Ioanna Skountzou

Emory University | EU · Department of Microbiology and Immunology

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) poses a clear threat to humanity. It has infected over 200 million and killed 4 million people worldwide, and infections continue with no end in sight. To control the pandemic, multiple effective vaccines have been developed, and global vaccinations are in progress. However, the virus...

Influenza A viruses (IAVs) pose a global health threat, contributing to hundreds of thousands of deaths and millions of hospitalizations annually. The two major surface glycoproteins of IAVs, hemagglutinin (HA) and neuraminidase (NA), are important antigens in eliciting neutralizing antibodies and protection against disease. However, NA is generall...

Neuraminidase (NA) is the second most abundant glycoprotein on the surface of influenza A viruses (IAV). Neuraminidase type 1 (NA1) based virus-like particles (VLPs) have previously been shown to protect against challenge with H1N1 and H3N2 IAV. In this study, we produced neuraminidase type 2 (NA2) VLPs derived from the sequence of the seasonal IAV...

Skin vaccination by microneedle (MN) patch simplifies the immunization process to increase access to vaccines for global health. Lyophilization has been widely used to stabilize vaccines and other biologics during storage, but is generally not compatible with the MN patch manufacturing processes. In this study, our goal was to develop a method to i...

Zika virus (ZIKV) has emerged as a serious health threat in the Americas and the Caribbean. ZIKV is transmitted by the bite of an infected mosquito, sexual contact, and blood transfusion. ZIKV can also be transmitted to the developing fetus in utero, in some cases resulting in spontaneous abortion, fetal brain abnormalities, and microcephaly. In ad...

While the majority of influenza-infected individuals show no or mild symptomatology, pregnant women are at higher risk of complications and infection-associated mortality. Although enhanced lung pathology and dysregulated hormones are thought to underlie adverse pregnancy outcomes following influenza infection, how pregnancy confounds long-term mat...

A severe consequence of adult Zika virus (ZIKV) infection is Guillain-Barré Syndrome (GBS), where autoreactive antibodies attack peripheral and central nervous systems (CNS) resulting in neuro-ocular pathology and fatal complications. During virally induced GBS, autoimmune brain demyelination and macular degeneration correlate with low virus neutra...

Zika virus (ZIKV) causes moderate to severe neuro-ocular sequelae, with symptoms ranging from conjunctivitis to Guillain-Barré Syndrome (GBS). Despite the international threat ZIKV poses, no licensed vaccine exists. As ZIKV and DENV are closely related, antibodies against one virus have demonstrated the ability to enhance the other. To examine if v...

Fetal placental macrophages (Hofbauer cells (HCs)) appear at 18 days post-conception and are maintained throughout pregnancy. Despite their identification more than a century ago, there are few studies characterizing the evolution of human HC phenotype through gestation. Using high-dimensional flow cytometry, we measured expression of CD68, CD80, C...

Vaccines prevent 2-3 million childhood deaths annually; however, low vaccine efficacy and the resulting need for booster doses create gaps in immunization coverage. In this translational study, we explore the benefits of extended release of licensed vaccine antigens into skin to increase immune responses after a single dose in order to design impro...

Placental immunity is dichotomous: tolerance of the semiallogenic fetus is balanced with limiting transmission of maternal pathogens. HCs are the major fetal immune cell at the placenta, but mechanisms responsible for maintaining immune homeostasis while preventing infection require elucidation. We determined the phenotype of human HCs throughout g...

In 2009, the H1N1 swine flu pandemic highlighted the vulnerability of pregnant women to influenza viral infection. Pregnant women infected with influenza A virus were at increased risk of hospitalization and severe acute respiratory distress syndrome (ARDS), which is associated with high mortality, while their newborns had an increased risk of pre-...

Zika virus (ZIKV) became a public health emergency of global concern in 2015 due to its rapid expansion from French Polynesia to Brazil, spreading quickly throughout the Americas. Its unexpected correlation to neurological impairments and defects, now known as congenital Zika syndrome, brought on an urgency to characterize the pathology and develop...

