Longitudinal viral shedding and antibody response characteristics of men with acute infection of monkeypox virus: a prospective cohort study.

Understanding of infection dynamics is important for public health measures against monkeypox virus (MPXV) infection. Herein, samples from multiple body sites and environmental fomites of 77 acute MPXV infections (HIV co-infection: N = 42) were collected every two to three days and used for detection of MPXV DNA, surface protein specific antibodies and neutralizing titers. Skin lesions show 100% positivity rate of MPXV DNA, followed by rectum (88.16%), saliva (83.78%) and oropharynx (78.95%). Positivity rate of oropharynx decreases rapidly after 7 days post symptom onset (d.p.o), while the rectum and saliva maintain a positivity rate similar to skin lesions. Viral dynamics are similar among skin lesions, saliva and oropharynx, with a peak at about 6 d.p.o. In contrast, viral levels in the rectum peak at the beginning of symptom onset and decrease rapidly thereafter. 52.66% of environmental fomite swabs are positive for MPXV DNA, with highest positivity rate (69.89%) from air-conditioning air outlets. High seropositivity against A29L (100%) and H3L (94.74%) are detected, while a correlation between IgG endpoint titers and neutralizing titers is only found for A29L. Most indexes are similar between HIV and Non-HIV participants, while HIV and rectitis are associated with higher viral loads in rectum.

Human neutralizing antibodies target a conserved lateral patch on H7N9 hemagglutinin head.

Avian influenza A virus H7N9 causes severe human infections with >30% fatality. Currently, there is no H7N9-specific prevention or treatment for humans. Here, from a 2013 H7N9 convalescent case in Hong Kong, we isolate four hemagglutinin (HA)-reactive monoclonal antibodies (mAbs), with three directed to the globular head domain (HA1) and one to the stalk domain (HA2). Two clonally related HA1-directed mAbs, H7.HK1 and H7.HK2, potently neutralize H7N9 and protect female mice from lethal H7N9/AH1 challenge. Cryo-EM structures reveal that H7.HK1 and H7.HK2 bind to a ß14-centered surface and disrupt the 220-loop that makes hydrophobic contacts with sialic acid on an adjacent protomer, thereby blocking viral entry. Sequence analysis indicates the lateral patch targeted by H7.HK1 and H7.HK2 to be conserved among influenza subtypes. Both H7.HK1 and H7.HK2 retain HA1 binding and neutralization capacity to later H7N9 isolates from 2016-2017, consistent with structural data showing that the antigenic mutations during this timeframe occur at their epitope peripheries. The HA2-directed mAb H7.HK4 lacks neutralizing activity but when used in combination with H7.HK2 moderately augments female mouse protection. Overall, our data reveal antibodies to a conserved lateral HA1 supersite that confer neutralization, and when combined with a HA2-directed non-neutralizing mAb, augment protection.

Neutralizing antibody responses and cellular responses against SARS-CoV-2 Omicron subvariants after mRNA SARS-CoV-2 vaccination in kidney transplant recipients.

Although the mRNA SARS-CoV-2 vaccine has improved the mortality rate in the general population, its efficacy against rapidly mutating virus strains, especially in kidney transplant recipients, remains unclear. We examined the anti-SARS-CoV-2 spike protein IgG antibody and neutralizing antibody titers and cellular immunity against B.1.1, BA.1, and BA.5 antigens in 73 uninfected kidney recipients and 16 uninfected healthy controls who received three doses of an mRNA SARS-CoV-2 vaccine. The IgG antibody titers were significantly lower in recipients than in healthy controls. Similarly, neutralizing antibody titers against three viral variants were significantly lower in recipients. When the virus was mutated, the neutralizing antibody titers decreased significantly in both groups. In cellular immunity analysis, the number of spike-specific CD8 + non-naïve T cells against three variants significantly decreased in recipients. Conversely, the frequency of spike-specific Th2 CD4 + T-cells in recipients was higher than that in healthy controls. Nineteen recipients and six healthy controls also received a bivalent omicron-containing booster vaccine, leading to increase IgG and neutralizing antibody titers in both groups. After that, eleven recipients and five healthy controls received XBB.1.5 monovalent vaccines, increasing the neutralizing antibody titers against not only XBB.1.5, but also EG.5.1 and BA.2.86 antigens in kidney recipients. Although kidney recipients did not gain sufficient immunity against Omicron BA.5 with the third dose of vaccine, humoral response against mutant SARS-CoV-2 lineages significantly increased after bivalent Omicron-containing booster vaccine and the XBB.1.5 monovalent vaccine. Therefore, it is important for kidney recipients to continue to administer updated vaccines.

