Assessing the Temporal Dynamics of the Lower Urinary Tract Microbiota and the Effects of Lifestyle

TLDR: Assessing the Clinical Significance of LUT Microbiota Variability Following Vaginal Intercourse and assessing the Ability of Urinary Isolates to Adhere to Urothelial Cells.

Abstract: xix CHAPTER I: INTRODUCTION AND LITERATURE REVIEW 1 The Human Microbiota 1 The Role of the Human Microbiota 2 Measuring the Microbiome 3 Analyzing Microbiome Data 5 “Urine Denial”: The Concept of Bladder Sterility 7 The Discovery of the Female Urinary Microbiota and Microbiome 9 Urine is Not Sterile: Now What? 13 Correlations with Health and Disease 16 Description of Urinary Bacterial Isolates 21 Other Urinary Microorganisms 25 Description of the Male Urinary Microbiota 25 Correlations with Health and Disease 28 Correlations with Lifestyle and Preliminary Measures of Stability 30 Differences between the Male and Female Urinary Microbiota 31 The Microbiome is Shaped by Environment 31 The GI Microbiota and Lifestyle 33 Description of GI Tract Physiology 33 Correlations between the GI Microbiota and Lifestyle 34 Temporal Dynamics of the GI Microbiota 36 The Vaginal Microbiota and Lifestyle 39 Description of Vaginal Tract Physiology 39 Correlations between the Vaginal Microbiota, BV Risk, and Lifestyle 40 Temporal Dynamics of the Vaginal Microbiota 41 UTI Risk and Lifestyle 44 Current Understanding of UTI and an Overview of Risk Factors 44 vii Probiotics and UTI Risk 45 Correlations between UTI Risk and Lifestyle 47 Correlations between UTI Risk and Urine Properties 53 Longitudinal Assessment of the Lower Urinary Tract Microbiota 54 Rational and Problems 55 What Specimen to Collect? 56 Preliminary Findings 63 Summary 64 CHAPTER II: MATERIALS AND METHODS 67 Overview of Clinical Studies 67 MAGnUM Study 68 ProFUM Study 70 PARTNER Study 76 Bacterial Identification Methods 80 Specimen Collection 80 Expanded Culture Protocol 81 16S Sequencing Protocol 82 Dipstick Urinalysis 85 Storage of Bacterial Isolates 85 Statistical Analyses 85 Whole Genome Sequencing 86 DNA Extraction and Library Prep 86 Assembly and Annotation 87 Bacterial Assays: Experimental Procedures 88 Bacterial Strains and Growth Conditions 88 Urothelial Cell Adherence Assays 93 Biofilm Assays 94 Growth Competition Assays 95 CHAPTER III: FEASIBILITY PROJECT: MAGnUM STUDY 96 Overview of MAGnUM Study 96 Longitudinal Assessment of the Microbiota of the Male LUT 96 Longitudinal Assessment of the Microbiota of the Female LUT 100 Summary 106 CHAPTER IV: EFFECT OF INTRINSIC LIFESTYLE FACTORS ON LUT MICROBIOTA 109 Overview of ProFUM Study 109 Screening Eligible Participants 109 viii ProFUM Specimen Integrity 112 Qualitative Description of the Longitudinal LUT Microbiota and Microbiome 115 Measuring the Overall Stability of the LUT Microbiota 122 Menstruation and LUT Microbiota Variability 125 Urine Properties and Constituents and LUT Microbiota Variability 138 Summary 143 CHAPTER V: EFFECT OF EXTRINSIC LIFESTYLE FACTORS ON LUT MICROBIOTA 147 Sexual Activity and LUT Microbiota Variability 147 Hygiene and LUT Microbiota Variability 159 Oral Probiotics and LUT Microbiota Variability 163 Summary 165 CHAPTER VI: MECHANISTIC DETERMINATION AND SIGNIFICANCE OF LUT MICROBIOTA VARIABILITY AND VAGINAL INTERCOURSE 168 Introduction and Rationale 168 Assessing the Clinical Significance of LUT Microbiota Variability Following Vaginal Intercourse 170 Effect of Urinary Isolates on UPEC Adherence to Urothelial Cells 170 Effect of Urinary Isolates on UPEC Biofilm Formation 172 Effect of Urinary Isolates on UPEC Growth 175 Assessing the Ability of Urinary Isolates to Adhere to Urothelial Cells 181 Assessing the Biological Significance of LUT Microbiota Variability Following Vaginal Intercourse 183 Overview of PARTNER Study 183 Description of Microbiota Findings 184 Measuring Clonal Relatedness of S. mitis Isolates 190 Summary 191 CHAPTER VII: DISCUSSION 194 Brief Summary 194 Discussion of Chapter III 194 Discussion of Chapter IV 199 Discussion of Chapter V 208 Discussion of Chapter VI 215 Significance and Implications 220 Future Directions 224