Differential modulation of Bordetella pertussis virulence genes as evidenced by DNA microarray analysis

Abstract

The production of most factors involved in Bordetella pertussis virulence is controlled by a two-component regulatory system termed BvgA/S. In the Bvg+ phase virulence-activated genes (vags) are expressed, and virulence-repressed genes (vrgs) are down-regulated. The expression of these genes can also be modulated by MgSO(4) or nicotinic acid. In this study we used microarrays to analyse the influence of BvgA/S or modulation on the expression of nearly 200 selected genes. With the exception of one vrg, all previously known vags and vrgs were correctly assigned as such, and the microarray analyses identified several new vags and vrgs, including genes coding for putative autotransporters, two-component systems, extracellular sigma factors, the adenylate cyclase accessory genes cyaBDE, and two genes coding for components of a type III secretion system. For most of the new vrgs and vags the results of the microarray analyses were confirmed by RT-PCR analysis and/or lacZfusions. The degree of regulation and modulation varied between genes, and showed a continuum from strongly BvgA/S-activated genes to strongly BvgA/S-repressed genes. The microarray analyses also led to the identification of a subset of vags and vrgs that are differentially regulated and modulated by MgSO(4) or nicotinic acid, indicating that these genes may be targets for multiple regulatory circuits. For example, the expression of bilA, a gene predicted to encode an intimin-like protein, was found to be activated by BvgA/S and up-modulated by nicotinic acid. Furthermore, surprisingly, in the strain analysed here, which produces only type 2 fimbriae, the fim3 gene was identified as a vrg, while fim2 was confirmed to be a vag.

Evaluation of the Role of the Bvg Intermediate Phase in Bordetella pertussis during Experimental Respiratory Infection

“…Therefore, taken together, our results demonstrate that bipA is a Bvg i -phase-specific gene. This conclusion confirms previous studies on B. bronchiseptica bipA (12,36) and B. pertussis bipA (14) but contradicts conclusions from other authors reporting either upregulation of bipA under Bvg Ϫ -phase conditions, thus regarding bipA as a vrg (2) or irregular transcriptional behaviors (16).…”

Species- and Strain-Specific Control of a Complex, Flexible Regulon by Bordetella BvgAS

“…Most known virulence factors and autotransporters that are present in the B. pertussis genome, including the TTSS, were Bvg activated by both strains, as was BB3242, encoding a putative toxin. Interestingly, fimX, fhaS, and dnt were Bvg activated only in Tohama I. fim3 was Bvg activated in GMT-1 but Bvg repressed in Tohama I, as previously described (24).…”

The BvgAS Regulon of Bordetella pertussis

“…Two hundred forty-five genes were identified as positively regulated by BvgA~P (Table S1B). Category 1 (Tables 2 and S2) contains well-known vags, such as virulence determinants and accessory factors, that have been identified by previous genetic and microarray studies (9,10,31,(33)(34)(35)(36); these include filamentous hemagglutinin/adhesin (fhaB and fhaC), adenylate cyclase toxins (cyaABCDE), fimbriae (fimABCD), pertactin (prn), pertussis toxins (ptxABDEC), and type 4 secretion system proteins (ptlABCDIEFGH). RT-qPCR confirmed the significant increase in RNA levels of several of these vags (Fig.…”

“…The two-component system BvgAS, comprised of the histidine kinase BvgS and the response regulator BvgA, is the master regulator of B. pertussis virulence (9,10). This system is coordinated by the intracellular level of phosphorylated BvgA (BvgA~P) (11,12).…”