Virology Commons^™

Identifying The Role Of Human Dna Polymerase Delta During Human Cytomegalovirus Genome Synthesis, Rachel Laree Dansereau

Dartmouth College Master’s Theses

Human cytomegalovirus (HCMV) is a large double stranded DNA virus and member of the Herpesviridae family. Like all herpesviruses, HCMV presents with characteristics of persistent, lifelong infections that transition between lytic and latent states. The replication of the viral genome is traditionally thought to occur by a rolling circle mechanism, but this model is supported by little evidence. It is known that HCMV actively recruits several host proteins to viral replication compartments (RCs), including proteins involved in the DNA damage response (DDR). The exact mechanisms of how these proteins are used and in what ways they help the virus remain …

Exploring The Application Of Wastewater Monitoring To Determine The Prevalence And Diversity Of Influenza On The Campus Of West Virginia University, Lakyn Renee Sanders

Graduate Theses, Dissertations, and Problem Reports (ETD)

Influenza viruses (IV) affect populations globally, with the World Health Organization (WHO) estimating that IV are responsible for an estimated 3-5 million infections and 290,000 – 650,000 deaths annually^1–3. More specifically, the 2019-2020 IV season is associated with an estimated 38 million illnesses and 22,000 deaths in the United States alone⁴. IV are rapidly spread between individuals through direct contact with infected individuals, contaminated surfaces, or aerosolized virus particles². Traditional IV infection monitoring relies on clinical screening of patients presenting influenza-like symptoms, commonly through the use of multi-target panel tests such as the BioFire …

Assessing The Efficiency Of Methotrexate (Mtx) As An Antiviral Drug Against Gammaherpes Virus Replication, Yennifer A. Gaspar Garcia

Honors Projects

It is estimated that ~15% of all cancers are caused by oncogenic virus infections. Two of the top seven cancer-causing human viruses are members of the gammaherpesvirus family: Epstein Barr Virus (EBV) and Kaposi’s Sarcoma Herpesvirus (KSHV). Our lab uses Murine Herpesvirus 68 (MHV-68), a mouse gammaherpesvirus with shares significant genetic homology to KSHV and EBV, as a model system to understand how gammaherpesviruses alter the metabolism of their host during lytic infection to promote their replication. We recently metabolically profiled MHV-68 infected host cells at various time points during the lytic infectious cycle. Our data showed nucleotide metabolism is …

Bibliometric Analysis Of Research Trends Related To Human Immunodeficiency Virus., Madyson E. Johnson, Kaitlyn R. Coffman, Valerie Rivas, Elizabeth Barrett

ATU Scholars Symposium

Human immunodeficiency virus poses major health burden across the globe including the United States of America and has its zoonotic origin from the Simian Immunodeficiency viruses of sooty mangabey monkey and chimpanzee in Africa. Epidemiological trends indicate that even though the mortality rates for HIV are falling globally, the rates of new cases have been rising since 2010 in several countries including the USA. An estimated 0.5% of the human population is currently infected with the virus with countries like USA, India, Nigeria and South Africa carrying the highest burden of the disease. Host receptors such as CD4 and coreceptors …

Elucidating The Mechanism Of Interleukin-17a Signaling In The Pathogenesis Of West Nile Virus, Farzana Nazneen

Dissertations

West Nile Virus (WNV), a neurotropic flavivirus transmitted by mosquitoes, is a leading cause of viral encephalitis in the United States and remains a significant public health concern. Unfortunately, no vaccine or therapeutic intervention is available against WNV infection. Previous studies, including ours, demonstrated that interleukin-17A (IL-17A) signaling increases the cytotoxic potential of CD8⁺ T cells to facilitate WNV and parasite clearance; however, the molecular mechanism is not understood. IL-17 receptor C (IL-17RC) is an obligatory co-receptor with IL-17 receptor A (IL-17RA) for signaling induced by IL-17A, IL-17A/F, and IL-17F. In this study, we found that IL-17RC deficient ( …