Immunogenicity evaluation of primary polio vaccination schedule with inactivated poliovirus vaccines and bivalent oral poliovirus vaccine.

BACKGROUND: To assess the immunogenicity of the current primary polio vaccination schedule in China and compare it with alternative schedules using Sabin or Salk-strain IPV (sIPV, wIPV). METHODS: A cross-sectional investigation was conducted at four sites in Chongqing, China, healthy infants aged 60-89 days were conveniently recruited and divided into four groups according to their received primary polio vaccination schedules (2sIPV + bOPV, 2wIPV + bOPV, 3sIPV, and 3wIPV). The sero-protection and neutralizing antibody titers against poliovirus serotypes (type 1, 2, and 3) were compared after the last dose. RESULTS: There were 408 infants completed the protocol. The observed seropositivity was more than 96% against poliovirus types 1, 2, and 3 in all groups. IPV-only groups induced higher antibody titers(GMT) against poliovirus type 2 (Median:192, QR: 96-384, P<0.05) than the "2IPV + bOPV" group. While the "2IPV + bOPV" group induced significantly higher antibody titers against poliovirus type 1 (Median:2048, QR: 768-2048, P<0.05)and type 3 (Median:2048, QR: 512-2048, P<0.05) than the IPV-only group. CONCLUSIONS: Our findings have proved that the two doses of IPV with one dose of bOPV is currently the best polio routine immunization schedule in China.

Functional and structural investigation of a broadly neutralizing SARS-CoV-2 antibody.

Since its emergence, SARS-CoV-2 has been continuously evolving, hampering the effectiveness of current vaccines against COVID-19. mAbs can be used to treat patients at risk of severe COVID-19. Thus, the development of broadly protective mAbs and an understanding of the underlying protective mechanisms are of great importance. Here, we isolated mAbs from donors with breakthrough infection with Omicron subvariants using a single-B cell screening platform. We identified a mAb, O5C2, which possesses broad-spectrum neutralization and antibody-dependent cell-mediated cytotoxic activities against SARS-CoV-2 variants, including EG.5.1. Single-particle analysis by cryo-electron microscopy revealed that O5C2 targeted an unusually large epitope within the receptor-binding domain of spike protein that overlapped with the angiotensin-converting enzyme 2 binding interface. Furthermore, O5C2 effectively protected against BA.5 Omicron infection in vivo by mediating changes in transcriptomes enriched in genes involved in apoptosis and interferon responses. Our findings provide insights into the development of pan-protective mAbs against SARS-CoV-2.

SARS-CoV-2 omicron BA.2.87.1 exhibits higher susceptibility to serum neutralization than EG.5.1 and JN.1.

As SARS-CoV-2 continues to spread and mutate, tracking the viral evolutionary trajectory and understanding the functional consequences of its mutations remain crucial. Here, we characterized the antibody evasion, ACE2 receptor engagement, and viral infectivity of the highly mutated SARS-CoV-2 Omicron subvariant BA.2.87.1. Compared with other Omicron subvariants, including EG.5.1 and the current predominant JN.1, BA.2.87.1 exhibits less immune evasion, reduced viral receptor engagement, and comparable infectivity in Calu-3 lung cells. Intriguingly, two large deletions (Δ15-26 and Δ136-146) in the N-terminal domain (NTD) of the spike protein facilitate subtly increased antibody evasion but significantly diminish viral infectivity. Collectively, our data support the announcement by the USA CDC that the public health risk posed by BA.2.87.1 appears to be low.

