EFFECT OF FULL VACCINATION AND POST-COVID OLFACTORY DYSFUNCTION IN RECOVERED COVID-19 PATIENT. A RETROSPECTIVE LONGITUDINAL STUDY WITH PROPENSITY MATCHING.

Background Symptoms after Coronavirus Disease (COVID-19) infection affect the quality of life of its survivor especially to the special senses including olfactory function. It is important to prevent the disability at an earlier stage. Vaccination as key prevention has been proven to be effective in reducing symptomatic disease and severity. However, the effects of vaccination on post COVID symptoms have not been evaluated. This study aimed to evaluate the possible protection of full vaccination and the occurrence of post-COVID olfactory dysfunction, specifically anosmia, and hyposmia in patients who were diagnosed with COVID-19. Method A longitudinal analysis using the retrospective cohort of the Indonesian patient-based Post-COVID-19 survey collected from July 2021 until December 2021, involving COVID-19 Patients confirmed by Real-Time Polymerase Chain Reaction (RT-PCR) and/or Antigen test. Variables including demography, comorbidities, health behavior, type of vaccine, symptoms, and treatment were collected through an online questionnaire based on the American Academy of Otolaryngology-Head and Neck Surgery (AAO-HNS). Participants were matched (1:1) using propensity matching score into two exposure statuses, infected 1)>14 days of full vaccination and 2)<14 days or incomplete or unvaccinated. The olfactory dysfunction was assessed two weeks and four weeks after negative conversion with PCR using a self-measured olfactory questionnaire (MOQ). The Generalized Estimating Equation (GEE) was performed to assess the effect of full vaccination on post-COVID-19 olfactory dysfunction. The Receiver Operating Characteristic determined the sensitivity and specificity of the cutoff value of the days from fully vaccinated to diagnosis and the olfactory dysfunction. Results A total of 442 participants were extracted from the cohort and inoculated with the inactivated viral vaccine (99.5%). The prevalence of olfactory dysfunction in two weeks was 9.95% and 5.43% after four weeks. Adjusted by other variables, people who were infected >14 days after being fully vaccinated had a 69% (adjusted Odds Ratio / aOR 0.31 95% CI 0.102-0.941) probability of developing olfactory dysfunction. Longer days of fully vaccinated to infection are associated with increased risk (aOR 1.012 95% CI 1.002-1.022 p-value 0.015). A cut-off of 88 days of full vaccination-to-diagnosis duration has Area Under Curve (AUC) of 0.693 (p=0.002), the sensitivity of 73.9%, and specificity of 63.3% in differentiating the olfactory dysfunction event in two weeks after COVID-19 with a crude odds ratio of 4.852 (95% CI 1.831-12.855 p=0.001) Conclusion After 14 days of full vaccination, the protective effect could reduce the chance of post-COVID olfactory dysfunction although a longer full vaccination-to-diagnosis duration increases the risk. It is important to consider a booster shot starting from 89 days after the last dose in those who received the inactivated viral regimen.

The TZM-bl Reporter Cell Line Expresses Kynureninase That Can Neutralize 2F5-like Antibodies in the HIV-1 Neutralization Assay

Induction of broadly neutralizing antibodies targeting ectodomain of the transmembrane (TM) glycoprotein gp41 HIV-1 provides a basis for the development of a universal anti-viral vaccine. The HeLa cell-derived TZM-bl reporter cell line is widely used for the estimation of lentiviruses neutralization by immune sera. The cell line is highly permissive to infection by most strains of HIV, SIV, and SHIV. Here we demonstrated that TZM-bl cells express a 48 kDa non-glycosylated protein (p48) recognized by broadly neutralizing monoclonal antibody (mAb) 2F5 targeting the ELDKWA (aa 669–674) epitope of gp41TM of HIV-1. A significant amount of p48 was found in the cell supernatant. The protein was identified as human kynureninase (KYNU), which has the ELDKWA epitope. The protein is further called “p48 KYNU”. The HIV-1 neutralization by mAb 2F5 and 4E10 in the presence of p48KYNU was tested on Jurkat and TZM-bl cells. It was demonstrated that p48KYNU reduces neutralization by 2F5-like antibodies, but it has almost no effect on mAb 4E10. Therefore, p48KYNU can attenuate HIV-1 neutralization by 2F5-like antibodies and hence create false-negative results. Thus, previously tested immune sera that recognized the ELDKWA-epitope and demonstrated a “weak neutralization” of HIV-1 in TZM-bl assay should be reevaluated.

