title: Haemophilus ducreyi Hfq contributes to virulence gene regulation as cells enter stationary phase.
abstract: To adapt to stresses encountered in stationary phase, Gram-negative bacteria utilize the alternative sigma factor RpoS. However, some species lack RpoS; thus, it is unclear how stationary-phase adaptation is regulated in these organisms. Here we defined the growth-phase-dependent transcriptomes of Haemophilus ducreyi, which lacks an RpoS homolog. Compared to mid-log-phase organisms, cells harvested from the stationary phase upregulated genes encoding several virulence determinants and a homolog of hfq. Insertional inactivation of hfq altered the expression of ~16% of the H. ducreyi genes. Importantly, there were a significant overlap and an inverse correlation in the transcript levels of genes differentially expressed in the hfq inactivation mutant relative to its parent and the genes differentially expressed in stationary phase relative to mid-log phase in the parent. Inactivation of hfq downregulated genes in the flp-tad and lspB-lspA2 operons, which encode several virulence determinants. To comply with FDA guidelines for human inoculation experiments, an unmarked hfq deletion mutant was constructed and was fully attenuated for virulence in humans. Inactivation or deletion of hfq downregulated Flp1 and impaired the ability of H. ducreyi to form microcolonies, downregulated DsrA and rendered H. ducreyi serum susceptible, and downregulated LspB and LspA2, which allow H. ducreyi to resist phagocytosis. We propose that, in the absence of an RpoS homolog, Hfq serves as a major contributor of H. ducreyi stationary-phase and virulence gene regulation. The contribution of Hfq to stationary-phase gene regulation may have broad implications for other organisms that lack an RpoS homolog.
  
  
  OBJECTIVE
  Pathogenic bacteria encounter a wide range of stresses in their hosts, including nutrient limitation; the ability to sense and respond to such stresses is crucial for bacterial pathogens to successfully establish an infection. Gram-negative bacteria frequently utilize the alternative sigma factor RpoS to adapt to stresses and stationary phase. However, homologs of RpoS are absent in some bacterial pathogens, including Haemophilus ducreyi, which causes chancroid and facilitates the acquisition and transmission of HIV-1. Here, we provide evidence that, in the absence of an RpoS homolog, Hfq serves as a major contributor of stationary-phase gene regulation and that Hfq is required for H. ducreyi to infect humans. To our knowledge, this is the first study describing Hfq as a major contributor of stationary-phase gene regulation in bacteria and the requirement of Hfq for the virulence of a bacterial pathogen in humans.
authors: Dharanesh
authors: D
authors: Gangaiah
authors: Maria
authors: M
authors: Labandeira-Rey
authors: Xinjun
authors: X
authors: Zhang
authors: Kate R
authors: KR
authors: Fortney
authors: Sheila
authors: S
authors: Ellinger
authors: Beth
authors: B
authors: Zwickl
authors: Beth
authors: B
authors: Baker
authors: Yunlong
authors: Y
authors: Liu
authors: Diane M
authors: DM
authors: Janowicz
authors: Barry P
authors: BP
authors: Katz
authors: Chad A
authors: CA
authors: Brautigam
authors: Robert S
authors: RS
authors: Munson
authors: Eric J
authors: EJ
authors: Hansen
authors: Stanley M
authors: SM
authors: Spinola
mesh_terms: D000328:Adult; D002602:Chancroid / Q000382:microbiology / Q000473:pathology; D005260:Female; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial; D055786:Gene Knockout Techniques; D006191:Haemophilus ducreyi / Q000235:genetics* / Q000254:growth & development* / Q000472:pathogenicity; D064368:Healthy Volunteers; D035001:Host Factor 1 Protein / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008297:Male; D008875:Middle Aged; D037521:Virulence Factors / Q000096:biosynthesis*
chemical_list: D035001:Host Factor 1 Protein; D037521:Virulence Factors