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Phi X 174, David J. Hall

Infectious Pathogens

Phi X 174 database number # 1CD3. Phi X 174 is a virus that infects bacteria and was the first DNA-based genome to be sequenced in 1977. This bacteriophage has a [+] circular single-stranded DNA genome. The assembly of the capsid has been successfully done in vitro and is the focus of many studies on self-assembly, drug delivery and nanotechnology.

Dengue Virus, David J. Hall

Infectious Pathogens

Dengue virus, database # 1THD Dengue fever, is an infectious tropical disease caused by the dengue virus and transmitted by mosquito. Symptoms include fever, headache, muscle and joint pains, and a characteristic skin rash that is similar to measles. In a small proportion of cases the disease develops into the life-threatening dengue hemorrhagic fever. Dengue is a positive strand RNA virus that is part of the flavivirus family. Dengue is an enveloped virus meaning it has a nucleocapsid protecting the RNA and the nucleocapsid is surrounded by lipids (hence the name enveloped).

Human Papilloma Virus 16, David J. Hall

Infectious Pathogens

Human Papilloma virus 16, database# 1DZL Infection by most papillomaviruses is either asymptomatic or causes small benign tumors, known as papillomas or warts (e.g. human papillomavirus HPV6 or HPV11). Papillomas caused by some types, however, such as human papillomaviruses 16 and 18, carry a risk of becoming cancerous. Human papilloma virus is a non-enveloped, double stranded, circular DNA virus.

Hepatitis B, David J. Hall

Infectious Pathogens

Hepatitis B, database # 2g33. Hepatitis B is an infectious inflammatory illness of the liver caused by the hepatitis B virus (HBV). Hepatitis B virus (HBV) is a member of the Hepadnavirus family.The virus particle, (virion) consists of an outer lipid envelope and an icosahedral nucleocapsid core composed of protein. These virions are 42 nM in diameter. The nucleocapsid encloses the viral DNA and a DNA polymerase that has reverse transcriptase activity.

Genetic Regulation Of Mucoidy In Pseudomonas Aeruginosa, T. Ryan Withers

Theses, Dissertations and Capstones

Cystic fibrosis is a genetic disorder that results from mutations in the CF transmembrane conductance regulator gene. These mutations cause a disruption in the chloride transport in mucosal tissues causing the accumulation of dehydrated mucus, and a decrease in the mucocilliary removal of environmental pathogens within the lungs. Additionally, the accumulation of dehydrated mucus within the lungs provides a hospitable environment for various bacteria, including the Gram-negative opportunistic pathogen Pseudomonas aeruginosa. P. aeruginosa uses the overproduction of a surface polysaccharide called alginate to form a biofilm to evade the host’s immunological defenses. The overproduction of alginate, often referred to as …

Identification Of Potential Lead Antimalarial Compounds From Marine Microbial Extracts, Abigail Carbonell

HIM 1990-2015

Malaria, caused by the parasite Plasmodium falciparum, has a long history as a global health threat. The vector-borne disease causes millions of deaths yearly, especially in developing countries with tropical climates that facilitate transmission. Compounding the problem is the emergence of drug-resistant strains due to overuse of outdated treatments. New compounds with antiplasmodial activity are needed to be developed as effective drugs against malaria. The hypothesis for this project is that marine microorganisms have a high likelihood of yielding novel antiplasmodial chemotypes because of their high diversity, which has not yet been explored for antimalarial development. In this project, microbes …

Molecular Architecture Of The Uncleaved Hiv-1 Envelope Glycoprotein Trimer, Youdong Mao, Luis Castillo-Menendez, Liping Wang, Christopher Gu, Alon Herschhorn, Anik Désormeaux, Andrés Finzi, Shi-Hua Xiang, Joseph Sodroski

