The adhesion and aggregation behaviors of Pseudomonas aeruginosa ATCC 10145.

by Woo Yiu Ho, Anthony.Thesis (M.Phil.)--Chinese University of Hong Kong, 1998.Includes bibliographical references (leaves 162-171).Abstract also in Chinese.Abstract --- p.iAcknowledgements --- p.iiiTable of Contents --- p.ivList of Figures --- p.ixList of Tables --- p.xiList of Abbreviations --- p.xiiChapter 1 --- INTRODUCTION --- p.1Chapter 1.1 --- Bacterial Adhesion and Aggregation --- p.1Chapter 1.1.1 --- Significance of Bacterial Adhesion Studies --- p.1Chapter 1.1.2 --- Definitions --- p.4Chapter 1.1.3 --- Colonization Process --- p.7Chapter 1.1.4 --- Specific and Nonspecific Interactions --- p.8Chapter 1.1.5 --- Models of Bacterial Adhesion and Aggregation Processes --- p.14Chapter 1.1.6 --- Experimental Systems in Adhesion Research --- p.16Chapter 1.1.7 --- Experimental Systems in Aggregation Research --- p.19Chapter 1.2 --- Pseudomonas aeruginosa --- p.21Chapter 1.2.1 --- General Description and Clinical Significance --- p.21Chapter 1.2.2 --- Adhesins of Pseudomonas aeruginosa --- p.22Chapter 1.2.3 --- "Alginate, Mucoidity, Biofilm Formation and Cystic Fibrosis" --- p.23Chapter 1.2.4 --- Lipopolysaccharides --- p.26Chapter 1.2.5 --- Pili --- p.29Chapter 1.2.6 --- Flagella --- p.30Chapter 1.2.7 --- Lectins --- p.31Chapter 1.2.8 --- Other Adhesins --- p.31Chapter 1.2.9 --- Rhamnolipids --- p.32Chapter 1.3 --- Current Study --- p.33Chapter 2 --- MATERIALS AND EQUIPMENT --- p.35Chapter 2.1 --- Bacterial Strain --- p.35Chapter 2.2 --- Solid Surfaces --- p.35Chapter 2.3 --- Chemicals --- p.36Chapter 2.4 --- Recipes --- p.38Chapter 2.5 --- Equipment --- p.38Chapter 3 --- METHODS --- p.40Chapter 3.1 --- Maintenance and Culturation --- p.40Chapter 3.1.1 --- Maintenance of Bacterial Strains --- p.40Chapter 3.1.2 --- Seed Culture Preparation --- p.40Chapter 3.1.3 --- Culturation in Defined Growth Media --- p.40Chapter 3.2 --- Bacterial Adhesion and Aggregation Assay Methods --- p.41Chapter 3.2.1 --- Bacterial Adhesion on Glass Assay --- p.41Chapter 3.2.2 --- Bacterial Adhesion on Plastic Assay --- p.44Chapter 3.2.3 --- Bacterial Adhesion under Shear Assay --- p.44Chapter 3.2.4 --- Bacterial Aggregation Examination by Adhesion on Glass Assay --- p.45Chapter 3.2.5 --- Bacterial Aggregation Examination by Top-agar Assay --- p.45Chapter 3.2.6 --- Bacterial Aggregation Examination by Epi-fluorescence Microscopy --- p.46Chapter 3.2.7 --- Bacterial Aggregation Screening Test --- p.46Chapter 3.3 --- Determination of the Effects of Various Factors on Adhesion and Aggregation --- p.47Chapter 3.3.1 --- Culturation Period --- p.47Chapter 3.3.2 --- Osmotic Shock during the Washing Procedure --- p.47Chapter 3.3.3 --- Growth Media --- p.48Chapter 3.3.4 --- Assay Conditions --- p.48Chapter 3.3.5 --- Cell Pretreatments --- p.48Chapter 3.4 --- Isolation of Aggregation-deficient Mutants --- p.49Chapter 3.5 --- Outer Membrane Protein Profiles --- p.50Chapter 3.5.1 --- Isolation of Outer Membrane Fraction --- p.50Chapter 3.5.2 --- Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis --- p.51Chapter 3.6 --- Determination of the Mobility of the Bacteria on Surfaces --- p.52Chapter 3.6.1 --- Subsurface Twitching Assay --- p.52Chapter 3.6.2 --- Soft-agar Swarm Assay --- p.53Chapter 3.7 --- Detection of Alginate Production --- p.53Chapter 3.7.1 --- Extraction of Alginate from Spent Growth Medium --- p.53Chapter 3.7.2 --- Releasing Cell Surface-associated Alginate --- p.54Chapter 3.8 --- Other Assay Methods --- p.55Chapter 3.8.1 --- Protein Assay --- p.55Chapter 3.8.2 --- Carbohydrate Determination --- p.55Chapter 3.8.3 --- Alginate Determination --- p.55Chapter 4 --- RESULTS --- p.57Chapter 4.1 --- Standardization of the Assays for Bacterial Adhesion and Aggregation --- p.57Chapter 4.1.1 --- Effects of Cell Density and Exposure Time on the Number of Adhered Bacteria Detected in Bacterial Adhesion on Glass Assay --- p.57Chapter 4.1.2 --- Characterization of Bacterial Aggregation by Different Examination Methods --- p.62Chapter 4.1.3 --- Effects of Culturation Period on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.67Chapter 4.1.4 --- Effects of Osmotic Shock during Washing on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.70Chapter 4.1.5 --- Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 as a Function of Time under the Standard Assay Condition --- p.71Chapter 4.1.6 --- Consistency of Bacterial Adhesion on Glass Assay --- p.74Chapter 4.2 --- Effects of Growth Media on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.77Chapter 4.3 --- Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 in Different Assay Media --- p.77Chapter 4.3.1 --- Effects of Various Buffers on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.77Chapter 4.3.2 --- Effects of pH