Application of Novel Delivery systems for ASFV antigens

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Presented by Waithaka Mwangi at the African Swine Fever Diagnostics, Surveillance, Epidemiology and Control Workshop, Nairobi, Kenya, 20-21 July 2011 …

Presented by Waithaka Mwangi at the African Swine Fever Diagnostics, Surveillance, Epidemiology and Control Workshop, Nairobi, Kenya, 20-21 July 2011

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  • 1. ASFV Diagnostics, Surveillance, Epidemiology and Control:Identification of Researchable Issues Targeted to the Endemic Areas within sub-Saharan Africa Hosted by BecA-ILRI and sponsored by CSIRO-AusAID Waithaka Mwangi Dept. of Veterinary Pathobiology College of Veterinary Medicine and Biomedical Sciences Texas A&M University
  • 2. • ASFV is a highly contagious pathogen that causes devastating hemorrhagic fever in pigs with ~100% case mortality rates.• It causes major economic losses, threatens food security, and limits pig production in affected countries.Goal:Develop a vaccine capable of induction of ASFV-specific protectiveimmunity.
  • 3. p32: immunogenic, implicated in virus internalization, antibody targetp54: Transmembrane, involved in virus particle maturation, antibody targetp72: Main component of the viral capsid, antibody and CTL targetPp220 and pp62 polyproteins:• Produce structural proteins that account for ~32% of the total protein virion mass and are the major components of the core shell• Indispensable for viral replication and production of viable virus
  • 4. Three synthetic codon-optimized chimeric genes generated: Designated saf1, saf2, and saf3.
  • 5. • Replication-incompetent adenovirus (Ad5): - Systemic and mucosal immunization • Bacillus subtilis: mucosal immunization
  • 6. Evaluate immunogenicity and protective efficacy of the lead vaccine candidates following intradermal or mucosalimmunization with recombinant adenovirus (rAd) or Bacillus (rBa), respectively, expressing saf1, saf2, and saf3.
  • 7. Immunization of pigs with adenovirus- or Bacillus-vectored ASFVchimeric antigens will confer systemic and/or mucosal immunity against ASFV Specific Aims:• Test whether intradermal or mucosal immunization of pigs with rAdSAF1-3 will confer protection against ASFV challenge.• Test whether mucosal immunization of pigs with rBaSAF1-3 will confer protection against mucosal ASFV challenge.
  • 8. Positive clone 1 2 3 4 5 6 7 8 9 10 11 Test cloneNegative control Mwangi, W., et al., 2011
  • 9. Generation of rAdenovirusA) B) C)Immunocytometric analysis of 293A cells infected with; A and B) rAdFMD virus; and C) control adenovirus. A and C) were probed with anti-FLAG AP-conjugated mAb, B was probed with an isotype-matched AP-conjugated mAb. Mwangi, W., et al 2011
  • 10. A) B)Immunization of calves with a single dose of the rAdFMD vaccine primed significant;A) FMD1-specific IFN-γ-secreting T cell responses; and B) FMD1-specific T cell proliferation, detectable in seven days. Mwangi, W., et al 2011
  • 11. 450 400 603 605 350# spot/10E5 cells 300 250 200 150 100 50 0 PHA O1Campos FMDV1 10 ug FMDV1 30 ug PBS Filgueira, M.P., et al., 2011
  • 12. IgA values: calculated as the S/P ratio = (sample – negative control)/(positive – negative control). Hargis, B.M., et al 2011
  • 13. Hydropathic profile of the SAF1 chimeric polypeptide
  • 14. A) B) C) D) Immunocytometric analysis of 293A cells transfected with: A) SAFI; B) SAFII; C) SAFIII expression constructs; and D) vector control The cells were probed with anti-FLAG AP-conjugated mAb. Mwangi, W., et al 2011
  • 15. • Recombinant SAFI-III proteins• Recombinant Adenovirus-SAFI-III• Recombinant B. subtilis-SAFI-IIIQuality control analysis
  • 16. Conduct dose-escalation immunization studies in pigs • Evaluate SAFI-III-specific immune responses • Evaluate recall responses upon boost - test sera for recognition of native ASFV antigens - test T cells for reactivity against ASF virus • identify dose required to induce optimal immune responses
  • 17. Conduct immunization studies in pigs; • Prime Evaluate SAFI-III-specific immune responses • Boost • Challenge • Protective index: Survival
  • 18. DC-targeted Control Mwangi, W., et al., 2011
  • 19. A 1 wk post-immunization 3 wks post-immunization 250 ∗ ∗p<0.001 500 ∗ ∗p<0.001IFN- γ + SFC/106 CD8-γ δ -PBMC IFN- γ + SFC/106 CD8-γ δ -PBMC 200 400 150 300 100 200 50 100 0 0 pCC98MSP1 pICMSP1 Vector pCC98MSP1 pICMSP1 Vector Mwangi, W., et al., 2011
  • 20. A 19 wks post-immunization 1 wk post-Boost ∗ ∗p<0.001 ∗ ∗p<0.001 Mwangi, W., et al., 2011
  • 21. Mwangi Lab: Jocelyn Bray, Shehnaz Lokhandwala, Ann-MarrieSurya Waghela Texas A&M University Luc Berghman, Texas A&M UniversityMariano Pérez- Filgueira, Instituto de Virología, CICVyA, INTA- Castelar, ArgentinaBilly M. Hargis University of ArkansasRichard Bishop ILRI in collaboration with DVS, Kenya and CINA-INIA, Valdeolmos, Spain