risk factors for antibody loss after hepatitis e virus natural infection and vaccination

266 Pages·2016·18.93 MB·English

by  David Reynolds

RISK FACTORS FOR ANTIBODY LOSS AFTER HEPATITIS E VIRUS NATURAL

INFECTION AND VACCINATION

by
Brittany Lynn Kmush, ScM

A dissertation submitted to Johns Hopkins University in conformity with the
requirements for the degree of Doctor of Philosophy.

Baltimore, Maryland
June, 2016

© 2016 Brittany Kmush
All Rights Reserved
ABSTRACT
Hepatitis E virus (HEV) is a vaccine-preventable emerging infectious disease causing 20
million infections in developing countries every year. In South East Asia, HEV causes yearly
outbreaks, with the majority of disease seen in adults, surprising for an enterically transmitted
pathogen. These outbreaks suggest that antibody persistence after exposure to HEV is not long
lasting. We revisited 170 subjects with a documented HEV infection from Bangladesh and China
and 97 subjects vaccinated with the HEV239 vaccine during a phase III trial in China to retest
their serum for anti-HEV antibodies 6 to 10 years after exposure. Overall, 22.1% (95%CI: 17.3-
27.6%) no longer had detectable antibodies at follow-up. Antibody loss was greater among the
naturally infected subjects compared to the vaccinated subjects, 24.1% (95%CI: 17.9-31.3%)
versus 18.6% (95%CI: 11.4-27.7%), although not statistically significant (p=0.292). Among all the
subjects, age at exposure was associated with antibody loss, with younger age increasing the
risk of antibody loss (RR: 0.87 per 10 years, 95% CI: 0.76-1.00, p=0.057). Among the subjects
from Bangladesh, each 10 year increase in age at infection decreased the risk of antibody loss by
50% across univariate and multivariate Poisson regression models (p<0.05). This age-dependent
antibody loss could partially explain cross-sectional sero-prevalence data from South East Asia
where children have reportedly low antibody prevalence. In multivariate models, factors that
increased the risk of exposure to HEV were generally associated with antibody persistence
among the naturally infected subjects. However, this pattern was not found in the vaccinated
subjects. This is one of the first studies to compare long term antibody persistence after HEV
exposure in both naturally infected and vaccinated individuals, exploring characteristics
associated with antibody persistence. The development of a successful, subunit vaccine has
increased the need to understand the duration of antibodies and protection after HEV infection
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and vaccination in order to implement the most cost effective disease control and vaccination
strategies.

PhD Advisor: Alain B. Labrique, Associate Professor
Department of International Health

Dissertation Readers: Kenrad E. Nelson, Professor
Department of Epidemiology

Christian Coles, Associate Professor
Department of International Health

Christopher Heaney, Assistant Professor
Department of Environmental Health Sciences

