Fig. 3: Intranasal vaccination of Syrian hamsters with a maximum dose of OTS-228 passage 15 and subsequent challenge infection.
a Experimental setup: Intranasal vaccination of 12 index hamsters with 106.0 TCID50/animal of OTS-228 P.15. Direct contact animals (n = 6) were co-housed 1 dpv. b Survival and (c) body weight were monitored daily, confirming no mortality or weight loss post-vaccination. d Shedding of OTS-228 vaccine virus genome was confirmed by RdRp (nsp12)-specific RT-qPCR of nasal wash samples, determining genome copies per mL (gc/mL). One of six direct contact animals tested positive on days 3 and 7. e Sera from 19 dpv were evaluated for SARS-CoV-2 RBD-specific antibodies by ELISA, confirming a humoral immune response in vaccinated animals and the genome-positive contact animal. f Virus neutralization test (VNT100): Some sera were tested for neutralizing antibodies against different VOCs (threshold > 1:128 starting dilution due to limited sample volume), showing positive results for WT (3/12), Alpha (4/12), and Delta (4/12). Three weeks post-vaccination, the vaccinated animals were challenged intranasally with a mixture of Alpha/Delta SARS-CoV-2 variants, and naïve direct contact animals were co-housed again one day post-challenge in a 1:1 setup. g Survival and (h) body weight were tracked until 14 dpc. None of the vaccinated animals showed mortality or weight loss, while two contact animals did not survived the infection. i Virus shedding: Viral genome copy numbers in nasal wash samples showed relatively high shedding on day 1 (mean 108.2 gc/mL) but a significant decline by day 5 (mean 104.7 gc/mL, a 3017-fold reduction). j Viral genome load was also determined in organ samples at 5 dpc and (k) 14 dpc, with only residual viral genome detectable in lung samples at both time points. l Lung evaluation for pneumonia-related atelectasis at 5 dpc showed no significant damage (Hematoxylin-eosin stained whole lung slide images (bar 2.5 mm), with detailed images showing perivascular (green arrow) and peribronchial inflammatory infiltrates (green arrowhead) (bar 100 µm)), and (m) the absence of SARS-CoV-2 N-protein antigen indicated a high level of lung protection. n Humoral immune response: ELISA measurements confirmed transmission of infection in three of the four remaining contact animals, based on binding of the SARS-CoV-2 RBD-domain of the spike protein. o Neutralizing antibodies: At 14 dpc, the neutralizing humoral immune response, tested in a VNT100, confirmed high titers of biologically relevant cross-reacting neutralizing antibodies in the vaccinated hamsters.
