Engineering an Optimal Y280-Lineage H9N2 Vaccine Strain by Tuning PB2 Activity
Abstract
:1. Introduction
2. Results
2.1. A Korean Y280-Lineage H9N2 Virus (SL20wt) Is Genetically Closely Related to Genotype S H9N2 Viruses in China and Different from Y439/Korea-Lineage Vaccine Strain
2.2. Recombinant Korean Y280-Lineage H9N2 Viruses Grow Well in ECEs Regardless of L226Q Mutation
2.3. MVV310PB2 Removes the Replication Capacity of PR8-Derived Recombinant Korean Y280-Lineage Virus in Mammalian Cells
2.4. L226Q Mutation Increases Mammalian Pathogenicity of Recombinant Korean Y280-Lineage Virus
2.5. The L226Q Mutation Reduces Susceptibility to Non-Specific Hemagglutination Inhibitors in Sera of SPF Chicken and Mouse
2.6. Monovalent Inactivated Oil Emulsion Vaccines Induce Very Low HI Antibody Titers to Heterogeneous Antigens
2.7. Bivalent Inactivated Oil Emulsion Vaccine Composed of Y439/Korea- and Korean Y280-Lineage H9N2 Viruses Compensates Skewed Humoral Immunity of Monovalent Vaccines
2.8. Comparison of Specific Neuraminidase Inhibiting Activities of Serum Samples from Monovalent and Bivalent Vaccine-Inoculated Chickens
3. Discussion
4. Materials and Methods
4.1. Viruses, Plasmids, Cells, and Eggs
4.2. RT-PCR, Sequencing, and Sequence Analysis
4.3. Generation of Recombinant Viruses by Site-Directed Mutagenesis and Reverse Genetics
4.4. Titration of Recombinant Viruses in ECEs
4.5. Growth Kinetics and Pathogenicity of Recombinant Viruses in Mammalian Hosts
4.6. Susceptibility Test of Recombinant Viruses to Non-Specific Inhibitors in SPF Chicken and Mouse Sera
4.7. Efficacy Test of Monovalent and Bivalent Inactivated Oil Emulsion Vaccines in Chickens
4.8. Statistical Analysis
5. Conclusions
6. Patents
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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HA | NA | |||||
---|---|---|---|---|---|---|
H3 Numbering | H9 Numbering | SL20wt | 01310 CE20 | SL20 Numbering | SL20wt | 01310 CE20 |
96 | 107 | L | M | 125 (122) | G | S |
133 | Q | L | 126 b | L | S | |
138 | T | R | 127 b | G | N | |
133 | 145 | S | N (NG) a | 187 b | K | R |
145 | 153 | D | G | 199 (196) | K | R |
156 | 164 | Q | H | 248 (245) | G | G |
158 | 166 | N | N (NG) a | 249 b | K | R |
160 | 168 | A | S | 253 (250) | R | R |
183 | 191 | N | H | 296 (293) | K | K |
189 | 197 | D | T | 344 (341) | R | R |
190 | 198 | V | E | 346 b | A | N |
192 | 200 | T | M | 356 (353) | N | Y |
205 | 213 | A | T | 360 b | I | V |
208 | 216 | E | D | 367 b | K | S |
216 | 224 | L | V | 368 (365) | E | K |
226 | 234 | L | Q | 400 (397) | S | N |
273 | 282 | K | N | 401 b | D | N |
274 | 283 | M | S | |||
275 | 285 | S | N | M2e | ||
276 | 287 | T | I | numbering | SL20 | 1310CE20 |
306 | 315 | S | P | 10 | L | P |
325 | 334 | S | A | 13 | T | N |
Virus | HA | NA | PB2 | PB1 | PA | NP | M | NS | EID50/mL(log10) a |
---|---|---|---|---|---|---|---|---|---|
SL20wt | SL20 | SL20 | SL20 | SL20 | SL20 | SL20 | SL20 | SL20 | 9.67 ± 0.29 |
rSL20(P) | SL20 | SL20 | PR8 | PR8 | PR8 | PR8 | PR8 | PR8 | 9.41 ± 0.12 |
rSL20(P)-L226Q | SL20-L226Q | SL20 | PR8 | PR8 | PR8 | PR8 | PR8 | PR8 | 9.42 ± 0.14 |
