SUP Program: M&I-HBCU SROP

Presentation Time: 9:00-9:20

Home University: North Carolina Central University

Research Mentor: Kristina De Paris, Microbiology and Immunology

Program: M&I-HBCU SROP

Research Title: Adjuvant Induced Gene Expression Rhesus Macaques

To reduce infant HIV infection rates by vaccination, the current study was designed to identify innate molecular signatures activated by two different vaccine regimens. Previously, we determined that infant rhesus macaques (RMs) vaccinated with an HIV envelope (Env) protein with a toll-like receptor (TLR) 7/8 adjuvant versus alum adjuvant had higher Env-specific antibody responses. Here, we selected two adjuvants, TLR7/8-based R848 or TLR3-based Ampligen, in an HIV vaccine regimen consisting of Modified Vaccinia Ankara (MVA) expressing HIV Env and HIV env protein.

Infant RMs were divided into four groups. Control RMs (Group A) received empty MVA intradermally (ID) and sublingually (SL) at weeks (wks) 0 and 4, and Ampligen or R848 only at wks 4 and 10, respectively. Group B and C RMs were immunized with MVA-Env (wks 0, 4) and HIV Env protein (wks 4, 10) in Ampligen or R848, respectively. One day after each immunization, 0.25mL of EDTA-anticoagulated blood was collected. Samples were stored in TRIzol until RNA extraction. Purified RNA was analyzed using the nCounter Nonhuman Primate Immunology Panel by NanoString to assess changes in gene expression. The data will be analyzed using the nCounter Analysis Software to identify specific biological pathways induced by the two distinct adjuvant. We will then correlate the different innate response to the vaccine-induced antibody responses to determine the optimal vaccine regimen.

Presentation Time: 1:55-2:15

Home University: North Carolina Central University

Research Mentor: Helen Lazear, Immunology and virology

Program: M&I-HBCU SROP

Research Title: The Power of Interferons: Interferons vs. Viruses

Nissi Ficklin
Email: nficklin@eagles.nccu.edu
Mentor’s information: Nick Catanzaro- nick_catanzaro@med.unc.edu (Helen Lazear lab)

The body is full of amazing things that it can do. One of which is fighting off viruses on its own. The innate immune system is the body’s first line of defense, and with the help of interferons this is possible. Interferons are cytokines that tell cells how to fight viruses. Interferons are glycoproteins that are produced by cells to respond to the virus, restrict the continuation of virus replication, and the spread of the virus before the adaptive response is developed. Type I interferons (IFN-α/β) and type III interferons (IFN-λ) are produced in response to viral infection and signal through the JAK/STAT pathway to induce the expression of interferon stimulated genes (ISGs). To test this hypothesis, we first designed and cloned single guide RNAs targeting candidate ISGs into the all-in-one expression vector, pLentiCRISPRv2. All constructs were verified by Sanger sequencing and introduced into an A549 human lung epithelial cell culture model. Cells are currently undergoing puromycin selection to obtain an edited population. Successful ISG knockouts will first be verified by western blot analysis. Zika virus replication will then be assessed in ISG knockout cells by performing multi-step growth curves and virus titrations. A better understanding of the innate immune response to viral infection will help develop novel antiviral therapeutics.

Presentation Time: 2:20-2:40

Home University: North Carolina Central University

Research Mentor: Peggy Cotter, Microbiology & Immunology

Program: M&I-HBCU SROP

Research Title: Burkholderia thaliandensis Contact-Dependent Growth Inhibition Competitions between strains E254 & E264

In nature, microbes use a variety of systems to compete for resources. Burkholderia thaliandensis uses a system called Contact-Dependent Growth Inhibition (CDI) system encoded by the bcpAIOB genes to compete with its neighbors. CDI systems function by delivering a toxin to a recipient cell which then dies unless it is protected by a cognate immunity protein, BcpI. To better understand CDI systems, I studied the CDI systems in two closely related B. thailandensis strains, E264 and E254. E264 has been extensively studied whereas E254 is a new strain to the lab that has a bcpAIOB locus ~97% identical to E264.

We measure the effectiveness of CDI systems through interbacterial competitions where we mark each strain with separate antibiotic resistances and compete them at a 1:1 ratio. Unexpectedly, we discovered that E254 has separate morphologies; both smooth (E254S) and rough (E254R) and that these behave differently in a competition. We also found that E264 lacking the bcpAIOB locus (ΔbcpAIOB) out-competes E254R by a significant margin indicating that the CDI system is not required for competition. Both E264 WT & E264 ΔbcpAIOB compete equally well with E254S. This suggests that either both CDI systems work equally well or that neither is being used. These data indicate that even closely related strains might use CDI systems differently and that further testing is needed.

Presentation Time: 2:45-3:05

Home University: Spelman College

Research Mentor: Michelle Mac, Microbiology and Immunology

Program: M&I-HBCU SROP

Research Title: Epigenetic Regulation of the HPV Viral Life Cycle

Human papillomavirus (HPV) is the most common viral sexually transmitted disease with the lifetime risk of contraction reaching over 80%. High-risk HPV types have been 09` with promoting cervical cancer as well as head and neck cancers. ​​HPVs are non-enveloped, small double-stranded DNA viruses with histone-associated genomes that infect the basal keratinocytes of the stratified epithelium. HPV viral DNA is established within the nucleus of the cell as episomes, then the viral genome is replicated and passed on to daughter cells. Because the HPV viral genome is histone-associated, there is an interest in how epigenetic regulation affects its viral life cycle. Epigenetic regulator SETD2 facilitates the repair of double-strand breaks (DSB) through trimethylation of H3k36me3 that allows LEDGF to bind to H3k36me3 and promote DSB resection. In response to DNA damage, LEDGF recruits the C-terminal-binding protein interacting protein (CtIP) nuclease, which promotes the resection of DSBs by binding to Rad51. The current study was designed to identify whether, CtIP, is needed for productive viral DNA replication by evaluating the effect of a knockdown of CtIP specific shRNA. It is hypothesized that the knockdown of CtIP will lead to reduced viral replication of HPV31+ cells.

Presentation Time: 4:50-5:10

Home University: North Carolina Central University

Research Mentor: Aaron Anselmo, Pharmaceutical sciences

Program: M&I-HBCU SROP

Research Title: Improving bacterial attachment to the skin

Therapeutic bacteria have recently received attention as a method to increase the health of the skin, as well as treat certain skin conditions like acne. However, therapeutic bacteria cannot colonize the skin in their natural state. To address this, we have developed a method to modify both the skin and the bacteria with complementary chemical groups that click together to improve bacterial attachment. Specifically, we conjugated azide to the bacterial surface and DBCO to the skin using NHS ester chemistry, which reacts with primary amines on both surfaces. First, we confirmed these modifications can successfully be applied to live bacteria using amine-coated 24 well plates. We analyzed bacterial attachment with microscopy and showed significantly increased attachment relative to controls. Next, we developed an ex vivo pig skin model to show that the skin can be successfully modified with DBCO. Finally, we used live human epidermal keratinocytes (HEKa) to see how the bacteria would attach to live cells. Data showed successful modification of the cells, as well as an increase in bacterial attachment after azide modification. Future work will analyze the viability of HEKa cells following bacterial attachment, as well as in vivo testing in a mouse model. This work demonstrates that bacteria and skin can be successfully modified to improve attachment, potentially enabling the use of therapeutic bacteria as a treatment for various skin conditions.