Manipulation Of Cellular Proliferating Cell Nuclear Antigen (Pcna) During Herpes Simplex Virus Type 1 Infection, Jessica Packard, Jill A. Dembowski

Electronic Theses and Dissertations

Herpes simplex virus type 1 (HSV-1) is a double stranded DNA virus that replicates within the nucleus of host cells. Using accelerated native isolation of proteins on nascent DNA (aniPOND), cellular proteins were found to associate with HSV-1 replication forks. One host factor of interest is proliferating cell nuclear antigen (PCNA), the DNA sliding clamp that tethers DNA polymerases and DNA damage response proteins to replicating cellular DNA. Although HSV-1 encodes its own DNA polymerase processivity factor (UL42), PCNA plays a key role in HSV-1 infection. We used commercially available PCNA inhibitors and short hairpin RNAs (shRNA) to investigate how …

Functional And Structural Analysis Of The Neimann-Pick Disease Type C Pathway To Include Caveolin-1, Anthony Michael Seat

Chemistry and Chemical Biology ETDs

Human disease is often thought of as an all or nothing prospect, either one has the disease or one does not. This does not bear out in clinical or personal experiences, instead demonstrating that disease occurs within a spectrum ranging from presumed unaffected to demonstrably and detrimentally affected.Neimann-Pick disease is one example of this spectrum look into diseased states, with multiple named versions of a phenotypically similar disease. We focus on Neimann-Picktype C (NPC), which is the result of a disruption in the efflux of cholesterol and sphingolipids from the endocytic pathway. NPC demonstrates this concept of a spectrum of …

Revolutionizing Our Understanding Of Specific Platelet-Pathogen Interactions, Hammodah Rawhi Hammodah Alfar

Theses and Dissertations--Molecular and Cellular Biochemistry

Aside from their classical role in maintaining hemostasis, platelets are considered first-line vascular guardians that influence both the innate and adaptive immune systems. This is mediated by either direct interactions with immune cells or through the release of pre-stored cytokines and chemokines upon interacting with pathogens such as viruses. While this establishes platelets as potential players in the response to pathogens, it remains unclear what platelets can do with pathogens and how important that is to immune responses, especially against viruses. Most systemic viremias, such as HIV-1, present with mild thrombocytopenia, which is mediated through several mechanisms such as a …

Role Of Autophagy-Related Genes In Lipid Transport For Efficient Replication Of Tombusviruses, Yuanrong Kang

Theses and Dissertations--Plant Pathology

Tombusviruses are positive-strand RNA viruses, which utilize numerous co-opted host factors and massive de-novo synthesized membrane resources to modify host membranes and form large viral replication organelles (VROs). Among plenty of host cellular processes, autophagy emerges as a potential membrane resource for VROs formation due to its ability to de-novo generate the double membrane structures, known as autophagosomes. Besides, as an important host defense pathway, autophagy is also a target for tombusviruses to prevent its antiviral activities. In this study, I identify the role of key autophagy related genes, Atg11, Atg8, and Atg2, which are involved in autophagosome formation, and …

Understanding Cellular Dynamics: Investigating A Yeast-Two Hybrid Protein-Protein Interaction, Riok3:Cse1l; Also By Investigating Trogocytosis Of Cd4+ T Cells From Rhog-/- Mice, Brittnee N. Crane

Graduate Student Theses, Dissertations, & Professional Papers

Rift Valley fever virus (RVFV) is a negative-sense virus that was first described in 1930 when it was isolated from sheep during an outbreak. In the last 25 years there has been a spread in the geographical range into the Middle East. The most susceptible animals are cattle, goats, sheep, and camels. The most overt symptom of livestock infection is that infected pregnant animals experience spontaneous abortion. Humans are susceptible as well and symptoms range from mild febrile illness to liver necrosis, blindness, hemorrhagic fever, and death. An important player in the cellular innate response to RVFV infections is the …

Potent Bile Acid Microbial Metabolites Modulate Clostridium Perfringens Virulence, Tahrir Jatli Alenezi, Ying Fu, Bilal Alrubaye, Thamer Alanazi, Ayidh Almansour, Hong Wang, Xiaolun Sun