rSL20(P)-310PB2 | SL20 | SL20 | 01310 | PR8 | PR8 | PR8 | PR8 | PR8 | 8.57 ± 0.46 * |
rSL20(P)-MVV310PB2 | SL20 | SL20 | 01310-MVV b | PR8 | PR8 | PR8 | PR8 | PR8 | 9.58 ± 0.14 |
rSL20-MVV310PB2 | SL20 | SL20 | 01310-MVV | SL20 | SL20 | SL20 | SL20 | SL20 | 9.33 ± 0.52 |
Recombinant Virus | Virus Isolation Rate | TCID50/0.1 mL a (log10) |
---|---|---|
rSL20(P) | 3/3 | 4.00 ± 0.25 |
rSL20(P)-L226Q | 3/3 | 5.00 ± 0.25 |
rSL20(P)-MVV310PB2 | 0/3 | 0.00 ± 0.00 |
Vaccine | Dose (EID50/mL, log10) | Weeks Post Vaccination (wpv) | HI Titer (GMT a) | ||
---|---|---|---|---|---|
rSL20(P) | rSL20(P)-L226Q | rSL20(P)-MVV310PB2 | |||
V-rSL20(P) | 9.25 | 3 wpv | 1195 (756–1633) | 256 (256–256) | 1195 (756–1633) |
4 wpv | 2303 (721–3884) | 288 (90–486) | 1195 (461–1929) | ||
V-rSL20(P)- L226Q | 8.75 | 3 wpv | 1741 (888–2594) | 256 (61–451) | 1075 (283–1867) |
4 wpv | 1638 (942–2355) | 307 (66–548) | 1331 (478–2184) | ||
V-rSL20(P)- MVV310PB2 | 9.00 | 3 wpv | 2560 (653–4467) | 371 (117–625) | 2304 (125–4483) |
4 wpv | 2048 (491–3605) | 666 (239–1092) | 1434 (737–2130) | ||
Negative control (allantoic fluid) | - | 3 wpv | <2 | <2 | <2 |
4 wpv | <2 | <2 | <2 |
Vaccine | Dose (EID50/mL, log10) | Weeks Post Vaccination (wpv) | GMT of HI Titer a | ||||
---|---|---|---|---|---|---|---|
Exp. 1 | Exp. 2 | ||||||
r310-NS28 | rSL20(P)-MVV310PB2 | ||||||
Exp. 1 | Exp. 2 | Exp. 1 | Exp. 2 | ||||
V-r310-NS28 | 8.75 | 9.25 | 2 wpv | NT | 74 (29–226) | NT b | 9 (3–33) |
3 wpv | 128 (23–716) | 294 † (143–604) | 111 (14–869) | 21 (7–67) | |||
4 wpv | 294 (41–2092) | 294 † (143–604) | 55 (7–434) | 42 (13–134) | |||
V-rSL20(P)-MVV310PB2 | 9.00 | 9.00 | 2 wpv | NT | <2 | NT | 294 (143–604) |
3 wpv | <2 | <2 | 2048 ‡ (866–4842) | 891 ‡ (434–1831) | |||
4 wpv | <2 | <2 | 3104 ‡ (1938–4973) | 446 (217–916) | |||
V-r310-NS28 + rSL20(P)-MVV310PB2 | 8.75 + 9.00 | 9.25 + 9.00 | 2 wpv | NT | 10.6 (4–28) | NT | 169 (35–806) |
3 wpv | 338 † (156–729) | 194.0 (90–419) | 2353 ‡ (1601–3457) | 388 (180–838) | |||
4 wpv | 194 (72–517) | 147.0 (72–302) | 1783 (867–3663) | 223 (87–572) | |||
Negative control (allantoic fluid) | - | - | 2 wpv | <2 | <2 | <2 | <2 |
3 wpv | <2 | <2 | <2 | <2 | |||
4 wpv | <2 | <2 | <2 | <2 |
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An, S.-H.; Hong, S.-M.; Song, J.-H.; Son, S.-E.; Lee, C.-Y.; Choi, K.-S.; Kwon, H.-J. Engineering an Optimal Y280-Lineage H9N2 Vaccine Strain by Tuning PB2 Activity. Int. J. Mol. Sci. 2023, 24, 8840. https://doi.org/10.3390/ijms24108840
An S-H, Hong S-M, Song J-H, Son S-E, Lee C-Y, Choi K-S, Kwon H-J. Engineering an Optimal Y280-Lineage H9N2 Vaccine Strain by Tuning PB2 Activity. International Journal of Molecular Sciences. 2023; 24(10):8840. https://doi.org/10.3390/ijms24108840
Chicago/Turabian StyleAn, Se-Hee, Seung-Min Hong, Jin-Ha Song, Seung-Eun Son, Chung-Young Lee, Kang-Seuk Choi, and Hyuk-Joon Kwon. 2023. "Engineering an Optimal Y280-Lineage H9N2 Vaccine Strain by Tuning PB2 Activity" International Journal of Molecular Sciences 24, no. 10: 8840. https://doi.org/10.3390/ijms24108840