Investigation on the possible application of a serological DIVA monitoring strategy when a rhvt-h5 vaccine is used to control Avian Influenza
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1 AVMA AAAP Convention 2016 August 7, San Antonio, Texas, USA Investigation on the possible application of a serological DIVA monitoring strategy when a rhvt-h5 vaccine is used to control Avian Influenza Yannick Gardin 1, Mieke Steensels 2, Benedicte Lambrecht 2, Vilmos Palya 3, John Elattrache 4, Stephanie Lesceu 5, Sjaak De Wit 6 1 Ceva Santé Animale, France, 2 Coda Cerva, Belgium, 3 Ceva Animal Health, Hungary, 4 Ceva Animal Health, United States, 5 IDvet, France, 6 GD Deventer, The Netherlands.
2 rhvt-h5* AI Vaccine Recombinant vector AI vaccine using HVT as vector with an insert encoding for the HA5 protein. Protects against clinical signs, reduces challenge virus shedding and spreading with reduction in the % of shedders and amount of virus shed. Used in some AI endemic Countries. With obvious technical and commercial success. Sometimes in combination with Killed AI vaccines, depending on the local situation. Technical and / or Regulatory requests for monitoring vaccination and virus circulation * Vectormune AI Ceva Animal Health
3 Monitoring AI vaccination AI VACCINATION Disease Prevention Farm scale Disease Control Territory scale Assessment of Protection «AI ENDEMIC» Countries ERADICATION Monitoring of Infection (DIVA) «AI FREE» Countries
4 Monitoring AI vaccination Administration Immune Coverage Response VACCINE REPLICATION PROTECTION On site check Deposition (blue dye) Transport Storage Preparation PCR Serology Field trials Challenge
5 Monitoring AI vaccination Administration Immune Coverage Response VACCINE REPLICATION PROTECTION On site check Deposition (blue dye) Transport Storage Preparation PCR Serology Field trials Challenge
6 Monitoring AI vaccination Administration Immune Coverage Response VACCINE REPLICATION PROTECTION On site check Deposition (blue dye) Transport Storage Preparation PCR Serology Field trials Challenge FACTORS OF VARIATION
7 rhvt-h5 serology rhvt-h5 induces anti-ha antibodies detectable by HI test provided the antigen used is homologous to the virus that donated the HA insert (clade 2.2 H5N1 HPAIV : A/Swan/Hungary/4999/2006) Antibody production is the consequence of both: - replication of the vector - expression of the insert Testing results may vary significantly depending on various factors with special regards to presence of MDA and time span between vaccination and testing. rhvt-h5 induces no anti-np antibodies
8 Antibody response to rhvt-h5 vaccine Antibody response w/wo MDA H5N1 homologous Ceva-GD antigen (Extract from study SCI SSIU Ceva Phylaxia)
9 Antibody response to rhvt-h5 vaccine Antibody response w/wo MDA H5N1 homologous Ceva-GD antigen (Extract from study SCI SSIU Ceva Phylaxia)
10 Antibody response to rhvt-h5 vaccine Antibody response w/wo MDA H5N1 homologous Ceva-GD antigen (Extract from study SCI SSIU Ceva Phylaxia)
