Baby's First Macrophage: Temporal Regulation of Hofbauer Cell Phenotype Influences Ligand-Mediated Innate Immune Responses across Gestation.

The importance of fetal placental macrophages (Hofbauer cell [HCs]) is underscored by their appearance 18 d postconception and maintenance through term; however, how human HCs evolve during healthy pregnancy and how microenvironment and ontogeny impact phenotype and function remain unknown. In this study, we comprehensively classify human HCs ex vivo, interrogate phenotypic plasticity, and characterize antiviral immune responses through gestation. Activated HCs were abundant in early pregnancy and decreased by term; molecular signatures emphasize inflammatory phenotypes early in gestation. Frequency of HCs with regulatory phenotypes remained high through term. Furthermore, term HCs exhibited blunted responses to stimulation, indicating reduced plasticity. IFN-λ1 is a key placental IFN that appeared less protective than IFN-α, suggesting a potential weakness in antiviral immunity. Ligand-specific responses were temporally regulated: we noted an absence of inflammatory mediators and reduced antiviral gene transcription following RIG-I activation at term despite all HCs producing inflammatory mediators following IFN-γ plus LPS stimulation. Collectively, we demonstrate sequential, evolving immunity as part of the natural history of HCs through gestation.

H1N1 influenza virus infection results in adverse pregnancy outcomes by disrupting tissue-specific hormonal regulation.

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 the Th1-Th2 shift. However, the underlying mechanisms which increase pregnant mothers' risk following influenza virus infection have not been fully elucidated. We used pregnant BALB/c mice during mid- to late gestation to determine the impact of a sub-lethal infection with A/Brisbane/59/07 H1N1 seasonal influenza virus on completion of gestation. Maternal and fetal health status was closely monitored and compared to infected non-pregnant mice. Severity of infection during pregnancy was correlated with premature rupture of amniotic membranes (PROM), fetal survival and body weight at birth, lung viral load and degree of systemic and tissue inflammation mediated by innate and adaptive immune responses. Here we report that influenza virus infection resulted in dysregulation of inflammatory responses that led to pre-term labor, impairment of fetal growth, increased fetal mortality and maternal morbidity. We observed significant compartment-specific immune responses correlated with changes in hormonal synthesis and regulation. Dysregulation of progesterone, COX-2, PGE2 and PGF2α expression in infected pregnant mice was accompanied by significant remodeling of placental architecture and upregulation of MMP-9 early after infection. Collectively these findings demonstrate the potential of a seasonal influenza virus to initiate a powerful pro-abortive mechanism with adverse outcomes in fetal health.

The safety, immunogenicity, and acceptability of inactivated influenza vaccine delivered by microneedle patch (TIV-MNP 2015): a randomised, partly blinded, placebo-controlled, phase 1 trial.

