilm-dependent mechanism. These results further indicate that AlgR aspartate 54 is not involved in its phenotypic switch from a repressor in non-mucoid cells to an activator in mucoid cells with respect to cyanide production.