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ACKNOWLEDGEMENTS

So many people contributed to the successful completion of this dissertation. I want to
acknowledge the “village” that guided and supported me.
Dr. Alain Labrique, my academic advisor, was my most significant mentor during my
time at Johns Hopkins. He guided me through both a master’s and doctorate degree, constantly
pushing me. He has treated me more like a colleague than a student and fostered a sense of
independence and confidence in myself and my research, preparing me for the next phase of my
journey. I would also like to thank the other members of my academic committees, Dr. Kenrad
Nelson, Dr. Chris Heaney and Dr. Chris Coles, for their advice, guidance, and encouragement.
This research would not have been possible without the support of the faculty and staff
at Xiamen University and the Jiangsu Centers for Disease Control Prevention. I would like to
acknowledge Dr. James Shih, whose persistence and encouragement gave me the opportunity
to travel to China and visit the site of one of the largest vaccine trials in history. Dr. Wu Ting not
only ensured that the day to day activities of the research were completed but also that I was
comfortable and supported in China. I also cannot thank Yu Huan and Shu Chen enough for
being my guides and friends during my time in China.
Another significant partner in this research was the icddr,b at both the Dhaka
Headquarters and Matlab Health Research Station. I want to thank Dr. K Zaman and everyone
at the Centre for Child and Adolescent Health. Dr. Zaman supported and guided me throughout
the entire research process. I also want to thank Dr. Asma Binte Aziz (also called Dr. Tuly) and
Subarna Barua for taking me under their wings and being my support system while I was in
Bangladesh.
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I also want to thank all of the faculty and staff at the JiVitA Maternal and Child Health
Research Project, even though the results from this site did not make it into the dissertation. Dr.
Saijuddin Shaikh was vital to this research, overseeing the day to day activities of this study.
Sucheta Mehra helped me navigate the IRBs in both Bangladesh and at JHSPH, a process which
proved much more difficult than I would have imagined. I also want to acknowledge Dr. Kerry
Schulze in the Center for Human Nutrition and all the staff in her lab. Dr. Schulze saw potential
in a first year master’s student and offered me a job, which opened countless doors for me. She
has been an invaluable mentor and role model for me throughout my time at JHSPH. Her
guidance was a major contributor to my growth over the years.
Last, but certainly not least, I would like to thank all of my friends and family,
particularly Johnny, my parents, and my in-laws, for all of their love and support through this
long process. From kind words of encouragement, to help with editing, to countless cups of
coffee (with the occasional glass of wine), this would not have been possible without you and I
cannot thank you enough!

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TABLE OF CONTENTS

List of Tables ......................................................................................................................... ix
List of Figures ......................................................................................................................... x

1. Introduction ....................................................................................................................... 1
A. Research Summary .................................................................................................................. 2
1. Overview of Research .......................................................................................................... 2
2. Research Timeline ................................................................................................................ 5
B. Collaborations .......................................................................................................................... 6
1. International Centre for Diarrheal Disease Research, Bangladesh (icddr,b) ....................... 6
2. National Institute of Diagnostics and Vaccine Development in Infectious Diseases
(NIDVD), School of Public Health, Xiamen University .............................................................. 6
C. Funding .................................................................................................................................... 8
D. Ethical Considerations ............................................................................................................. 9
1. Review and Approval ........................................................................................................... 9
2. Study Overview .................................................................................................................... 9
3. Study Procedures ............................................................................................................... 10

2. Research Context ............................................................................................................. 14
A. Health Indicators ................................................................................................................... 15
1. Overview of health in Bangladesh ..................................................................................... 15
2. Overview of health in China ............................................................................................... 17
B. Research Sites and Previous Studies ..................................................................................... 19
1. Matlab Health Research Station ........................................................................................ 19
2. HEV239 Vaccine Trial ......................................................................................................... 20
C. Overview of the Hepatitis E Vaccine and Current Recommendations .................................. 22

3. Literature Review ............................................................................................................. 24
A. Epidemiology of Hepatitis E ................................................................................................... 25
1. Genotype 1 and 2 Endemic Areas ...................................................................................... 25
2. Genotype 3 and 4 Endemic Areas ...................................................................................... 30
B. Virology .................................................................................................................................. 34
C. Immune response to infection ............................................................................................... 36
1. Overview of Immune System ............................................................................................. 36
2. Immune Response to HEV infection .................................................................................. 39
D. Clinical Presentation .............................................................................................................. 40
E. Prevention and Control .......................................................................................................... 43
1. Overview of Vaccination Theory and Methodology .......................................................... 43
2. GSK Vaccine Candidate ...................................................................................................... 44
3. Innovax Vaccine Candidate ................................................................................................ 45
F. Persistence of Antibodies ....................................................................................................... 46
1. Antibody Duration and Protection after infection ............................................................. 46
2. Antibody Duration and Protection after Vaccination ........................................................ 50