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 influenza. METHODS: The TIV-MNP 2015 study was a randomised, partly blinded, placebo-controlled, phase 1, clinical trial at Emory University that enrolled non-pregnant, immunocompetent adults from Atlanta, GA, USA, who were aged 18-49 years, naive to the 2014-15 influenza vaccine, and did not have any significant dermatological disorders. Participants were randomly assigned (1:1:1:1) to four groups and received a single dose of inactivated influenza vaccine (fluvirin: 18 µg of haemagglutinin per H1N1 vaccine strain, 17 µg of haemagglutinin per H3N2 vaccine strain, and 15 µg of haemagglutinin per B vaccine strain) (1) by microneedle patch or (2) by intramuscular injection, or received (3) placebo by microneedle patch, all administered by an unmasked health-care worker; or received a single dose of (4) inactivated influenza vaccine by microneedle patch self-administered by study participants. A research pharmacist prepared the randomisation code using a computer-generated randomisation schedule with a block size of 4. Because of the nature of the study, participants were not masked to the type of vaccination method (ie, microneedle patch vs intramuscular injection). Primary safety outcome measures are the incidence of study product-related serious adverse events within 180 days, grade 3 solicited or unsolicited adverse events within 28 days, and solicited injection site and systemic reactogenicity on the day of study product administration through 7 days after administration, and secondary safety outcomes are new-onset chronic illnesses within 180 days and unsolicited adverse events within 28 days, all analysed by intention to treat. Secondary immunogenicity outcomes are antibody titres at day 28 and percentages of seroconversion and seroprotection, all determined by haemagglutination inhibition antibody assay. The trial is completed and registered with ClinicalTrials.gov, number NCT02438423. FINDINGS: Between June 23, 2015, and Sept 25, 2015, 100 participants were enrolled and randomly assigned to a group. There were no treatment-related serious adverse events, no treatment-related unsolicited grade 3 or higher adverse events, and no new-onset chronic illnesses. Among vaccinated groups (vaccine via health-care worker administered microneedle patch or intramuscular injection, or self-administered microneedle patch), overall incidence of solicited adverse events (n=89 vs n=73 vs n=73) and unsolicited adverse events (n=18 vs n=12 vs n=14) were similar. Reactogenicity was mild, transient, and most commonly reported as tenderness (15 [60%] of 25 participants [95% CI 39-79]) and pain (11 [44%] of 25 [24-65]) after intramuscular injection; and as tenderness (33 [66%] of 50 [51-79]), erythema (20 [40%] of 50 [26-55]), and pruritus (41 [82%] of 50 [69-91]) after vaccination by microneedle patch application. The geometric mean titres were similar at day 28 between the microneedle patch administered by a health-care worker versus the intramuscular route for the H1N1 strain (1197 [95% CI 855-1675] vs 997 [703-1415]; p=0·5), the H3N2 strain (287 [192-430] vs 223 [160-312]; p=0·4), and the B strain (126 [86-184] vs 94 [73-122]; p=0·06). Similar geometric mean titres were reported in participants who self-administered the microneedle patch (all p>0·05). The seroconversion percentages were significantly higher at day 28 after microneedle patch vaccination compared with placebo (all p<0·0001) and were similar to intramuscular injection (all p>0·01). INTERPRETATION: Use of dissolvable microneedle patches for influenza vaccination was well tolerated and generated robust antibody responses. FUNDING: National Institutes of Health.

Programming the magnitude and persistence of antibody responses with innate immunity.

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, activates dendritic cells via multiple TLRs to stimulate proinflammatory cytokines. Triggering specific combinations of TLRs in dendritic cells can induce synergistic production of cytokines, which results in enhanced T-cell responses, but its impact on antibody responses remain unknown. Learning the critical parameters of innate immunity that program such antibody responses remains a major challenge in vaccinology. Here we demonstrate that immunization of mice with synthetic nanoparticles containing antigens plus ligands that signal through TLR4 and TLR7 induces synergistic increases in antigen-specific, neutralizing antibodies compared to immunization with nanoparticles containing antigens plus a single TLR ligand. Consistent with this there was enhanced persistence of germinal centres and of plasma-cell responses, which persisted in the lymph nodes for >1.5 years. Surprisingly, there was no enhancement of the early short-lived plasma-cell response relative to that observed with single TLR ligands. Molecular profiling of activated B cells, isolated 7 days after immunization, indicated that there was early programming towards B-cell memory. Antibody responses were dependent on direct triggering of both TLRs on B cells and dendritic cells, as well as on T-cell help. Immunization protected completely against lethal avian and swine influenza virus strains in mice, and induced robust immunity against pandemic H1N1 influenza in rhesus macaques.

Skin immunization with influenza vaccines.

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 immunity is limited, and results in the need for annual revaccination. The immunogenicity of the present split or subunit vaccines is also lower than that observed with whole inactivated virus, and the vaccines are not very effective in high risk groups such as the young or the elderly. Vaccine coverage is incomplete, due in part to concerns about the use of hypodermic needles for delivery. Alternative approaches for vaccination are being developed which address many of these concerns. Here we review new approaches which focus on skin immunization, including the development of needle-free delivery systems which use stable dry formulations and induce stronger and longer-lasting immune responses.

