Mouse-adapted SARS-CoV-2 viruses and methods of use thereof

1. A recombinant SARS-CoV-2 virus particle comprising

a spike protein, wherein the spike protein comprises the amino acid sequence: MFVFLVLLPLVSSQCVNLTTRTQLPPAYTNSFTRGVYYPDKVFRSSVLHSTQDLFLPFFS NVTWFHAIHVSGTNGTKRFDNPVLPFNDGVYFASTEKSNIIRGWIFGTTLDSKTQSLLIV NNATNVVIKVCEFQFCNDPFLGVYYHKNNKSWMESEFRVYSSANNCTFEYVSQPFLMD LEGKQGNFKNLREFVFKNIDGYFKIYSKHTPINLVRDLPQGFSALEPLVDLPIGINITRFQT LLALHRSYLTPGDSSSGWTAGAAAYYVGYLQPRTFLLKYNENGTITDAVDCALDPLSET KCTLKSFTVEKGIYQTSNFRVQPTESIVRFPNITNLCPFGEVFNATRFASVYAWNRKRISN CVADYSVLYNSASFSTFKCYGVSPTKLNDLCFTNVYADSFVIRGDEVRQIAPGQTGKIA DYNYKLPDDFTGCVIAWNSNNLDSKVGGNYNYLYRLFRKSNLKPFERDISTEIYQAGST PCNGVEGFNCYFPLQSYGFQPTNGVGYQPYRVVVLSFELLHAPATVCGPKKSTNLVKN KCVNFNFNGLTGTGVLTESNKKFLPFQQFGRDIADTTDAVRDPQTLEILDITPCSFGGVS VITPGTNTSNQVAVLYQDVNCTEVPVAIHADQLTPTWRVYSTGSNVFQTRAGCLIGAEH VNNSYECDIPIGAGICASYQTQTNSPRRARSVASQSIIAYTMSLGAENSVAYSNNSIAIPT NFTISVTTEILPVSMTKTSVDCTMYICGDSTECSNLLLQYGSFCTQLNRALTGIAVEQDK NTQEVFAQVKQIYKTPPIKDFGGFNFSQILPDPSKPSKRSFIEDLLFNKVTLADAGFIKQY GDCLGDIAARDLICAQKFNGLTVLPPLLTDEMIAQYTSALLAGTITSGWTFGAGAALQIP FAMQMAYRFNGIGVTQNVLYENQKLIANQFNSAIGKIQDSLSSTASALGKLQDVVNQN AQALNTLVKQLSSNFGAISSVLNDILSRLDKVEAEVQIDRLITGRLQSLQTYVTQQLIRAA EIRASANLAATKMSECVLGQSKRVDFCGKGYHLMSFPQSAPHGVVFLHVTYVPAQEKN FTTAPAICHDGKAHFPREGVFVSNGTHWFVTQRNFYEPQIITTDNTFVSGNCDVVIGIVN NTVYDPLQPELDSFKEELDKYFKNHTSPDVDLGDISGINASVVNIQKEIDRLNEVAKNLN ESLIDLQELGKYEQYIKWPWYIWLGFIAGLIAIVMVTIMLCCMTSCCSCLKGCCSCGSCC KFDEDDSEPVLKGVKLHYT (SEQ ID NO:2) or an amino acid sequence at least 90% identical thereto, wherein either said amino acid sequence comprises a substitution of Q498Y and P499T.

2. An isolated nucleic acid molecule encoding the SARS-CoV-2 virus particle of claim 1 .

3. An isolated nucleic acid molecule encoding the SARS-CoV-2 virus particle of claim 1 , wherein the nucleic acid molecule comprises one or more of the following nonsynonymous nucleotide substitutions, wherein the numbering is based on the reference nucleotide sequence of SEQ ID NO:1:

G9479T, C9491T, A12678G, A12884G, C23039A, C9438T, A11847G, A12159G, T27221C, C23054T, A23056C, C23057A, C23059G.

4. The isolated nucleic acid molecule of claim 3 , further comprising one or more synonymous nucleotide substitution(s).

5. The isolated nucleic acid molecule of claim 4 , wherein the synonymous nucleotide substitution is T15102A.

6. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:8.

7. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:9.

8. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:10.

9. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:11.

10. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:12.

11. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:15.

12. The isolated nucleic acid molecule of claim 3 , wherein the nucleic acid molecule comprises the nucleotide sequence of SEQ ID NO:16, 17, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39 or 40.

13. A population of SARS-CoV-2 particles comprising the SARS-CoV-2 virus particle of claim 1 .

14. A composition comprising the SARS-CoV-2 virus particle of claim 1 in a pharmaceutically acceptable carrier.

15. A composition comprising the isolated nucleic acid molecule of claim 2 in a pharmaceutically acceptable carrier.

16. A method of producing an immune response to a coronavirus in a subject, comprising administering to the subject an effective amount of the SARS-CoV-2 virus particle of claim 1 .

17. The recombinant SARS-CoV-2 virus particle of claim 1 , wherein the spike protein amino acid sequence further comprises the substitution Q493K.

18. The isolated nucleic acid molecule of claim 2 , wherein the nucleic acid molecule comprises a nucleotide sequence of SEQ ID NO:15.

19. The isolated nucleic acid molecule of claim 2 , wherein the nucleic acid molecule encodes

a substitution of Q493K in the spike protein amino acid sequence, and further encodes

a SARS-CoV-2 nsp4 protein comprising a T285I, G309C and/or H313Y substitution(s);

a SARS-CoV-2 nsp7 protein comprising a K2R substitution;

a SARS-CoV-2 nsp8 protein comprising a E23G and/or K196R substitution;

a SARS-CoV-2 nsp9 protein comprising a T67A substitution; and/or

a SARS-CoV-2 ORF6 protein comprising a F7S substitution,

wherein the numbering corresponds to the amino acid sequences of SEQ ID NO:3 (nsp4 protein), SEQ ID NO:4 (nsp7 protein), SEQ ID NO:5 (nsp8 protein), SEQ ID NO:6 (nsp9 protein) or SEQ ID NO:7 (ORF6 protein), respectively.

20. The isolated nucleic acid molecule of claim 19 , wherein the nucleic acid molecule further encodes

a SARS-CoV-2 nsp4 protein comprising the amino acid sequence of SEQ ID NO:3, wherein said amino acid sequence comprises a T285I substitution;

a SARS-CoV-2 nsp7 protein comprising the amino acid sequence of SEQ ID NO:4, wherein said amino acid sequence comprises a K2R substitution;

a SARS-CoV-2 nsp8 protein comprising the amino acid sequence of SEQ ID NO:5, wherein said amino acid sequence comprises a E23G substitution; and

a SARS-CoV-2 ORF6 protein comprising the amino acid sequence of SEQ ID NO:7, wherein said amino acid sequence comprises a F7S substitution.

21. The isolated nucleic acid molecule of claim 20 , wherein the nucleic acid molecule comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:8, 9, 10, 11, 12, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, and 40.

22. The isolated nucleic acid molecule of claim 20 , wherein the nucleic acid molecule comprises a nucleotide sequence selected from the group consisting of SEQ ID NO:8 and SEQ ID NO:30.