The role of IBV proteins in protection: cellular immune responses. COST meeting WG2 + WG3 Budapest, Hungary, 2015

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1 The role of IBV proteins in protection: cellular immune responses COST meeting WG2 + WG3 Budapest, Hungary,

2 Presentation include: Laboratory results Literature summary Role of T cells in response to IBV vaccination and protection Role of innate immunity in response to IBV Summary

3 A. Laboratory results: Cellular immune response to IBV structural proteins S1, N and M 3

4 Experimental design Antigens S1, N and M IBV structural proteins were expressed in E.coli Positive control: H120 Challenge strain: M41 Immunization protocol: SPF chicks were immunized three times by eyedrop at 1, 14 and 28 days of age. 4

5 Experimental scheme: Vaccination Spleen cells culture Stimulation with antigen ELISPOT for cells secreting interferon γ 5

6 in vitro stimulation by M41 Fig. 2. INDEX IFN secreting-cells vaccine H120 M41 6

7 Fig. 3. N Vaccination with N induce high rate of IFNγ-SECRETING SPLENOCYTES 7

8 Fig. 4. Stimulator- S1 INDEX IFN secreting-cells Vaccine ללא חיסון H120 S1 S1+LT N+LT LT S Vaccination with S induce high rate of IFNγ-secreting splenocytes 8

9 Table 1. Immune responses and resistance to challenge Antigen H120 S1 N M PBS IFNɣ PRODUCING CELLS ANTIBODIES CHALLENGE RESISTANCE

10 Conclusions The possibility to induce a T cell immune response following non injected (and no adjuvant) vaccination with recombinant proteins was demonstrated

11 Conclusions The possibility to induce a T cell immune response following non injected (and no adjuvant) vaccination with recombinant proteins was demonstrated Available T cell epitopes on S and N

12 Conclusions The possibility to induce a T cell immune response following non injected (and no adjuvant) vaccination with recombinant proteins was demonstrated Available T cell epitopes on S and N No induction following immunization or stimulation with rm protein

13 Conclusions The possibility to induce a T cell immune response following non injected (and no adjuvant) vaccination with recombinant proteins was demonstrated Available T cell epitopes on S and N No induction following immunization or stimulation with rm protein Specific response to each protein

14 Conclusions The possibility to induce a T cell immune response following non injected (and no adjuvant) vaccination with recombinant proteins was demonstrated Available T cell epitopes on S and N No induction following immunization or stimulation with rm protein Specific response to each protein Birds vaccinated by H120 (whole virus) respond to stimulation by recombinant proteins

15 Conclusions The possibility to induce a T cell immune response following non injected (and no adjuvant) vaccination with recombinant proteins was demonstrated Available T cell epitopes on S and N No induction following immunization or stimulation with rm protein Specific response to each protein Birds vaccinated by H120 (whole virus) respond to stimulation by recombinant proteins Only partial protection was achieved following recombinant proteins immunization

16 B. Literature summary The role of T lymphocytes and innate immune response in protection 16

17 IBV immunological characteristics *Low immunogenicity *Range of protection mainly homologous strains *Long term of development and implementation of a protective immune response *Risk of recombination of attenuated vaccine with field virulent strains 17

18 Role of T cells in response to IBV vaccination and protection 18

19 1. As related to antibody response Antibody level is challenge dependent and increase dramatically following challenge, CD8+ respond to vaccine and elevate faster following challenge. (Vet Immunol. Immunopath. 148: , 2012) 25 19

20 1. As related to antibody response Antibody level is challenge dependent and increase dramatically following challenge, CD8+ respond to vaccine and elevate faster following challenge. (Vet Immunol. Immunopath. 148: , 2012) 25 A vaccine composed of fowlpox virus expressing S1 and IFNɣ induce less antibodies but faster elimination of virus as compared to the attenuated vaccine. (Vaccine 23: 7046, 2009) 37 20

21 2. Protection CD8+ responses (as tested by secretion of IFNɣ and granzyme) following vaccination were found in correlation with protection, including: decreased score of tracheal lesions and viral load, as well as protective memory (Viral Immunol. 26: , 2013) 21 (Viral Immunol. 27: , 2014) 30 21

