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Porcine Immunoglobulin Fc Fused P30/P54 Protein of African Swine Fever Virus Displaying on Surface of S. cerevisiae Elicit Strong Antibody Production in Swine

Abstract

African swine fever virus (ASFV) infects domestic pigs and European wild boars with strong, hemorrhagic and high mortality. The primary cellular targets of ASFV is the porcine macrophages. Up to now, no commercial vaccine or effective treatment available to control the disease. In this study, three recombinant Saccharomyces cerevisiae (S. cerevisiae) strains expressing fused ASFV proteins-porcine Ig heavy chains were constructed and the immunogenicity of the S. cerevisiae-vectored cocktail ASFV feeding vaccine was further evaluated. To be specific, the P30-Fcγ and P54-Fcα fusion proteins displaying on surface of S. cerevisiae cells were produced by fusing the Fc fragment of porcine immunoglobulin IgG1 or IgA1 with p30 or p54 gene of ASFV respectively. The recombinant P30-Fcγ and P54-Fcα fusion proteins expressed by S. cerevisiae were verified by Western blotting, flow cytometry and immunofluorescence assay. Porcine immunoglobulin Fc fragment fused P30/P54 proteins elicited P30/P54-specific antibody production and induced higher mucosal immunity in swine. The absorption and phagocytosis of recombinant S. cerevisiae strains in IPEC-J2 cells or porcine alveolar macrophage (PAM) cells were significantly enhanced, too. Here, we introduce a kind of cheap and safe oral S. cerevisiae-vectored vaccine, which could activate the specific mucosal immunity for controlling ASFV infection.

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References

  1. Argilaguet JM, Pérez-Martín E, López S, Goethe M, Rodríguez F (2013) BacMam immunization partially protects pigs against sublethal challenge with African swine fever virus. Antiviral Res 98:61–65

    CAS  PubMed  Google Scholar 

  2. Barderas MG, Rodríguez F, Gómez-Puertas P, Avilés M, Beitia F, Alonso C, Escribano JM (2001) Antigenic and immunogenic properties of a chimera of two immunodominant African swine fever virus proteins. Arch Virol 146:1681–1691

    CAS  PubMed  Google Scholar 

  3. Chen X, Yang J, Ji Y, Okoth E, Liu B, Li X, Yin H, Zhu Q (2016) Recombinant Newcastle disease virus expressing African swine fever virus protein 72 is safe and immunogenic in mice. Virol Sin 31:150–159

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Chen P, Zhang L, Chang N, Shi P, Gao T, Zhang L, Huang J (2018) Preparation of virus-like particles for porcine circovirus type 2 by yeast fab assembly. Virus Genes 54:246–255

    CAS  PubMed  Google Scholar 

  5. Colgrove GS, Haelterman EO, Coggins L (1969) Pathogenesis of African swine fever in young pigs. Am J Vet Res 30:1343–1359

    CAS  PubMed  Google Scholar 

  6. Crowe J, Dbeli H, Gentz R, Hochuli E, Henco K (1994) 6xHis-Ni-NTA chromatography as a superior technique in recombinant protein expressiod/purification. Methods Mol Biol 31:371–387

    CAS  PubMed  Google Scholar 

  7. Czajkowsky DM, Hu J, Shao Z, Pleass RJ (2012) Fc-fusion proteins: new developments and future perspectives. EMBO Mol Med 4:1015–1028

    CAS  PubMed  PubMed Central  Google Scholar 

  8. De Smet R, Allais L, Cuvelier CA (2014) Recent advances in oral vaccine development. Hum Vaccin Immunother 10:1309–1318

    PubMed  PubMed Central  Google Scholar 

  9. Doytchinova IA, Flower DR (2007) VaxiJen: a server for prediction of protective antigens, tumour antigens and subunit vaccines. BMC Bioinform 8:4

    Google Scholar 

  10. Frommel D, Hong R (1970) Studies on human immunoglobulin G Fc sub-fragments. Structural requirements for biological expression. Biochim Biophys Acta 200:113–124

