Intranasal Inoculation Research Articles

A Novel Organized Nasopharynx-Associated Lymphoid Tissue in Teleosts That Expresses Molecular Markers Characteristic of Mammalian Germinal Centers.

Nasal immunity is an ancient and conserved arm of the mucosal immune system in vertebrates. In teleost fish, we previously reported the presence of a nasopharynx-associated lymphoid tissue (NALT) characterized by scattered immune cells located in the trout olfactory lamellae. This diffuse NALT mounts innate and adaptive immune responses to nasal infection or vaccination. In mammals, lymphoid structures such as adenoids and tonsils support affinity maturation of the adaptive immune response in the nasopharyngeal cavity. These structures, known as organized NALT (O-NALT), have not been identified in teleost fish to date, but their evolutionary forerunners exist in sarcopterygian fish. In this study, we report that the rainbow trout nasal cavity is lined with a lymphoepithelium that extends from the most dorsal opening of the nares to the ventral nasal cavity. Within the nasal lymphoepithelium we found lymphocyte aggregates called O-NALT in this study that are composed of ∼ 56% CD4+, 24% IgM+, 16% CD8α+, and 4% IgT+ lymphocytes and that have high constitutive aicda mRNA expression. Intranasal (i.n.) vaccination with live attenuated infectious hematopoietic necrosis virus triggers expansions of B and T cells and aicda expression in response to primary i.n. vaccination. IgM+ B cells undergo proliferation and apoptosis within O-NALT upon prime but not boost i.n. vaccination. Our results suggest that novel mucosal microenvironments such as O-NALT may be involved in the affinity maturation of the adaptive immune response in early vertebrates.

Effects of prenatal dietary rumen-protected choline supplementation during late gestation on calf growth, metabolism, and vaccine response.

The objective of this study was to examine the effects of prenatal supplementation and dose of rumen-protected choline (RPC) on neonatal calf growth, metabolism, and vaccine response. Parous Holstein cows were blocked by calving month and randomly assigned within block to receive 45 g/d of RPC [20.4 g/d of choline ions (CHOL45), n = 19], 30 g/d of RPC [13.6 g/d of choline ions (CHOL30), n = 22], or no RPC (CON, n = 19) as a top-dress, starting 24 d before expected calving. Calf body weights were recorded for the first 3 wk of life. All calves were fed colostrum replacer (300 g of IgG) at birth, and apparent efficiency of IgG absorption was calculated. On d 1, 7, 14, and 21, blood samples were taken to quantify plasma reactive oxygen and nitrogen species, antioxidant potential, haptoglobin, nonesterified fatty acids (NEFA), β-hydroxybutyrate, and glucose. Calves received an intranasal vaccine at birth, and nasal secretions were collected on d 0, 7, 10, 14, and 21 to quantify bovine respiratory syncytial virus-specific IgA. Data were analyzed using linear mixed models including the fixed effects of treatment, time (when applicable), calf sex, and prepartum dam data (-24 d) along with interactions. Treatment did not affect calf body weight, β-hydroxybutyrate, or glucose concentrations. For apparent efficiency of IgG absorption, treatment interacted with the dam's prepartum body condition score. Where the dam's body condition score was ≤3.25, IgG absorption was reduced in calves born from CHOL45 dams as compared with calves from either CHOL30 or CON dams. Calves from CHOL30 dams had a lesser oxidative stress index (OSi; reactive oxygen and nitrogen species/antioxidant potential) than calves from CON dams. Haptoglobin concentrations were less in heifer calves from CHOL45 dams as compared with heifers from CON dams. The dam's prepartum NEFA concentration interacted with treatment. When dam NEFA was minimal, calves from CHOL45 and CHOL30 dams had greater or tended to have greater NEFA, respectively. Conversely, when dam NEFA was greater, calves from CHOL30 and CHOL45 dams had lesser or tended to have lesser NEFA than calves from CON dams, respectively. For vaccine response, treatment interacted with the dam's prepartum OSi. Among calves born from dams with a greater OSi, calves from CHOL45 and CHOL30 dams had lesser bovine respiratory syncytial virus-specific IgA concentrations in nasal secretions as compared with CON. Prenatal RPC supplementation during late gestation affected IgG absorption, neonatal calf metabolism, and vaccine response with some effects dependent on the dam's prepartum parameters.