11 Experiment at IZSVe Materials and methods 10 SPF chickens vaccinated at d1 with rhvt-h5 Challenge 28 dpv with H5N1 HPAIV - A/Chicken/Bangladesh/11RS / EID 50 / bird 100μl Allantoic Fluid Choanal slit & cloacal swabs tested by RRT-PCR + virus isolation for RRT-PCR (+) samples - on 2, 4, 6 & 8 dpi Serum samples taken 27 dpv & 14 dpi, tested by: - HI test using 2 antigens: - homologous to the vaccine (co/cr) - identical to the challenge virus (ch/ba) - NP compet. ELISA (ID Screen Influenza A ab IDVet) (Bonfante at al. Trials at IZSVe, 2012)
12 Experiment at IZSVe Detection of infected birds using NP ELISA Chicken ID RRT-PCR (Ct) Days post infection Post vaccination Post infection css cs css cs css cs css cs co/cr ch/ba co/cr ch/ba 34 Neg Neg Neg Neg Neg Neg Neg Neg Neg 35 Neg Neg Neg Neg Neg Neg Neg Neg Neg 37 Neg Neg Neg Neg Neg Neg Neg Neg 2 Neg 7 5 Neg 38 Neg Neg 32.8* Neg Neg Neg Neg Neg 5 4 Pos 39 Neg Neg Neg Neg Neg Neg Neg Neg Neg 60 Neg Neg Neg Neg Neg Neg Neg Neg Pos 76 Neg Neg Neg Neg Neg Neg Neg Neg Neg 77 Neg Neg Neg Neg Neg Neg Neg Neg 3 Neg 6 4 Neg 99 Neg Neg Neg Neg Neg Neg Neg Neg Neg 148 Neg Neg Neg Neg Neg Neg Neg Neg 3 Neg 7 4 Pos GMT css: choanal slit swab co/cr: vaccine related antigen cs: cloacal swab ch/ba: challenge virus antigen * : positive virus isolation GMT: Geometric mean titer HI titers (Log2) NP-ELISA (Bonfante at al. Trials at IZSVe, 2012)
13 Experiment at IZSVe Detection of infected birds using NP ELISA Chicken ID RRT-PCR (Ct) Days post infection Post vaccination Post infection css cs css cs css cs css cs co/cr ch/ba co/cr ch/ba 34 Neg Neg Neg Neg Neg Neg Neg Neg Neg 35 Neg Neg Neg Neg Neg Neg Neg Neg Neg 37 Neg Neg Neg Neg Neg Neg Neg Neg 2 Neg 7 5 Neg 38 Neg Neg 32.8* Neg Neg Neg Neg Neg 5 4 Pos 39 Neg Neg Neg Neg Neg Neg Neg Neg Neg 60 Neg Neg Neg Neg Neg Neg Neg Neg Pos 76 Neg Neg Neg Neg Neg Neg Neg Neg Neg 77 Neg Neg Neg Neg Neg Neg Neg Neg 3 Neg 6 4 Neg 99 Neg Neg Neg Neg Neg Neg Neg Neg Neg 148 Neg Neg Neg Neg Neg Neg Neg Neg 3 Neg 7 4 Pos GMT css: choanal slit swab co/cr: vaccine related antigen cs: cloacal swab ch/ba: challenge virus antigen * : positive virus isolation GMT: Geometric mean titer HI titers (Log2) NP-ELISA (Bonfante at al. Trials at IZSVe, 2012)
14 Experiment at IZSVe Detection of infected birds using NP ELISA Chicken ID RRT-PCR (Ct) Days post infection Post vaccination Post infection css cs css cs css cs css cs co/cr ch/ba co/cr ch/ba 34 Neg Neg Neg Neg Neg Neg Neg Neg Neg 35 Neg Neg Neg Neg Neg Neg Neg Neg Neg 37 Neg Neg Neg Neg Neg Neg Neg Neg 2 Neg 7 5 Neg 38 Neg Neg 32.8* Neg Neg Neg Neg Neg 5 4 Pos 39 Neg Neg Neg Neg Neg Neg Neg Neg Neg 60 Neg Neg Neg Neg Neg Neg Neg Neg Pos 76 Neg Neg Neg Neg Neg Neg Neg Neg Neg 77 Neg Neg Neg Neg Neg Neg Neg Neg 3 Neg 6 4 Neg 99 Neg Neg Neg Neg Neg Neg Neg Neg Neg 148 Neg Neg Neg Neg Neg Neg Neg Neg 3 Neg 7 4 Pos GMT css: choanal slit swab co/cr: vaccine related antigen cs: cloacal swab ch/ba: challenge virus antigen * : positive virus isolation GMT: Geometric mean titer HI titers (Log2) NP-ELISA 3+/10 (Bonfante at al. Trials at IZSVe, 2012)