ZBTB32 is an early repressor of the CIITA and MHC class II gene expression during B cell differentiation to plasma cells.

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 plasma cell differentiation model. ZBTB32 activity occurred at a time when B lymphocyte-induced maturation protein-1 (Blimp-1), the regulator of plasma cell fate and suppressor of CIITA, was minimally induced. Ectopic expression of ZBTB32 suppressed CIITA and I-A gene expression in B cells. Short hairpin RNA depletion of ZBTB32 in a plasma cell line resulted in re-expression of CIITA and I-A. Compared with conditional Blimp-1 knockout and wild-type B cells, B cells from ZBTB32/ROG-knockout mice displayed delayed kinetics in silencing CIITA during ex vivo plasma cell differentiation. ZBTB32 was found to bind to the CIITA gene, suggesting that ZBTB32 directly regulates CIITA. Lastly, ZBTB32 and Blimp-1 coimmunoprecipitated, suggesting that the two repressors may ultimately function together to silence CIITA expression. These results introduce ZBTB32 as a novel regulator of MHC-II gene expression and a potential regulatory partner of Blimp-1 in repressing gene expression.

Targeting the skin for microneedle delivery of influenza vaccine.

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, including metal and dissolving microneedle patches that have demonstrated promising results. Additionally we analyze the immunological basis for microneedle skin immunization and targeting of the skin's dense population of antigen presenting cells, their role, characterization, and function. Additionally we analyze the importance of inflammatory signaling in the skin after microneedle delivery.

A wild boar cathelicidin peptide derivative inhibits severe acute respiratory syndrome coronavirus-2 and its drifted variants.

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 continues to mutate. Even when full vaccine coverage is achieved, vaccine-resistant mutants will likely emerge, thus requiring new annual vaccines against drifted variants analogous to influenza. A complimentary solution to this problem could be developing antiviral drugs that inhibit SARS CoV-2 and its drifted variants. Host defense peptides represent a potential source for such an antiviral as they possess broad antimicrobial activity and significant diversity across species. We screened the cathelicidin family of peptides from 16 different species for antiviral activity and identified a wild boar peptide derivative that inhibits SARS CoV-2. This peptide, which we named Yongshi and means warrior in Mandarin, acts as a viral entry inhibitor. Following the binding of SARS-CoV-2 to its receptor, the spike protein is cleaved, and heptad repeats 1 and 2 multimerize to form the fusion complex that enables the virion to enter the cell. A deep learning-based protein sequence comparison algorithm and molecular modeling suggest that Yongshi acts as a mimetic to the heptad repeats of the virus, thereby disrupting the fusion process. Experimental data confirm the binding of Yongshi to the heptad repeat 1 with a fourfold higher affinity than heptad repeat 2 of SARS-CoV-2. Yongshi also binds to the heptad repeat 1 of SARS-CoV-1 and MERS-CoV. Interestingly, it inhibits all drifted variants of SARS CoV-2 that we tested, including the alpha, beta, gamma, delta, kappa and omicron variants.

Immunity to pre-1950 H1N1 influenza viruses confers cross-protection against the pandemic swine-origin 2009 A (H1N1) influenza virus.

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 that influenza strains that circulated 50-60 y ago might provide cross-protection against the swine-origin 2009 H1N1 influenza virus. To test this, we determined the ability of representative H1N1 influenza viruses that circulated in the human population from 1930 to 2000, to induce cross-reactivity to and cross-protection against the pandemic swine-origin H1N1 virus, A/California/04/09. We show that exposure of mice to the 1947 virus, A/FM/1/47, or the 1934 virus, A/PR/8/34, induced robust cross-protective immune responses and these mice were protected against a lethal challenge with mouse-adapted A/California/04/09 H1N1 virus. Conversely, we observed that mice exposed to the 2009 H1N1 virus were protected against a lethal challenge with mouse-adapted 1947 or 1934 H1N1 viruses. In addition, exposure to the 2009 H1N1 virus induced broad cross-reactivity against H1N1 as well as H3N2 influenza viruses. Finally, we show that vaccination with the older H1N1 viruses, particularly A/FM/1/47, confers protective immunity against the 2009 pandemic H1N1 virus. Taken together, our data provide an explanation for the decreased susceptibility of the elderly to the 2009 H1N1 outbreak and demonstrate that vaccination with the pre-1950 influenza strains can cross-protect against the pandemic swine-origin 2009 H1N1 influenza virus.

