Fig. 1.

Respiratory administration of Spike trimer vaccine protects against lethal challenge. A) Schematic showing the vaccination and challenge schedule in female K18 hACE2 mice. Mice were infected intranasally with 1.875x105 TCID SARS-CoV-2 isolate SARS-CoV-2/01/human/2020/SWE. Animals were euthanized after losing > 20% of their body mass or when other symptoms became severe. Half of the surviving animals were euthanized at 10 dpi and the other half at 11 dpi. B) Kaplan-Meier survival curve of the control and vaccinated groups. Vaccinated groups are labelled with dose of Spike trimer and the route of administration; s.c., subcutaneous; i.n., intranasal; i.t., intratracheal. The groups included the following number of animals: control n = 8, low dose s.c. n = 7, high dose s.c. n = 7, low dose i.n. n = 8, high dose i.n. n = 8, low dose i.t. n = 7, and high dose i.t. n = 5. Statistical analysis by log-rank (Mantel-Cox) test where *** indicates P < 0.001. C) SARS-CoV-2 RNA levels in BAL fluid and lung tissue at termination. 1 mL of BAL fluid for each mouse was collected at the day of euthanasia. Of this fluid, 90 μl was subjected to RNA extraction and SARS-CoV-2 E gene RT-qPCR. Ct values are plotted for each vaccine group on the left y axis. A section of lung tissue was harvested at termination for RNA and 5 ng of RNA was subsequently analysed by RT-qPCR for E gene copy number (shown on right y axis). D) Histopathology scoring of perivascualar inflammatory cell infiltration in striatum-level meninges tissue from the brains of the challenged mice at termination. E) Anti-Spike S1 IgG ELISA for pre-challenge serum samples, two weeks after second immunisation. Serum was analysed for IgG against the Spike domain from SARS-CoV-2 (Founder 2020 strain first detected in Wuhan, China). F) As E, except anti-Spike S1 IgA was analysed G) Anti-Spike S1 IgA was analysed from BAL fluid harvested at termination. BAL was diluted 1:30 and the optical density (OD) is shown for individual mice. H) Neutralizing antibodies from BAL fluid at termination. BAL was then diluted beginning at 1-in-4 (LoD) and tested for neutralization of SARS- CoV-2 infection of Vero E6 cells. Results for each individual are plotted.