Recombinant African Swine Fever Virus Arm/07/CBM/c2 Lacking CD2v and A238L Is Attenuated and Protects Pigs against Virulent Korean Paju Strain
Abstract
:1. Introduction
2. Materials and Methods
2.1. Design and Generation of CRISPR-Cas9 Vectors for the Deletion of EP402R and A238L Genes from the Virulent African Swine Fever Virus (ASFV) Strain Arm/07/CBM/c2
2.2. Generation of Recombinant Virus Arm-ΔCD2v-GFP-ΔA238L-Cherry by CRISPR-Cas9 Technology
2.3. Isolation of Recombinant Viruses from Wild-Type Viruses by Plaque Isolation
2.4. Viral DNA Extraction for NGS Analysis
2.5. Illumina Sequencing and Data Analysis
2.6. Viral Growth Kinetics
2.7. RT-qPCR Assay
2.8. Determination of cGAS-STING Pathway Activation by Western Blot
2.9. Animal Experiment
- (1)
- Ethics Statement for Use of Animals
- (2)
- Immunization and Challenge with ASFV Korean isolate
- (3)
- Samples collection and assessment of Clinical signs
2.10. ASFV Real-Time qPCR
2.11. ELISA and Neutralization Assay
3. Results
3.1. Generation of ASFV Recombinant Virus Arm-ΔCD2v-GFP-ΔA238L-mCherry by CRISPR-Cas9 Technology
3.2. Sequence Characterization of Recombinant Virus Arm-ΔCD2v-ΔA238L
3.3. Growth of Arm-ΔCD2v-ΔA238L In Vitro Is Similar to the Growth of Wild-Type Arm/07/CBM/c2
3.4. IFN Regulation In Vitro as a Signature of Attenuation of Arm-ΔCD2v-ΔA238L
3.5. Arm-ΔCD2v-ΔA238L Is Attenuated In Vivo
3.6. Arm-ΔCD2v-ΔA238L Protects against a Virulent Challenge of Genotype II ASFV Korean Paju Strain
3.7. Viremia and Virus Shedding Assessment in the Vaccinated Animals
3.8. Specific Antibodies against ASFV Are Detected in Vaccinated Animals before and after Challenge with ASFV Virulent Korean Paju Strain
4. Discussion
5. Conclusions
6. Patents
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
- Blome, S.; Franzke, K.; Beer, M. African swine fever-A review of current knowledge. Virus Res. 2020, 287, 198099. [Google Scholar] [CrossRef]
- Revilla, Y.; Perez-Nunez, D.; Richt, J.A. African Swine Fever Virus Biology and Vaccine Approaches. Adv. Virus Res. 2018, 100, 41–74. [Google Scholar] [CrossRef] [PubMed]
- Vinuela, E. African swine fever virus. Curr. Top. Microbiol. Immunol. 1985, 116, 151–170. [Google Scholar] [CrossRef]
- Sauter-Louis, C.; Forth, J.H.; Probst, C.; Staubach, C.; Hlinak, A.; Rudovsky, A.; Holland, D.; Schlieben, P.; Goldner, M.; Schatz, J.; et al. Joining the club: First detection of African swine fever in wild boar in Germany. Transbound. Emerg. Dis. 2021, 68, 1744–1752. [Google Scholar] [CrossRef]
- Ge, S.; Li, J.; Fan, X.; Liu, F.; Li, L.; Wang, Q.; Ren, W.; Bao, J.; Liu, C.; Wang, H.; et al. Molecular Characterization of African Swine Fever Virus, China, 2018. Emerg. Infect. Dis. 2018, 24, 2131–2133. [Google Scholar] [CrossRef] [PubMed]
- Gonzales, W.; Moreno, C.; Duran, U.; Henao, N.; Bencosme, M.; Lora, P.; Reyes, R.; Nunez, R.; De Gracia, A.; Perez, A.M. African swine fever in the Dominican Republic. Transbound. Emerg. Dis. 2021, 68, 3018–3019. [Google Scholar] [CrossRef] [PubMed]
- Bosch-Camos, L.; Lopez, E.; Rodriguez, F. African swine fever vaccines: A promising work still in progress. Porc. Health Manag. 2020, 6, 17. [Google Scholar] [CrossRef]