Immunization by vaccine-coated microneedle arrays protects against lethal influenza virus challenge.

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 produced and coated with inactivated influenza virus A/PR/8/34 (IIV). As much as 10 microg of viral proteins could be coated onto an array of 5 microneedles, and the coated IIV was delivered into skin at high efficiency within minutes. The coated MNs were used to immunize mice in comparison with conventional intramuscular injection at the same dose. Analysis of immune responses showed that a single immunization with IIV-coated MNs induced strong antibody responses against influenza virus, with significant levels of hemagglutination inhibition activities (>1:40), which were comparable to those induced by conventional intramuscular immunization. Moreover, mice immunized by a single dose of IIV coated on MNs were effectively protected against lethal challenge by a high dose of mouse-adapted influenza virus A/PR/8/34. These results show that MNs are highly effective as a simple method of vaccine delivery to elicit protective immune responses against virus infection.

Serological memory and long-term protection to novel H1N1 influenza virus after skin vaccination.

BACKGROUND: 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. METHODS: BALB/c mice received a single dose of 5 µg inactivated A/California/04/09 virus via coated metal microneedles (MN) applied to skin or via subcutaneous injection. RESULTS: MN or subcutaneous vaccination elicited similar serum IgG and hemagglutination inhibition titers and 100% protection against lethal viral challenge 6 weeks after vaccination. Six months after vaccination, the subcutaneous group exhibited a 60% decrease in functional antibody titers and extensive lung inflammation after challenge with 10 × LD(50) of homologous virus. In contrast, the MN group maintained high functional antibody titers and IFN-γ levels, inhibition of viral replication, and no signs of lung inflammation after challenge. MN vaccination conferred complete protection against lethal challenge, whereas subcutaneous vaccination induced only partial protection. These findings were further supported by high numbers of bone marrow plasma cells and spleen antibody-secreting cells detected in the MN group. CONCLUSIONS: A single skin vaccination with MN induced potent long-lived immunity and improved protection against the 2009 H1N1 influenza virus, compared with subcutaneous injection.

A Novel Neuraminidase Virus-Like Particle Vaccine Offers Protection Against Heterologous H3N2 Influenza Virus Infection in the Porcine Model.

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 generally ignored in the formulation and development of influenza vaccines. In this study, we evaluate the immunogenicity and efficacy against challenge of a novel NA virus-like particles (VLPs) vaccine in the porcine model. We developed an NA2 VLP vaccine containing the NA protein from A/Perth/16/2009 (H3N2) and the matrix 1 (M1) protein from A/MI/73/2015, formulated with a water-in-oil-in-water adjuvant. Responses to NA2 VLPs were compared to a commercial adjuvanted quadrivalent whole inactivated virus (QWIV) swine IAV vaccine. Animals were prime boost vaccinated 21 days apart and challenged four weeks later with an H3N2 swine IAV field isolate, A/swine/NC/KH1552516/2016. Pigs vaccinated with the commercial QWIV vaccine demonstrated high hemagglutination inhibition (HAI) titers but very weak anti-NA antibody titers and subsequently undetectable NA inhibition (NAI) titers. Conversely, NA2 VLP vaccinated pigs demonstrated undetectable HAI titers but high anti-NA antibody titers and NAI titers. Post-challenge, NA2 VLPs and the commercial QWIV vaccine showed similar reductions in virus replication, pulmonary neutrophilic infiltration, and lung inflammation compared to unvaccinated controls. These data suggest that anti-NA immunity following NA2 VLP vaccination offers comparable protection to QWIV swine IAV vaccines inducing primarily anti-HA responses.