4. Two generations of “Gold Standards:” The impact of a decade in hepatitis E virus testing
innovation on population seroprevalence ............................................................................. 54
A. Abstract .................................................................................................................................. 55
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B. Key Words .............................................................................................................................. 55
C. Introduction ........................................................................................................................... 56
D. Methods ................................................................................................................................ 57
E. Results .................................................................................................................................... 58
F. Discussion ............................................................................................................................... 59
G. Acknowledgments ................................................................................................................. 61
H. Conflicts of Interest ............................................................................................................... 61
I. Funding ................................................................................................................................... 61

5. Prevalence of and Risk Factors for Antibody Loss after Hepatitis E Virus Infection in Rural
Bangladesh .......................................................................................................................... 62
A. Abstract .................................................................................................................................. 63
B. Key Words .............................................................................................................................. 64
C. Introduction ........................................................................................................................... 65
D. Methods ................................................................................................................................ 67
1. Participant Selection .......................................................................................................... 67
2. Hepatitis E Virus Antibody Testing ..................................................................................... 68
3. Statistical Methods ............................................................................................................ 70
E. Results .................................................................................................................................... 73
F. Discussion ............................................................................................................................... 77
G. Acknowledgements ............................................................................................................... 81
H. Conflicts of Interest ............................................................................................................... 82
I. Funding ................................................................................................................................... 82

6. Long Term Antibody Persistence after Hepatitis E Infection and Vaccination in China ......... 83
A. Abstract .................................................................................................................................. 84
B. Key Words .............................................................................................................................. 84
C. Introduction ........................................................................................................................... 85
D. Methods ................................................................................................................................ 87
1. Participant Selection and Enrollment ................................................................................ 87
2. Hepatitis E Virus Antibody Testing ..................................................................................... 88
3. Statistical Methods ............................................................................................................ 89
E. Results .................................................................................................................................... 92
1. Naturally Infected .............................................................................................................. 92
2. Vaccinated .......................................................................................................................... 93
3. Overall ................................................................................................................................ 95
F. Discussion ............................................................................................................................... 95
G. Acknowledgements ............................................................................................................... 98
H. Conflicts of Interest ............................................................................................................... 98
I. Funding ................................................................................................................................... 98

7. Risk Factors for Antibody Loss after Hepatitis E Virus Infection and Vaccination: Results of a
Multi-site Cohort Study ...................................................................................................... 100
A. Abstract ................................................................................................................................ 101
B. Key Words ............................................................................................................................ 101
C. Introduction ......................................................................................................................... 102
D. Methods .............................................................................................................................. 105
1. Participant Selection ........................................................................................................ 105
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2. Hepatitis E Virus Antibody Testing ................................................................................... 108
3. Statistical Methods .......................................................................................................... 109
E. Results .................................................................................................................................. 113
1. Follow-up ......................................................................................................................... 113
2. Univariate Analysis ........................................................................................................... 114
3. Multivariate Analysis ........................................................................................................ 116
F. Discussion ............................................................................................................................. 118
G. Acknowledgements ............................................................................................................. 123
H. Conflicts of Interest ............................................................................................................. 123
I. Funding ................................................................................................................................. 123

8. Final Discussion .............................................................................................................. 125
A. Conclusions .......................................................................................................................... 126
B. Remaining questions ............................................................................................................ 130
C. Limitations ........................................................................................................................... 134
D. Strengths.............................................................................................................................. 137

References Part I: Tables .................................................................................................... 139
References Part II: Figures .................................................................................................. 161
References Part III: Appendices .......................................................................................... 175
Appendix A: HEV Exposure Assessment Questionnaire-Combined (For Analysis Purposes Only)
................................................................................................................................................. 176
Appendix B: HEV Exposure Assessment Questionnaire-Matlab (English) ............................... 191
Appendix B1: HEV Exposure Assessment Questionnaire-Matlab (Bangla) .............................. 202
Appendix C: HEV Exposure Assessment Questionnaire-China (English).................................. 214
Appendix C1: HEV Exposure Assessment Questionnaire-China (Mandarin Chinese) ............. 222
References Part IV: Literature Cited .................................................................................... 231