    CAS  PubMed  Google Scholar 

  11. Fujita Y, Katahira S, Ueda M, Tanaka A, Okada H, Morikawa Y, Fukuda H, Kondo A (2002) Construction of whole-cell biocatalyst for xylan degradation through cell-surface xylanase display in Saccharomyces cerevisiae. J Mol Catal B Enzym 17:189–195

    CAS  Google Scholar 

  12. Galindo I, Cuesta-Geijo MA, Hlavova K, Muñoz-Moreno R, Barrado-Gil L, Dominguez J, Alonso C (2015) African swine fever virus infects macrophages, the natural host cells, via clathrin- and cholesterol-dependent endocytosis. Virus Res 200:45–55

    CAS  PubMed  Google Scholar 

  13. Gallardo C, Mwaengo DM, Macharia JM, Arias M, Taracha EA, Soler A, Okoth E, Martín E, Kasiti J, Bishop RP (2009) Enhanced discrimination of African swine fever virus isolates through nucleotide sequencing of the p54, p72, and pB602L (CVR) genes. Virus Genes 38:85

    CAS  PubMed  Google Scholar 

  14. Gamkrelidze M, Dąbrowska K (2014) T4 bacteriophage as a phage display platform. Arch Microbiol 196:473–479

    CAS  PubMed  PubMed Central  Google Scholar 

  15. Gietz RD, Woods RA (2002) Transformation of yeast by lithium acetate/single-stranded carrier DNA/polyethylene glycol method. Methods Enzymol 350:87–96

    CAS  PubMed  Google Scholar 

  16. Guo J, Li F, He Q, Jin H, Liu M, Li S, Hu S, Xiao Y, Bi D, Li Z (2016) Neonatal Fc receptor-mediated IgG transport across porcine intestinal epithelial cells: potentially provide the mucosal protection. DNA Cell Biol 35:301–309

    CAS  PubMed  Google Scholar 

  17. Jankovich JK, Chapman D, Hansen DT, Robida MD, Loskutov A, Craciunescu F, Borovkov A, Kibler K, Goatley L, King K (2018) Immunisation of pigs by DNA prime and recombinant vaccinia virus boost to identify and rank African swine fever virus immunogenic and protective proteins. J Virol 92:e02219-17

    Google Scholar 

  18. Jazayeri JA, Carroll GJ (2008) Fc-based cytokines: prospects for engineering superior therapeutics. BioDrugs 22:11–26

    CAS  PubMed  Google Scholar 

  19. Kim HJ, Kim HJ (2016) Yeast as an expression system for producing virus-like particles: what factors do we need to consider? Lett Appl Microbiol 64:111–123

    PubMed  Google Scholar 

  20. Kiyono H, Azegami T (2015) The mucosal immune system: from dentistry to vaccine development. Proc Jpn Acad Ser B Phys Biol Sci 91:423–439

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Klemm P, Schembri MA (2000) Fimbrial surface display systems in bacteria: from vaccines to random libraries. Microbiology 146:3025–3032

    CAS  PubMed  Google Scholar 

  22. Leibundgut-Landmann S, Gross O, Robinson MJ, Osorio F, Slack EC, Tsoni SV, Schweighoffer E, Tybulewicz V, Brown GD, Ruland J, Reis e Sousa C (2007) Syk- and CARD9-dependent coupling of innate immunity to the induction of T helper cells that produce interleukin 17. Nat Immunol 8:630–638

    CAS  PubMed  Google Scholar 

  23. Liu N, Feng WU, Yi-Bing XU, Wang Q, Guo CH, Huang YM (2011) Expression of the ORF2 gene of porcine circovirus type 2 in yeast. China Anim Husb Vet Med 38:80–83 (in Chinese)

    CAS  Google Scholar 

  24. Lubisi BA, Bastos ADS, Dwarka RM, Vosloo W (2005) Molecular epidemiology of African swine fever in East Africa. Arch Virol 150:2439–2452