Mosaic RBD nanoparticle elicits immunodominant antibody responses across sarbecoviruses.

Nanoparticle vaccines displaying mosaic receptor-binding domains (RBDs) or spike (S) from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) or other sarbecoviruses are used in preparedness against potential zoonotic outbreaks. Here, we describe a self-assembling nanoparticle using lumazine synthase (LuS) as the scaffold to display RBDs from different sarbecoviruses. Mosaic nanoparticles induce sarbecovirus cross-neutralizing antibodies comparable to a nanoparticle cocktail. We find mosaic nanoparticles elicit a B cell receptor repertoire using an immunodominant germline gene pair of IGHV14-3:IGKV14-111. Most of the tested IGHV14-3:IGKV14-111 monoclonal antibodies (mAbs) are broadly cross-reactive to clade 1a, 1b, and 3 sarbecoviruses. Using mAb competition and cryo-electron microscopy, we determine that a representative IGHV14-3:IGKV14-111 mAb, M2-7, binds to a conserved epitope on the RBD, largely overlapping with the pan-sarbecovirus mAb S2H97. This suggests mosaic nanoparticles expand B cell recognition of the common epitopes shared by different clades of sarbecoviruses. These results provide immunological insights into the cross-reactive responses elicited by mosaic nanoparticles against sarbecoviruses.

A novel immunofluorescent test system for SARS-CoV-2 detection in infected cells.

Highly variable pandemic coronavirus SARS-CoV-2, which causes the hazardous COVID-19 infection, has been persistent in the human population since late 2019. A prompt assessment of individual and herd immunity against the infection can be accomplished by using rapid tests to determine antiviral antibody levels. The microneutralization assay (MN) is one of the most widely used diagnostic methods that has been proposed to assess the qualitative and quantitative characteristics of virus-specific humoral immunity in COVID-19 convalescents or vaccine recipients. However, some aspects of the assay, such as sensitivity and time cost, need improvement. Here, we developed an express test, which may be potentially used in clinical practice for the assessment of serum-caused SARS-CoV-2 inhibition in infected cell cultures. It implies the detection and counting of coronaviral fluorescent-forming units (FFU) and includes two sequentially used developing components: biotinylated mouse monoclonal antibodies against the recombinant N protein of SARS-CoV-2 (B.1) and the recombinant EGFP-streptavidin fusion protein. Due to the universal specificity of the antibodies, our analytical tool is suitable for the detection of various strains of SARS-CoV-2 when determining both the infectious titer of viruses and the titer of serum virus-neutralizing antibodies. The developed two-component test system is characterized by high sensitivity, a reduced number of analytic stages and low assay cost, as well as by flexibility, since it may be modified for detection of other pathogens using the appropriate antibodies.

Distinct T cell responsiveness to different COVID-19 vaccines and cross-reactivity to SARS-CoV-2 variants with age and CMV status.

Introduction: Accumulating evidence indicates the importance of T cell immunity in vaccination-induced protection against severe COVID-19 disease, especially against SARS-CoV-2 Variants-of-Concern (VOCs) that more readily escape from recognition by neutralizing antibodies. However, there is limited knowledge on the T cell responses across different age groups and the impact of CMV status after primary and booster vaccination with different vaccine combinations. Moreover, it remains unclear whether age has an effect on the ability of T cells to cross-react against VOCs. Methods: Therefore, we interrogated the Spike-specific T cell responses in healthy adults of the Dutch population across different ages, whom received different vaccine types for the primary series and/or booster vaccination, using IFNÉ£ ELISpot. Cells were stimulated with overlapping peptide pools of the ancestral Spike protein and different VOCs. Results: Robust Spike-specific T cell responses were detected in the vast majority of participants upon the primary vaccination series, regardless of the vaccine type (i.e. BNT162b2, mRNA-1273, ChAdOx1 nCoV-19, or Ad26.COV2.S). Clearly, in the 70+ age group, responses were overall lower and showed more variation compared to younger age groups. Only in CMV-seropositive older adults (>70y) there was a significant inverse relation of age with T cell responses. Although T cell responses increased in all age groups after booster vaccination, Spike-specific T cell frequencies remained lower in the 70+ age group. Regardless of age or CMV status, primary mRNA-1273 vaccination followed by BNT162b2 booster vaccination showed limited booster effect compared to the BNT162b2/BNT162b2 or BNT162b2/mRNA-1273 primary-booster regimen. A modest reduction in cross-reactivity to the Alpha, Delta and Omicron BA.1, but not the Beta or Gamma variant, was observed after primary vaccination. Discussion: Together, this study shows that age, CMV status, but also the primary-booster vaccination regimen influence the height of the vaccination-induced Spike-specific T cell response, but did not impact the VOC cross-reactivity.

