Analysis of Human B Cell Development and Antibody Responses Following Immunization with a Live Attenuated Cholera Vaccine Open Access

Adekunle, Oluwaseyi (Summer 2021)

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B cells are important mediators of protection that provide immunity against infectious pathogens like Vibrio cholerae. Both infection and the inactivated oral cholera vaccines (OCV) generate humoral responses, however infection provides durable immunity and the OCV do not. To better understand the immune factors that are important for the generation and maintenance of cholera immunity, we utilized a live attended cholera vaccine as a model of V. cholerae exposure. First, we examined the kinetics of serum and plasmablasts responses following vaccination. We observed that the vaccine induced durable antibody titers that were maintained one year after vaccination. We found that the plasmablast responses were primarily specific for the two immunodominant antigens lipopolysaccharide (LPS) and cholera toxin (CT). The acute responses displayed preferential immunoglobulin isotype usage, with LPS specific cells being largely IgM or IgA producing, while CT responses were predominantly IgG. CCR9 was highly expressed on vaccine induced plasmablasts, especially on the LPS specific cells, suggesting a role in migration to the gastrointestinal tract. Finally, using single cell approaches we observed that the LPS specific plasmablasts also displayed hallmarks of ongoing affinity maturation. Next, we examined the properties of LPS specific antibodies and how immunoglobulin isotype and subclass effects their functional attributes. To approach this, we utilized a panel of monoclonal antibodies that we expressed in different immunoglobulin backbones. We then characterized the impact of these changes on their binding and functional capabilities. We observed that isotype had a profound impact by increasing the low-affinity antibodies binding and functional properties. We found that pentameric IgM, dimeric IgA, and monomeric IgA were more potent than their IgG counterparts at inhibiting motility. Finally, by analyzing the F(ab) versions of these antibodies, we showed that LPS crosslinking was essential for motility inhibition. Collectively, these findings provide key insights into cholera specific B cells development, the role of mucosal homing, and mechanisms of antibody mediated immunity. We believe that these findings will help inform future vaccine development by highlighting aspects of the response that are associated with protection and raise important questions on the relationship between peripheral and mucosal humoral responses. 

Table of Contents

Abstract iii

Chapter 1: Introduction. 1

B cell Immune responses. 2

Early B Cell Development 2

Activation of B cells. 4

Germinal Center Reaction. 6

Antibody Secreting Cells. 7

Memory B cells. 9

Response to non-protein antigens. 10

Antibodies, functional response of B cells. 11

Intestinal B cell responses. 13

Cholera. 16

Pathogenesis of V. cholerae. 16

Immune response to cholera disease. 18

Mechanisms of protection. 22

Live attenuated vaccine. 24

Whole cell killed vaccines. 26

Summary. 30

Chapter 2: Longitudinal analysis of human humoral responses after vaccination with a live attenuated V. cholerae vaccine. 32

Abstract 33

Introduction. 34

Results. 36

Discussion. 44

Materials and Methods. 52

Acknowledgements. 56

Figures and legend. 57

Chapter 3: Gut homing plasmablasts induced by Vibrio cholerae vaccination exhibit hallmarks of affinity maturation. 72

Abstract 73

Introduction. 74

Results. 75

Discussion. 82

Materials and Methods. 89

Acknowledgements. 94

Figures. 95

Chapter 4: Immunoglobulin isotype and epitope potently influence the functional properties of Vibrio cholerae O-specific polysaccharide antibodies. 111

Abstract 113

Introduction. 115

Results. 119

Discussion. 129

Materials and Methods. 135

Figures and Legends. 147

Chapter 5: Discussion. 167

Summary. 168

Future Directions. 170

Conclusion. 175

References. 176

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