Intranasal Sendai virus-based SARS-CoV-2 vaccine using a mouse model.

The coronavirus disease 2019 (COVID-19) epidemic remains worldwide. The usefulness of the intranasal vaccine and boost immunization against severe acute respiratory syndrome-related coronavirus (SARS-CoV-2) has recently received much attention. We developed an intranasal SARS-CoV-2 vaccine by loading the receptor binding domain of the S protein (S-RBD) of SARS-CoV-2 as an antigen into an F-deficient Sendai virus vector. After the S-RBD-Fd antigen with trimer formation ability was intranasally administered to mice, S-RBD-specific IgM, IgG, IgA, and neutralizing antibody titers were increased in serum or bronchoalveolar lavage fluid for 12 weeks. Furthermore, in mice that received a booster dose at week 8, a marked increase in neutralizing antibodies in the serum and bronchoalveolar lavage fluid was observed at the final evaluation at week 12, which neutralized the pseudotyped lentivirus expressing the SARS-CoV-2 spike protein, indicating the usefulness of the Sendai virus-based SARS-CoV-2 intranasal vaccine.

Effect of colonisation with Neisseria lactamica on cross-reactive anti-meningococcal B-cell responses: a randomised, controlled, human infection trial

Pharyngeal colonisation by the commensal bacterium Neisseria lactamica inhibits colonisation by Neisseria meningitidis and has an inverse epidemiological association with meningococcal disease. The mechanisms that underpin this relationship are unclear, but could involve the induction of cross-reactive immunity. In this study, we aimed to evaluate whether colonisation with N lactamica induces N lactamica-specific B-cell responses that are cross-reactive with N meningitidis. In this randomised, placebo-controlled, human infection trial at University Hospital Southampton Clinical Research Facility (Southampton, UK), healthy adults aged 18-45 years were randomly assigned (2:1) to receive intranasal inoculation with either 105 colony-forming units of N lactamica in 1 mL phosphate-buffered saline (PBS) or 1 mL PBS alone. Participants and researchers conducting participant sampling and immunological assays were masked to allocation. The primary endpoint was the frequency of circulating N lactamica-specific plasma cells and memory B cells after N lactamica inoculation (day 7-28) compared with baseline values (day 0), measured using enzyme-linked immunospot assays. The secondary endpoint was to measure the frequency of N meningitidis-specific B cells. In a second study, we measured the effect of duration of N lactamica colonisation on seroconversion by terminating carriage at either 4 days or 14 days with single-dose oral ciprofloxacin. The studies are now closed to participants. The trials are registered with ClinicalTrials.gov, NCT03633474 and NCT03549325. Of 50 participants assessed for eligibility between Sept 5, 2018, and March 3, 2019, 31 were randomly assigned (n=20 N lactamica, n=11 PBS). Among the 17 participants who were colonised with N lactamica, the median baselines compared with peak post-colonisation N lactamica-specific plasma-cell frequencies (per 105 peripheral blood mononuclear cells) were 0·0 (IQR 0·0-0·0) versus 5·0 (1·5-10·5) for IgA-secreting plasma cells (p<0·0001), and 0·0 (0·0-0·0) versus 3·0 (1·5-9·5) for IgG-secreting plasma cells (p<0·0001). Median N lactamica-specific IgG memory-B-cell frequencies (percentage of total IgG memory B cells) increased from 0·0024% (0·0000-0·0097) at baseline to 0·0384% (0·0275-0·0649) at day 28 (p<0·0001). The frequency of N meningitidis-specific IgA-secreting and IgG-secreting plasma cells and memory B cells also increased signficantly in participants who were colonised with N lactamica. Upper respiratory tract symptoms were reported in ten (50%) of 20 participants who were inoculated with N lactamica and six (55%) of 11 participants who were inoculated with PBS (p>0·99). Three additional adverse events (two in the N lactamica group and one in the PBS group) and no serious adverse events were reported. In the second study, anti-N lactamica and anti-N meningitidis serum IgG titres increased only in participants who were colonised with N lactamica for 14 days. Natural immunity to N meningitidis after colonisation with N lactamica might be due to cross-reactive adaptive responses. Exploitation of this microbial mechanism with a genetically modified live vector could protect against N meningitidis colonisation and disease. Wellcome Trust, Medical Research Council, and NIHR Southampton Biomedical Research Centre.