Poultry Science Faculty Publications and Presentations

Clostridium perfringens is a versatile pathogen, inducing diseases in the skin, intestine (such as chicken necrotic enteritis (NE)), and other organs. The classical sign of NE is the foul smell gas in the ballooned small intestine. We hypothesized that deoxycholic acid (DCA) reduced NE by inhibiting C. perfringens virulence signaling pathways. To evaluate the hypothesis, C. perfringens strains CP1 and wild-type (WT) HN13 and its mutants were cultured with different bile acids, including DCA and isoallolithocholic acid (isoalloLCA). Growth, hydrogen sulfide (H₂S) production, and virulence gene expression were measured. Notably, isoalloLCA was more potent in reducing growth, H …

Analyzing Pseudomonas Aeruginosa With Bacteriophage Tags Using Photoacoustic Flow Cytometry, Jennifer C. Schinke

Electronic Theses and Dissertations

The number of daily bacterial infections is climbing and the CDC explains that this is due to the antibiotic-resistant threat in the United States. Finding a faster way of bacterial identification is necessary as it currently takes 1-4 days for a medical lab to culture and identify bacteria. Photoacoustic flow cytometry (PAFC) can be used as an alternative method resulting in swift identification within an hour (Edgar, 2019). Pseudomonas aeruginosa, cell line PA01, will be coated in up to a few hundred red dyed phages making it detectible by the photoacoustic flow cytometry system. Bacteriophages (phages) are viruses that …

The Role Of Adenovirus Serotype 5 E4 11k In The Relocalization Of The Cellular P Body Proteins, Kasey A. Karen, La`Quita Randolph, Kevin Neubrecht, Heather Vincent

Georgia Journal of Science

Human adenoviruses are a useful tool to understand basic cellular biology in addition to viral infections. Historically, cellular splicing was first discovered in adenovirus, but other cellular processes, such as double-strand break repair and aggresome formation, have been further elucidated through adenoviral infection. The adenovirus protein E4 11k has been shown to disrupt cytoplasmic processing bodies (p bodies), which are not well-understood but are involved in mRNA metabolism. Several p body proteins were found to be reorganized in the cytoplasm with adenovirus serotype 5 (Ad5) able to cause the colocalization of these p body proteins with aggresomes. The p body …

Development Of A Novel Fluorescent Reporter Baculovirus Vector For Testing Promoters In Lepidopterans, Daniel Howard

All Theses

Testing gene expression patterns is an important process in learning more about an organism. The standard methods of rtPCR and RNASeq provide highly detailed data on specific expression patterns, but can be resource consuming if many different samples including genes or organisms must be tested. A potential alternative for studying expression is the use of a reporter system carried by a vector system that encodes fluorescent proteins under the control of promoters of interest. Many lepidopterans (moths) are pestiferous and better understanding of gene expression levels in lepidopterans, both endogenous and exogenous including from their viruses, would be beneficial. Baculoviruses …

Proposing An Rna Interference (Rnai)-Based Treatment For Human Immunodeficiency Virus (Hiv) By Analyzing The Post-Transcriptional Gene Targeting Of Sars-Cov-2, Hepatitis C Virus, And A549 Lung Cancer Cells, Arjun Jagdeesh

Undergraduate Research Posters

Human Immunodeficiency Virus (HIV) is a retrovirus that infects CD4+ T cell lymphocytes in humans, leading to the development of Acquired Immunodeficiency Syndrome (AIDS) if left untreated. While current treatment methods, including antiretroviral combination treatments, effectively limit HIV replication, HIV can evade these treatments due to its high mutation rate. Long-term antiretroviral treatment can also be toxic to patients, meaning patients would benefit from a new mechanism of HIV treatment. RNA interference (RNAi) is an antiviral pathway found in mammals, plants, and insects that involves a small-interfering RNA that is incorporated into a protein complex called the RNA-induced Silencing Complex …