Neuraminidase is one of the two surface glycoproteins of influenza A and B viruses. It has enzymatic activity that cleaves terminal sialic acid from glycans, and that activity is essential at several points in the virus life cycle. While neuraminidase is a major target for influenza antivirals, it is largely ignored in vaccine development. Current...

The widely used influenza subunit vaccine would benefit from increased protection rates in vulnerable populations. Skin immunization by microneedle (MN) patch can increase vaccine immunogenicity, as well as increase vaccination coverage due to simplified administration. To further increase immunogenicity, we used granulocyte-macrophage colony stimu...

Influenza virus causes life-threatening infections in pregnant women and their newborns. Immunization during pregnancy is the most effective means of preventing maternal and infant mortality/morbidity; however, influenza vaccination rates of pregnant women remain under 50%. Furthermore, the availability of vaccines in low-resource populations is li...

Increased susceptibility to influenza virus infection during pregnancy has been attributed to immunological changes occurring before and during gestation in order to “tolerate” the developing fetus. These systemic changes are most often characterized by a suppression of cell-mediated immunity and elevation of humoral immune responses referred to as...

Infection and pregnancy impact cytokine expression in a compartment-specific manner. Cytokine and chemokine expression in serum, lungs, placenta and fetuses was determined via Bio-Rad 23-plex Luminex Assay. The effect of infection on cytokine and chemokine expression is represented as fold-change of the average values of infected mice over average...

Serum chemokine and cytokine levels 4 days post-infection. Serum was collected from pregnant and non-pregnant infected and non-infected mice 4 d.pi. (16 days post mating), and cytokine and chemokine expression was quantified via Bio-Rad 23-plex Luminex Assay. The shaded fold-differences are significant (p<0.05). * RANTES is the only chemokine that...

Placental and fetal chemokine and cytokine levels 4 days post-infection. Placentae and fetuses were collected from pregnant and non-pregnant infected and non-infected mice 4 d.p.i. (E16 gestation), and cytokine and chemokine expression was quantified in homogenized lysates via Bio-Rad 23-plex Luminex Assay. The shaded fold-differences are significa...

Viral titer in the lungs affects expression of progesterone and PGF2α in a compartment-specific manner. Hormone expression was quantified in sera, lung and placental lysates in pregnant infected mice with ELISA, and viral load was quantified with plaque assay in MDCK-derived cells. Virus titer detected in the lungs was plotted against corresponding...

Lung chemokine and cytokine levels 4 days post-infection. Lungs were collected from pregnant and non-pregnant infected and non-infected mice 4 d.p.i. (E16 gestation), and cytokine and chemokine expression was quantified in homogenized lysates via Bio-Rad 23-plex Luminex Assay. The shaded fold-differences are significant (p≤0.05). P values were dete...

Background Microneedle patches provide an alternative to conventional needle-and-syringe immunisation, and potentially offer improved immunogenicity, simplicity, cost-effectiveness, acceptability, and safety. We describe safety, immunogenicity, and acceptability of the first-in-man study on single, dissolvable microneedle patch vaccination against...

Nature Communications 7: Article number: 11826 (2016); Published: 7 June 2016; Updated: 16 August 2016 The original version of this Article contained an error in the spelling of the author Frances Eun-Hyung Lee that was incorrectly given as Francis Eun-Hyung Lee. This has now been corrected in both the PDF and HTML versions of the Article.

Maternal and neonatal tetanus claim tens of thousands lives every year in developing countries, but could be prevented by hygienic practices and improved immunization of pregnant women. This study tested the hypothesis that skin vaccination can overcome the immunologically transformed state of pregnancy and enhance protective immunity to tetanus in...