Nebraska Center for Virology: Faculty Publications

The human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer, a membrane-fusing machine, mediates virus entry into host cells and is the sole virusspecific target for neutralizing antibodies. Binding the receptors, CD4 and CCR5/CXCR4, triggers Env conformational changes from the metastable unliganded state to the fusion-active state. We used cryo-electron microscopy to obtain a 6-Å structure of the membranebound, heavily glycosylated HIV-1 Env trimer in its uncleaved and unliganded state. The spatial organization of secondary structure elements reveals that the unliganded conformations of both gp120 and gp41 subunits differ from those induced by receptor binding. The gp120 trimer association domains, which …

Variations In The Biological Functions Of Hiv-1 Clade C Envelope In A Shiv-Infected Rhesus Macaque During Disease Progression, For Yue Tso, Levon Abrahamyan, Shiu-Lok Hu, Ruth M. Ruprecht, Charles Wood

Nebraska Center for Virology: Faculty Publications

A better understanding of how the biological functions of the HIV-1 envelope (Env) changes during disease progression may aid the design of an efficacious anti-HIV-1 vaccine. Although studies from patient had provided some insights on this issue, the differences in the study cohorts and methodology had make it difficult to reach a consensus of the variations in the HIV-1 Env functions during disease progression. To this end, an animal model that can be infected under controlled environment and reflect the disease course of HIV-1 infection in human will be beneficial. Such an animal model was previously demonstrated by the infection …

Kaposi’S Sarcoma-Associated Herpesvirus Transactivator Rta Induces Cell Cycle Arrest In G0/G1 Phase By Stabilizing And Promoting Nuclear Localization Of P27Kip, Pankaj Kumar, Charles Wood

Nebraska Center for Virology: Faculty Publications

The Kaposi’s sarcoma-associated herpesvirus (KSHV) immediate-early gene, replication, and transcription activator (KRta) is a key viral protein that serves as the master regulator for viral lytic replication. In this study, we investigated the role of K-Rta in cell cycle regulation and found that the expression of K-Rta in doxycycline (Dox)-inducible BJAB cells induced cell cycle arrest in G₀/G₁ phase. Western blot analysis of key cell cycle regulators revealed that K-Rta-mediated cell cycle arrest was associated with a decrease in cyclin A and phosphorylated Rb (pS807/pS811) protein levels, both markers of S phase progression, and an increase in …

Low-Abundance Resistant Mutations In Hiv-1 Subtype C Antiretroviral Therapy-Naïve Individuals As Revealed By Pyrosequencing, Sandra Gonzalez, Damien C. Tully, Clement Gondwe, Charles Wood

Nebraska Center for Virology: Faculty Publications

Given the recent scale-up of antiretroviral therapy (ART) in sub-Saharan Africa, we sought to determine how often and at what levels do drug-resistant mutant variants exist in ART-naïve HIV subtype C infected individuals. Samples from 10 ART-naïve Zambian individuals were subjected to ultra-deep pyrosequencing (UDPS) to characterize the frequency of low-abundance drug resistance mutations in the pol gene. Low-abundance clinically relevant variants were detected for nucleoside reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs) and protease inhibitors (PIs) in eight of the ten subjects. Intermediate to high-level resistance was predicted for the majority of NRTIs. Mutations conferring resistance to …

Clade C Hiv-1 Isolates Circulating In Southern Africa Exhibit A Greater Frequency Of Dicysteine Motif-Containing Tat Variants Than Those In Southeast Asia And Cause Increased Neurovirulence, Vasudev R. Rao, Ujjwal Neogi, Joshua S, Talboom, Ligia Padilla, Mustafizur Rahman, Cari Fritz-French, Sandra Gonzalez-Ramirez, Anjali Verma, Charles Wood, Ruth M. Ruprecht, Udaykumar Ranga, Tasnim Azim, Vinayaka R. Prasad

