Document Type thesis Author Name Arora, Bhupinder S URN etd-0826103-011111 Title Detection of polysaccharides on a bacterial cell surface using Atomic Force Microscopy Degree MS Department Chemical Engineering Advisors Prof. Terri A Camesano, Advisor Prof. Ravindra Datta, Department Head Keywords Leuconostoc mesenteroides NIRC1542 Atomic Force Microscope Pseudomonas putida KT2442 Adhesion Date of Presentation/Defense 2003-08-26 Availability unrestricted
Bacteria during the course of their life undergo a lot of developments on their surface. The changes that occur inside a cell result in the production of a variety of biopolymers on the cell surface. These polysaccharides have been found to play a major role in deciding the adhesive or repulsive nature of a bacterial cell. Based on the application the adhesive nature of a cell sometimes needs to be manipulated such that bacteria are required to have higher adhesions for bioremediation applications and in the case of bioreactors bacteria must not stick to walls to avoid fouling. In order to control adhesions of a cell to a variety of substrates, knowledge of the polysaccharides present on its surface is needed.
Therefore the goal of the present study is to detect the sugars present on the surface of Pseudomonas putida KT2442 using Atomic force microscopy and to relate properties of the polysaccharides to bacterial adhesion. Previous experiments suggested that cellulose and other sugars were produced by Pseudomonas putida KT2442.
Thus the cells were grown to late exponential phase and treated with cellulase to degrade any cellulose, if present, on the surface of the cells. Control experiments were done on untreated cells and cells that were not treated with cellulase but were centrifuged, since centrifugation is a part of the cellulase treatment and may also affect the bacterial surface. An appropriate (Steric) fitting model for the atomic force microscope (AFM) approach curves was applied to calculate the height and density of the polymer brush layer present on the cell surface. There was a decrease in the density of the polymer brush and increase in the height of the brush upon treatment with cellulase. Centrifugation alone did not affect the approach curves. From looking at the retraction curves it verified the results got from the approach curves and indicated stretching out of the polymer brush to greater distances after the treatment with cellulase.
Another batch of cells was treated with dextranase to check for the presence of dextran on the cell surface. Dextranase treated cells behaved identical to the control cells, suggesting that dextran is not one of the polysaccharides present on the bacterial surface. No change was observed in retraction curves data for dextranase treated and untreated cells.
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