Supplementary Figures 1 - 3 and Supplementary Table 1

The 2009 H1N1 flu pandemic demonstrated that pregnant women infected with influenza were at risk for severe respiratory distress and premature-rupture-of-membranes (PROM), leading to high incidence of hospitalization, preterm births and small for gestation age (SGA) neonates. We utilize a syngeneic BALB/c pregnant mouse model which recapitulates cl...

This study tested the hypothesis that optimized microneedle patch formulations can stabilize trivalent subunit influenza vaccine during long-term storage outside the cold chain and when exposed to potential stresses found during manufacturing and storage. Formulations containing combinations of trehalose/sucrose, sucrose/arginine, and arginine/hept...

Systemic use of epidermal growth factor receptor inhibitors (EGFRIs) has been shown to alter MHC expression and that of several chemokines, and to enhance immune cell recruitment into human skin. We hypothesized that EGFRIs may have value as cutaneous immune response modifiers, and determined the effects of topical application of an irreversible EG...

Prevention of seasonal influenza epidemics and pandemics relies on widespread vaccination coverage to induce protective immunity. In addition to a good antigenic match with the circulating viruses, the effectiveness of individual strains represented in the trivalent vaccines depends on their immunogenicity. In this study, we evaluated the immunogen...

Skin has gained substantial attention as a vaccine target organ due to its immunological properties, which include a high density of professional antigen presenting cells (APCs). Previous studies have demonstrated the effectiveness of this vaccination route not only in animal models but also in adults. Young children represent a population group th...

Detection of immunoglobulin M (IgM) antibodies has long been used as an important diagnostic tool for identifying active viral infections, but their relevance in later stages has not been clearly defined in vivo. In this study, we followed the kinetics, longevity, and function of influenza virus-specific IgM antibodies for 2 years following subleth...

Cutaneous vaccination with microneedle patches offers several advantages over more frequently used approaches for vaccine delivery, including improved protective immunity. However, the involvement of specific APC subsets and their contribution to the induction of immunity following cutaneous vaccine delivery is not well understood. A better underst...

Problems with existing influenza vaccines include the strain specificity of the immune response, resulting in the need for frequent reformulation in response to viral antigenic drift. Even in years when the same influenza strains are prevalent, the duration of immunityduration of immunity is limited, and results in the need for annual revaccination...

Influenza infection represents a major socioeconomic burden worldwide. Skin represents a new target that has gained much attention in recent years for delivery of influenza vaccine as an alternative to the conventional intramuscular route of immunization. In this review we describe different microneedle vaccination approaches used in vivo, includin...

CIITA and MHC class II expression is silenced during the differentiation of B cells to plasma cells. When B cell differentiation is carried out ex vivo, CIITA silencing occurs rapidly, but the factors contributing to this event are not known. ZBTB32, also known as repressor of GATA3, was identified as an early repressor of CIITA in an ex vivo plasm...

Recent studies have demonstrated the effectiveness of vaccine delivery to the skin by vaccine-coated microneedles; however there is little information on the effects of adjuvants using this approach for vaccination. Here we investigate the use of TLR ligands as adjuvants with skin-based delivery of influenza subunit vaccine. BALB/c mice received 1...

Flagellin, a bioactive Toll-like receptor (TLR) 5 ligand, may trigger the innate immunity that in turn is important for subsequent adaptive immune responses. In the present study, the adjuvant effects of the monomeric and polymeric forms of Salmonella flagellin (mFliC and pFliC, respectively) were examined in specific-pathogen free (SPF) chickens i...

Influenza infection represents a major socio-economic burden worldwide. Novel delivery methods can render influenza vaccination easier and more acceptable by the public, and importantly confer protection equal or superior to that induced by conventional systemic administration. An attractive target for vaccine delivery is the skin. Recent studies h...

Unlabelled: Microneedle patches (MN) provide a novel method of vaccine delivery to the skin with the objective of targeting the large network of resident antigen-presenting cells to induce an efficient immune response. Our previous reports demonstrated that cutaneous delivery of inactivated influenza virus-coated MN to mice protects against lethal...