Detection of Airborne Influenza A and SARS-CoV-2 Virus Shedding following Ocular Inoculation of Ferrets.

Despite reports of confirmed human infection following ocular exposure with both influenza A virus (IAV) and SARS-CoV-2, the dynamics of virus spread throughout oculonasal tissues and the relative capacity of virus transmission following ocular inoculation remain poorly understood. Furthermore, the impact of exposure route on subsequent release of airborne viral particles into the air has not been examined previously. To assess this, ferrets were inoculated by the ocular route with A(H1N1)pdm09 and A(H7N9) IAVs and two SARS-CoV-2 (early pandemic Washington/1 and Delta variant) viruses. Virus replication was assessed in both respiratory and ocular specimens, and transmission was evaluated in direct contact or respiratory droplet settings. Viral RNA in aerosols shed by inoculated ferrets was quantified with a two-stage cyclone aerosol sampler (National Institute for Occupational Safety and Health [NIOSH]). All IAV and SARS-CoV-2 viruses mounted a productive and transmissible infection in ferrets following ocular inoculation, with peak viral titers and release of virus-laden aerosols from ferrets indistinguishable from those from ferrets inoculated by previously characterized intranasal inoculation methods. Viral RNA was detected in ferret conjunctival washes from all viruses examined, though infectious virus in this specimen was recovered only following IAV inoculation. Low-dose ocular-only aerosol exposure or inhalation aerosol exposure of ferrets to IAV similarly led to productive infection of ferrets and shedding of aerosolized virus. Viral evolution during infection was comparable between all inoculation routes examined. These data support that both IAV and SARS-CoV-2 can establish a high-titer mammalian infection following ocular exposure that is associated with rapid detection of virus-laden aerosols shed by inoculated animals. IMPORTANCE Documented human infection with influenza viruses and SARS-CoV-2 has been reported among individuals wearing respiratory protection in the absence of eye protection, highlighting the capacity of these respiratory tract-tropic viruses to exploit nonrespiratory routes of exposure to initiate productive infection. However, comprehensive evaluations of how ocular exposure may modulate virus pathogenicity and transmissibility in mammals relative to respiratory exposure are limited and have not investigated multiple virus families side by side. Using the ferret model, we show that ocular exposure with multiple strains of either coronaviruses or influenza A viruses leads to an infection that results in shedding of detectable aerosolized virus from inoculated animals, contributing toward onward transmission of both viruses to susceptible contacts. Collectively, these studies support that the ocular surface represents a susceptible mucosal surface that, if exposed to a sufficient quantity of either virus, permits establishment of an infection which is similarly transmissible as that following respiratory exposure.

Intranasal inoculation of IFN-λ resolves SARS-CoV-2 lung infection via the rapid reduction of viral burden and improvement of tissue damage.