- Wu, K.; Liu, J.; Wang, L.; Fan, S.; Li, Z.; Li, Y.; Yi, L.; Ding, H.; Zhao, M.; Chen, J. Current State of Global African Swine Fever Vaccine Development under the Prevalence and Transmission of ASF in China. Vaccines 2020, 8, 531. [Google Scholar] [CrossRef]
- Tran, X.H.; Phuong, L.T.T.; Huy, N.Q.; Thuy, D.T.; Nguyen, V.D.; Quang, P.H.; Ngon, Q.V.; Rai, A.; Gay, C.G.; Gladue, D.P.; et al. Evaluation of the Safety Profile of the ASFV Vaccine Candidate ASFV-G-DeltaI177L. Viruses 2022, 14, 896. [Google Scholar] [CrossRef]
- Abkallo, H.M.; Hemmink, J.D.; Oduor, B.; Khazalwa, E.M.; Svitek, N.; Assad-Garcia, N.; Khayumbi, J.; Fuchs, W.; Vashee, S.; Steinaa, L. Co-Deletion of A238L and EP402R Genes from a Genotype IX African Swine Fever Virus Results in Partial Attenuation and Protection in Swine. Viruses 2022, 14, 2024. [Google Scholar] [CrossRef] [PubMed]
- Borca, M.V.; Ramirez-Medina, E.; Silva, E.; Vuono, E.; Rai, A.; Pruitt, S.; Holinka, L.G.; Velazquez-Salinas, L.; Zhu, J.; Gladue, D.P. Development of a Highly Effective African Swine Fever Virus Vaccine by Deletion of the I177L Gene Results in Sterile Immunity against the Current Epidemic Eurasia Strain. J. Virol. 2020, 94, 1–15. [Google Scholar] [CrossRef]
- Chen, W.; Zhao, D.; He, X.; Liu, R.; Wang, Z.; Zhang, X.; Li, F.; Shan, D.; Chen, H.; Zhang, J.; et al. A seven-gene-deleted African swine fever virus is safe and effective as a live attenuated vaccine in pigs. Sci. China Life Sci. 2020, 63, 623–634. [Google Scholar] [CrossRef]
- Ding, M.; Dang, W.; Liu, H.; Xu, F.; Huang, H.; Sunkang, Y.; Li, T.; Pei, J.; Liu, X.; Zhang, Y.; et al. Combinational Deletions of MGF360-9L and MGF505-7R Attenuated Highly Virulent African Swine Fever Virus and Conferred Protection against Homologous Challenge. J. Virol. 2022, 96, e0032922. [Google Scholar] [CrossRef]
- Gallardo, C.; Sanchez, E.G.; Perez-Nunez, D.; Nogal, M.; de Leon, P.; Carrascosa, A.L.; Nieto, R.; Soler, A.; Arias, M.L.; Revilla, Y. African swine fever virus (ASFV) protection mediated by NH/P68 and NH/P68 recombinant live-attenuated viruses. Vaccine 2018, 36, 2694–2704. [Google Scholar] [CrossRef]
- Gladue, D.P.; Ramirez-Medina, E.; Vuono, E.; Silva, E.; Rai, A.; Pruitt, S.; Espinoza, N.; Velazquez-Salinas, L.; Borca, M.V. Deletion of the A137R Gene from the Pandemic Strain of African Swine Fever Virus Attenuates the Strain and Offers Protection against the Virulent Pandemic Virus. J. Virol. 2021, 95, e0113921. [Google Scholar] [CrossRef]
- Monteagudo, P.L.; Lacasta, A.; Lopez, E.; Bosch, L.; Collado, J.; Pina-Pedrero, S.; Correa-Fiz, F.; Accensi, F.; Navas, M.J.; Vidal, E.; et al. BA71DeltaCD2: A New Recombinant Live Attenuated African Swine Fever Virus with Cross-Protective Capabilities. J. Virol. 2017, 91, 1–17. [Google Scholar] [CrossRef]
- O’Donnell, V.; Holinka, L.G.; Gladue, D.P.; Sanford, B.; Krug, P.W.; Lu, X.; Arzt, J.; Reese, B.; Carrillo, C.; Risatti, G.R.; et al. African Swine Fever Virus Georgia Isolate Harboring Deletions of MGF360 and MGF505 Genes Is Attenuated in Swine and Confers Protection against Challenge with Virulent Parental Virus. J. Virol. 2015, 89, 6048–6056. [Google Scholar] [CrossRef] [PubMed]