Nebraska Center for Virology: Faculty Publications

Background: HIV-1 Clade C (Subtype C; HIV-1C) is responsible for greater than 50% of infections worldwide. Unlike clade B HIV-1 (Subtype B; HIV-1B), which is known to cause HIV associated dementia (HAD) in approximately 15% to 30% of the infected individuals, HIV-1C has been linked with lower prevalence of HAD (0 to 6%) in India and Ethiopia. However, recent studies report a higher prevalence of HAD in South Africa, Zambia and Botswana, where HIV-1C infections predominate. Therefore, we examined whether Southern African HIV-1C is genetically distinct and investigated its neurovirulence. HIV-1 Tat protein is a viral determinant of neurocognitive dysfunction. …

Global Neuroaids Roundtable, Jeymohan Joseph, Cristian L. Achim, Michael J. Boivin, Bruce J. Brew, David B. Clifford, Deborah A. Colosi, Ronald J. Ellis, Robert K. Heaton, Amadou Gallo-Diop, Igor Grant, Georgette D. Kanmogne, Mahendra Kumar, Scott Letendre, Thomas D. Marcotte, Avindra Nath, Carlos A. Pardo, Robert H. Paul, Lynn Pulliam, Kevin Robertson, Walter Royal Iii, Ned Sacktor, Pasiri Sithinamsuwan, Davey M. Smith, Victor Valcour, Brian Wigdahl, Charles Wood

Nebraska Center for Virology: Faculty Publications

In May 2012, the Division of AIDS Research at the National Institute of Mental Health (NIMH) organized the “Global NeuroAIDS Roundtable” in conjunction with the 11th International Symposium on Neurovirology and the 2012 Conference on HIV in the Nervous System. The meeting was held in New York, NY, USA and brought together NIMH-funded investigators who are currently working on projects related to the neurological complications of AIDS (NeuroAIDS) in Africa, Asia, Eastern Europe, and Latin America in order to provide an opportunity to share their recent findings and discuss the challenges encountered within each country. The major goals of the …

Extracellular Proteases Are Key Mediators Of Staphylococcus Aureus Virulence Via The Global Modulation Of Virulence-Determinant Stability, Stacey L. Kolar, J. Antonio Ibarra, Frances E. Rivera, Joe M. Mootz, Jessica E. Davenport, Stanley M. Stevens Jr., Alexander R. Horswill, Lindsey N. Shaw

Molecular Biosciences Faculty Publications

Staphylococcus aureus is a highly virulent and successful pathogen that causes a diverse array of diseases. Recently, an increase of severe infections in healthy subjects has been observed, caused by community-associated methicillin-resistant S. aureus (CA-MRSA). The reason for enhanced CA-MRSA virulence is unclear; however, work suggests that it results from hypersecretion of agr-regulated toxins, including secreted proteases. In this study, we explore the contribution of exo-proteases to CA-MRSA pathogenesis using a mutant lacking all 10 enzymes. We show that they are required for growth in peptide-rich environments, serum, in the presence of antimicrobial peptides (AMPs), and in human blood. …

Cpg And Interleukin-15 Synergize To Enhance Ifn-Gamma Production By Activated Cd8(+) T Cells, Dustin Cobb, Sigi Guo, Ronald B. Smeltz

Bioelectrics Publications

Interleukin-15 (IL-15) regulates the development and maintenance of memory CD8(+) T cells. Paradoxically, we previously reported that IL-15 could enhance CD8(+) T-cell responses to IL-12, a proinflammatory cytokine required for optimal priming of effector CD8(+) T cells. To expand the physiological relevance of these findings, we tested IL-15 for its ability to enhance T-cell responses to bacterial CpG. Expectedly, CpG enhanced the production of IFN-gamma by CD8(+) T cells polyclonally activated with anti-CD3. However, addition of IL-15 to CpG-stimulated cultures led to a striking increase in IFN-gamma production. The effect of CpG and IL-15 was also evident with CD8(+) T …

Hypoxia Inducible Factor 1 Alpha (Hif-1a): A Major Regulator Of Placental Development, Renee E. Albers