The innate immune responses of upper airway could further our understanding toward antiviral strategies against SARS-CoV-2. We characterize the potential of interferon (IFN)-λ as an innate immune inducer for the rapid clearance of SARS-CoV-2 in the lung and the therapeutic efficacy of intranasal inoculation of IFN-λ to resolve acute lung infection. Syrian golden hamsters were infected with SARS-CoV-2 and the dynamics of SARS-CoV-2 infection depending on IFN-λ inoculation were tested. SARS-CoV-2-infected Syrian golden hamsters exhibited a significant decrease in body weight and high viral mRNA level at 3 days post-infection (dpi). Although viral replication was reduced completely from 7 dpi, the pathologic findings remained prominent until 14 dpi in the lung of hamsters. The transcription of IFN-λ was significantly induced in response to SARS-CoV-2 infection with the increase of IFN-stimulated genes. Intranasal inoculation of IFN-λ restricted SARS-CoV-2 replication in the lungs of infected completely from 3 dpi with markedly reduction of inflammatory cytokines. The transcriptional phenotypes were altered to the direction of damage repair and tissue remodeling in the lungs of SARS-CoV-2-infected hamsters following intranasal inoculation of IFN-λ, which improved SARS-CoV-2-caused lung damage. Collectively, our findings suggest that IFN-λ might be a potent innate immune inducer in the lung and intranasal inoculation of IFN-λ resolves SARS-CoV-2 infection with rapid viral clearance and improvement of lung damage.

Intranasal Vaccination with rePcrV Protects against Pseudomonas aeruginosa and Generates Lung Tissue-Resident Memory T Cells.

Tissue-resident memory T (TRM) cells are immune sentinels that bear a key role in the local immune system and rapidly respond to infection. Our previous studies showed that mucosal immunization via intranasal pathways was more effective than intramuscular route. However, the mechanism of enhanced protective immunity remains unclear. Here, we formulated a Pseudomonas aeruginosa vaccine composed of type III secretion protein PcrV from P. aeruginosa and curdlan adjuvant and then administered by the intranasal route. Flow cytometry and immunofluorescence staining showed that the ratio of CD44+CD62L-CD69+CD4+ TRM cells induced by this vaccine was significantly increased, and IL-17A production was notably enhanced. Further analysis revealed that vaccinated mice can protect against the P. aeruginosa challenge even after administration with FTY720 treatment. What is more, our results showed that CD4+ TRM might be involved in the recruitment of neutrophils and provided partial protection against Pseudomonas aeruginosa. Taken together, these data demonstrated that CD4+ TRM cells were elicited in lung tissues after immunization with rePcrV and contributed to protective immunity. Furthermore, it provided novel strategies for the development of vaccines for P. aeruginosa and other respiratory-targeted vaccines.

&lt;i&gt;In situ&lt;/i&gt; gels as a modern method of intranasal vaccine delivery

The continuous emergence of new pathogens and the evolution of microbial drug resistance make it absolutely necessary to develop innovative, effective vaccination strategies. Use of nasal vaccination can increase convenience, safety, cause both local and systemic immune reactions. Intranasal administration nevertheless has a number of shortcomings that can be overcome by using the latest achievements of pharmaceutical science. One of the aspects of such solution may be the use of systems for the production of intranasal vaccines in situ polymer compositions that provide a directed sol-gel transition controlled by the physiological conditions of the nasal cavity. At the same time, the gelation of the administered dose in contact with the nasal mucosa involves prolonged exposure of the drug at the injection site, greater mucoadhesion, counteraction to mucociliary clearance, modified and more complete release. A number of both foreign and domestic manufacturers produces polymers such as chitosan, gums, polyoxyethylene and polyoxypropylene block copolymers (poloxamers, proxanols), carbomers. For effective pharmaceutical development of new intranasal IBD delivery systems corresponding to the QbD concept, not only the knowledge of the range of excipients is necessary, but also simple, accessible, and reproducible methods for determining indicators that define the critical parameters of such delivery systems. In accordance with the conducted scientific search, the main indicators of standardization of in situ intranasal systems were identified: temperature and time of gel formation, gel strength, rheological characteristics, mucoadhesion, release, nasal mucociliary clearance time.

Intranasal therapeutic vaccine containing HBsAg and HBcAg for patients with chronic hepatitis B; 18months follow-up results of phase IIa clinical study.