- O’Donnell, V.; Risatti, G.R.; Holinka, L.G.; Krug, P.W.; Carlson, J.; Velazquez-Salinas, L.; Azzinaro, P.A.; Gladue, D.P.; Borca, M.V. Simultaneous Deletion of the 9GL and UK Genes from the African Swine Fever Virus Georgia 2007 Isolate Offers Increased Safety and Protection against Homologous Challenge. J. Virol. 2017, 91, e01760-16. [Google Scholar] [CrossRef] [PubMed]
- Reis, A.L.; Abrams, C.C.; Goatley, L.C.; Netherton, C.; Chapman, D.G.; Sanchez-Cordon, P.; Dixon, L.K. Deletion of African swine fever virus interferon inhibitors from the genome of a virulent isolate reduces virulence in domestic pigs and induces a protective response. Vaccine 2016, 34, 4698–4705. [Google Scholar] [CrossRef] [PubMed]
- Reis, A.L.; Goatley, L.C.; Jabbar, T.; Sanchez-Cordon, P.J.; Netherton, C.L.; Chapman, D.A.G.; Dixon, L.K. Deletion of the African Swine Fever Virus Gene DP148R Does Not Reduce Virus Replication in Culture but Reduces Virus Virulence in Pigs and Induces High Levels of Protection against Challenge. J. Virol. 2017, 91, e01428-17. [Google Scholar] [CrossRef]
- Zhang, J.; Zhang, Y.; Chen, T.; Yang, J.; Yue, H.; Wang, L.; Zhou, X.; Qi, Y.; Han, X.; Ke, J.; et al. Deletion of the L7L-L11L Genes Attenuates ASFV and Induces Protection against Homologous Challenge. Viruses 2021, 13, 255. [Google Scholar] [CrossRef] [PubMed]
- Gladue, D.P.; Borca, M.V. Recombinant ASF Live Attenuated Virus Strains as Experimental Vaccine Candidates. Viruses 2022, 14, 878. [Google Scholar] [CrossRef]
- Perez-Nunez, D.; Castillo-Rosa, E.; Vigara-Astillero, G.; Garcia-Belmonte, R.; Gallardo, C.; Revilla, Y. Identification and Isolation of Two Different Subpopulations Within African Swine Fever Virus Arm/07 Stock. Vaccines 2020, 8, 625. [Google Scholar] [CrossRef]
- Powell, P.P.; Dixon, L.K.; Parkhouse, R.M. An IkappaB homolog encoded by African swine fever virus provides a novel mechanism for downregulation of proinflammatory cytokine responses in host macrophages. J. Virol. 1996, 70, 8527–8533. [Google Scholar] [CrossRef]
- Revilla, Y.; Callejo, M.; Rodriguez, J.M.; Culebras, E.; Nogal, M.L.; Salas, M.L.; Vinuela, E.; Fresno, M. Inhibition of nuclear factor kappaB activation by a virus-encoded IkappaB-like protein. J. Biol. Chem. 1998, 273, 5405–5411. [Google Scholar] [CrossRef]
- Tait, S.W.; Reid, E.B.; Greaves, D.R.; Wileman, T.E.; Powell, P.P. Mechanism of inactivation of NF-kappa B by a viral homologue of I kappa b alpha. Signal-induced release of i kappa b alpha results in binding of the viral homologue to NF-kappa B. J. Biol. Chem. 2000, 275, 34656–34664. [Google Scholar] [CrossRef] [PubMed]
- Miskin, J.E.; Abrams, C.C.; Goatley, L.C.; Dixon, L.K. A viral mechanism for inhibition of the cellular phosphatase calcineurin. Science 1998, 281, 562–565. [Google Scholar] [CrossRef]
- Granja, A.G.; Nogal, M.L.; Hurtado, C.; Del Aguila, C.; Carrascosa, A.L.; Salas, M.L.; Fresno, M.; Revilla, Y. The viral protein A238L inhibits TNF-alpha expression through a CBP/p300 transcriptional coactivators pathway. J. Immunol. 2006, 176, 451–462. [Google Scholar] [CrossRef]