Browse all Theses and Dissertations

Hypoxia-inducible factor-1 alpha (HIF-1a) is a critical component of the cellular oxygen-sensing machinery and is essential for placental formation and embryonic survival. In this study, we promoted prolonged expression of HIF-1a, by using a form that is insensitive to oxygen, denoted as CA-HIF-1a. In order to have continual placental specific expression of the CA-HIF-1a, lentiviral infection was performed on embryos at the blastocyst stage of development and transferred into pseudo-pregnant mothers. Placental analysis was performed via in situ hybridization on embryonic day 14.5 (E14.5) to determine the effects of CA-HIF-1a prolonged expression. Data indicate that prolonged activity of CA-HIF-1a restricted …

Expression Of An Epitope Tagged Tarp Effector In Chlamydia Trachomatis, Brenda Nguyen

HIM 1990-2015

Previous studies performed on Chlamydia trachomatis have demonstrated how these obligate intracellular microbes invade host cells through the utilization of secreted effector proteins. One secreted effector called Tarp (translocated actin recruiting protein) is implicated in cytoskeleton rearrangements that promote bacterial entry into the host cell. The focus of our study is to create a plasmid that carries the tarP gene that when transcribed and translated from within Chlamydia trachomatis will generate a c-Myc epitope tagged Tarp. The tag will be used in future studies to track the progression of the protein through the infectious process and will allow us to …

Understanding The Role Of Plasmodium Falciparum Vamp8 Snare Homologue, Katherine Ferreira

HIM 1990-2015

Malaria is one of the worlds most deadly infectious diseases and results in almost a million deaths each year, largely in children under the age of five in Sub-Saharan Africa. Outside Africa, malaria is responsible for a large number of cases in the Amazon rainforest of Brazil, Middle East, and in some areas of Asia [37]. According to the World Health Organization, there was an estimated 655, 000 deaths from malaria in 2012. Malaria is caused by a eukaryotic Apicomplexan parasite, Plasmodium, which has three distinct life cycles occurring in the midgut of the female Anopheles mosquito, the liver of …

Equine Serum Antibody Responses To Streptococcus Equi And Streptococcus Zooepidemicus, Rafaela De Negri

Theses and Dissertations--Veterinary Science

Streptococcus zooepidemicus (Sz) and Streptococcus equi (Se) share 98% DNA sequence homology, but display different pathogenic properties. Infection by one organism does not cross-protect against the other. To better understand pathogenic differences between these organisms and gain information about which proteins are expressed in horses infected experimentally with Se, intrauterine Sz or naturally with respiratory Sz we compared antibody specificities of convalescent sera using ELISA. These comparisons were based on sets of 8 and 14 immunoreactive recombinant proteins of Se strain CF32 and Sz strain NC78, respectively. Sera from donkeys that were previously naturally affected with strangles and later developed …

Type I Interferon Activation Of Natural Killer (Nk) Cells By Cytomegalovirus (Cmv) And Their Interaction With Dendritic (Dc) And Nkt Cells, Howaida A. Oulad Abdelati

Browse all Theses and Dissertations

In our current study the roles of natural killer (NK) cells in regulation of the acute phase of murine cytomegalovirus (MCMV) infection were demonstrated. NK cells utilize perforin and gamma interferon to kill MCMV infected cells. Activation of NK cells is controlled by the balance between inhibitory and activation receptors and modulated by cytokines produced from the infected cells. Type I interferons are produced mainly by plasmacytoid dendritic cells (PDCs). Type I interferons have a substantial role in enhancing NK cells cytotoxic activity and NK cells proliferation and maintain cell survival through STAT mediated signaling pathway. IL-12 produced by dendritic …

Exploitation Of The Bacteriophage-Derived Peptidase Chapk, Pierre-Mehdi Hadbi

Theses

This project investigated an anti-staphylococcal bacteriophage-derived peptidoglycan hydrolase enzyme, namely a cysteine-histidine amydohydrolase/peptidase (CHAP_k). The study focused firstly on optimizing the production of recombinant CHAP_k, which was previously cloned in an E. coli expression system, and sets out the resulting optimal Standard Operating Procedures (SOPs) for production, purification and determination of concentration and activity of the resulting protein stock. A typical yield resulting from the investigation was 300pg of CHAP_k from one litre of E.coli culture. The activity of the purified enzyme was typically in the region of 190U nmol ’. Secondly, the project investigated …

