Vibrio parahaemolyticus distinguishes between life in a liquid environment and life on a surface. Growth on a surface induces differentiation to a swarmer cell type. Movement of the swarmer cell across a solid surface, or swarming, results in progressive spreading of the bacterial colony and colonization of that surface. Colonization of surfaces plays a role in the pathogenesis of many organisms, and the study of the alternate cell-surface characteristics resulting from surface-dependent gene expression may provide insight for determining virulence factors. Recognition of surfaces involves detection of physical signals peculiar for that circumstance and transduction of that information to the swarmer cell gene system (laf). We know that the polar flagellum acts as a tactile organelle, and that surface recognition involves sensing forces that restrict movement of the polar flagellum. The primary objective of this research will be to investigate the mechanism by which the polar flagellum. The primary objective of this research will be to investigate the mechanism by which the polar flagellum acts as a sensor and conveys information to the laf gene system. Initial work will concentrate on the physical and genetic characterization of the sensor, i.e. the polar flagellar machinery. The structure of the polar flagellum is complex and needs to be defined: it is composed of heterologous subunits and is sheathed by an extension of the outer membrane. Other participating genes in the polar motility system (fla, che, and mot) will be identified, mapped, and cloned by using transposon mutagenesis. Localized mutagenesis of cloned genes in Escherichia coli and gene replacement of mutant for wild-type alleles in V. parahaemolyticus will be used to genetically dissect key fla and che loci. Energization of the motor will be examined. Also, selections and screens have been designed to look directly for laf specific regulatory genes. On identification of the parts and some of the circuitry of the sensory machinery, questions will be asked about the interaction of specific components of the fla and laf gene systems. For example, is a specific flagellin subunit also a repressor of the laf system? And is the direction of polar flagellar rotation critical to laf gene expression? Hypotheses will be tested by using fusions that couple laf gene transcription to reporter genes.
