Construction Of A ?mbaA?potD1 Mutant And The Effects On Biofilm Formation In Vibrio Cholerae

Vibrio cholerae is a bacterial species identified as the causative agent of the diseasecholera. Similar to other aquatic bacteria, V. cholerae forms complex communities ofmicroorganisms bounded by a polysaccharide matrix, known as biofilms. Biofilms allowplanktonic bacteria to become immobilized within environments of favorable conditions bypermitting adherence to surfaces. This matrix-encased microbial community providesprotection against environmental stressors to ensure survival. Polyamines, or small cationicmolecules essential to cell processes, are among the many environmental signals that act inregulating biofilm formation.V. cholerae cells contain two systems, the NspS/MbaA signaling complex and thePotABCD1 ATP-binding cassette transporter, that function to detect and import thesepolyamines. The objective of this project was to determine the interplay between these twosystems in regulating biofilm formation by investigating whether PotD1 and MbaA inhibitbiofilm formation additively. To accomplish this, I explored the effects of knocking out theMbaA and PotD1 proteins. I constructed a ?mbaA?potD1 double mutant bacterial strain.Through colony PCR and gel electrophoresis, I was able to confirm the construction of thismutant. I then performed biofilm assays with the newly constructed mutant, the wild-type,and ?mbaA and ?potD1 single mutants. The results showed that the ?mbaA?potD1 mutantdid not form more biofilm than the single mutants, contradicting the hypothesized effects.Performing further biofilm assays with this double mutant and collecting data over varioustime points will allow our lab to further characterize the role of the PotABCD1 transportersystem and the NspS/MbaA complex in the regulation of biofilm formation.