    CAS  PubMed  Google Scholar 

  25. Ludtke JJ, Sololoff AV, Wong SC, Zhang G, Wolff JA (2007) In vivo selection and validation of liver-specific ligands using a new T7 phage peptide display system. Drug Deliv 14:357–369

    CAS  PubMed  Google Scholar 

  26. Luther NJ, Udeama PG, Majiyagbe KA, Shamaki D, Owolodun OA (2008) Polymerase chain reaction (PCR) detection of the genome of African swine fever virus (ASFV) from natural infection in a Nigerian baby warthog (Phacochoereus aethiopicus). Niger Vet J. https://doi.org/10.4314/nvj.v28i2.3559

    Article  Google Scholar 

  27. Luzio NRD, Williams DL (1978) Protective effect of glucan against systemic Staphylococcus aureus septicemia in normal and leukemic mice. Infect Immun 20:804–810

    PubMed  PubMed Central  Google Scholar 

  28. Markova N, Kussovski V, Drandarska I, Nikolaeva S, Georgieva N, Radoucheva T (2003) Protective activity of Lentinan in experimental tuberculosis. Int Immunopharmacol 3:1557–1562

    CAS  PubMed  Google Scholar 

  29. Mendez-Fernandez YV, Pogulis RP, Block MS, Johnson AJ, Kuhns ST, Allen KS, Hansen MJ, Pease LR (2010) Enhanced binding of low-affinity antibodies interacting simultaneously with targeted cell surface molecules and Fc receptor. Tissue Antigens 60:515–525

    Google Scholar 

  30. Negri DRM, Riccomi A, Pinto D, Vendetti S, Rossi A, Cicconi R, Ruggiero P, Giudice GD, Magistris MTD (2010) Persistence of mucosal and systemic immune responses following sublingual immunization. Vaccine 28:4175–4180

    CAS  PubMed  Google Scholar 

  31. Niwa M, Maruyama H, Fujimoto T, Dohi K, Maruyama IN (2000) Affinity selection of cDNA libraries by λ phage surface display. Gene 256:229–236

    CAS  PubMed  Google Scholar 

  32. Ohmit SE, Victor JC, Rotthoff JR, Teich ER, Truscon RK (2006) Prevention of antigenically drifted influenza by inactivated and live attenuated vaccines. N Engl J Med 355:2513–2522

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Olson EJ, Standing JE, Griego-Harper N, Hoffman OA, Limper AH (1996) Fungal beta-glucan interacts with vitronectin and stimulates tumor necrosis factor alpha release from macrophages. Infect Immun 64:3548–3554

    CAS  PubMed  PubMed Central  Google Scholar 

  34. Oura CAL, Denyer MS, Takamatsu H, Parkhouse RME (2005) In vivo depletion of CD8+ T lymphocytes abrogates protective immunity to African swine fever virus. J Gen Virol 86:2445–2450

    CAS  PubMed  Google Scholar 

  35. Parisotto G, Souza JSD, Ferrão MF, Furtado JC, Molz RF (1998) The African swine fever virus proteins p54 and p30 are involved in two distinct steps of virus attachment and both contribute to the antibody-mediated protective immune response. Virology 243:461–471

    Google Scholar 

  36. Rock DL (2017) Challenges for African swine fever vaccine development—“… perhaps the end of the beginning”. Vet Microbiol 206:52–58

    CAS  PubMed  Google Scholar 

  37. Ruiz-Gonzalvo F, Rodríguez F, Escribano JM (1996) Functional and immunological properties of the baculovirus-expressed hemagglutinin of African swine fever virus. Virology 218:285–289

    CAS  PubMed  Google Scholar 

  38. Sanna G, Dei GS, Bacciu D, Angioi PP, Giammarioli M, De Mia GM, Oggiano A (2017) Improved strategy for molecular characterization of African swine fever viruses from sardinia, based on analysis of p30, CD2V and I73R/I329L variable regions. Transbound Emerg Dis 64:1280–1286

    CAS  PubMed  Google Scholar 

  39. Sivelle C, Sierocki R, Ferreira-Pinto K, Simon S, Maillere B, Nozach H (2018) Fab is the most efficient format to express functional antibodies by yeast surface display. Mabs 10:720–729