HBsAg loss with anti-HBs acquisition is considered a functional cure and ideal treatment goal for patients with CHB. Our group have reported the efficacy of therapeutic vaccine with HBsAg and HBcAg (NASVAC) by intranasal and subcutaneous injection. In this study, we investigated the safety and efficacy of newly developed CVP-NASVAC, which contained NASVAC with mucoadhesive carboxyl vinyl polymer (CVP) in the dedicated device. A single dose, open-label, phase IIa clinical trial of CVP-NASVAC was conducted. Patients with CHB treated with nucleoside/nucleotide analogs (NAs) and HBV carriers not undergoing anti-HBV treatment were enrolled. CVP-NASVAC was injected through the nose for, in total, 10 times. Participants were followed-up for 18months, and their HBsAg reduction and anti-HBs induction assessed as endpoints. Among the patients with CHB treated with NAs (n=27) and HBV carriers without NAs (n=36), 74.1% and 75.0% exhibited reductions in their baseline HBsAg, and the mean reductions were -0.1454 log10 IU/ml (p<0.05) and -0.2677 log10 IU/ml (p<0.05), respectively. Anti-HBs antibody was detected in 40.7% and 58.3% of patients treated with and without NAs, respectively. Six of 71 (9.5%) patients were functionally cured after the CVP-NASVAC treatment. Anti-HBs induction and HBsAg reduction was observed after CVP-NASVAC treatment in some patients with CHB. The CVP-NASVAC is a safe treatment, which might expect to achieve functional cure for patients with CHB.

An intranasal vaccine targeting the receptor binding domain of SARS-CoV-2 elicits a protective immune response

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the pathogen responsible for COVID-19, has caused an ongoing worldwide pandemic. Due to the rapid emergence of variants of concern (VOCs), novel vaccines and vaccination strategies are urgently needed. We developed an intranasal vaccine consisting of the SARS-CoV-2 receptor binding domain (RBD) fused to the antibody Fc fragment (RBD-Fc). RBD-Fc could induce strong humoral immune responses via intranasal vaccination. Notably, this immunogen could efficiently induce IgG and IgA and establish mucosal immunity in the respiratory tract. The induced antibodies could efficiently neutralize wild-type SARS-CoV-2 and currently identified SARS-CoV-2 VOCs, including the Omicron variant. In a mouse model, intranasal immunization could provide complete protection against a lethal SARS-CoV-2 challenge. Unfortunately, the limitation of our study is the small number of animals used in the immune response analysis. Our results suggest that recombinant RBD-Fc delivered via intranasal vaccination has considerable potential as a mucosal vaccine that may reduce the risk of SARS-CoV-2 infection.

A replication-deficient H9N2 influenza virus carrying H5 hemagglutinin conferred protection against H9N2 and H5N1 influenza viruses in mice

H5N1 and H9N2 influenza viruses have been reported to cause human infections and are believed to have pandemic potential. The vaccine is an effective tool to prevent influenza virus infection. However, inactivated influenza vaccines sometimes result in low antigenicity as result leads to generating of incomplete immune protection in the form of low cellular and humoral immunity. While the low temperature adapted, traditional live attenuated influenza vaccine (LAIV) is associated with the potential risk to revert to a virulent phenotype, there appears an essential need for an alternative potent methodology to design and develop influenza vaccines with substantial safety and efficacy which may confer solid protection against H9N2 or H5N1 influenza virus infections. In the present study, a replication-deficient recombinant influenza virus, WM01ma-HA(H5), expressing hemagglutinin (HA) of both H9N2 and H5N1 subtypes was developed. The chimeric gene segment expressing HA(H5), was designed using the sequence of an open reading frame (ORF) of HA adopted from A/wild duck/Hunan/021/2005(H5N1)(HN021ma) which was flanked by the NA packaging signals of mouse-adapted strain A/Mink/Shandong/WM01/2014(H9N2)(WM01ma). Due to the absence of ORF of structural protein NA, the replication of this engineered H9N2 influenza viruses WM01ma-HA(H5) was hampered in vitro and in vivo but was well competent in MDCK cells stably expressing the NA protein of WM01ma. Intranasal vaccination of mice with WM01ma-HA(H5) stimulated robust immune response without any clinical signs and conferred complete protection from infection by H5N1 or H9N2 subtype influenza viruses.