22 2. Protection CD8+ responses (as tested by IFNɣ and granzyme levels) following vaccination were found in correlation with protection, including: decreased score of tracheal lesions and viral load, as well as protective memory (Viral Immunol. 26: , 2013) 21 (Viral Immunol. 27: , 2014) 30 Vaccination with S2 (no neutralizing antibody epitopes) protect against dis similar S1. This may indicate the importance of CTL in protection against IBV (Avian Dis. 58: 83 89, 2014) 29 22

23 2. Protection CD8+ responses (as tested by IFNɣ and granzyme levels) following vaccination were found in correlation with protection, including: decreased score of tracheal lesions and viral load, as well as protective memory (Viral Immunol. 26: , 2013) 21 (Viral Immunol. 27: , 2014) 30 Vaccination with S2 (no neutralizing antibody epitopes) protect against dis similar S1. This may indicate the importance of CTL in protection against IBV (Avian Dis. 58: 83 89, 2014) 29 Following adoptive transfer: IBV primed CD8+ protect chicks from acute infection. (Dev. Comp. Immunol. 24: , 2000) 22 23

24 3. Timing of response CD8+ eliminate IBV at the first stages of infection and humoral response at later stages (J. Vet. Med. Sci. 62:397, 2000) 26 (Dev. Comp. Immunol. 24: , 2000) 22 24

25 4. T cell epitopes T cell epitopes were identified on S and N IBV structural proteins, but not M (Dev. Comp. IMMUNOL. 24: , 2000; Arch. Virol. 150: , 2005) 25

26 CTL response is MHC haplotype dependent

27 Avian present and respond differently to MHCI epitopes according to their MHC class I and II haplotypes. In a commercial flock a variety of haplotypes exist Meaning: The response to vaccine differ among birds. = Partial effectivenes of a vaccine may be due to many variants of the virus but also a result of diversity in haplotypes in a flock. 27

28 Influence of MHC haplotype on response to IBV Mortality following challenge was significantly influenced by the MHC (B) haplotype of genetic lines The B*15 haplotype were resistant in contrast to chickens possessing the B*13 or B*21 haplotypes (Avian Pathol. 33:605, 2004) 27

29 Haplotype-dependent innate responses Innate immune response (e.g. NO production following stimulation) was higher in B19 haplotype as compared to B2 and was in correlation with resistance to IBV challenge Poultry Sci 93: 830,

30 ROLE OF INNATE IMMUNITY IN RESPONSE TO IBV 30

31 Innate response to IBV Following IBV infection: Down regulation of innate immune genes (TLR 3, IL 1β, IFNɣ) Initiation of transcription of those genes and recruitment of macrophages Delay in response> inadequate for protection and associated with IBV replication and histological changes Possible mechanism for down regulation of host innate immune molecules (studied for other coronaviruses): disruption of downstream signaling cascades by viral proteins (e.g. nucleocapsid N). Virology

32 Innate immune memory Epigenetic control of myeloid cell differentiation, identity and function. Nat. Rev. Immunol. 15:7 17, Towards a better understanding of host defense mechanisms Current Opinion in Immunology 29:1 7, 2014

33 SUMMARY Proposed model Vet. Immunol. IMMUNOPATH. 121: , 2008 IBV Recognition by TLR etc. cytokines CD4 + T cell activation Th 1? Th2 CD8 + B (antibodies) 33

34 SUMMARY The immune response to IBV include innate, CD8+ and antibody responses at this order Immunological acquired protection involve both: CTL and antibodies Whereas CTL based vaccine is less virus variability dependent, it is haplotype dependent Innate immunity have a major role inresponse to IBV

35 Proposals for directions in vaccine development A. Broad range vaccine Partially effective vaccines may contribute to the emergence of new IBV strains by mutations and selections (Avian Dis. 56:501, 2012; Avian dis. 58: 102, 2014; and others). Broad range vaccine, consisting on several representatives of genotypes may decrease the escape of mutants and variants

36 B. VACCINES TARGETING CD8+ The consequences of an immune response are critically dependent on class of immunity generated. One major junction is the determination whether antigen predominantly generates Th1 or Th2 cells. Targeting toward Th1 may elevate CD8+

37 ADVANTAGE independent on variability and structural epitopes as respond to linear epitopes, can cross react with distinct strains. DISADVANTAGE MHC haplotype dependent. In our study (not presented) it was found that only limited IBV epitopes can be detected by farm chicken. As a consequence of the scattering of haplotypes in the commercial flocks, only part of the birds will become protected 37

38 Thank you 38

Natalia Taborda Vanegas. Doc. Sci. Student Immunovirology Group Universidad de Antioquia

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