Original antigenic sin responses to influenza viruses.

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, produce Abs against older viral strains at the expense of responses to novel, protective antigenic determinants. This exacerbates the severity of the current infection. This blind spot of the immune system and the redirection of responses to the "original Ag" rather than to novel epitopes were described fifty years ago. Recent reports have questioned the existence of this phenomenon. Hence, we revisited this issue to determine the extent to which original antigenic sin is induced by variant influenza viruses. Using two related strains of influenza A virus, we show that original antigenic sin leads to a significant decrease in development of protective immunity and recall responses to the second virus. In addition, we show that sequential infection of mice with two live influenza virus strains leads to almost exclusive Ab responses to the first viral strain, suggesting that original antigenic sin could be a potential strategy by which variant influenza viruses subvert the immune system.

Bivalent vaccination with NA1 and NA2 neuraminidase virus-like particles is protective against challenge with H1N1 and H3N2 influenza A viruses in a murine model.

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 A/Perth/16/2009. Intramuscular vaccination of mice with NA2 VLPs induced high anti-NA serum IgG levels capable of inhibiting NA activity. NA2 VLP vaccination protected against mortality in a lethal A/Hong Kong/1/1968 (H3N2) virus challenge model, but not against lethal challenge with A/California/04/2009 (H1N1) virus. However, bivalent vaccination with NA1 and NA2 VLPs demonstrated no antigenic competition in anti-NA IgG responses and protected against lethal challenge with H1N1 and H3N2 viruses. Here we demonstrate that vaccination with NA VLPs is protective against influenza challenge and supports focusing on anti-NA responses in the development of future vaccination strategies.

Fabrication of microneedle patches with lyophilized influenza vaccine suspended in organic solvent.

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 incorporate lyophilized inactivated H1N1 influenza vaccine into MN patches during manufacturing by suspending freeze-dried vaccine in anhydrous organic solvent during the casting process. Using a casting formulation containing chloroform and polyvinylpyrrolidone, lyophilized influenza vaccine maintained activity during manufacturing and subsequent storage for 3 months at 40 °C. Influenza vaccination using these MN patches generated strong immune responses in a murine model. This manufacturing process may enable vaccines and other biologics to be stabilized by lyophilization and administered via a MN patch.

The amphibian peptide Yodha is virucidal for Zika and dengue viruses.

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 adults, ZIKV infection has been correlated with Guillain-Barre syndrome. Despite the public health threat posed by ZIKV, neither a vaccine nor antiviral drugs for use in humans are currently available. We have identified an amphibian host defense peptide, Yodha, which has potent virucidal activity against ZIKV. It acts directly on the virus and destroys Zika virus particles within 5 min of exposure. The Yodha peptide was effective against the Asian, African, and South American Zika virus strains and has the potential to be developed as an antiviral therapeutic in the fight against Zika virus. The peptide was also effective against all four dengue virus serotypes. Thus, Yodha peptide could potentially be developed as a pan-therapeutic for Zika and dengue viruses.

Transcutaneous immunization with influenza vaccines.

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 ethanol-water hydration and stripping, which allows for large vaccine molecules or even particulate antigens to gain access to the skin's immune cells. The use of toxin adjuvants such as cholera or heat-labile toxins was demonstrated to enhance the immunogenicity of vaccine antigens, probably due to their stimulatory effects on immune cells. Oleic acid or retinoic acid, known as permeation enhancers or immune modulators, were found to increase immune responses to inactivated whole-influenza viral vaccines. The further development of more effective delivery systems and nontoxic adjuvants is needed to enhance the efficacy of this approach to vaccination.

Zika virus-induced neuro-ocular pathology in immunocompetent mice correlates with anti-ganglioside autoantibodies.