Brief Curriculum Vitae ........................................................................................................ 253

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LIST OF TABLES

Table 1: Description of the four HEV genotypes that infect humans .......................................... 140
Table 2: Comparison of the two anti-HEV immune-assays in rural Bangladesh (n=1009) (2004-
2005). ................................................................................................................................... 141
Table 3: Comparison of WRAIR anti-HEV total Ig assay with Wantai anti-HEV IgG assay by age
group in participants from rural Bangladesh for the 1009 participants that had both tests
completed (2004-2005). ...................................................................................................... 142
Table 4: Summary of Participants included in Hepatitis E Antibody Persistence Study (2015). . 143
Table 5: Comparison between Revisited and Lost to Follow Up Participants in Hepatitis E
Antibody Persistence Study in Matlab, Bangladesh (2015). ................................................ 144
Table 6: Comparison of the two anti-HEV immune-assays in rural Bangladesh from banked sera
in participants in rural Bangladesh (2004-2005). ................................................................. 145
Table 7: Demographic Characteristics by Antibody Persistence status in Rural Bangladesh(2015).
............................................................................................................................................. 146
Table 8: Results of Univariate and Multivariate Poisson Regression Models for Risk Factors for
antibody loss after HEV Infection. ....................................................................................... 148
Table 9: Comparison between Revisited and Lost to Follow Up in in Hepatitis E Antibody
Persistence Study, Dongtai, China (2015). ........................................................................... 150
Table 10: Demographic Characteristics by Antibody Persistence and Vaccination Status in
Dongtai, China (2015). ......................................................................................................... 151
Table 11: Demographic and Exposure Risk Factors for loss of HEV antibodies at follow-up for the
entire cohort in Dongtai, China (n=168) (2015). .................................................................. 154
Table 12: Results of Univariate and Multivariate Poisson Regression Models for Risk Factors for
antibody loss after HEV exposure in Dongtai, China (2015). ............................................... 156
Table 13: Demographic and Exposure Risk Factors for loss of HEV antibodies at follow-up (2015).
............................................................................................................................................. 157
Table 14: Self-reported Illness Symptoms within the Last 10 years by Antibody Persistence Status
(2015). .................................................................................................................................. 159
Table 15: Results of Multivariate Poisson Regression Models for Risk Factors for antibody
persistence after HEV exposure (2015). .............................................................................. 160

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LIST OF FIGURES

Figure 1: Map of the Matlab Study Area showing icddr,b and government service areas. ........ 162
Figure 2: Map of Dongtai County, Jiangsu Province, China. ........................................................ 163
Figure 3: The hepatitis E virus (HEV) genome. ............................................................................. 164
Figure 4: Course of acute hepatitis E virus (HEV) infection. ........................................................ 165
Figure 5: Cumulative Hazard of Hepatitis E. ................................................................................ 166
Figure 6: Cumulative incidence of hepatitis E. ............................................................................. 167
Figure 7: IgG anti-HEV levels induced by natural infection. ........................................................ 168
Figure 8: Population anti-HEV seroprevalence by age in Matlab, Bangladesh, a comparison of the
WRAIR (n = 1,025) and Wantai (n = 1,009) assays (2004-2005). ......................................... 169
Figure 9: Cohort Selection and Follow-up Diagram. .................................................................... 170
Figure 10: Box Plot of Age by antibody persistence status in rural Bangladesh (2015). ............. 171
Figure 11: Antibody loss by age group in rural Bangladesh 2015 (n=100). ................................. 172
Figure 12: Observed and predicted anti-HEV negative percentage by age-group (n=1009)....... 173
Figure 13: Conceptual Model of Antibody Persistence to Hepatitis E Virus (HEV), among those
already with HEV antibodies, from either vaccination or infection. ................................... 174

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