Assessment Of Motility And Hemolytic Activity In Clinical Isolates Of Acinetobacter Baumannii From University Of Kentucky Hospital, Lexington, Ky, Amber R. Stanton

Online Theses and Dissertations

Acinetobacter baumannii is a pathogen rising in notoriety worldwide due to outbreaks linked to multi-drug resistant strains. Research is currently focused on identifying virulence factors, which may contribute to increased ability to cause human disease, such as hemolytic activity and surface motility. The aim of this study was to determine the presence of these two virulent traits in clinical isolates. Forty-eight clinical isolates were recovered from University of Kentucky hospital in Lexington, KY. No hemolytic activity was observed for any of the isolates. Evidence of surface motility was observed in 13 isolates. The brand and concentration of media used allowed …

Amp-Activated Protein Kinase Knockdown In Labyrinthine Trophoblast Cells Results In Altered Morphology And Function, Erica Ashton Kayleigh Carey

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The placenta is a transient organ that develops upon initiation of pregnancy and is essential for fetal development and survival. The rodent placenta consists of three layers with predominantly analogous cell types in the human placenta. The labyrinth layer, which lies closest to the fetus, facilitates nutrient and waste exchange between mother and baby. Abnormalities of the placenta may occur as a result of cellular stress and have been associated with the pregnancy-associated disorders, intrauterine growth restriction and placental insufficiency. AMP-activated protein kinase (AMPK) is a master sensor of cellular stress and changes in AMPK expression or activation could be …

The Effect Of Hsv-1 Infection On Differentiated And Polarized U937 Cells, Allolo Dreiwish Aldreiwish

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Herpes simplex virus type 1 (HSV-1) challenges the host immune system through several mechanisms (Frey, et al., 2009). In vitro, U937 cells (human macrophage-like precursor cell line) are not susceptible to HSV-1 infection when they are not differentiating (Lopez-Guerrero and Alonso, 1997). Differentiation of these cells resistance can abrogates their resistance to HSV-1 (Tenney and Morahan, 1991). In this study, we examined the effect of HSV-1 infection on differentiated and polarized U937 cells. U937 cells are differentiated to M0 cells.

Then, M0 cells are polarized to distinct phenotypes, M1 or M2. M1 are proinflammatory macrophages while M2 are anti-inflammatory cells. …

Establishment Of A Quiscence Herpes Simplex Type 1 Infection In L929 Fibroblasts And Neuro-2a Cells By A Nucleoside Analogue Acyclovir, Noura Shaklawoon

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Herpes simplex type 1 (HSV-1) quiescent infection was established in L929 cells, murine fibroblasts, and in Neuro-2A (N2A) cells, mouse neuroblastoma, by treating them with the nucleoside analogue acyclovir (ACV) 24 hours before infection. Subsequent release of virus from the non-productive state was accomplished by treating the cells with the histone deacetylase inhibitor trichostatin A (TSA). Treatment of both L929 and N2A cell lines with ACV 24 hours before infection induced protection from HSV-1 cytopathic effect. A quiescent state was confirmed by absence of virus plaques when supernatant fluids from ACV-treated HSV-1 infected cultures were titered on Vero cell monolayers. …

N-Glycosylation, Localization And Trafficking Of Endogenous Nkcc1 In Cos7 Cells, Richa Singh

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The Na+K+2Cl^- co-transporter-1 (NKCC1) effects the electroneutral movement of Na+, K+ and 2Cl^- ions across plasma membranes. NKCC1 is considered a widely distributed, highly N-glycosylated ion transporter involved in the regulation of the intracellular chloride concentration ([Cl^-]i), cell volume and salt secretion. NKCC1 locates in the plasma membrane of all cells studied so far, particularly on the basolateral side of most polarized cells. However, the majority of immunoreactive NKCC1 distributes in intracellular compartments of unknown nature whereas a small proportion of the transporter appears to be located in the plasma membrane. Here, we identified and characterized …