- Granja, A.G.; Nogal, M.L.; Hurtado, C.; Vila, V.; Carrascosa, A.L.; Salas, M.L.; Fresno, M.; Revilla, Y. The viral protein A238L inhibits cyclooxygenase-2 expression through a nuclear factor of activated T cell-dependent transactivation pathway. J. Biol. Chem. 2004, 279, 53736–53746. [Google Scholar] [CrossRef]
- Granja, A.G.; Sabina, P.; Salas, M.L.; Fresno, M.; Revilla, Y. Regulation of inducible nitric oxide synthase expression by viral A238L-mediated inhibition of p65/RelA acetylation and p300 transactivation. J. Virol. 2006, 80, 10487–10496. [Google Scholar] [CrossRef] [PubMed]
- Granja, A.G.; Perkins, N.D.; Revilla, Y. A238L inhibits NF-ATc2, NF-kappa B, and c-Jun activation through a novel mechanism involving protein kinase C-theta-mediated up-regulation of the amino-terminal transactivation domain of p300. J. Immunol. 2008, 180, 2429–2442. [Google Scholar] [CrossRef]
- Granja, A.G.; Sanchez, E.G.; Sabina, P.; Fresno, M.; Revilla, Y. African swine fever virus blocks the host cell antiviral inflammatory response through a direct inhibition of PKC-theta-mediated p300 transactivation. J. Virol. 2009, 83, 969–980. [Google Scholar] [CrossRef]
- Borca, M.V.; Kutish, G.F.; Afonso, C.L.; Irusta, P.; Carrillo, C.; Brun, A.; Sussman, M.; Rock, D.L. An African swine fever virus gene with similarity to the T-lymphocyte surface antigen CD2 mediates hemadsorption. Virology 1994, 199, 463–468. [Google Scholar] [CrossRef] [PubMed]
- Rodriguez, J.M.; Yanez, R.J.; Almazan, F.; Vinuela, E.; Rodriguez, J.F. African swine fever virus encodes a CD2 homolog responsible for the adhesion of erythrocytes to infected cells. J. Virol. 1993, 67, 5312–5320. [Google Scholar] [CrossRef]
- Kay-Jackson, P.C.; Goatley, L.C.; Cox, L.; Miskin, J.E.; Parkhouse, R.M.E.; Wienands, J.; Dixon, L.K. The CD2v protein of African swine fever virus interacts with the actin-binding adaptor protein SH3P7. J. Gen. Virol. 2004, 85, 119–130. [Google Scholar] [CrossRef] [PubMed]
- Perez-Nunez, D.; Garcia-Urdiales, E.; Martinez-Bonet, M.; Nogal, M.L.; Barroso, S.; Revilla, Y.; Madrid, R. CD2v Interacts with Adaptor Protein AP-1 during African Swine Fever Infection. PLoS ONE 2015, 10, e0123714. [Google Scholar] [CrossRef] [PubMed]
- Chaulagain, S.; Delhon, G.A.; Khatiwada, S.; Rock, D.L. African Swine Fever Virus CD2v Protein Induces beta-Interferon Expression and Apoptosis in Swine Peripheral Blood Mononuclear Cells. Viruses 2021, 13, 1480. [Google Scholar] [CrossRef] [PubMed]
- Hemmink, J.D.; Khazalwa, E.M.; Abkallo, H.M.; Oduor, B.; Khayumbi, J.; Svitek, N.; Henson, S.P.; Blome, S.; Keil, G.; Bishop, R.P.; et al. Deletion of the CD2v Gene from the Genome of ASFV-Kenya-IX-1033 Partially Reduces Virulence and Induces Protection in Pigs. Viruses 2022, 14, 1917. [Google Scholar] [CrossRef]
- Teklue, T.; Wang, T.; Luo, Y.; Hu, R.; Sun, Y.; Qiu, H.J. Generation and Evaluation of an African Swine Fever Virus Mutant with Deletion of the CD2v and UK Genes. Vaccines 2020, 8, 763. [Google Scholar] [CrossRef]
- Borca, M.V.; Carrillo, C.; Zsak, L.; Laegreid, W.W.; Kutish, G.F.; Neilan, J.G.; Burrage, T.G.; Rock, D.L. Deletion of a CD2-like gene, 8-DR, from African swine fever virus affects viral infection in domestic swine. J. Virol. 1998, 72, 2881–2889. [Google Scholar] [CrossRef] [PubMed]