In vivo fluorescence imaging of mice immunized via the skin using coated MN or intramuscular injection. The kinetics of antigen trafficking were evaluated in mice immunized with 20 µg of Alexa 488-labeled OVA using the PANSEE panoramic imaging system. Images were captured at 10 min (d0), day 1 (d1), day 2 (d2), day 3 (d3), and day 5 (day 5) of mice...

Supplemental Table 1: Primer sequences used in this paper Primer sequences for genotyping, qRT-PCR, 5’RACE, and cloning. Underlined sequences in cloning primers indicates restriction enzyme recognition sequences.

Supplemental Fig. 2. 5’fRACE sequence results from splenic dendritic cells Sequences obtained from 5’RACE analysis are shown relative to the predicted transcriptional start sites for each promoter. Colored lines represent transcribed sequence and thicker lines represent translated sequence. The number of transcripts sequenced in each mouse line are...

Supplemental Fig. 3. CIITA mice form antibody secreting B cells in response to immunization with influenza Mice were immunized and serially bled as described in materials and methods. HAI titer and IgG, IgG1, or IgG2a concentration is plotted for each mouse and each time point.

Supplemental Fig. 1. CIITA regulated genes do not require CIITA isoform I for expression A. Splenic dendritic cells and B. Macrophages treated with IFN-γ 24 h were analyzed by qRT-PCR for the indicated genes identified previously as being regulated by CIITA. Ct values were normalized to the values for actin mRNA and graphed as fold over wild-type....

Three distinct promoters control the master regulator of major histocompatibility complex (MHC) class II expression, class II transactivator (CIITA), in a cell type-specific manner. Promoter I (pI) CIITA, expressed primarily by dendritic cells (DCs) and macrophages, expresses a unique isoform that contains a caspase-recruitment domain (CARD). The a...

A major goal in influenza vaccine development is induction of serological memory and cellular responses to confer long-term protection and limit virus spread after infection. Here, we investigate induction of long-lived immunity against the 2009 H1N1 virus after skin vaccination. BALB/c mice received a single dose of 5 μg inactivated A/California/0...

The emergence of the swine-origin 2009 influenza pandemic illustrates the need for improved vaccine production and delivery strategies. Skin-based immunization represents an attractive alternative to traditional hypodermic needle vaccination routes. Microneedles (MNs) can deliver vaccine to the epidermis and dermis, which are rich in antigen-presen...

Many successful vaccines induce persistent antibody responses that can last a lifetime. The mechanisms by which they do so remain unclear, but emerging evidence indicates that they activate dendritic cells via Toll-like receptors (TLRs). For example, the yellow fever vaccine YF-17D, one of the most successful empiric vaccines ever developed, activa...

The 2009 pandemic H1N1 influenza pandemic demonstrated the global health threat of reassortant influenza strains. Herein, we report a detailed analysis of plasmablast and monoclonal antibody responses induced by pandemic H1N1 infection in humans. Unlike antibodies elicited by annual influenza vaccinations, most neutralizing antibodies induced by pa...

The recent swine-origin H1N1 pandemic illustrates the need to develop improved procedures for rapid production of influenza vaccines. One alternative to the current egg-based manufacture of influenza vaccine is to produce a hemagglutinin (HA) subunit vaccine using a recombinant expression system with the potential for high protein yields, ease of c...

The 2009 H1N1 influenza virus outbreak is the first pandemic of the twenty-first century. Epidemiological data reveal that of all the people afflicted with H1N1 virus, <5% are over 51 y of age. Interestingly, in the uninfected population, 33% of those >60 y old have pre-existing neutralizing Abs against the 2009 H1N1 virus. This finding suggests th...

