Transmission Ecology and Zoonotic Potential of the Emerging Tick-borne Heartland virus in Georgia Restricted; Files Only

Abstract

Heartland virus (HRTV) is an emerging tick-borne bunyavirus that has been associated with over sixty severe and fatal human infections since its discovery in Missouri in 2009. HRTV is understudied and little is known about its transmission ecology, persistence in the environment, and true burden of infection and disease across the US. These knowledge gaps make it difficult to characterize the threat HRTV poses to humans.

 

Applying a cross-scales approach to understanding key components of HRTV dynamics, this dissertation begins to address unknowns in HRTV knowledge in Georgia by: 1) estimating the distribution of Amblyomma americanum on a fine spatial scale, 2) evaluating phylogenetic relationships between collected HRTV-positive tick samples and reference sequences, and 3) developing and applying an assay to assess the seroprevalence of HRTV in Georgia residents.

 

Specifically, Chapter 1 of this dissertation provides an introduction to the importance of studying vector-borne diseases, highlighting current knowledge on HRTV and Amblyomma americanum.

 

Chapter 2 focuses on characterizing the ecologic niche of Amblyomma americanum across the state highlighting areas of increased probability of encountering these ticks. Through development of a logistic regression model of important climate, elevation, and vegetation factors based on field sampling from forty-three natural areas, we project the probability of finding these ticks across the state at a ~1 km resolution. Areas with the highest probability include a band of central to south-central Georgia and the coastal region.

 

Chapter 3 examines a hotspot of HRTV in central Georgia through identification and full viral genome sequencing of HRTV-positive ticks across five years. Phylogenetic analyses reveal strong genetic conservation across time with samples >99.7% related across years. Additional HRTV-positive ticks were detected in 2023 ~175 km away but still show high genetic similarity to the other Georgia-collected samples. Reference sequences downloaded from GenBank representing human infections and ticks from across the US also assort geographically, highlighting spatial clustering across the country. This phylogenetic analysis provides preliminary evidence implicating small geographic foci of HRTV as dominating transmission ecology.

 

Chapter 4 pivots to understanding the current HRTV burden of infection in human populations, examining seropositivity in Georgia residents. We optimize and establish a new HRTV IgG ELISA to screen human sera to detect infections in Georgia. After screening 201 randomly selected serum samples from the Atlanta area, no seropositivity was detected. This assay will be applied to other regions in ongoing and future seroprevalence investigations in Georgia.

 

Finally, Chapter 5 unifies findings from each chapter, discussing strengths, limitations, future directions, and overall conclusions to the work.

Table of Contents

Chapter 1. Introduction

1.1 The importance of studying vector-borne disease

1.2 Emerging tick-borne arboviruses and Heartland virus

1.3 Heartland virus transmission ecology

Figure 1.1. Theorized HRTV transmission cycle.

1.4 Modeling Amblyomma americanum distribution

Table 1.1. List of significant climate predictors in Amblyomma americanum habitat suitability modeling

1.5 Heartland virus phylogenetics and comparison with Dabie bandavirus

1.6 Seroprevalence studies in Heartland virus

1.7 Heartland virus in Georgia

Figure 1.2. Tick sampling sites in central Georgia 2018 and 2019

1.8 Ticks in Georgia

1.9. Dissertation Overview

Chapter 2: Mapping the Distribution of Amblyomma americanum in Georgia, USA

2.1 Abstract

2.2 Introduction

2.3 Methods

2.4 Results

Figure 2.1. Density of A. americanum at each site per 100 m² sampled

Figure 2.2. Percent of transects where A. americanum was found, split by habitat in each ecoregion

Table 2.1. Model predictors and coefficients for best logistic regression model

Table 2.2. Model performance

Figure 2.3. Probability of A. americanum occurrence across Georgia

Figure 2.4. Estimated ecological niche of A. americanum in Georgia

2.5 Discussion

2.6 Conclusions

2.7 Supplemental Material

Supplemental Table S2.1. Table of collinearity (VIF) in the training set

Supplemental Figure S2.1. Predicted effect of bioclimatic variable isothermality

Supplemental Figure S2.2. Map of isothermality (bc3/bioclim3) across Georgia

Chapter 3: Phylogenetic Evidence for Geographic Clustering and Persistence of Heartland Virus in a Transmission Hot-spot in Georgia

3.1 Abstract

3.2 Introduction

3.3 Methods

3.4 Results

Table 3.1. Species and life stage of ticks collected during field sampling from 2021-2023

Table 3.2. Heartland positive pool characteristics

Figure 3.1. Map of collection sites visited from 2021-2023

Table 3.3. Pairwise genetic distances between HRTV sequences in Georgia

Table 3.4: Pairwise genetic distances between all HRTV sequences in the US

Figure 3.2: Maximum-likelihood phylogenetic trees of HRTV and US map of sequence locations

3.5 Discussion

3.6 Conclusions

3.7 Supplemental Material

Supplemental Table S3.1. Extraction procedure

Supplemental Table S3.2. Heartland RT-qPCR assay information

Supplemental Table S3.3. Tick actin RT-qPCR assay information

Supplemental Table S3.4. Custom primer sequences for the HRTV amplicon primer set

Supplemental Figure S3.1. Temporal residual histogram of HRTV L segments

Chapter 4: Development and Implementation of a Heartland Virus IgG Enzyme-linked Immunosorbent Assay for Seroprevalence Screening

4.1 Abstract

4.2 Introduction

4.3 Methods

Figure 4.1. Heartland sandwich IgG ELISA diagram

Figure 4.2. Example plate map

4.4 Results

Table 4.1. Titration trial dilution results

Table 4.2. Final titration plate trial

Table 4.3. Negative control serum results

Figure 4.3. Histogram of human HRTV IgG optical density results

Table 4.4. Human serum potential positives readings

4.5 Discussion

4.6 Conclusions

4.7 Supplemental Material

Supplemental Table S4.1. Plate read for monoclonal (mAb 2BB5) antibody titration trial

Supplemental Table S4.2. Plate read for secondary conjugated antibody titration trial

Supplemental Table S4.3. Plate read for HRTV positive control serum and HRTV antigen trial

Supplemental Table S4.4. Plate read for COPE serum negative control trial

Supplemental Table S4.5. Vitamin D testing results for samples 1-13

Chapter 5. Discussion

5.1 Summary of findings

5.2 Strengths and limitations

5.3 Future Directions

5.4 Conclusions

6 References

About this Dissertation

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School	Laney Graduate School
Department	Environmental Health
Degree	Ph.D.
Submission	Dissertation
Language	English
Research Field	Health Sciences, Public Health Environmental Health Biology, Ecology
关键词	Field work Phylogenetics ELISA Vector-borne disease Lone star tick Heartland virus Serology Species distribution map
Committee Chair / Thesis Advisor	Vazquez-Prokopec, Gonzalo, Emory University Piantadosi, Anne, Emory University
Committee Members	Kitron, Uriel, Emory University Collins, Matthew, Emory University

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