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 neutralization and elevated antibody-mediated infection among Fcγ-R bearing cells. The use of interferon-deficient mice for ZIKV studies limits elucidation of antibody-dependent enhancement (ADE) and long-term pathology (≥120 days), due to high lethality post-infection. Here we used immunocompetent BALB/c mice, which generate robust humoral immune responses, to investigate long-term impacts of ZIKV infection. A high infectious dose (1x106 FFU per mouse) of ZIKV was administered intravenously. Control animals received a single dose of anti-IFNAR blocking monoclonal antibody and succumbed to lethal neurological pathology within 13 days. Immunocompetent mice exhibited motor impairment such as arthralgia, as well as ocular inflammation resulting in retinal vascular damage, and corneal edema. This pathology persisted 100 days after infection with evidence of chronic inflammation in immune-privileged tissues, demyelination in the hippocampus and motor cortex regions of the brain, and retinal/corneal hyperplasia. Anti-inflammatory transcriptional responses were tissue-specific, likely contributing to differential pathology in these organs. Pathology in immunocompetent animals coincided with weakly neutralizing antibodies and increased ADE among ZIKV strains (PRVABC59, FLR, and MR766) and all Dengue virus (DENV) serotypes. These antibodies were autoreactive to GBS-associated gangliosides. This study highlights the importance of longevity studies in ZIKV infection and confirms the role of anti-ganglioside antibodies in ZIKV-induced neuro-ocular disease.

Pregnancy Downregulates Plasmablast Metabolic Gene Expression Following Influenza Without Altering Long-Term Antibody Function.

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 maternal anti-influenza immunity remains to be elucidated. Previously, we linked seasonal influenza infection to clinical observations of adverse pregnancy outcomes, enhanced lung and placental histopathology, and reduced control of viral replication in lungs of infected pregnant mothers. Here, we expand on this work and demonstrate that lower infectious doses of the pandemic A/California/07/2009 influenza virus generated adverse gestational outcomes similar to higher doses of seasonal viruses. Mice infected during pregnancy demonstrated lower hemagglutination inhibition and neutralizing antibody titers than non-pregnant animals until 63 days post infection. These differences in humoral immunity suggest that pregnancy impacts antibody maturation mechanisms without alterations to B cell frequency or antibody secretion. This is further supported by transcriptional analysis of plasmablasts, which demonstrate downregulated B cell metabolism and post-translational modification systems only among pregnant animals. In sum, these findings corroborate a link between adverse pregnancy outcomes and severe pathology observed during pandemic influenza infection. Furthermore, our data propose that pregnancy directly confounds humoral responses following influenza infection which resolves post-partem. Additional studies are required to specify the involvement of plasmablast metabolism with early humoral immunity abnormalities to best guide vaccination strategies and improve our understanding of the immunological consequences of pregnancy.

Cutaneous vaccination ameliorates Zika virus-induced neuro-ocular pathology via reduction of anti-ganglioside antibodies.

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 vaccination can confer robust, long-term protection against ZIKV, preventing neuro-ocular pathology and long-term inflammation in immune-privileged compartments, BALB/c mice received two doses of unadjuvanted inactivated whole ZIKV vaccine (ZVIP) intramuscularly (IM) or cutaneously with dissolving microneedle patches (MNP). MNP immunization induced significantly higher B and T cell responses compared to IM vaccination, resulting in increased antibody titers with greater avidity for ZPIV as well as increased numbers of IFN-γ, TNF-α, IL- and IL-4 secreting T cells. When compared to IM vaccination, antibodies generated by cutaneous vaccination demonstrated greater neutralization activity, increased cross-reactivity with Asian and African lineage ZIKV strains (PRVABC59, FLR, and MR766) and Dengue virus (DENV) serotypes, limited ADE, and lower reactivity to GBS-associated gangliosides. MNP vaccination effectively controlled viremia and inflammation, preventing neuro-ocular pathology. Conversely, IM vaccination exacerbated ocular pathology, resulting in uncontrolled, long-term inflammation. Importantly, neuro-ocular pathology correlated with anti-ganglioside antibodies implicated in demyelination and GBS. This study highlights the importance of longevity studies in ZIKV immunization, and the need of exploring alternative vaccination platforms to improve the quality of vaccine-induced immune responses.