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.