SARS-CoV-2 Omicron XBB.1 variant was isolated from respiratory tract specimens of laboratory-confirmed COVID-19 patient in Hong Kong. Other clinical isolates of SARS-CoV-2 have been described previously13,14. VeroE6-TMPRSS2 cells were cultured until confluency. Viruses were inoculated the next day and incubated at 37 °C for 5 days. Viral supernatant was harvested and inactivated with paraformaldehyde to a final concentration of 0.2% at 4 °C for 5 more days. Virions were concentrated and purified as described5. Purified inactivated SARS-CoV-2 containing 3 μg of protein was mixed at 1:1 ratio with ImjectTM Alum Adjuvant (Thermo Scientific, 77161) before injected intramuscularly into Syrian golden hamsters. A booster dose was given after 14 days.

ELISA assays for SARS-CoV-2 spike receptor binding domain (RBD) or nucleocapsid, surrogate virus neutralization test (sVNT) using cPassTM SARS-CoV-2 Neutralization Antibody Detection Kit, and live virus microneutralization assay (LVMNA) were carried out as previously described15.

All protocols for animal experiments have been approved by the Committee on the Use of Live Animals in Teaching and Research (CULATR) of the University of Hong Kong (Reference code: CULATR 5754-21). They were also performed in accordance with the standard operating procedures of Biosafety Level 3 animal facilities. Male Syrian golden hamsters aged 4-6 weeks old were used. At day 99 post-vaccination, hamsters anesthetized with ketamine (200 mg/kg) and xylazine (10 mg/kg) intraperitoneally were intranasally challenged with 105 PFU (in 50 μL) of SARS-CoV-2 Omicron XBB.1 subvariant. At 2 days post-infection (dpi), hamsters were sacrificed via intraperitoneal injection of dorminal (200 μL) for virological and histopathological evaluation. RT-qPCR and Western blotting were performed as described5,15. Primers used are listed in Supplementary Table 1.

Statistical significance was assessed by unpaired two-tailed Student’s t test or one-way analysis of variance (ANOVA) with Welch’s correction and Dunnett’s post-hoc test using Prism version 8.4.3.

Further information on research design is available in the Nature Research Reporting Summary linked to this article.