15 Experiment 1 at Coda-Cerva Materials and methods 2 x 10 SPF chickens vaccinated at d1 with rhvt-h5 Challenge at 28 dpv with H5N81 HPAIV - A/Turkey/Germany-MV/R 2472/2014 (H5N8 GE) EID 50 or 10 6 EID 50 / bird Oropharyngeal & cloacal swabs tested by RRT-PCR - on 2, 5, 9 & 14 dpi Serum samples taken 27 dpv & 14 dpi, tested by: - HI test using 2 antigens: - homologous to the vaccine - identical to the challenge virus (H5N8 GE) - NP compet. ELISA (ID Screen Influenza A ab IDVet) (Steensels et al. Coda Cerva, 9th ISAI, Athens, GA, USA, 2015)
16 Experiment 1 at Coda-Cerva Clinical protection % Survival dpi No vaccination - 10^5 EID50/dose H5N8 GE rhvt-h5-10^5 EID50/dose H5N8 GE No vaccination - 10^6 EID50/dose H5N8 GE rhvt-h5-10^6 EID50/dose H5N8 GE (Steensels et al. Coda Cerva, 9th ISAI, Athens, GA, USA, 2015)
17 Experiment 1 at Coda-Cerva Viral excretion post challenge (Steensels et al. Coda Cerva, 9th ISAI, Athens, GA, USA, 2015)
18 Experiment 1 at Coda-Cerva HI Antibody Responses (Steensels et al. Coda Cerva, 9th ISAI, Athens, GA, USA, 2015)
19 Experiment 1 at Coda-Cerva Detection of infected birds using NP ELISA 0+/10 1+/10 (Steensels et al. Coda Cerva, 9th ISAI, Athens, GA, USA, 2015)
20 Experiment 2 at Coda-Cerva Materials and methods 2 x 15 SPF chickens vaccinated at d1 with rhvt-h5 Challenge at 4 woa with H5N81 HPAIV - A/Turkey/Germany-MV/R 2472/2014 (H5N8 GE) EID 50 / bird Oropharyngeal & cloacal swabs tested by RRT-PCR - on 2, 5, 9 & 14 dpi Serum samples taken 27 dpv & 14, 21 & 28 dpi, tested by: - HI test with 2 antigens: - homologous to the vaccine - identical to the challenge virus (H5N8 GE) - NP compet. ELISA (ID Screen Influenza A ab IDVet) (Ceva Coda Cerva unpublished data, 2015)
21 Experiment 2 at Coda-Cerva Viral excretion post challenge (Ceva Coda Cerva unpublished data, 2015)
22 Experiment 2 at Coda-Cerva HI Antibody Responses (Ceva Coda Cerva unpublished data, 2015)
23 Experiment 2 at Coda-Cerva Detection of infected birds using Indirect NP ELISA 4+/8 (Ceva Coda Cerva unpublished data, 2015)
24 Experiment 2 at Coda-Cerva Detection of infected birds using Competition NP ELISA 4-5+/10 (Ceva Coda Cerva unpublished data, 2015)
25 Conclusions Vaccination against AI (H5) using rhvt-h5 vaccine can be monitored for the vaccine «take» using the HI test (as well as some commercial ELISAs). In case of challenge, vaccination with rhvt-h5 vaccine limits viral invasion with : - a reduced % of infected birds, as detected by RRT-PCRs - a hardly detectable increase in HI titres against homologous antigen - a clear but limited increase in HI titres against the challenge virus used as antigen. - the rise of a variable percentage of birds becoming positive when tested with NP ELISA.
26 Conclusions Vaccination with rhvt-h5 vaccine does not prevent implementation of a DIVA monitoring strategy and an eradication plan. More field experience is required to determine the optimum number of samples to be taken, as well as time intervals between samplings. An improvement in the sensitivity of the NP ELISA will help implementing such an action plan.
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