Novel Approaches For The Eradication Of Hiv Latently Infected Cells, Sally Al Ali

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The development of a suitable experimental cell model to study HIV latency in primary cells could have a massive effect on the current approaches to eradicate virus in latently infected cells. The main proposal of this paper is to develop an in vitro HIV cell model that represents HIV latency in vivo, then to create a more effective viral vector in order to target HIV reservoirs. For this goal, a directed evolution method is suggested to be used in order to mutate the AAV cap gene to generate a recombinant AAV vector that is capable of infecting primary resting CD4+ …

Conversion Of The U937 Monocyte Into "Macrophage-Like" Populations Exhibiting M1 Or M2 Characteristics, Bradley M. Sharp

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The U937 cell line is an oncogenic human monocyte cell line. These monocytes have the potential of differentiating into either macrophages or dendritic cells (Lawrence et al., 2011). This differentiation pattern depends on the characteristics of the tissue microenvironment (Kigerl et al., 2009). PMA (Phorbol 12-Myristate 13-Acetate) is a phorbol ester capable of transforming monocytic cells toward the macrophage pathway. Upon treatment with PMA, U937 cells under-go a series of morphological and functional changes. Traditionally monocytic cell lines are used as a model of macrophage function, because current human macrophage cell lines require a T-cell conditioned growth medium and contact …

Hpai H5n1: A Global Pandemic Concern, With Implications For Pandemic Preperation And Public Health Policy, Lauren M. Koontz

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Research on highly pathogenic avian influenza A (HPAI) H5N1 has gained much attention in recent years due to its devastating impact within the bird population. While transmission to humans is rare, infections are fatal in more than half of the cases. One of the main concerns among the scientific community is the ability for this virus to mutate in a way much like seasonal influeanza that would allow it to be transmitted efficiently among the human population. Two researchers were able to separately create mutant isolates of H5N1 influenza A virus that could be transmitted via the respiratory route between …

Towards Defining The Chloroviruses: A Genomic Journey Through A Genus Of Large Dna Viruses, Adrien Jeanniard, David D. Dunigan, James Gurnon, Irina V. Agarkova, Ming Kang, Jason Vitek, Garry Duncan, O William Mcclung, Megan Larsen, Jean-Michel Claverie, James L. Van Etten, Guillaume Blanc

Nebraska Center for Virology: Faculty Publications

Background: Giant viruses in the genus Chlorovirus (family Phycodnaviridae) infect eukaryotic green microalgae. The prototype member of the genus, Paramecium bursaria chlorella virus 1, was sequenced more than 15 years ago, and to date there are only 6 fully sequenced chloroviruses in public databases. Presented here are the draft genome sequences of 35 additional chloroviruses (287 – 348 Kb/319 – 381 predicted protein encoding genes) collected across the globe; they infect one of three different green algal species. These new data allowed us to analyze the genomic landscape of 41 chloroviruses, which revealed some remarkable features about these viruses. …

Evaluation Of Higher Plant Virus Resistance Genes In The Green Alga, Chlorella Variabilis Nc64a, During The Early Phase Of Infection With Paramecium Bursaria Chlorella Virus-1, Janet M. Rowe, David D. Dunigan, Guillaume Blanc, James R. Gurnon, Yuannan Xia, James L. Van Etten

Nebraska Center for Virology: Faculty Publications

With growing industrial interest in algae plus their critical roles in aquatic systems, the need to understand the effects of algal pathogens is increasing. We examined a model algal host–virus system, Chlorella variabilis NC64A and virus, PBCV-1. C. variabilis encodes 375 homologs to genes involved in RNA silencing and in response to virus infection in higher plants. Illumina RNA-Seq data showed that 325 of these homologs were expressed in healthy and early PBCV-1 infected (≤60 min) cells. For each of the RNA silencing genes to which homologs were found, mRNA transcripts were detected in healthy and infected cells. C. variabilis …