B-269. Characterization of Bordetella bronchiseptica Strains Requiring Elevated CO2 Levels for Induction of Bvg+ Phase Genes

M. Liu1, S. Moghadam1, C. A. Cummings1, S. Doulatov1, D. Fujiyama1, A. Jani2, E. Pineda1, J. D. Cherry1, M. Forsyth2, I. Schroeder1, P. A. Cotter2, J. F. Miller1;
1Univ. of California, Los Angeles, CA, 2Univ. of California, Santa Barbara, CA.

Bordetella brochiseptica strain JC100, unlike other common Bordetella strains, requires growth in the presence of high CO2 concentration (5%) for expression of characteristics typical of Bvg+ (virulent) phase Bordetella. A recent survey of B. bronchiseptica isolates identified additional strains that also require 5% CO2 for full level expression of Bvg+ phase genes, and these strains fell into two phylogenetically distinct clades (complex I and complex IV). Swapping the bvgAS loci between JC100 and RB50, a B. bronchiseptica strain that does not require 5% CO2 for expression of Bvg+ phase phenotypes, demonstrted that the CO2 phenotype was not linked to bvgAS. Introduction of the constitutively active bvgS-C3 allele into the bvgAS locus of JC100 did not eliminate the requirement for 5% CO2 for expression of Bvg+ phase phenotypes. RT-PCR analysis using JC100 and RB50 grown in batch fermentation cultures in which only the CO2 concentration was varried confirmed that CO2, and not other potential variables such as pH, was the relevant signal. Global expression analysis using Bordetella microarrays indicated that, in the absence of 5% CO2, known Bvg- phase genes such as the flagellar and chemotaxis genes were expressed in JC100 and known Bvg+ phase genes were not. Under these same conditions, Bvg+ phase genes were expressed in RB50. We have screened over 60,000 transposon mutagenized JC100 colonies but did not identify any mutants that displayed Bvg+ phase characteristics without 5% CO2. In addition, we have observed that RB50 has enhanced FHA and adenylate cyclase expression under 5% CO2. On the basis of these and other data, we hypothesize the existence of a CO2-responsive factor in B. bronchiseptica that intersects with the BvgS-BvgA phosphorelay at a point downstream from signal recognition by BvgS.