In a recent study published in npj Vaccines, a group of researchers demonstrated that a single-dose intranasal application of a modified Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strain ∆3678 effectively induces robust immune responses, protecting mice against various forms of the virus, suggesting its potential as an efficient mucosal vaccine.


Over three years, SARS-CoV-2, the instigator of the coronavirus disease 2019 (COVID-19) pandemic, has claimed over 6.9 million lives globally. Despite the expedited creation of four Food and Drug Administration (FDA)-authorized vaccines, their efficacy is threatened by emerging, more transmissible variants known for immune evasion.

The current vaccines, delivered via injection, can potentially provoke insufficient respiratory tract immune reactions, especially against these variants. Intranasal immunization can stimulate robust local and systemic immunity, offering enhanced protection.

Further research is essential due to the ongoing global health threat posed by SARS-CoV-2 variants and the limited respiratory tract immune responses induced by current parenterally administered vaccines, underscoring the need for an optimized intranasal vaccine that effectively bolsters both mucosal and systemic immunity.

About the study

The researchers cultivated Vero-E6 cells from African green monkeys and Vero-E6-TMPRSS2 cells, maintaining them at 37°C with 5% CO2.

They used infectious clones of SARS-CoV-2, ∆3678, and the BA.5 variant. For animal studies, female K18-hACE2 C57BL/6J mice, aged 8-10 weeks, were infected intranasally  (i.n.) with either the WT WA1 or ∆3678 viruses or mock-infected.

On day 28 post-initial infection, they were challenged with the WT WA1 or BA.5 virus. The researchers monitored the mice for weight changes and health status, euthanizing those losing over 20% of their body weight.

The University of Texas Medical Branch’s Institutional Animal Care and Use Committee sanctioned all experiments in an Animal Biosafety Level 3 (ABSL3) environment. Researchers processed lung lobes for plaque assays and utilized centrifugation for clarification.

They employed the Enzyme-Linked Immunosorbent Assay (ELISA) with SARS-CoV-2 Receptor-Binding Domain (RBD) protein for antibody detection. B cell Enzyme-Linked ImmunoSPOT (ELISPOT) assays adhered to known protocols. Lung leukocytes or splenocytes were also exposed to SARS-CoV-2 peptides for intracellular cytokine staining using specific antibodies.

Lung tissues were subjected to RNA extraction and analyzed using the nCounter Pro System, with results visualized in R 4.1.2. Statistical evaluations of viral loads, cytokines, antibodies, and cell responses were performed using Prism software, employing non-paired Student’s t-tests for p-values.

Study results

In the present study, researchers employed K18-hACE2 mice to explore the potential of the ∆3678  virus as a vaccine against SARS-CoV-2, inspired by previous research. The mice received intranasal immunization with this attenuated strain, with groups administered wild-type SARS-CoV-2 USA-WA1/2020 or a placebo serving as controls.

Remarkably, all mice immunized with the ∆3678  virus thrived without signs of weight loss or ill health 28 days after vaccination, contrasting with some fatalities and weight loss in mice exposed to the wild-type virus.

When the researchers analyzed immune responses, they discovered both the ∆3678  and wild-type virus evoked a Th1-skewed response in the lungs. The ∆3678 group exhibited notable T cell activation, especially in the heightened presence and activity of CD4+ and CD8+ T cells producing Interferon Gamma (IFNγ).

Although both groups showed robust Immunoglobulin A (IgA)+ B cell responses, the ∆3678  groups were slightly diminished. Interestingly, both groups exhibited comparable SARS-CoV-2-specific IgA or IgG antibodies, with similar patterns observed in the spleen, indicating a systemic Th1-prone response.

The team then assessed the vaccine’s protective efficacy, exposing the immunized mice to a high dose of wild-type SARS-CoV-2, and observed that mice previously inoculated with either the ∆3678 or wild-type virus exhibited neither traces of the virus in their lungs or trachea nor weight loss following the challenge, contrasting with the control group.

Notably, even when challenged with the Omicron BA.5 variant, the ∆3678 group demonstrated no detectable viral presence, highlighting the vaccine’s broad protective potential.

Further analyses post-challenge revealed an enhanced immune response in ∆3678-immunized mice, particularly in the increased numbers of activated T cells in the lungs and spleen. Additionally, both vaccinated groups exhibited strong antibody responses.

Researchers analyzed lung tissue gene expression to decipher the immunity mechanisms, uncovering diminished inflammatory signaling in vaccinated groups and identifying genes linked to long-lasting lung-resident memory T-cell responses.

These findings collectively suggest the ∆3678 strain, though highly attenuated, effectively stimulates comprehensive immune responses in mice, even those previously exposed to SARS-CoV-2. The vaccine appears to confer solid protection against diverse SARS-CoV-2 strains, including the Omicron variant.

The promise shown by intranasal administration of this live-attenuated vaccine candidate in mice warrants further exploration, especially considering the potential for enhanced and sustained mucosal immunity.

However, its efficacy in humans requires further investigation. The profound insights gained from this study underline the ∆3678 virus as a compelling candidate for future SARS-CoV-2 vaccines, potentially impacting both human and veterinary medicine.

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