Date of Award

2025

Document Type

Thesis

Degree Name

Master of Science (MS)

Program

Biomedical Sciences

Track

Molecular and Systems Pharmacology

Research Advisor

Colleen Jonsson

Committee

Amber Smith; Michael Whitt

Keywords

Hantavirus, Next Generation Sequencing, Vaccines, Vesicular stomatitis virus pseudotype system, Virus characterization, Virus phenotypes

Abstract

The genus Orthohantavirus, is a group of tripartite, negative-sense RNA viruses of which some members are the etiologic agents for hemorrhagic fever with renal syndrome (HFRS) or orthohantavirus cardiopulmonary syndrome (HCPS) in the Old and New Worlds, respectively. Sampling of wild rodents for these viruses has increased our knowledge of their host range and continues to lead to the discovery of new orthohantaviruses species and strains. HFRS and HPS have case fatality rates of up to 15% and 40%, respectively. And in some years, morbidity rates of up to 100,000 cases can be reached, globally. The epidemiological evidence of human-to-human transmission of the South American orthohantavirus, Andes virus (ANDV), suggests the pandemic potential of orthohantaviruses. Orthohantaviruses persist in their rodent hosts for their lifetime without apparent disease although fecundity has been noted. Ecological studies have shown there is a higher odds of male rodent infection than female rodent infection, and epidemiological studies show this likelihood in humans too. While literature suggest sex-based differences, the mechanism driving this is not fully understood. We also do not know how sex might influence the wildlife epidemiology and spread to humans In this thesis, I present an amplicon-based, multiplex RT-PCR and next-generation sequencing strategy that I used to characterize the Juquitiba virus (JUQV) harbored by Oligoryzomys, from rodent field samples collected in Paraguay. Using Bayesian inference and using the sequences that I and others from the Jonsson laboratory obtained, I estimated spatial and temporal evolutionary parameters using BEAST. The results suggest that the JUQV population circulating in rodents in Paraguay is under-sampled. The JUQV population characteristics of the viral genome differ and cluster by geographical location. The BEAST program estimated that the S segments of JUQV have higher substitution and mutation rates than the M segment and that the ancestor of the S segment is more recent (2004) than that of the M segment (1950). Moreover, I found that my analyses suggest that the founder virus is located in the Caaguazú Department of Paraguay. Further sampling will be necessary to verify this proposition. Pseudotyping studies of the JUQV glycoprotein (GP) using the Vesicular Stomatitis Virus (VSV) system were used to investigate the phenotypic consequences of several mutations that showed a negative selecting pressure using a DataMonkey Server that examined all JUQV GP sequences available (unpublished results of Dr. Brianna Spruill-Harrell). Preliminary studies suggest that Q292H and V504I reduced the number of cells infected by each pseudotype when compared to wild (WT) VSV pseudotype, however, only V504I VSV pseudotypes led to a statistically significant reduction in the number of cells. Lastly, I present an additional research approach to accelerate primer design so as to obtain a full reference genome of novel orthohantaviruses. In these studies, I used both vRNA and cRNA primers to amplify the segment from rodent tissue and identify an initial reference genome which is presumably composed of vRNA, cRNA and mRNA. This research will help to accelerate primer design to study the vRNA which is in the virion and transmitted from animal to animal or animal to human. Finally in this thesis, I led review of the research efforts directed toward vaccine development for the prevention of orthohantavirus infection. This research shows that, overall, translation to clinical effectiveness of and pan-protection by vaccine candidates in the human population remains the biggest challenge.

ORCID

0009-0009-8907-7218

DOI

10.21007/etd.cghs.2025.0698

Download

Available for download on Friday, June 05, 2026

Included in

Virology Commons

Share

COinS