Long-Term Pulmonary Dysfunction by Hyperoxia Exposure during Severe Viral Lower Respiratory Tract Infection in Mice.

Viral-induced lower respiratory tract infection (LRTI), mainly by respiratory syncytial virus (RSV), causes a major health burden among young children and has been associated with long-term respiratory dysfunction. Children with severe viral LRTI are frequently treated with oxygen therapy, hypothetically posing an additional risk factor for pulmonary sequelae. The main goal of this study was to determine the effect of concurrent hyperoxia exposure during the acute phase of viral LRTI on long-term pulmonary outcome. As an experimental model for severe RSV LRTI in infants, C57Bl/6J mice received an intranasal inoculation with the pneumonia virus of mice J3666 strain at post-natal day 7, and were subsequently exposed to hyperoxia (85% O2) or normoxia (21% O2) from post-natal day 10 to 17 during the acute phase of disease. Long-term outcomes, including lung function and structural development, were assessed 3 weeks post-inoculation at post-natal day 28. Compared to normoxic conditions, hyperoxia exposure in PVM-inoculated mice induced a transient growth arrest without subsequent catchup growth, as well as a long-term increase in airway resistance. This hyperoxia-induced pulmonary dysfunction was not associated with developmental changes to the airway or lung structure. These findings suggest that hyperoxia exposure during viral LRTI at young age may aggravate subsequent long-term pulmonary sequelae. Further research is needed to investigate the specific mechanisms underlying this alteration to pulmonary function.

Cross-protection against African swine fever virus upon intranasal vaccination is associated with an adaptive-innate immune crosstalk.

African swine fever virus (ASFV) is causing a worldwide pandemic affecting the porcine industry and leading to important global economic consequences. The virus causes a highly lethal hemorrhagic disease in wild boars and domestic pigs. Lack of effective vaccines hampers the control of virus spread, thus increasing the pressure on the scientific community for urgent solutions. However, knowledge on the immune components associated with protection is very limited. Here we characterized the in vitro recall response induced by immune cells from pigs intranasally vaccinated with the BA71ΔCD2 deletion mutant virus. Vaccination conferred dose-dependent cross-protection associated with both ASFV-specific antibodies and IFNγ-secreting cells. Importantly, bulk and single-cell transcriptomics of blood and lymph node cells from vaccinated pigs revealed a positive feedback from adaptive to innate immunity. Indeed, activation of Th1 and cytotoxic T cells was concomitant with a rapid IFNγ-dependent triggering of an inflammatory response characterized by TNF-producing macrophages, as well as CXCL10-expressing lymphocytes and cross-presenting dendritic cells. Altogether, this study provides a detailed phenotypic characterization of the immune cell subsets involved in cross-protection against ASFV, and highlights key functional immune mechanisms to be considered for the development of an effective ASF vaccine.

Intranasal Exposure to Rift Valley Fever Virus Live-Attenuated Strains Leads to High Mortality Rate in Immunocompetent Mice.

Rift Valley fever virus (RVFV) is a pathogenic arthropod-borne virus that can cause serious illness in both ruminants and humans. The virus can be transmitted by an arthropod bite or contact with contaminated fluids or tissues. Two live-attenuated veterinary vaccines-the Smithburn (SB) and Clone 13 (Cl.13)-are currently used during epizootic events in Africa. However, their residual pathogenicity (i.e., SB) or potential of reversion (i.e., Cl.13) causes important adverse effects, strongly limiting their use in the field. In this study, we infected immunocompetent mice with SB or Cl.13 by a subcutaneous or an intranasal inoculation. Interestingly, we found that, unlike the subcutaneous infection, the intranasal inoculation led to a high mortality rate. In addition, we detected high titers and viral N antigen levels in the brain of both the SB- and Cl.13-infected mice. Overall, we unveil a clear correlation between the pathogenicity and the route of administration of both SB and Cl.13, with the intranasal inoculation leading to a stronger neurovirulence and higher mortality rate than the subcutaneous infection.