Parenteral Antioxidant Supplementation at Birth Improves the Response to Intranasal Vaccination in Newborn Dairy Calves

Newborn calves experience oxidative stress throughout the first month of their life, which is known to decrease lymphocyte functions relevant to vaccine responsiveness. Thus, this study aimed to determine the extent to which parenteral antioxidant supplementation given at birth improves the response to an intranasal viral vaccine in the first month of life of newborn dairy calves. For this, 21 calves were randomly assigned at birth to one of two commercially available antioxidant micronutrient supplements or a placebo group receiving 0.9% sterile saline (n = 7/group). Serum and nasal secretion samples were collected before administration of treatments and an intranasal vaccine against respiratory viruses (bovine herpesvirus type 1, bovine syncytial respiratory virus, and parainfluenza 3), and once weekly for the first four weeks of age. Systemic redox balance was determined in serum. Immunoglobulin A specific for bovine herpesvirus 1 and bovine syncytial respiratory virus was quantified in nasal secretions as a proxy to intranasal vaccine responsiveness. Our results showed that parenteral administration of antioxidants at birth improved calves’ redox balance. Additionally, calves receiving antioxidant supplementation had higher concentrations of immunoglobulin A in their nasal secretions than calves in the control group. Thus, we conclude that supplementation of calves with antioxidants at birth could be a practical strategy to improve intranasal vaccine response. Future larger studies should evaluate the extent to which this increased mucosal response to intranasal vaccination could result in decreased calf morbidity and mortality.

Epigallocatechin-3-Gallate as a Novel Vaccine Adjuvant

Vaccine adjuvants from natural resources have been utilized for enhancing vaccine efficacy against infectious diseases. This study examined the potential use of catechins, polyphenolic materials derived from green tea, as adjuvants for subunit and inactivated vaccines. Previously, catechins have been documented to have irreversible virucidal function, with the possible applicability in the inactivated viral vaccine platform. In a mouse model, the coadministration of epigallocatechin-3-gallate (EGCG) with influenza hemagglutinin (HA) antigens induced high levels of neutralizing antibodies, comparable to that induced by alum, providing complete protection against the lethal challenge. Adjuvant effects were observed for all types of HA antigens, including recombinant full-length HA and HA1 globular domain, and egg-derived inactivated split influenza vaccines. The combination of alum and EGCG further increased neutralizing (NT) antibody titers with the corresponding hemagglutination inhibition (HI) titers, demonstrating a dose-sparing effect. Remarkably, EGCG induced immunoglobulin isotype switching from IgG1 to IgG2a (approximately &gt;64–700 fold increase), exerting a more balanced TH1/TH2 response compared to alum. The upregulation of IgG2a correlated with significant enhancement of antibody-dependent cellular cytotoxicity (ADCC) function (approximately 14 fold increase), providing a potent effector-mediated protection in addition to NT and HI. As the first report on a novel class of vaccine adjuvants with built-in virucidal activities, the results of this study will help improve the efficacy and safety of vaccines for pandemic preparedness.

584. Phase 1 Placebo-Controlled Trial of COVI-VAC™, an Intranasal, Live Attenuated COVID-19 Vaccine

Background COVI-VACTM is an intra-nasal live-attenuated SARS-COV-2 synthetic viral vaccine being developed for the prevention of COVID-19. COVI-VAC is attenuated through deletion of the furin cleavage site and introduction of 283 silent deoptimizing mutations that maintain viral amino acid sequence but result in significant attenuation due to slow translation in the human host cell. Notably, COVI-VAC includes all viral antigens and is not limited to spike. COVI-VAC has demonstrated attenuation, immunogenicity and single dose protection in both Syrian golden hamster and non-human primate models. Methods 48 healthy young adults were enrolled in an inpatient quarantine setting to one of 3 dose escalating cohorts and randomized to COVI-VAC or saline placebo given as nose drops, as a single 0.5mL dose or 2 doses 28 days apart. Endpoints included solicited and unsolicited adverse events, serum cytokines, viral shedding and sequence stability, mucosal and serum antibody responses and IFN ELISpot. Subjects will be followed for 1 year for late safety events and durability of immune response. Results Dosing is complete. There has been no trend in solicited reactogenicity events, and all unsolicited adverse events reported to date have been mild. There have been no SAEs or Grade 3 or 4 events. Vaccine virus from anonymized subjects was shed at levels lower than that likely to result in onward transmission, and the deoptimized sequence of the shed virus remained unchanged compared to the original vaccine sequence. Unblinded data including immunogenicity will be available prior to the IDWeek meeting. Conclusion COVI-VAC appears safe and well tolerated in healthy young adults. Vaccination resulted in minimal viral shedding without sequence instability. Safety and shedding data supports continued development in a wider Phase 2/3 population. Disclosures Sybil Tasker, MD, MPH, FIDSA, Codagenix Inc (Employee, Shareholder) Daryl Bendel, MD, Codagenix Inc (Scientific Research Study Investigator) Melissa Bevan, MD, Codagenix Inc (Scientific Research Study Investigator) Steffen Mueller, PhD, Codagenix Inc (Board Member, Employee, Shareholder) Anna Kushnir, PHD, Codagenix Inc (Employee) Brandon Londt, PhD, Codagenix Inc (Other Financial or Material Support, contracted lab services) J. Robert Coleman, PhD, Codagenix Inc. (Board Member, Employee, Shareholder)

