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- All Subjects: Herpes simplex virus
- Creators: Varsani, Arvind
- Member of: ASU Electronic Theses and Dissertations
- Resource Type: Text
Description
Globally, about two-thirds of the population is latently infected with herpes simplex virus type 1 (HSV-1). HSV-1 is a large double stranded DNA virus with a genome size of ~150kbp. Small defective genomes, which minimally contain an HSV-1 origin of replication and packaging signal, arise naturally via recombination during viral DNA replication. These small defective genomes can be mimicked by constructing a bacterial plasmid containing the HSV-1 origin of replication and packaging signal, transfecting these recombinant plasmids into mammalian cells, and infecting with a replicating helper virus. The absence of most viral genes in the amplicon vector allows large pieces of foreign DNA (up to 150kbp) to be incorporated. The HSV-1 amplicon is replicated and packaged by the helper virus to form HSV-1 particles containing the amplicon DNA. We constructed a novel HSV-1 amplicon vector system containing lambda phage-derived attR sites to facilitate insertion of transgenes by Invitrogen Gateway recombination. To demonstrate that the amplicon vectors work as expected, we packaged the vector constructs expressing Emerald GFP using the replication-competent helper viruses OK-14 or HSV-mScartlet-I-UL25 in Vero cells and demonstrate that the vector stock can subsequently transduce and express Emerald GFP. In further work, we will insert transgenes into the amplicon vector using Invitrogen Gateway recombination to study their functionality.
ContributorsVelarde, Kimberly (Author) / Hogue, Ian B (Thesis advisor) / Manfredsson, Fredric (Committee member) / Sandoval, Ivette (Committee member) / Varsani, Arvind (Committee member) / Arizona State University (Publisher)
Created2021
Description
Alpha herpesviruses are a family of neuroinvasive viruses that infect multiplevertebrate species. Alpha herpesviruses are responsible for human and livestock infections, most notably Herpes Simplex Virus (HSV), Varicella Zoster virus (VZV), and Pseudorabies Virus (PRV). PRV is a potent swine virus that can infect other mammals, and results in lethal encephalitis that can be devastating to livestock and of great financial expense to farmers. HSV, types 1 and 2, and VZV are widespread throughout the global human population, with estimates of the HSV-1 burden at about 60% of people worldwide. The hallmark of alpha herpesvirus infection is a persistent, lifelong infection that can reactivate throughout the lifespan of the host. Currently, the precise mechanisms of how these viruses undergo intracellular trafficking to emerge from the infected cell in epithelial tissues is not well understood. Many insights have been made with PRV in animal neurons, both in culture systems and animal models, about the viral genes and host factors involved in these processes. However, understanding of these mechanisms, and the interplay between viral and host proteins, in the human pathogen HSV-1 is even more lacking. Using recombinant fluorescent virus strains of HSV-1 and Total Internal Reflection Microscopy to image the transport of mature viral progeny in epithelial cells, it was determined that the egress of HSV-1 uses constitutive cellular secretory pathways. Specifically, the viral progeny traffic from the trans-Golgi network to the site of exocytosis at the plasma membrane via Rab6a secretory vesicles. This work will contribute to the understanding of how alpha herpesviruses complete their lifecycles in host cells, particularly at the sites where infection initially occurs and can spread to a new organism. Knowledge of these processes may lead to the development of therapeutics or prophylactics to reduce the burden of these viruses.
ContributorsBergeman, Melissa Hope (Author) / Hogue, Ian B (Thesis advisor) / Hogue, Brenda (Committee member) / Roberson, Robert (Committee member) / Varsani, Arvind (Committee member) / Arizona State University (Publisher)
Created2023