Resolving The Repression Pathway Of Virulence Gene Hila In Salmonella, Alexandra King, Lon Chubiz Phd, Brenda Pratte, Lauren Daugherty

Undergraduate Research Symposium

Salmonella is a relatively abundant, virulent species of bacteria that is most known for spreading gastrointestinal diseases through food. These illnesses result in approximately 1.35 million infections, including over 25,000 hospitalizations each year, in the U.S. alone (CDC.gov). As antibiotic resistance becomes an increasingly urgent public health problem, the importance of developing alternative treatment methods is only becoming more crucial. One of the genes responsible for this virulence is known as hilA. HilA is the main transcriptional regulator of Salmonella Pathogenicity Island-1 gene (UniProt). SPI-1 plays an important role in the invasion of Salmonella into epithelial cells. The proteins encoded …

Conformational Flexibility And Local Frustration In The Functional States Of The Sars-Cov-2 Spike B.1.1.7 And B.1.351 Variants: Mutation-Induced Allosteric Modulation Mechanism Of Functional Dynamics And Protein Stability, Gennady M. Verkhivker

Mathematics, Physics, and Computer Science Faculty Articles and Research

Structural and functional studies of the SARS-CoV-2 spike proteins have recently determined distinct functional states of the B.1.1.7 and B.1.351 spike variants, providing a molecular framework for understanding the mechanisms that link the effect of mutations with the enhanced virus infectivity and transmissibility. A detailed dynamic and energetic analysis of these variants was undertaken in the present work to quantify the effects of different mutations on functional conformational changes and stability of the SARS-CoV-2 spike protein. We employed the efficient and accurate coarse-grained (CG) simulations of multiple functional states of the D614G mutant, B.1.1.7 and B.1.351 spike variants to characterize …

Alternative Splicing Regulates The Innate Immune Response To Viral Infection, Luke A. White

Graduate Student Theses, Dissertations, & Professional Papers

Rift Valley fever virus (RVFV) is a mosquito-borne RNA virus that infects humans and livestock in sub-Saharan Africa and the Arabian peninsula, causing disease ranging from a mild flu-like illness to liver damage, blindness, hemorrhagic fever, death, and, especially in livestock animals, high rates of abortive pregnancies. There is no approved vaccine for RVFV, and as a disease with a high rate of spread that causes severe illness, it is listed as a Category A pathogen by the USA CDC. A better understanding of RVFV’s molecular virology will be instrumental to combating RVFV as climate change causes its mosquito host …

Investigation Of An Alternative Protocol For The Production Of Sars-Cov-2 Antigenic Proteins, Nichole Ninaltowski

USF Tampa Graduate Theses and Dissertations

With the COVID-19 pandemic showing no signs of slowing down, large-scale antigenic protein production is still needed for surveillance using serologic assays. From screening to vaccines to biotherapeutics, being able to produce the proteins for these assays is essential; however, the current gold standard method for producing SARS-CoV-2 spike proteins is prohibitively expensive for most research groups.

Alternative methods of transfecting mammalian cells to produce recombinant proteins that are relatively inexpensive have been used for years. Unlike the expensive, commercially available lipid-based methods, other established methods such as polyethyleneimine (PEI), are considerably easier, and cheaper to meet the needs of …

Role Of Endocytic Machinery Regulators In Egfr Traffic And Viral Entry, Insha Mushtaq

Theses & Dissertations

STUDY 1: Role of endocytic regulator EHD1 and its binding partner RUSC2 in EGFR traffic

Abstract

Epidermal growth factor receptor (EGFR) is a prototype receptor tyrosine kinase and an oncoprotein in many solid tumors. Cell surface display of EGFR is essential for cellular responses to its ligands. While post activation endocytic trafficking of EGFR has been well elucidated, little is known about mechanisms of basal/pre-activation surface display of EGFR. Here, we identify a novel role of the endocytic regulator EHD1 and a potential EHD1 partner, RUSC2, in cell surface display of EGFR. EHD1 and RUSC2 colocalize with EGFR in vesicular/tubular …