- Borca, M.V.; O’Donnell, V.; Holinka, L.G.; Risatti, G.R.; Ramirez-Medina, E.; Vuono, E.A.; Shi, J.; Pruitt, S.; Rai, A.; Silva, E.; et al. Deletion of CD2-like gene from the genome of African swine fever virus strain Georgia does not attenuate virulence in swine. Sci. Rep. 2020, 10, 494. [Google Scholar] [CrossRef] [PubMed]
- Ran, F.A.; Hsu, P.D.; Wright, J.; Agarwala, V.; Scott, D.A.; Zhang, F. Genome engineering using the CRISPR-Cas9 system. Nat. Protoc. 2013, 8, 2281–2308. [Google Scholar] [CrossRef]
- Garcia-Escudero, R.; Vinuela, E. Structure of African swine fever virus late promoters: Requirement of a TATA sequence at the initiation region. J. Virol. 2000, 74, 8176–8182. [Google Scholar] [CrossRef] [PubMed]
- Bolger, A.M.; Lohse, M.; Usadel, B. Trimmomatic: A flexible trimmer for Illumina sequence data. Bioinformatics 2014, 30, 2114–2120. [Google Scholar] [CrossRef]
- Schmieder, R.; Edwards, R. Quality control and preprocessing of metagenomic datasets. Bioinformatics 2011, 27, 863–864. [Google Scholar] [CrossRef]
- Langmead, B.; Salzberg, S.L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 2012, 9, 357–359. [Google Scholar] [CrossRef]
- Cingolani, P.; Platts, A.; Wang le, L.; Coon, M.; Nguyen, T.; Wang, L.; Land, S.J.; Lu, X.; Ruden, D.M. A program for annotating and predicting the effects of single nucleotide polymorphisms, SnpEff: SNPs in the genome of Drosophila melanogaster strain w1118; iso-2; iso-3. Fly 2012, 6, 80–92. [Google Scholar] [CrossRef] [PubMed]
- Wick, R.R.; Judd, L.M.; Gorrie, C.L.; Holt, K.E. Unicycler: Resolving bacterial genome assemblies from short and long sequencing reads. PLoS Comput. Biol. 2017, 13, e1005595. [Google Scholar] [CrossRef] [PubMed]
- Silva, G.G.; Dutilh, B.E.; Matthews, T.D.; Elkins, K.; Schmieder, R.; Dinsdale, E.A.; Edwards, R.A. Combining de novo and reference-guided assembly with scaffold_builder. Source Code Biol. Med. 2013, 8, 23. [Google Scholar] [CrossRef]
- Nadalin, F.; Vezzi, F.; Policriti, A. GapFiller: A de novo assembly approach to fill the gap within paired reads. BMC Bioinform. 2012, 13 (Suppl. 14), S8. [Google Scholar] [CrossRef]
- King, K.; Chapman, D.; Argilaguet, J.M.; Fishbourne, E.; Hutet, E.; Cariolet, R.; Hutchings, G.; Oura, C.A.; Netherton, C.L.; Moffat, K.; et al. Protection of European domestic pigs from virulent African isolates of African swine fever virus by experimental immunisation. Vaccine 2011, 29, 4593–4600. [Google Scholar] [CrossRef]
- Sunwoo, S.Y.; Perez-Nunez, D.; Morozov, I.; Sanchez, E.G.; Gaudreault, N.N.; Trujillo, J.D.; Mur, L.; Nogal, M.; Madden, D.; Urbaniak, K.; et al. DNA-Protein Vaccination Strategy Does Not Protect from Challenge with African Swine Fever Virus Armenia 2007 Strain. Vaccines 2019, 7, 12. [Google Scholar] [CrossRef] [PubMed]
- Perez-Nunez, D.; Sunwoo, S.Y.; Sanchez, E.G.; Haley, N.; Garcia-Belmonte, R.; Nogal, M.; Morozov, I.; Madden, D.; Gaudreault, N.N.; Mur, L.; et al. Evaluation of a viral DNA-protein immunization strategy against African swine fever in domestic pigs. Vet. Immunol. Immunopathol. 2019, 208, 34–43. [Google Scholar] [CrossRef] [PubMed]