Designing boosting immunogens for HIV-1 vaccine development.

Inducing HIV-1 broadly neutralizing antibodies (bnAbs) through vaccination poses exceptional challenges. In this issue of Cell Host & Microbe, Wiehe and colleagues report the elicitation of affinity-matured bnAbs in knock-in mice through boosting immunogen vaccination, which selects for key improbable mutations.

Method for quantifying the Pasteurella multocida antigen adsorbed on aluminum hydroxide adjuvant in swine atrophic rhinitis vaccine.

Swine atrophic rhinitis is a disease caused by Pasteurella multocida and Bordetella bronchiseptica that affects pigs. Inactivated vaccines containing the toxins produced by Pasteurella multocida and Bordetella bronchiseptica have been widely used for the prevention of swine atrophic rhinitis. The efficacy of a vaccine is correlated with the amount of antigen present; however, the protective toxin of P. multocida bound to aluminum hydroxide, which is used as an adjuvant, can hinder the monitoring of the antigen concentration in the vaccine. This study assessed the applicability of a dot immunoassay as an antigen quantification method using monoclonal antibodies. This quantification method was able to detect the antigen with high specificity and sensitivity even when the antigen was bound to the adjuvant, and its application to vaccine products revealed a correlation between the amount of antigen present in the vaccine and the neutralizing antibody titers induced in pigs. The antigen quantification method presented in this study is a simple and sensitive assay capable of quantifying the amount of antigen present in a vaccine that can be used as an alternative quality control measure.

Vaccination in twin pregnancies: comparison between immunization before conception and during pregnancy.

To evaluate the development of neutralizing Anti-Spike Protein IgG (Anti-S-IgG) during twin pregnancies before conception vs. during pregnancy. In this prospective study, three blood samples were collected from pregnant women and subjected to anti-S-IgG immunodiagnostics. The patient's medical records, including vaccination and PCR test results, were collected from the hospital's electronic database. Age-matched non-pregnant women were used as a control group. We enrolled 83 women with twin pregnancies. 49 women were vaccinated before conception, 21 women were vaccinated during pregnancy, and 13 were not vaccinated. Of the 13 women who weren't vaccinated, three became positive during pregnancy, and all three were severely ill. By contrast, in women who were vaccinated during or before pregnancy, COVID-19 infection during pregnancy caused only mild symptoms. A ten-fold lower level of neutralizing Anti-S-IgG in the 3rd trimester was observed in healthy women who were vaccinated before conception and remained healthy until discharge from the hospital after delivery 1605 (IQR: 763-2410) compared to the healthy women who were vaccinated during pregnancy 152 AU/mL (IQR: 54-360). This difference was higher among women who were infected by COVID-19 (as verified by a positive PCR test). The third-trimester level of neutralizing Ant-S-IgG in the infected group was 4770 AU/mL (4760-6100) in infected women vaccinated before conception compared to those vaccinated during pregnancy who had 70 AU/mL (IQR: 20-170) (p < 0.001). In women vaccinated at 13-16 weeks gestation, neutralizing Anti-S-IgG at 20-22 weeks went up to 372 AU/mL (IQR: 120-1598) but rapidly dropped to 112 AU/mL (IQR: 54-357) at 28-30 weeks, (p < 0.001), a faster decline than in women vaccinated at a median 22 weeks before conception. Being infected by COVID-19 before conception was linked to having low Anti-S-IgG levels during pregnancy, whereas being infected by COVID-19 during pregnancy led to a very high response in the 3rd trimester. In twin pregnancies, significantly lower neutralizing Anti-S-IgG levels were observed in women vaccinated during pregnancy compared to those vaccinated before conception, whether infected or not infected by COVID-19. A full course of vaccination before conception is recommended.Trial registration. ClinicalTrials.gov Protocol Registration and Results System (PRS) Receipt Release Date: October 4, 2021. https://clinicaltrials.gov/ ID: NCT04595214.

