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Results reveal that the signal transduction system component ArcB of Haemophilus influenzae although lacks the PAS domain, involved in signal reception of a variety of sensory proteins, still is capable of mediating signal transduction in response to growth conditions.

The Bordetella global virulence regulator bvg, along with iron, was found to decrease the manufacture of alcaligin by Bordetella bronchiseptica RB50 and GP1SN. This ability is strongly related to Bordetella phylogenetic lineage and with host species from which the organisms were taken. These findings are in conflict with past experiments showing that the production of alcaligin by B. bronchiseptica MBORD846 and B. pertussis was only affected by iron but not by bvg.

Evaluation of the motility control facilitated by the Bordetella virulence control locus, bvgAS, and the variation in the genetic circuity helps study the function of negative control by BvgAS in the environment of Bordetella in a host. frlAB, the regulatory locus that is essential for flagellar organelle synthesis is negatively regulated by BvgAS. frl expression pattern reversal stimulates motility in the Bvg+ growth phase. Reduction in the colonization by Bordetella bronchisptica due to the ectopic expression of frl at the time of onset of respiratory infection reveals that both positive and negative regulation by BvgAS are essential during colonization.

The Bordetella flaA gene, which encodes flagellin, belongs to a class of genes termed vrg, which are negatively regulated by the bvgAS locus. The mechanism of flaA gene regulation by BvgAS was investigated. Northern blot and primer extension analysis showed that the flaA gene is regulated by BvgAS at the level of transcription. Furthermore, analysis of the motility phenotype of Bordetella bronchiseptica indicates that the Bordetella virulence control system regulates the transcriptional control of flaA.

The effect of the bvgAS locus on motility and flagellar synthesis in Bordetella bronchiseptica was examined. The bvgAS locus is involved in the regulation of various virulence factors in response to environmental signals. Mutations involving the entire bvgAS locus or the bvgS gene resulted in a motile phenotype, which also arises in response to environmental signals which modulate the Bvg system. Electron microscopy provided evidence that suggests regulation occurs at a step prior to flagellum assembly, with some transcriptional control involved.

Mariner-family transposable elements are active in a wide variety of organisms and are becoming increasingly important genetic tools in species lacking sophisticated genetics. The Himar1 element, isolated from the horn fly, Haematobia irritans, is active in Escherichia coli when expressed appropriately. We used this fact to devise a genetic screen for hyperactive mutants of Himar1 transposase that enhance overall transposition from [approximately equals] 4- to 50-fold as measured in an E. coli assay. Purified mutant transposases retain their hyperactivity, although to a lesser degree, in an in vitro transposition assay. Mutants like those described herein should enable sophisticated analysis of the biochemistry of mariner transposition and should improve the use of these elements as genetic tools, both in vivo and in vitro.

Nontypeable Haemophilus influenzae (NTHi) is a major cause of community acquired pneumonia and exacerbation of chronic obstructive pulmonary disease. A current effort in NTHi vaccine development has focused on generating humoral responses and has been greatly impeded by antigenic variation among the numerous circulating NTHi strains. In this study, we showed that pulmonary immunization of mice with killed NTHi generated broad protection against lung infection by different strains. While passive transfer of immune antibodies protected only against the homologous strain, transfer of immune T cells conferred protection against both homologous and heterologous strains. Further characterization revealed a strong Th17 response that was cross-reactive with different NTHi strains. Responding Th17 cells recognized both cytosolic and membrane-associated antigens, while immune antibodies preferentially responded to surface antigens and were highly strain specific. We further identified several conserved proteins recognized by lung Th17 cells during NTHi infection. Two proteins yielding the strongest responses were tested as vaccine candidates by immunization of mice with purified proteins plus an adjuvant. Immunization induced antigen-specific Th17 cells that recognized different strains and, upon adoptive transfer, conferred protection. Furthermore, immunized mice were protected against challenge with not only NTHi strains but also a fully virulent, encapsulated strain. Together, these results show that the immune mechanism of cross-protection against pneumonia involves Th17 cells, which respond to a broad spectrum of antigens, including those that are highly conserved among NTHi strains. These mechanistic insights suggest that inclusion of Th17 antigens in subunit vaccines offers the advantage of inducing broad protection and complements the current antibody-based approaches. [ABSTRACT FROM AUTHOR]