- Walczak, M.; Szczotka-Bochniarz, A.; Zmudzki, J.; Juszkiewicz, M.; Szymankiewicz, K.; Niemczuk, K.; Perez-Nunez, D.; Liu, L.; Revilla, Y. Non-Invasive Sampling in the Aspect of African Swine Fever Detection-A Risk to Accurate Diagnosis. Viruses 2022, 14, 1756. [Google Scholar] [CrossRef]
- Gallardo, C.; Soler, A.; Rodze, I.; Nieto, R.; Cano-Gomez, C.; Fernandez-Pinero, J.; Arias, M. Attenuated and non-haemadsorbing (non-HAD) genotype II African swine fever virus (ASFV) isolated in Europe, Latvia 2017. Transbound. Emerg. Dis. 2019, 66, 1399–1404. [Google Scholar] [CrossRef] [PubMed]
- Garcia-Belmonte, R.; Perez-Nunez, D.; Pittau, M.; Richt, J.A.; Revilla, Y. African Swine Fever Virus Armenia/07 Virulent Strain Controls Interferon Beta Production through the cGAS-STING Pathway. J. Virol. 2019, 93, e02298-18. [Google Scholar] [CrossRef] [PubMed]
- Leitao, A.; Cartaxeiro, C.; Coelho, R.; Cruz, B.; Parkhouse, R.M.E.; Portugal, F.C.; Vigario, J.D.; Martins, C.L.V. The non-haemadsorbing African swine fever virus isolate ASFV/NH/P68 provides a model for defining the protective anti-virus immune response. J. Gen. Virol. 2001, 82, 513–523. [Google Scholar] [CrossRef] [PubMed]
- Abkallo, H.M.; Svitek, N.; Oduor, B.; Awino, E.; Henson, S.P.; Oyola, S.O.; Mwalimu, S.; Assad-Garcia, N.; Fuchs, W.; Vashee, S.; et al. Rapid CRISPR/Cas9 Editing of Genotype IX African Swine Fever Virus Circulating in Eastern and Central Africa. Front. Genet. 2021, 12, 733674. [Google Scholar] [CrossRef] [PubMed]
- Borca, M.V.; Rai, A.; Ramirez-Medina, E.; Silva, E.; Velazquez-Salinas, L.; Vuono, E.; Pruitt, S.; Espinoza, N.; Gladue, D.P. A Cell Culture-Adapted Vaccine Virus against the Current African Swine Fever Virus Pandemic Strain. J. Virol. 2021, 95, e0012321. [Google Scholar] [CrossRef]
- Krug, P.W.; Holinka, L.G.; O’Donnell, V.; Reese, B.; Sanford, B.; Fernandez-Sainz, I.; Gladue, D.P.; Arzt, J.; Rodriguez, L.; Risatti, G.R.; et al. The progressive adaptation of a georgian isolate of African swine fever virus to vero cells leads to a gradual attenuation of virulence in swine corresponding to major modifications of the viral genome. J. Virol. 2015, 89, 2324–2332. [Google Scholar] [CrossRef]
- Yanez, R.J.; Rodriguez, J.M.; Nogal, M.L.; Yuste, L.; Enriquez, C.; Rodriguez, J.F.; Vinuela, E. Analysis of the complete nucleotide sequence of African swine fever virus. Virology 1995, 208, 249–278. [Google Scholar] [CrossRef]
- Olesen, A.S.; Kodama, M.; Lohse, L.; Accensi, F.; Rasmussen, T.B.; Lazov, C.M.; Limborg, M.T.; Gilbert, M.T.P.; Botner, A.; Belsham, G.J. Identification of African Swine Fever Virus Transcription within Peripheral Blood Mononuclear Cells of Acutely Infected Pigs. Viruses 2021, 13, 2333. [Google Scholar] [CrossRef] [PubMed]
- Neilan, J.G.; Lu, Z.; Kutish, G.F.; Zsak, L.; Lewis, T.L.; Rock, D.L. A conserved African swine fever virus IkappaB homolog, 5EL, is nonessential for growth in vitro and virulence in domestic swine. Virology 1997, 235, 377–385. [Google Scholar] [CrossRef]
- Salguero, F.J.; Gil, S.; Revilla, Y.; Gallardo, C.; Arias, M.; Martins, C. Cytokine mRNA expression and pathological findings in pigs inoculated with African swine fever virus (E-70) deleted on A238L. Vet. Immunol. Immunopathol. 2008, 124, 107–119. [Google Scholar] [CrossRef]