Intervention of CXCL5 attenuated neuroinflammation and promoted neurological recovery after traumatic brain injury.

Neuroinflammation after traumatic brain injury (TBI) exhibits a strong correlation with neurological impairment, which is a crucial target for improving the prognosis of TBI patients. The involvement of CXCL5/CXCR2 signaling in the regulation of neuroinflammation in brain injury models has been documented. Therefore, the effects of CXCL5 on post-TBI neuroinflammation and its potential mechanisms need to be explored. Following TBI, C57BL/6 mice were administered intraperitoneal injections of a CXCL5 neutralizing antibody (Nab-CXCL5) (5 mg/kg, 2 times/day). Subsequently, the effects on neuroinflammation, nerve injury, and neurological function were assessed. Nab-CXCL5 significantly reduced the release of inflammatory factors, inhibited the formation of inflammatory microglia and astrocytes, and reduced the infiltration of peripheral immune cells in TBI mice. Additionally, this intervention led to a reduction in neuronal impairment and facilitated the restoration of sensorimotor abilities, as well as improvements in learning and memory functions. Peripheral administration of the Nab-CXCL5 to TBI mice could suppress neuroinflammation, reduce neurological damage, and improve neurological function. Our data suggest that neutralizing antibodies against CXCL5 (Nab-CXCL5) may be a promising agent for treating TBI.

Original COVID-19 priming regimen impacts the immunogenicity of bivalent BA.1 and BA.5 boosters.

Waning antibody responses after COVID-19 vaccination combined with the emergence of the SARS-CoV-2 Omicron lineage led to reduced vaccine effectiveness. As a countermeasure, bivalent mRNA-based booster vaccines encoding the ancestral spike protein in combination with that of Omicron BA.1 or BA.5 were introduced. Since then, different BA.2-descendent lineages have become dominant, such as XBB.1.5, JN.1, or EG.5.1. Here, we report post-hoc analyses of data from the SWITCH-ON study, assessing how different COVID-19 priming regimens affect the immunogenicity of bivalent booster vaccinations and breakthrough infections (NCT05471440). BA.1 and BA.5 bivalent vaccines boosted neutralizing antibodies and T-cells up to 3 months after boost; however, cross-neutralization of XBB.1.5 was poor. Interestingly, different combinations of prime-boost regimens induced divergent responses: participants primed with Ad26.COV2.S developed lower binding antibody levels after bivalent boost while neutralization and T-cell responses were similar to mRNA-based primed participants. In contrast, the breadth of neutralization was higher in mRNA-primed and bivalent BA.5 boosted participants. Combined, our data further support the current use of monovalent vaccines based on circulating strains when vaccinating risk groups, as recently recommended by the WHO. We emphasize the importance of the continuous assessment of immune responses targeting circulating variants to guide future COVID-19 vaccination policies.

Heterologous prime-boost vaccination drives early maturation of HIV broadly neutralizing antibody precursors in humanized mice.

