A molecular and genetic analysis of the requirement of SadB for modulating surface-associated behaviors in Pseudomonas aeruginosa PA14

Pseudomonas aeruginosa is one of the model organisms used to study the developmental process of biofilm formation in Gram-negative bacteria. Recently, a genetic approach {lcub}O'Toole, 1998 #404{rcub} was taken to study biofilm formation in P. aeruginosa and has unearthed numerous determinants which have shaped and defined our understanding of the steps involved in biofilm formation. This preliminary work has provided us with a collection of tools that has enabled us to delve deeper into mechanisms underlying this form of microbial development.; This thesis describes the characterization of the sadB gene and the protein it encodes (SadB) as they relate to surface-associated behaviors. Microscopic studies were used to show that SadB is required for the transition from reversible to irreversible attachment in P. aeruginosa during biofilm formation. Biochemical techniques were used to localize SadB to the cytoplasm. Structure function analysis has shown that all but the last ∼40 amino acids are required for SadB function.; SadB was also shown to modulate the surface-associated swarming behavior that P. aeruginosa exhibits on semi-solid surfaces. Biochemical and genetic approaches were utilized to demonstrate that rhamnolipid surfactants are required to modulate swarming and that this modulation is dependent on the function of SadB. We have also shown that hamnolipid surfactants are required for channel maintenance in biofilm formation, suggesting a link between swarming motility and biofilm formation.; A genetic analysis of the involvement of SadB in biofilm formtion and swarming motility has revealed that SadB is involved in modulating flagellar function via the flagellar stator complex and chemotaxis cluster IV of P. aeruginosa. Thus observation has lead to a model whereby P. aeruginosa initiates surface contact by it flagellated pole. Upon sensing this interaction, cells can either return to the planktonic population or engage in surface associated behaviors such as biofilm formation and swarming motility, thus implicating SadB as a molecular switch involved in executing the cell's decision to progress to a sessile or a motile surface-associated lifestyle.