Intranasal nanoemulsion adjuvanted S-2P vaccine demonstrates protection in hamsters and induces systemic, cell-mediated and mucosal immunity in mice.

With the rapid progress made in the development of vaccines to fight the SARS-CoV-2 pandemic, almost >90% of vaccine candidates under development and a 100% of the licensed vaccines are delivered intramuscularly (IM). While these vaccines are highly efficacious against COVID-19 disease, their efficacy against SARS-CoV-2 infection of upper respiratory tract and transmission is at best temporary. Development of safe and efficacious vaccines that are able to induce robust mucosal and systemic immune responses are needed to control new variants. In this study, we have used our nanoemulsion adjuvant (NE01) to intranasally (IN) deliver stabilized spike protein (S-2P) to induce immunogenicity in mouse and hamster models. Data presented demonstrate the induction of robust immunity in mice resulting in 100% seroconversion and protection against SARS-CoV-2 in a hamster challenge model. There was a significant induction of mucosal immune responses as demonstrated by IgA- and IgG-producing memory B cells in the lungs of animals that received intranasal immunizations compared to an alum adjuvanted intramuscular vaccine. The efficacy of the S-2P/NE01 vaccine was also demonstrated in an intranasal hamster challenge model with SARS-CoV-2 and conferred significant protection against weight loss, lung pathology, and viral clearance from both upper and lower respiratory tract. Our findings demonstrate that intranasal NE01-adjuvanted vaccine promotes protective immunity against SARS-CoV-2 infection and disease through activation of three arms of immune system: humoral, cellular, and mucosal, suggesting that an intranasal SARS-CoV-2 vaccine may play a role in addressing a unique public health problem and unmet medical need.

Long-Lasting Protection Induced by a Polyanhydride Nanovaccine against Respiratory Syncytial Virus in an Outbred Mouse Model.

Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infections in children. In humans, natural infection with RSV affords only partial long-term protection from reinfection, and there is no licensed RSV vaccine currently available. We have developed a new vaccine candidate, termed RSVNanoVax, composed of polyanhydride nanoparticles encapsulating the RSV prefusion F protein and a CpG 1668 oligodeoxynucleotide adjuvant. We recently reported that vaccination of inbred BALB/c mice with RSVNanoVax induced both RSV-specific cellular and humoral immunity, which provided protection from viral replication and RSV-induced disease. To further assess the efficacy of RSVNanoVax, here, we utilized outbred Swiss Webster mice to examine vaccine efficacy in a more genetically diverse population. Following intranasal prime-boost vaccination with RSVNanoVax, Swiss Webster mice exhibited robust titers of systemic RSV F-directed IgG antibodies and RSV F-directed IgA within the lungs and nasal passages that were sustained out to at least 1 year post-vaccination. Serum antibodies maintained robust neutralizing activity against both RSV A and B strains. Following RSV challenge, vaccinated Swiss Webster mice exhibited rapid viral clearance from the lungs. Overall, our results indicate that RSVNanoVax represents a promising RSV vaccine candidate capable of providing long-term protection and immunity in a genetically diverse population. IMPORTANCE Respiratory syncytial virus (RSV) infection causes thousands of infections and deaths in children and elderly adults each year. Research in this field is of great importance as there remains no licensed vaccine to prevent RSV infections. We developed a novel vaccine candidate, RSVNanoVax, utilizing the RSV prefusion F protein encapsulated in polyanhydride nanoparticles. Here, we show that the intranasal delivery of RSVNanoVax protected outbred mice from viral replication within the lungs when challenged with RSV out to 1 year post-vaccination. Additionally, RSV-specific antibody responses were generated in both the serum and lung tissue and sustained long-term. These results demonstrate that our vaccine is an encouraging candidate for driving long-term protection in the lungs in a genetically diverse population.

