Bordetella PlrSR regulatory system controls BvgAS activity and virulence in the lower respiratory tract.

Bacterial pathogens coordinate virulence using two-component regulatory systems (TCS). The Bordetella virulence gene (BvgAS) phosphorelay-type TCS controls expression of all known protein virulence factor-encoding genes and is considered the "master virulence regulator" in Bordetella pertussis, the causal agent of pertussis, and related organisms, including the broad host range pathogen Bordetella bronchiseptica We recently discovered an additional sensor kinase, PlrS [for persistence in the lower respiratory tract (LRT) sensor], which is required for B. bronchiseptica persistence in the LRT. Here, we show that PlrS is required for BvgAS to become and remain fully active in mouse lungs but not the nasal cavity, demonstrating that PlrS coordinates virulence specifically in the LRT. PlrS is required for LRT persistence even when BvgAS is rendered constitutively active, suggesting the presence of BvgAS-independent, PlrS-dependent virulence factors that are critical for bacterial survival in the LRT. We show that PlrS is also required for persistence of the human pathogen B. pertussis in the murine LRT and we provide evidence that PlrS most likely functions via the putative cognate response regulator PlrR. These data support a model in which PlrS senses conditions present in the LRT and activates PlrR, which controls expression of genes required for the maintenance of BvgAS activity and for essential BvgAS-independent functions. In addition to providing a major advance in our understanding of virulence regulation in Bordetella, which has served as a paradigm for several decades, these results indicate the existence of previously unknown virulence factors that may serve as new vaccine components and therapeutic or diagnostic targets.

Chemical biology applied to the study of bacterial pathogens.

In recent years, chemical biology and chemical genomics have been increasingly applied to the field of microbiology to uncover new potential therapeutics as well as to probe virulence mechanisms in pathogens. The approach offers some clear advantages, as identified compounds (i) can serve as a proof of principle for the applicability of drugs to specific targets; (ii) can serve as conditional effectors to explore the function of their targets in vitro and in vivo; (iii) can be used to modulate gene expression in otherwise genetically intractable organisms; and (iv) can be tailored to a narrow or broad range of bacteria. This review highlights recent examples from the literature to illustrate how the use of small molecules has advanced discovery of novel potential treatments and has been applied to explore biological mechanisms underlying pathogenicity. We also use these examples to discuss practical considerations that are key to establishing a screening or discovery program. Finally, we discuss the advantages and challenges of different approaches and the methods that are emerging to address these challenges.

Small-molecule inhibitors of toxT expression in Vibrio cholerae.

UNLABELLED: Vibrio cholerae, a Gram-negative bacterium, infects humans and causes cholera, a severe disease characterized by vomiting and diarrhea. These symptoms are primarily caused by cholera toxin (CT), whose production by V. cholerae is tightly regulated by the virulence cascade. In this study, we designed and carried out a high-throughput chemical genetic screen to identify inhibitors of the virulence cascade. We identified three compounds, which we named toxtazin A and toxtazin B and B', representing two novel classes of toxT transcription inhibitors. All three compounds reduce production of both CT and the toxin-coregulated pilus (TCP), an important colonization factor. We present evidence that toxtazin A works at the level of the toxT promoter and that toxtazins B and B' work at the level of the tcpP promoter. Treatment with toxtazin B results in a 100-fold reduction in colonization in an infant mouse model of infection, though toxtazin A did not reduce colonization at the concentrations tested. These results add to the growing body of literature indicating that small-molecule inhibitors of virulence genes could be developed to treat infections, as alternatives to antibiotics become increasingly needed. IMPORTANCE: V. cholerae caused more than 580,000 infections worldwide in 2011 alone (WHO, Wkly. Epidemiol. Rec. 87:289-304, 2012). Cholera is treated with an oral rehydration therapy consisting of water, glucose, and electrolytes. However, as V. cholerae is transmitted via contaminated water, treatment can be difficult for communities whose water source is contaminated. In this study, we address the need for new therapeutic approaches by targeting the production of the main virulence factor, cholera toxin (CT). The high-throughput screen presented here led to the identification of two novel classes of inhibitors of the virulence cascade in V. cholerae, toxtazin A and toxtazins B and B'. We demonstrate that (i) small-molecule inhibitors of virulence gene production can be identified in a high-throughput screen, (ii) targeting virulence gene production is an effective therapeutic strategy, and (iii) small-molecule inhibitors can uncover unknown layers of gene regulation, even in well-studied regulatory cascades.