A protective HIV vaccine will likely need to induce broadly neutralizing antibodies (bnAbs). Vaccination with the germline-targeting immunogen eOD-GT8 60mer adjuvanted with AS01B was found to induce VRC01-class bnAb precursors in 97% of vaccine recipients in the IAVI G001 phase 1 clinical trial; however, heterologous boost immunizations with antigens more similar to the native glycoprotein will be required to induce bnAbs. Therefore, we designed core-g28v2 60mer, a nanoparticle immunogen to be used as a first boost after eOD-GT8 60mer priming. We found, using a humanized mouse model approximating human conditions of VRC01-class precursor B cell diversity, affinity, and frequency, that both protein- and mRNA-based heterologous prime-boost regimens induced VRC01-class antibodies that gained key mutations and bound to near-native HIV envelope trimers lacking the N276 glycan. We further showed that VRC01-class antibodies induced by mRNA-based regimens could neutralize pseudoviruses lacking the N276 glycan. These results demonstrated that heterologous boosting can drive maturation toward VRC01-class bnAb development and supported the initiation of the IAVI G002 phase 1 trial testing mRNA-encoded nanoparticle prime-boost regimens.

Measles Foci Reduction Neutralization Test (FRNT).

The plaque reduction neutralization test (PRNT) and the enzyme-linked immunosorbent assay (ELISA) are both widely used to assess immunity to infectious diseases such as measles, but they use two different measurement principles: ELISA measures the ability of antibodies to bind to virus components, while the PRNT detects the aptitude of antibodies to prevent the infection of a susceptible cell. As a result, detection of measles virus (MV) neutralizing antibodies is the gold standard for assessing immunity to measles. However, the assay is laborious and requires experience and excellent technical skills. In addition, the result is only available after several days. Therefore, the classical PRNT is not suitable for high-throughput testing. By using an immunocolorimetric assay (ICA) to detect MV-infected cells, the standard PRNT has been developed into a focus reduction neutralization test (FRNT). This assay is faster and has improved specificity. The FRNT described here is extremely useful when immunity to measles virus needs to be assessed in patients with a specific medical condition, such as immunocompromised individuals in whom presumed residual immunity needs to be assessed. The FRNT is not generally recommended for use with large numbers of specimens, such as in a seroprevalence study.

Cross-Reactive Antibody Responses to Coronaviruses Elicited by SARS-CoV-2 Infection or Vaccination.

BACKGROUND: The newly emerged SARS-CoV-2 possesses shared antigenic epitopes with other human coronaviruses. We investigated if COVID-19 vaccination or SARS-CoV-2 infection may boost cross-reactive antibodies to other human coronaviruses. METHODS: Prevaccination and postvaccination sera from SARS-CoV-2 naïve healthy subjects who received three doses of the mRNA vaccine (BioNTech, BNT) or the inactivated vaccine (CoronaVac, CV) were used to monitor the level of cross-reactive antibodies raised against other human coronaviruses by enzyme-linked immunosorbent assay. In comparison, convalescent sera from COVID-19 patients with or without prior vaccination history were also tested. Pseudoparticle neutralization assay was performed to detect neutralization antibody against MERS-CoV. RESULTS: Among SARS-CoV-2 infection-naïve subjects, BNT or CV significantly increased the anti-S2 antibodies against Betacoronaviruses (OC43 and MERS-CoV) but not Alphacoronaviruses (229E). The prevaccination antibody response to the common cold human coronaviruses did not negatively impact the postvaccination antibody response to SARS-CoV-2. Cross-reactive antibodies that binds to the S2 protein of MERS-CoV were similarly detected from the convalescent sera of COVID-19 patients with or without vaccination history. However, these anti-S2 antibodies do not possess neutralizing activity in MERS-CoV pseudoparticle neutralization tests. CONCLUSIONS: Our results suggest that SARS-CoV-2 infection or vaccination may potentially modulate population immune landscape against previously exposed or novel human coronaviruses. The findings have implications for future sero-epidemiological studies on MERS-CoV.

Safety, immunogenicity and protective effect of sequential vaccination with inactivated and recombinant protein COVID-19 vaccine in the elderly: a prospective longitudinal study.