- Boinas, F.S.; Hutchings, G.H.; Dixon, L.K.; Wilkinson, P.J. Characterization of pathogenic and non-pathogenic African swine fever virus isolates from Ornithodoros erraticus inhabiting pig premises in Portugal. J. Gen. Virol. 2004, 85, 2177–2187. [Google Scholar] [CrossRef]
- Xie, Z.; Liu, Y.; Di, D.; Liu, J.; Gong, L.; Chen, Z.; Li, Y.; Yu, W.; Lv, L.; Zhong, Q.; et al. Protection Evaluation of a Five-Gene-Deleted African Swine Fever Virus Vaccine Candidate Against Homologous Challenge. Front. Microbiol. 2022, 13, 902932. [Google Scholar] [CrossRef]
- Petrovan, V.; Rathakrishnan, A.; Islam, M.; Goatley, L.C.; Moffat, K.; Sanchez-Cordon, P.J.; Reis, A.L.; Dixon, L.K. Role of African Swine Fever Virus Proteins EP153R and EP402R in Reducing Viral Persistence in Blood and Virulence in Pigs Infected with BeninDeltaDP148R. J. Virol. 2022, 96, e0134021. [Google Scholar] [CrossRef]
- Gladue, D.P.; O’Donnell, V.; Ramirez-Medina, E.; Rai, A.; Pruitt, S.; Vuono, E.A.; Silva, E.; Velazquez-Salinas, L.; Borca, M.V. Deletion of CD2-Like (CD2v) and C-Type Lectin-Like (EP153R) Genes from African Swine Fever Virus Georgia-9GL Abrogates Its Effectiveness as an Experimental Vaccine. Viruses 2020, 12, 1185. [Google Scholar] [CrossRef]
- Koltsova, G.; Koltsov, A.; Krutko, S.; Kholod, N.; Tulman, E.R.; Kolbasov, D. Growth Kinetics and Protective Efficacy of Attenuated ASFV Strain Congo with Deletion of the EP402 Gene. Viruses 2021, 13, 1259. [Google Scholar] [CrossRef] [PubMed]
Oligos to Detect: | |
---|---|
GFP (Recombinant virus) | 5′-ACATGGTCCTGCTGGAGTTC-3′ 5′-GCCTGAAATACCAGAAAGAGAAGAC-3′ |
EP402R (Parental virus) | 5′-CCATTAAGCATCATAATTGGGATAAC-3′ 5′-GCCTGAAATACCAGAAAGAGAAGAC-3′ |
mCherry (Recombinant virus) | 5′-CATAACCAATTGCCCATCCTC-3′ 5′-AACTCCTTGATGATGGCCAT-3′ |
A238L (Parental virus) | 5′-CATAACCAATTGCCCATCCTC-3′ 5′-GGAGTTAGTCAAATCTCTTAACCATG-3′ |
Vaccinated Animals | Non-Vaccinated Animals | ||||||||
---|---|---|---|---|---|---|---|---|---|
dpv | #55 | #56 | #57 | #58 | #65 | #66 | #67 | #68 | |
0 | 39.00 | 38.25 | 39.18 | 38.79 | - | - | - | - | |
3 | ND | ND | ND | 37.32 | - | - | - | - | |
5 | ND | ND | ND | ND | - | - | - | - | |
7 | ND | 39.15 | ND | 35.25 | - | - | - | - | |
10 | 37.08 | 39.81 | 36.69 | 41.10 | - | - | - | - | |
14 | 37.18 | ND | 38.05 | ND | - | - | - | - | |
21 | ND | ND | ND | ND | - | - | - | - | |
28 | ND | ND | ND | ND | - | - | - | - | |
dpc | Significance (vaccinated vs. non-vaccinated | ||||||||
0 | ND | ND | ND | ND | ND | ND | ND | ND | - |
3 | ND | 35.26 | 38.97 | 39.11 | 21.25 | 23.14 | 24.86 | 39.34 | ** |
5 | 39.61 | ND | 37.17 | 37.30 | 20.41 | 21.86 | 22.42 | 36.56 | *** |
7 | ND | ND | 37.83 | 40.18 | 19.91 | 21.32 | 22.28 | 22.98 | **** |
10 | 36.28 | ND | 36.49 | 35.74 | 24.87 | 20.56 | ** | ||
14 | ND | ND | 38.77 | 38.59 | |||||
21 | ND | ND | 35.60 | 38.83 |
Nasal Swabs | Fecal Swabs | |||||
---|---|---|---|---|---|---|
dpv | Vaccinated (Mean) | Non-Vaccinated (Mean) | Signifcance Vaccinate vs. Non-Vaccinated | Vaccinated (Media) | Non-Vaccinated (Media) | Signifcance |
