Application of Novel Delivery systems for ASFV antigens
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
• 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.
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
Three synthetic codon-optimized chimeric genes generated: Designated saf1, saf2, and saf3.
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.
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.
Positive clone 1 2 3 4 5 6 7 8 9 10 11 Test cloneNegative control Mwangi, W., et al., 2011
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
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
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
IgA values: calculated as the S/P ratio = (sample – negative control)/(positive – negative control). Hargis, B.M., et al 2011
Hydropathic profile of the SAF1 chimeric polypeptide
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
• Recombinant SAFI-III proteins• Recombinant Adenovirus-SAFI-III• Recombinant B. subtilis-SAFI-IIIQuality control analysis
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
Conduct immunization studies in pigs; • Prime Evaluate SAFI-III-specific immune responses • Boost • Challenge • Protective index: Survival
A 19 wks post-immunization 1 wk post-Boost ∗ ∗p<0.001 ∗ ∗p<0.001 Mwangi, W., et al., 2011
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