Vibrio parahaemolyticus (Vp) is a Gram-negative, halophilic, motile bacterium that is ubiquitous in the marine environment. Vp is responsible for seafood-borne illnesses andsevere infections can result in sepsis and possibly death in immunocompromised individuals. Vp expresses two different types of flagella, a polar flagellum that is required for swimming motility, and lateral flagella that are required for swarming motility. Other Vibrio spp only express polar flagella, and generally polar flagella are involved in Vibrio-mediated virulence, so the Vp polar flagellum is the focus of this study. The Vp polar flagellar filament is composed of four flagellin subunits, FlaABCD, with an additional two flagellin subunits FlaEF being secreted outside the cell. To dissect the roles of the individual flagellins to Vp polar flagellar synthesis, we are constructing Vp strains lacking the various flagellin genes, and then evaluating motility of the mutants.  A Vp strain was constructed that lacks three of the six flagellins (DflaEDB), and this strain maintained near wildtype levels of motility.  This indicates that FlaACF are sufficient for polar flagellar synthesis and motility.   We are currently constructing a deletion of flaF, using PCR, splicing by overlap extension, and Golden Gate assembly.  This conjugatable plasmid will be used to construct a DflaF ΔflaEDB Vp mutant strain, and then evaluate its motility. Our goal is to identify the minimum flagellin(s) that are required for Vp polar flagellar synthesis, and then study their localization within the filament.   These studies will contribute to understanding flagellar-mediated pathogenicity of Vibrio species.