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. 2021 Oct 27;12:6197. doi: 10.1038/s41467-021-26499-y

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a Design of the study. Nine-to-ten-week-old female BALB/c mice were used. Group 1–5 (n = 8) were vaccinated with unadjuvanted NDV-HXP-S at 1 µg, 0.3 µg, 0.1 µg, 0.03 µg, and 0.01 µg per mouse, respectively. Group 6 and 7 (n = 8) were vaccinated with 0.1 µg of NDV-HXP-S with 10 or 30 µg of CpG 1018 per mouse, respectively. Group 8 and 9 (n = 8) were vaccinated with 0.03 µg of NDV-HXP-S with 10 or 30 µg of CpG 1018 per mouse, respectively. Group 10 (n = 8) was vaccinated with 1 µg of WT NDV as the negative control. The vaccine was administered via the intramuscular (I.M.) route at D0 and D21. Blood was collected at D21 and D43. Mice were sensitized with Ad5-hACE2 at D45 and challenged with 10⁵ PFU of the USA-WA1/2020 strain. b Spike-specific serum IgG. Antibodies in post-prime (D21) and post-boost (D43) sera were measured by ELISAs. Geometric mean titer (GMT) represented by the area under the curve (AUC) was graphed, with geometric standard deviation (SD) as error bars. c Neutralizing activity of serum antibodies. Post-boost sera from group 1, group 7, and group 10 were pooled within each group and tested in neutralization assays against the USA-WA1/2020 strain (WT), the B.1.351 variant, and B.1.1.7 variant in technical duplicate. Serum dilutions inhibiting 50% of the infection (ID₅₀) were plotted. (LoD: limit of detection; LoD = 1:20; An ID₅₀ = 1:10 was assigned to negative samples). d Viral load in the lungs. Lungs of a subset of animals (n = 4) from each group were collected on day 2 and day 5 post challenge. The whole lungs were homogenized in 1 mL of PBS. Viral titers were measured by plaque assay on Vero E6 cells and plotted as GMT of PFU/mL (LoD = 50 PFU/mL; a titer of 25 PFU/mL was assigned to negative samples). The error bars represent geometric SD. Statistical difference was analyzed by ordinary one-way ANOVA corrected for Dunnett’s multiple comparisons test (****p < 0.0001). (blue: unadjuvanted groups; red: 0.1 µg adjuvanted with CpG 1018; green: 0.03 µg adjuvanted with CpG 1018).