on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.78Chapter 4.3.3 --- Effects of Various Electrolytes on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.81Chapter 4.3.4 --- Concentration Effects of Monovalent and Divalent Cations on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.88Chapter 4.3.5 --- Concentration Effects of Phosphate Buffers on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.93Chapter 4.3.6 --- Concentration Effects of Ammonium Sulfate and Cyclohexylammonium Sulfate on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.96Chapter 4.3.7 --- Effects of Cation Chelation on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.99Chapter 4.3.8 --- Effects of Sugars on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.100Chapter 4.3.9 --- Effects of Amino Acids on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.101Chapter 4.4 --- Adhesion and Aggregation after Pretreatments of the Cells --- p.103Chapter 4.4.1 --- Effects of Protease Treatments on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.103Chapter 4.4.2 --- Effects of Externally Added Proteins on Adhesion and Aggregation of Pronase-treated Cells --- p.107Chapter 4.4.3 --- Effects of Acid or Base Treatments on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.108Chapter 4.4.4 --- Effects of Heat Treatment on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.108Chapter 4.4.5 --- Effects of Extensive Washing on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.110Chapter 4.5 --- Isolation and Growth Characteristics of Aggregation-deficient Mutants --- p.111Chapter 4.6 --- Comparisons of the Adhesion and Aggregation Characters of Pseudomonas aeruginosa ATCC 10145 and Mutant 9 --- p.115Chapter 4.6.1 --- Under Standard Condition --- p.115Chapter 4.6.2 --- On Different Surfaces and in Different Electrolytes --- p.115Chapter 4.6.3 --- Under Shear --- p.118Chapter 4.6.4 --- Adhesion and Aggregation of Combined Suspensions of Pseudomonas aeruginosa ATCC 10145 and Mutant 9 --- p.122Chapter 4.7 --- Characterization of the Cell Surface Properties of Pseudomonas aeruginosa ATCC 10145 and Mutant 9 --- p.125Chapter 4.7.1 --- Outer Membrane Protein Profiles --- p.125Chapter 4.7.2 --- Pili-elicited Twitching Mobility --- p.125Chapter 4.7.3 --- Mobility Due to Flagella --- p.128Chapter 4.7.4 --- Production of Alginate --- p.128Chapter 5 --- DISCUSSIONS --- p.130Chapter 5.1 --- Choice of the Materials --- p.130Chapter 5.2 --- Development of the Assay Methods --- p.130Chapter 5.2.1 --- Development of the Procedures for Bacterial Adhesion Assays --- p.130Chapter 5.2.2 --- Development of the Assay Methods for Bacterial Aggregation --- p.132Chapter 5.2.3 --- Standardization of the Assays --- p.133Chapter 5.2.4 --- Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 as a Function of Time under the Standard Assay Condition --- p.134Chapter 5.2.5 --- Consistency of Bacterial Adhesion on Glass Assay --- p.135Chapter 5.2.6 --- Limits of Bacterial Adhesion on Glass Assay --- p.135Chapter 5.3 --- Effects of Growth Media on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.135Chapter 5.4 --- Effects of Various Chemicals in the Assay Media on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.136Chapter 5.4.1 --- Effects of Electrolytes on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.137Chapter 5.4.2 --- Effects of Aggregation on Adhesion --- p.140Chapter 5.4.3 --- Effects of Cyclohexylammonium Sulfate and Ammonium Sulfate on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.141Chapter 5.4.4 --- Effects of Sugars and Amino Acids on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.143Chapter 5.5 --- Effects of Various Cell-surface Modifications on Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 --- p.144Chapter 5.6 --- Isolation and Growth Characteristics of Aggregation-deficient Mutants --- p.146Chapter 5.7 --- Comparisons of the Adhesion and Aggregation Characters of Pseudonomas aeruginosa ATCC 10145 and Mutant 9 --- p.147Chapter 5.7.1 --- Adhesion and Aggregation of Pseudonomas aeruginosa ATCC 10145 and Mutant 9 on Different Surfaces In Different Electrolytes --- p.147Chapter 5.7.2 --- Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 and Mutant 9 Under Shear --- p.147Chapter 5.7.3 --- Adhesion and Aggregation of Combined Suspensions of Pseudomonas aeruginosa ATCC 10145 and Mutant 9 --- p.148Chapter 5.8 --- Characterization of the Cell Surface Properties of Pseudomonas aeruginosa ATCC 10145 and Mutant 9 --- p.148Chapter 5.9 --- General Discussions --- p.151Chapter 6 --- APPENDIX --- p.154Chapter 6.1 --- Visual Examination of Adhesion and Aggregation of Pseudomonas aeruginosa ATCC 10145 on Glass --- p.154Chapter 6.2 --- Fractal Analysis of Bacterial Aggregates --- p.154Chapter 7 --- REFERENCES --- p.16

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