Comparative Transcriptomics Analyses of a Vero Cell Line in Suspension Versus Adherent Culture Conditions

The Vero cell line is the most used continuous cell line for viral vaccine manufacturing. Its anchorage-dependent use renders scaling-up challenging and operations very labor intensive which affects cost effectiveness. Thus, efforts to adapt Vero cells to suspension cultures have been invested but hurdles such as the long doubling time and low cell viability remain to be addressed. In this study, building on the recently published Vero cell line annotated genome, a functional genomics analysis of the Vero cells adapted to suspension is performed to better understand the genetic and phenotypic switches at play during the adaptation of Vero cells from anchorage-dependent to suspension cultures. Results show a downregulation of the epithelial to mesenchymal transition (EMT) pathway, highlighting the dissociation between the adaptation to suspension process and EMT. Surprisingly, an upregulation of cell adhesion components is observed, notably the CDH18 gene, the cytoskeleton pathway, and the extracellular pathway. Moreover, a downregulation of the glycolytic pathway are balanced by an upregulation of the asparagine metabolism pathway, promoting cell adaptation to nutrient deprivation. A downregulation of the adherens junctions and the folate pathways alongside with the FYN gene are possible explanations behind the currently observed low cell viability and long doubling time.

Avian Cell Line DuckCelt®-T17 Is an Efficient Production System for Live-Attenuated Human Metapneumovirus Vaccine Candidate Metavac®

The development of a live-attenuated vaccine (LAV) for the prevention of human metapneumovirus (HMPV) infection is often hampered by the lack of highly efficient and scalable cell-based production systems that support eventual global vaccine production. Avian cell lines cultivated in suspension compete with traditional cell platforms used for viral vaccine manufacture. We investigated whether the DuckCelt®-T17 avian cell line (Vaxxel), previously described as an efficient production system for several influenza strains, could also be used to produce a new HMPV LAV candidate (Metavac®, SH gene-deleted A1/C-85473 HMPV). To that end, we characterized the operational parameters of MOI, cell density, and trypsin addition to achieve the optimal production of Metavac®, and demonstrated that the DuckCelt®-T17 cell line is permissive and well-adapted to the production of the wild-type A1/C-85473 HMPV and the Metavac® vaccine candidate. Moreover, our results confirmed that the LAV candidate produced in DuckCelt®-T17 cells conserves its advantageous replication properties in LLC-MK2 and 3D-reconstituted human airway epithelium models, and its capacity to induce efficient neutralizing antibodies in a BALB/c mouse model. Our results suggest that the DuckCelt®-T17 avian cell line is a very promising platform for the scalable in-suspension serum-free production of the HMPV-based LAV candidate Metavac®.

Large-Scale Microcarrier Culture of Chinese Perch Brain Cell for Viral Vaccine Production in a Stirred Bioreactor

Mandarin fish (Siniperca chuatsi) is one of the important cultured fish species in China. Infectious spleen and kidney necrosis virus (ISKNV) and Siniperca Chuatsi rhabdovirus (SCRV) have hindered the development of mandarin fish farming industry. Vaccination is the most effective method for control of viral diseases, however viral vaccine production requires the large-scale culture of cells. Herein, a suspension culture system of Chinese perch brain cell (CPB) was developed on Cytodex 1 microcarrier in a stirred bioreactor. Firstly, CPB cells were cultured using Cytodex 1 microcarrier in 125 mL stirring flasks. With the optimum operational parameters, CPB cells grew well, distributed uniformly, and could fully cover the microcarriers. Then, CPB cells were digested with trypsin and expanded step-by-step with different expansion ratios from the 125 mL stirring bottle to a 500 mL stirring bottle, and finally to a 3-L bioreactor. Results showed that with an expansion ratio of 1:3, we achieved a high cell density level (2.25 × 106 cells/mL) with an efficient use of the microcarriers, which also confirmed the data obtained from the 125 mL stirring flask. Moreover, obvious cytopathic effects (CPE) were observed in the suspended CPB cells post-infection with ISKNV and SCRV. This study provided a large-scale culture system of CPB cells for virus vaccine production.