    CAS  PubMed  PubMed Central  Google Scholar 

  40. Song L, Liu Y, Chen J (2010) Inorganic binding peptide-mediated immobilization based on baculovirus surface display system. J Basic Microbiol 50:457–464

    CAS  PubMed  Google Scholar 

  41. Stedman AC (2017) BCG as a vaccine vehicle to deliver porcine immunity to African swine fever virus. Dissertation, University of Surrey

  42. Vaughn DE, Bjorkman PJ (1997) High-affinity binding of the neonatal Fc receptor to its IgG ligand requires receptor immobilization. Biochemistry 36:9374–9380

    CAS  PubMed  Google Scholar 

  43. Wang SC, Bligh SWA, Zhu CL, Shi SS, Wang ZT, Hu ZB, Crowder J, Branford-White C, Vella C (2008) Sulfated β-glucan derived from oat bran with potent anti-HIV activity. J Agric Food Chem 56:2624–2629

    CAS  PubMed  Google Scholar 

  44. Wozniak-Knopp G, Stadlmayr G, Perthold JW, Stadlbauer K, Woisetschläger M, Sun H, Rüker F (2017) Designing Fcabs: well-expressed and stable high affinity antigen-binding Fc fragments. Protein Eng Des Sel 30:1–15

    Google Scholar 

  45. Woźniakowski G, Frączyk M, Kowalczyk A, PomorskaMól M, Niemczuk K, Pejsak Z (2017) Polymerase cross-linking spiral reaction (PCLSR) for detection of African swine fever virus (ASFV) in pigs and wild boars. Sci Rep 7:42903

    PubMed  PubMed Central  Google Scholar 

  46. Yoshida M, Masuda A, Kuo TT, Kobayashi K, Claypool SM, Takagawa T, Kutsumi H, Azuma T, Lencer WI, Blumberg RS (2007) IgG transport across mucosal barriers by neonatal Fc receptor for IgG and mucosal immunity. Springer Semin Immunopathol 28:397–403

    Google Scholar 

  47. Zhang G, Peng Y, Schoenlaub L, Elliott A, Mitchell W, Zhang Y (2013) Formalin-inactivated Coxiella burnetii phase I vaccine-induced protection depends on B cells to produce protective IgM and IgG. Infect Immun 81:2112–2122

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Zhao D, Liu R, Zhang X, Li F, Wang J, Zhang J, Liu X, Wang L, Zhang J, Wu X, Guan Y, Chen W, Wang X, He X, Bu Z (2019) Replication and virulence in pigs of the first African swine fever virus isolated in China. Emerg Microb Infect 8:438–447

    CAS  Google Scholar 

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Acknowledgements

This work was supported by the National Key Research and Development Program of China (2018YFD0500500).

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Authors

Contributions

JH conceived and designed the experiments. CC, DH, JS, Z T, LZ, M Z and KT performed the experiments. CC analyzed the data. JH contributed reagents/materials/analysis tools. CC and JH wrote the paper.

Corresponding author

Correspondence to Jinhai Huang.

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The authors declare that they have no conflict of interest.

Animal and Human Rights Statement

The whole study was approved by the Administrative Committee on Animal Welfare of the Institute of Radiation Medicine, Academy of Medical Sciences, China (Laboratory Animal Ethical and Welfare Committee, permit number IRM-DWLL-2019020). All institutional and national guidelines for the care and use of laboratory animals were followed.

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Chen, C., Hua, D., Shi, J. et al. Porcine Immunoglobulin Fc Fused P30/P54 Protein of African Swine Fever Virus Displaying on Surface of S. cerevisiae Elicit Strong Antibody Production in Swine. Virol. Sin. 36, 207–219 (2021). https://doi.org/10.1007/s12250-020-00278-3

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Keywords

  • African swine fever virus (ASFV)
  • S. cerevisiae
  • Porcine immunoglobulin Fc
  • P30-Fcγ/P54-Fcα fusion proteins