Unadjuvanted intranasal spike vaccine elicits protective mucosal immunity against sarbecoviruses.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has highlighted the need for vaccines that not only prevent disease but also prevent transmission. Parenteral vaccines induce robust systemic immunity but poor immunity at the respiratory mucosa. We developed a vaccine strategy that we call "prime and spike," which leverages existing immunity generated by primary vaccination (prime) to elicit mucosal immune memory within the respiratory tract by using unadjuvanted intranasal spike boosters (spike). We show that prime and spike induces robust resident memory B and T cell responses, induces immunoglobulin A at the respiratory mucosa, boosts systemic immunity, and completely protects mice with partial immunity from lethal SARS-CoV-2 infection. Using divergent spike proteins, prime and spike enables the induction of cross-reactive immunity against sarbecoviruses.

A rational strategy for the maintenance of antiviral immunity to new SARS-CoV-2 strains

New variants of SARS-CoV-2 such as Omicron BA.2, BA.4/5, BA.2.12.1 and BA 2.75 are characterized by higher infectivity and the ability to escape virus-neutralizing antibodies against previous coronavirus variants. The S-trimer of BA.2 and its phylogenetic derivatives are characterized by a predominant Up-conformation, which facilitates the interaction with ACE2 on target cells and promotes the resistance to neutralizing antibodies. The immunity acquired from the infection with earlier strains is non-sterile for both early and later strains; the booster systemic immunization does not significantly affect the effectiveness of antiviral immunity, and its feasibility is currently being questioned. Studies of the mucosal immune response have shown that intranasal immunization with adenovirus vaccines provides more pronounced protective immunity than systemic reimmunization does. A promising approach is the creation of multivalent inhaled next generation vaccines containing immunoadjuvants that activate B- and T-cell mucosal immunity. Currently, a large number of intranasal vaccines are undergoing phase I/II trials, while the preclinical and preliminary clinical results indicate that this method of vaccination provides a better mucosal immune response at the entry site of the virus than systemic immunization does. This strategy may provide a long-term immune protection against the currently existing and yet unknown new strains of SARS-CoV-2.

Protection from COVID-19 with a VSV-based vaccine expressing the spike and nucleocapsid proteins.

Successful vaccine efforts countering the COVID-19 pandemic are centralized around the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) protein as viral antigen and have greatly reduced the morbidity and mortality associated with COVID-19. Since the start of this pandemic, SARS-CoV-2 has evolved resulting in new variants of concern (VOC) challenging the vaccine-established immunologic memory. We show that vaccination with a vesicular stomatitis virus (VSV)-based vaccine expressing the SARS-CoV-2 S plus the conserved nucleocapsid (N) protein was protective in a hamster challenge model when a single dose was administered 28 or 10 days prior to challenge, respectively. In this study, only intranasal vaccination resulted in protection against challenge with multiple VOC highlighting that the addition of the N protein indeed improved protective efficacy. This data demonstrates the ability of a VSV-based dual-antigen vaccine to reduce viral shedding and protect from disease caused by SARS-CoV-2 VOC.

Nasal transmission of equine parvovirus hepatitis.

Equine parvovirus hepatitis (EqPV-H) is highly prevalent and causes subclinical to fatal hepatitis, which can occur in outbreaks. Whereas iatrogenic transmission is well documented, the mode of horizontal transmission is not known. The virus is shed in nasal, oral and fecal secretions, and PO transmission has been reported in a single horse. Investigate the efficiency of PO and nasal transmission of EqPV-H in a larger cohort. Prospective experimental transmission study. Eleven EqPV-H-negative horses were inoculated with 5 × 106 genome equivalents EqPV-H. Serum PCR and serology for EqPV-H were performed weekly and monthly, respectively. Horses first were inoculated PO, and then intranasally 8 weeks later. No horse became viremic or seroconverted within 8 weeks after PO inoculation. After intranasal inoculation, 5 horses became viremic within 6 to 12 weeks and seroconverted within 10 to 19 weeks. After a period without monitoring from 12 to 19 weeks postinoculation, another 5 horses were found to be viremic at 19 to 22 weeks. The second set of 5 horses could have been infected by horizontal transmission from the first 5 because of cohousing. We demonstrated that EqPV-H can be transmitted nasally. The prolonged eclipse phase before detectable viremia indicates biosecurity measures to control spread could be impractical.