The safety and efficacy of COVID-19 vaccines in the elderly, a high-risk group for severe COVID-19 infection, have not been fully understood. To clarify these issues, this prospective study followed up 157 elderly and 73 young participants for 16 months and compared the safety, immunogenicity, and efficacy of two doses of the inactivated vaccine BBIBP-CorV followed by a booster dose of the recombinant protein vaccine ZF2001. The results showed that this vaccination protocol was safe and tolerable in the elderly. After administering two doses of the BBIBP-CorV, the positivity rates and titers of neutralizing and anti-RBD antibodies in the elderly were significantly lower than those in the young individuals. After the ZF2001 booster dose, the antibody-positive rates in the elderly were comparable to those in the young; however, the antibody titers remained lower. Gender, age, and underlying diseases were independently associated with vaccine immunogenicity in elderly individuals. The pseudovirus neutralization assay showed that, compared with those after receiving two doses of BBIBP-CorV priming, some participants obtained immunological protection against BA.5 and BF.7 after receiving the ZF2001 booster. Breakthrough infection symptoms last longer in the infected elderly and pre-infection antibody titers were negatively associated with the severity of post-infection symptoms. The antibody levels in the elderly increased significantly after breakthrough infection but were still lower than those in the young. Our data suggest that multiple booster vaccinations at short intervals to maintain high antibody levels may be an effective strategy for protecting the elderly against COVID-19.

Enhanced potency of an IgM-like nanobody targeting conserved epitope in SARS-CoV-2 spike N-terminal domain.

Almost all the neutralizing antibodies targeting the receptor-binding domain (RBD) of spike (S) protein show weakened or lost efficacy against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged or emerging variants, such as Omicron and its sub-variants. This suggests that highly conserved epitopes are crucial for the development of neutralizing antibodies. Here, we present one nanobody, N235, displaying broad neutralization against the SARS-CoV-2 prototype and multiple variants, including the newly emerged Omicron and its sub-variants. Cryo-electron microscopy demonstrates N235 binds a novel, conserved, cryptic epitope in the N-terminal domain (NTD) of the S protein, which interferes with the RBD in the neighboring S protein. The neutralization mechanism interpreted via flow cytometry and Western blot shows that N235 appears to induce the S1 subunit shedding from the trimeric S complex. Furthermore, a nano-IgM construct (MN235), engineered by fusing N235 with the human IgM Fc region, displays prevention via inducing S1 shedding and cross-linking virus particles. Compared to N235, MN235 exhibits varied enhancement in neutralization against pseudotyped and authentic viruses in vitro. The intranasal administration of MN235 in low doses can effectively prevent the infection of Omicron sub-variant BA.1 and XBB in vivo, suggesting that it can be developed as a promising prophylactic antibody to cope with the ongoing and future infection.

A universal recombinant adenovirus type 5 vector-based COVID-19 vaccine.

A universal recombinant adenovirus type-5 (Ad5) vaccine against COVID19 (Ad-US) was constructed, and immunogenicity and broad-spectrum of Ad5-US were evaluated with both intranasal and intramuscular immunization routes. The humoral immune response of Ad5-US in serum and bronchoalveolar lavage fluid were evaluated by the enzyme-linked immunosorbent assay (ELISA), recombinant vesicular stomatitis virus based pseudovirus neutralization assay, and angiotensin-converting enzyme-2 (ACE2) -binding inhibition assay. The cellular immune response and Th1/Th2 biased immune response of Ad5-US were evaluated by the IFN-γ ELISpot assay, intracellular cytokine staining, and Meso Scale Discovery (MSD) profiling of Th1/Th2 cytokines. Intramuscular priming followed by an intranasal booster with Ad5-US elicited the broad-spectrum and high levels of IgG, IgA, pseudovirus neutralizing antibody (PNAb), and Th1-skewing of the T-cell response. Overall, the adenovirus type-5 vectored universal SARS-CoV-2 vaccine Ad5-US was successfully constructed, and Ad5-US was highly immunogenic and broad spectrum. Intramuscular priming followed by an intranasal booster with Ad5-US induced the high and broad spectrum systemic immune responses and local mucosal immune responses.