Influenza prophylaxis would benefit from a vaccination method enabling simplified logistics and improved immunogenicity without the dangers posed by hypodermic needles. Here we introduce dissolving microneedle patches for influenza vaccination using a simple patch-based system that targets delivery to skin's antigen-presenting cells. Microneedles w...

The respiratory illnesses caused by influenza virus can be dramatically reduced by vaccination. The current trivalent inactivated influenza vaccine is effective in eliciting systemic virus-specific antibodies sufficient to control viral replication. However, influenza protection generated after parenteral immunization could be improved by the induc...

H5N1 influenza A virus remains a global threat since its outbreak in 1997. Its re-emergence in 2003 spread from Asia to Europe, the Middle-East and Africa, raising concerns of a possible pandemic. While stockpiling of H5N1 prepandemic vaccines has been initiated, strain prediction is difficult due to the emergence of distinct clades/subclades. Anti...

Bacterial flagellins are potent inducers of innate immune responses in the mouse lung because they bind to TLR5 expressed on the apical surfaces of airway epithelial cells. TLR engagement leads to the initiation of a signaling cascade that results in a pro-inflammatory response with subsequent up-regulation of several cytokines and leads to adaptiv...

Transcutaneous immunization (TCI) is a novel vaccination route involving the topical application of vaccine antigens on the skin. The skin is an attractive site for vaccination because it is rich in various antigen-capturing immune cells. The outer skin barrier can be overcome through the use of mild chemical and/or physical treatments, including e...

Most immune responses follow Burnet's rule in that Ag recruits specific lymphocytes from a large repertoire and induces them to proliferate and differentiate into effector cells. However, the phenomenon of "original antigenic sin" stands out as a paradox to Burnet's rule of B cell engagement. Humans, upon infection with a novel influenza strain, pr...

Influenza prophylaxis would benefit from a simple method to administer influenza vaccine into skin without the need for hypodermic needles. In this study, solid metal microneedle arrays (MNs) were investigated as a system for cutaneous vaccine delivery using influenza virus antigen. The MNs with 5 monument-shaped microneedles per array were produce...

Influenza is a contagious disease caused by a pathogenic virus, with outbreaks all over the world and thousands of hospitalizations and deaths every year. Due to virus antigenic drift and short-lived immune responses, annual vaccination is required. However, vaccine coverage is incomplete, and improvement in immunization is needed. The objective of...

Influenza virus is one of the common causes of serious respiratory illness. Typically, the virus causes seasonal epidemics of disease; during a typical season 36,000 deaths and 200,000 hospitalizations are observed, with up to 1.5 million deaths worldwide, which make it the largest cause of death in the US from vaccine-preventable disease. The vacc...

We have designed a membrane-anchored form of the Toll-like receptor 5 ligand flagellin, the major proinflammatory determinant of enteropathogenic Salmonella, which was found to be glycosylated and expressed on cell surfaces. A chimeric influenza virus-like particle (cVLP) vaccine candidate containing A/PR8/34 (H(1)N(1)) hemagglutinin (HA), matrix p...

The rapid spread of human immunodeficiency virus (HIV) worldwide makes it a high priority to develop an effective vaccine. Since live attenuated or inactivated HIV is not likely to be approved as a vaccine due to safety concerns, HIV virus like particles (VLPs) offer an attractive alternative because they are safe due to the lack of a viral genome....

Extensive glycosylation and variable loops of the HIV envelope protein (Env) are reported to shield some neutralizing epitopes. Here, we investigated the immunogenicity of mutated HIV Envs presented in virus-like particles (VLPs). We immunized mice with simian human immunodeficiency virus (SHIV) VLPs containing mutant HIV Env with reduced glycosyla...

SER virus is a type 5 parainfluenza virus that does not exhibit syncytium formation, in contrast to most other paramyxoviruses. This property has been attributed, at least in part, to the presence of an extension of the cytoplasmic tail (CT) of the SER F protein, as truncations or mutations of this region resulted in enhanced fusion. In this study...