The Gut and Vaginal Microbial Communities during Pregnancy in Patients With and Without Urogenital Infections Restricted; Files Only

Abstract

Background: The gut and vaginal microbiome both change over the course of a pregnancy and have been associated with many pregnancy complications. Pregnant individuals are more susceptible to urinary tract infections (UTIs), bacterial vaginosis (BV), and chlamydia infection. It is currently unclear if there are differences in the microbiome or the collection of antimicrobial resistance (AMR) genes for pregnant individuals who develop urogenital infections versus those who don’t.

Purpose: The dissertation aims to examine (1) how bacterial species change during pregnancy for those with and without urogenital infections, and (2) how AMR genes change during pregnancy after antibiotic treatment for urogenital infections.

Methods: A subset of the data from the Emory University African American Vaginal, Oral, and Gut Microbiome in Pregnancy Cohort Study were sent for metagenomic sequencing (238 patients, rectal and vaginal swabs at 8-14 weeks & 24 - 30 weeks pregnancy). Taxonomic assignment was done with the metaphlan2 software tool, and AMR genes were detected with the AMR Finder Plus tool. 16S rRNA data from the same samples had taxonomic assignment with the PECAN and DADA2 tools. Associations between the microbiome and urogenital infections were analyzed with a linear decomposition model. Differences in the frequency of AMR genes was analyzed with a chi-squared test for independence. hAMRoaster, a new bioinformatics tool, was created to compare the performance of different AMR gene processing pipelines.

Results: Collectively, this dissertation finds that the gut and vaginal microbial communities are not significantly impacted by urogenital infections or their treatment. Specific microbes and AMR genes tend to be increased in those who developed urogenital infections compared to those who did not, however, these differences do not persist for the entire pregnancy. These findings should reassure most patients that being diagnosed with a urogenital infection and receiving antibiotic therapy for that infection will not have a significant, detrimental impact on their microbiome overall during pregnancy.

Table of Contents

Chapter 1: Introduction ……………………………………………………………………1

What Is a Microbiome? ……………………………………………………………..1

Gut Microbiome Sampling ………………………………………………………….3

Vaginal Microbiome Sampling ……………………………………………………..4

Bioinformatic Approaches to Microbiome Data …………………………………5

The Microbiome in Health …………………………………………………………..8

Outline of Dissertation ……………………………………………………………12

Summary …………………………………………………………………………….19

Figure 1 ……………………………………………………………………..20

Figure 2 ……………………………………………………………………..21

References ……………………………………………………………………..22

Chapter 2: The Impact of Urogenital Infections on the Gut and Vaginal Microbiome in Pregnancy ………………………………………………………………………………….32

Abstract …………………………………………………………………………….32

Introduction …………………………………………………………………………34

Methods …………………………………………………………………………….35

Results ………………………………………………………………………………38

Discussion …………………………………………………………………………..40

Conclusion ………………………………………………………………………….45

Figure 1: Number of Species Identified per Sample by MetaPhlAn2 ………46

Figure 2: Prevalence of BV, UTI, and Chlamydia Diagnosis in This Sample ..47

Table 1: Comparison of Total Number of Species Identified by Different Bioinformatic Tools …………………………………………………………..49

Table 2: Significant p Values ………………………………………………...50

Table 3: Significant Taxa from Aim 1 Analysis ………………………………51

References ……………………………………………………………………………58

Chapter 3: hAMRoaster: A Tool for Comparing Performance of AMR Gene Detection Software ……………………………………………………………………………………..63

Abstract ………………………………………………………………………………63

Introduction ………………………………………………………………………….65

Methods ………………………………………………………………………………67

Results ………………………………………………………………………………..71

Figure 1: Schematic l Methods ………………………………………………80

Figure 2: Antimicrobial Resistance (AMR) Genes Detected By Software Tools by Drug Class …………………………………………………………………81

Figure 3: Sensitivity of Software Tools for Detection of Antimicrobial Resistance (AMR) Genes Across Coverage Levels …………………………………84

Figure 4: Percent Detection of Unknown Antimicrobial (AMR) Resistance Genes Across Coverage ……………………………………………………………86

Figure 5: Agreement (Cohen’s Kappa) Values between Tools across Coverage Levels Calculated in R Using the Kappa2 Function ……………………….87

Table 1A: Clinical Isolates Included in the High Resistance Simulated Community..... 88

Table 1B: Clinical Isolates Included in the Low Resistance Simulated Community.... 89

Table 2: Tools Identified from Search Methods with the Selection Criteria and Whether They Subsequently Worked or Not...90

Table 3A: Summary Statistics for the High Resistance Data from hAMRoaster.....94

Table 3B: Summary Statistics for the Low Resistance Data from hAMRoaster.....99

Supplementary Text 1 ……………………………………………………………..105

Supplementary Table 1 ………………………………………………………106

References …………………………………………………………………………...107

Chapter 4: Frequency of Antimicrobial Resistance Genes in a Pregnancy Cohort for People with and without Urogenital Infections during Pregnancy ……………………………..113

Abstract …………………………………………113

Introduction ………………………………………………………………………….115

Methods ……………………………………………………………………………..117

Results ………………………………………………………………………………119

Discussion ………………………………………………………………………….121

Future Directions ……………………………………………………………………123

Limitations …………………………………………………………………………...124

Conclusion …………………………………………………………………………..125

Figure 1: Alpha Diversity of Vaginal and Rectal Samples for all Groups …126

Figure 2: Total Number of AMR Genes per Drug Class across Body Sites ..127

Table 1: Vaginal Infection Rates in This Cohort ……………………………129

Table 2: Standardized Residual Values from Chi Squared ………………….130

Supplementary Table 1: Clinical Isolates Used for Simulated Data ………..131

Supplementary Table 2: Preliminary Data Results from hAMRonization to Compare AMR Tool Performance …………………………………………132

Supplementary Table 3: hAMRoaster Output for Preliminary Data Analysis...133

References ………………………………………………………………………….134

 

Chapter 5: Closing Remarks ………………………………………………………………137

Introduction ………………………………………………………………………….137

Chapter 2: The Impact of Urogenital Infections on the Gut and Vaginal Microbiome in Pregnancy …………………………………………………………………………….137

Chapter 3: hAMRoaster: A Tool for Comparing Performance of AMR Gene Detection Software ……………………………………………………………………………… 138

Chapter 4: Frequency of Antimicrobial Resistance Genes in a Pregnancy Cohort for Those with and without Urogenital Infections During Pregnancy ……………………139

Discussion …………………………………………………………………………….140

Limitations ……………………………………………………………………………142

Implications for Future Research and Practice ……………………………………….142

Table 1: Key Findings ………………………………………………………..144

References ……………………………………………………………………………145

About this Dissertation

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School	Laney Graduate School
Department	Nursing
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Health Sciences, Nursing Biology, Microbiology
Stichwort	pregnancy microbiome bioinformatics AMR antibiotics
Committee Chair / Thesis Advisor	Timothy Read, Emory University Vicki Hertzberg, Emory University
Committee Members	Anne Lang Dunlop, Emory University Michelle Wright, The University of Texas at Austin

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