0 | ND | ND | ||||
3 | ND | ND | ||||
5 | ND | ND | ||||
7 | ND | 40 ± 0 | ||||
10 | ND | ND | ||||
14 | 38.2 ± 0.043 | 39.645 ± 0.5 | ||||
21 | 39.785 ± 0 | 39.08 ± 0.73 | ||||
28 | ND | 40 ± 0 | ||||
dpc | ||||||
0 | ND | ND | - | ND | 39.97 ± 0.065 | - |
3 | ND | 39.76 ± 0 | - | 38.22 ± 0 | 35.57 ± 2.28 | - |
5 | ND | 33.64 ± 2.44 | * | 38.22 ± 0 | 33.56 ± 5.58 | - |
7 | ND | 27.23 ± 4.03 | **** | ND | 29.36 ± 0.8 | ** |
10 | 39.73 ± 0.045 | 26.87 ± 4.29 | **** | 39.15 ± 0 | 32.27 ± 2.87 | * |
14 | ND | 29.35 ± 2.96 | **** | 38.71 ± 0.51 | - | |
21 | ND | - | ND | - |
Samples | Vaccinated Animals | Non-Vaccinated Animals | Significance (Vaccinated vs. Non-Vaccinated) | ||||||
---|---|---|---|---|---|---|---|---|---|
#55 | #56 | #57 | #58 | #65 | #66 | #67 | #68 | ||
Tonsil | 33.99 | 36.03 | 24.03 | 33.22 | 16.75 | 18.04 | 18.96 | 18.48 | ** |
Man * LN | 36.59 | ND | 32.26 | 39.93 | 18.27 | 19.08 | 16.98 | 19.20 | *** |
SC * LN | 34.23 | 38.93 | 31.53 | 34.01 | 19.21 | 18.44 | 21.02 | 19.60 | *** |
GH * LN | 40.40 | 37.51 | 34.34 | ND | 18.28 | 21.37 | 19.33 | 22.15 | *** |
Renal LN | 38.11 | ND | 30.74 | ND | 18.98 | 21.45 | 18.46 | 20.74 | ** |
Mes LN | 30.99 | ND | 34.85 | 38.21 | 18.09 | 24.54 | 17.60 | 24.25 | ** |
Spleen | ND | 37.61 | 33.55 | 38.22 | 16.86 | 17.79 | 20.06 | 18.83 | *** |
Lung | ND | ND | 28.00 | 35.06 | 16.85 | 19.50 | 18.83 | 21.07 | ** |
Heart | 40.41 | 38.19 | 29.06 | 35.07 | 21.81 | 22.39 | 22..02 | 23.77 | *** |
Liver | 38.81 | ND | 33.08 | 38.07 | 16.44 | 18.92 | 18.69 | 18.07 | *** |
Kidney | ND | 40.13 | 34.56 | 35.55 | 21.06 | 22.16 | 22.09 | 23.61 | *** |
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Pérez-Núñez, D.; Sunwoo, S.-Y.; García-Belmonte, R.; Kim, C.; Vigara-Astillero, G.; Riera, E.; Kim, D.-m.; Jeong, J.; Tark, D.; Ko, Y.-S.; et al. Recombinant African Swine Fever Virus Arm/07/CBM/c2 Lacking CD2v and A238L Is Attenuated and Protects Pigs against Virulent Korean Paju Strain. Vaccines 2022, 10, 1992. https://doi.org/10.3390/vaccines10121992
Pérez-Núñez D, Sunwoo S-Y, García-Belmonte R, Kim C, Vigara-Astillero G, Riera E, Kim D-m, Jeong J, Tark D, Ko Y-S, et al. Recombinant African Swine Fever Virus Arm/07/CBM/c2 Lacking CD2v and A238L Is Attenuated and Protects Pigs against Virulent Korean Paju Strain. Vaccines. 2022; 10(12):1992. https://doi.org/10.3390/vaccines10121992
Chicago/Turabian StylePérez-Núñez, Daniel, Sun-Young Sunwoo, Raquel García-Belmonte, Chansong Kim, Gonzalo Vigara-Astillero, Elena Riera, Dae-min Kim, Jiyun Jeong, Dongseob Tark, Young-Seung Ko, and et al. 2022. "Recombinant African Swine Fever Virus Arm/07/CBM/c2 Lacking CD2v and A238L Is Attenuated and Protects Pigs against Virulent Korean Paju Strain" Vaccines 10, no. 12: 1992. https://doi.org/10.3390/vaccines10121992
APA StylePérez-Núñez, D., Sunwoo, S. -Y., García-Belmonte, R., Kim, C., Vigara-Astillero, G., Riera, E., Kim, D. -m., Jeong, J., Tark, D., Ko, Y. -S., You, Y. -K., & Revilla, Y. (2022). Recombinant African Swine Fever Virus Arm/07/CBM/c2 Lacking CD2v and A238L Is Attenuated and Protects Pigs against Virulent Korean Paju Strain. Vaccines, 10(12), 1992. https://doi.org/10.3390/vaccines10121992