Entry And Replication Of Negative-Strand Rna Viruses, Kerri Boggs

Theses and Dissertations--Molecular and Cellular Biochemistry

Hendra virus (HeV) and human metapneumovirus (HMPV) are negative-sense, singled-stranded RNA viruses. The paramyxovirus HeV is classified as a biosafety level 4 pathogen due to its high fatality rate and the lack of a human vaccine or antiviral treatment. HMPV is a widespread pneumovirus that causes respiratory tract infections which are particularly dangerous for young children, immunocompromised individuals, and the elderly. Like HeV, no vaccines or therapies are available to combat HMPV infections. These viruses fuse their lipid envelopes with a cell to initiate infection. Blocking cell entry is a promising approach for antiviral development, and many vaccines are designed …

Characterization Of The Influences Of Human Cytomegalovirus Glycoprotein O (Go) Expression On Gh/Gl Complexes Assembly And Its Polymorphisms On Cell-Free And Cell-To-Cell Spread, And Antibody Neutralization., Le Z. Day

Graduate Student Theses, Dissertations, & Professional Papers

Human cytomegalovirus (HCMV) is widely spread throughout the world and immunocompromised individuals can suffer severe diseases from HCMV infection. Once the infection is established, HCMV can spread through the body and infect many major somatic cell types. The glycoproteins H and L (gH/gL) on HCMV envelope can be bound by either gO or the UL128-131 proteins to form complexes gH/gL/gO and gH/gL/UL128-131 that are critical for viral entry and spread, and these two complexes are important targets of neutralizing antibodies. Strains of HCMV vary considerably in the levels of gH/gL/gO and gH/gL/UL128-131. gO is one of the most diverse loci …

Cyclophilin A Enhances Hiv-1 Reverse Transcription In Human Microglial Cells, Zachary Michael Ingram

Graduate Theses/Dissertations

Parenchymal microglia represent a susceptible cell type to HIV infection and contribute to HIV Associated Neurocognitive Disorders (HAND). Currently, HIV host-protein interactions in microglia are understudied, but relevant to the design of antiviral drugs. HIV replication events rely on host and viral proteins to evade an immune response while improve replication success. Post-fusion the HIV capsid is released into the cytoplasm and begins trafficking towards the nucleus. During transit viral RNA is transcribed to DNA through reverse transcription (RT). In addition, the HIV capsid that protects the reverse transcription complex disassembles in a step termed uncoating. Once the pre-integration complex …

Kinetics Of Hiv-1 Uncoating In C20 Microglial Cells, Melanie Anne Taylor

Graduate Theses/Dissertations

Uncoating is a poorly understood yet required step of HIV-1 replication that is defined as the disassembly of the viral capsid structure. The goal of this project is to characterize uncoating in C20 microglial cells. These cells are a natural target of HIV-1 that are infected to establish latent viral reservoirs and HIV-associated neurological disorders. A stable C20 cell line that expresses TRIM-CypA was established to study the kinetics of uncoating with the CsA washout assay. The expression of TRIM-CypA was confirmed by western blot and the functionality of the protein was confirmed by a viral infectivity assay. Using this …

Genome Sequences Of Three Cruciviruses Found In The Willamette Valley (Oregon), Ignacio De La Higuera, Ellis L. Torrance, Alyssa A. Pratt, George Kasun, Amberlee Maluenda, Kenneth M. Stedman

Biology Faculty Publications and Presentations

Cruciviruses are single-stranded DNA (ssDNA) viruses whose genomes suggest the possibility of gene transfer between DNA and RNA viruses. Many crucivirus genome sequences have been found in metagenomic data sets, although no crucivirus has been isolated. Here, we present the complete genome sequences of three cruciviruses recovered from environmental samples from Oregon.

Engineering Yeast To Evaluate Human Proteins Involved In Selective Rna Packaging During Hiv Particle Production, Ryan M. Bitter

Master's Theses

Despite recent advances in antiretroviral therapy, nearly 37 million people continue to live with human immunodeficiency virus (HIV). Basic and applied research on the assembly of HIV could be enhanced by using a genetically tractable organism, such as yeast, rather than mammalian cells. While previous studies showed that expression of the HIV Gag polyprotein in Saccharomyces cerevisiae spheroplasts resulted in the production of virus-like particles (VLPs), many questions regarding the utility of yeast in HIV assembly remain uninvestigated. Here, we report use of S. cerevisiae for both the production of VLPs with selectively packaged RNA and to evaluate the human …

Mechanisms Adopted By Dengue-2 Viruses To Induce Autophagy In Mammalian Cells, Sounak Ghosh Roy

Dissertations, Theses, and Capstone Projects

Dengue, the most rapidly spreading flavivirus, threatens to affect almost half of the human global population. We previously showed that dengue-2 protects canine kidney cells (MDCK) from cytotoxic chemicals. We showed, independently, that cell protection, as well as viral replication and maturation, are positively regulated by PI3K-dependent autophagy. However, we had not identified the specific pathway that induces autophagy in infected cells. The current study explores the role of a specific branch of the endoplasmic reticulum (ER) stress-mediated Unfolded Protein Response (UPR), the PERK/eIF2α/ATF4 pathway in the induction of autophagy by …

Arthropod Evs Mediate Dengue Virus Transmission Through Interaction With A Tetraspanin Domain Containing Glycoprotein Tsp29fb, Ashish Vora, Wenshuo Zhou, Berlin Londono-Renteria, Michael Woodson, Michael B. Sherman, Tonya M. Collpitts, Girish Neelakanta, Hameeda Sultana

Biological Sciences Faculty Publications

Dengue virus (DENV) is a mosquito-borne flavivirus that causes dengue fever in humans, worldwide. Using in vitro cell lines derived from Aedes albopictus and Aedes aegypti, the primary vectors of DENV, we report that DENV2/DENV3-infected cells secrete extracellular vesicles (EVs), including exosomes, containing infectious viral RNA and proteins. A full-length DENV2 genome, detected in arthropod EVs, was infectious to naïve mosquito and mammalian cells, including human-skin keratinocytes and blood endothelial cells. Cryo-electron microscopy showed mosquito EVs with a size range from 30 to 250 nm. Treatments with RNase A, Triton X-100, and 4G2 antibody-bead binding assays showed that infectious …

Restoration Of Phage Growth On A Non-Permissive Host By Bypassing Transcription Termination Signals, Millicent Ronkainen

Mahurin Honors College Capstone Experience/Thesis Projects

RNA polymerase is the central enzyme in all gene expression. The rpoCY75N mutation in the zinc-binding domain of the β’ subunit of E. coli RNA polymerase blocks a unique RNA-based mechanism of transcription antitermination utilized by bacteriophage HK022 and its relatives. Here, we describe the characterization of mutant phage, orc0368, which overcomes the rpoCY75N mutation. The orc0368 genome varies from the wild type phage genome by 4 single base pair mutations. Three of these mutations were not characterized because they occur in intergenic regions but the fourth was chosen for study because of its location between a series …

Hiv Suppresses Cervical Neutrophil Infiltration In Women With Normal Or Abnormal Pap Smears, Mika Caplan, Peter C. Angeletti, Duan Loy, Kandali Samwel, Daniela Gonzalez, Crispin Chisanga, Julius Mwaiselage, Asafu Munema, Brenda Bashekera, Kessy Goodwin, John Dustin Loy, John T. West, Charles Wood

UCARE: Research Products

Human immunodeficiency virus (HIV) infection of CD4+ T cells results in a weakened immune system due to decreased white blood cells, particularly multi-lobed neutrophils (neutropenia) and other granulocytes.¹ The purpose of this study was to determine whether a correlation exists between HPV/HIV status and presence of neutrophils. To test this, we used a computer software program (QuPath) to analyze neutrophil infiltration seen in pap smears of both HIV+ and HIV- patients from samples collected in Tanzania from three different sites: Bagamoyo, Chalinze, and Dar es Salaam. The software was used to quantify neutrophils per image based on the size …