how nd vaccines stimulate bird’s immune system and ... · bird’s immune system and...
TRANSCRIPT
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How ND vaccines stimulate bird’s immune system and
immunological specificity of rHVT-F vector vaccine
F.Rauw
Avian Virology and Immunology Unit
28-30th April 2015 CEVA Poultry Vaccinology Sumit_Budapest, Hungary
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Primary Lymphoid organs in chicken
o The Bursa of Fabricius (BF) :
• This organ is unique to bird
• Regulates the development of the humoral compartment of the immune system
• Essential for the development of B-cells (≈ bone marrow in mammals) (Ig gene diversification and amplification). Only 5% of the total cell numbers survive to form the mature B-cell population. This minority of cells start to emigrate out of bursa around hatching
o The Thymus :
• Regulates the development of the cellular compartment of the immune system
• Essential for the development of T-cells (mainly TH). In chicken, all T-cells originate in thymus and they emigrate to form the peripheral cell pool
o Bursa of Fabricius and thymus regress with onset of sexual maturity.
Thymus
Bursa
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Peripheral Lymphoid organs in chicken
o Chickens have no lymph nodes (≈ the primary site of antigen presentation in mammals).
o Spleen : important for antigen processing and antibody production after hatching. After the involution of the thymus and the bursa, spleen becomes as a primary organ, supplier of lymphoid cells for others peripheral organs.
o Others peripheral organs : lymphoid tissues associated to the harderian gland and to the digestive, respiratory and reproductive tracts
o Day-old chick = fully immunocompetent but not fully immunomature (takes 2 weeks)
Thymus
Bursa
Harderian gland (HG)
Spleen
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Two types of immune responses
o Innate - Natural - Non-specific - Not acquired immunity • Epithelial barriers (skin and mucous membranes)
• Phagocytic cells : granulocytes, macrophages, dendritic cells (DTC)
• Natural killer cells (NK )
• Plasma proteins
Very fast, not specific to pathogens and lack of memory
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Two types of immune responses
o Innate - Natural - Non-specific - Not acquired immunity • Epithelial barriers (skin and mucous membranes)
• Phagocytic cells : granulocytes, macrophages, dendritic cells (DTC)
• Natural killer cells (NK )
• Plasma proteins
Very fast, not specific to pathogens and lack of memory
o Adaptive - Specific - Acquired immunity • B-cells : immunoglobulins (IgG , IgM and IgA)
• T-cells : cytokines (IFN-g, Il-2, IL-4, IL-13 …)
Specificity , diversity and memory
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Bridging
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Adaptative immunity : 5 phases
o Properties : Specificity , diversity and memory
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Adaptative immune response in chickens
Chickens respond to vaccination or infection by developing humoral, cellular and mucosal immune responses specific to antigens
At systemic level :
• Humoral immunity
• Cell-mediated immunity
At local level :
• Mucosal immunity : • Antibody-mediated immunity • Cell-mediated immunity
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Systemic and local immune response in chickens
Local immunity in the digestive tract
Local immunity in the respiratory tract
Local immunity in the head
Humoral immunity
Cell-mediated immunity
Local immunity in the reproductive tract
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Adaptative immune response in chickens
Chickens respond to vaccination or infection by developing humoral, cellular and mucosal immune responses specific to antigens
How is it induced ? How is it acting ? How is it measurable ? Which types of results ?
At systemic level :
• Humoral immunity
• Cell-mediated immunity
At local level :
• Mucosal immunity : • Antibody-mediated immunity • Cell-mediated immunity
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Immune response in chickens
Chickens respond to vaccination or infection by developing humoral, cellular and mucosal immune responses specific to antigens
o Humoral immunity : • B-cells
• Measured by detection of specific antibodies in sera (HI, seroneutralisation, ELISAs)
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Humoral immunity : antibody production
7/05/2015 11
IgM IgG
B
Antigen-specific binding
Primary response (vaccination /infection )
Plasmocytes
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Humoral immunity : antibody production
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IgM IgG
B
Secondary response (boost or infection post-vaccination)
Primary response (vaccination /infection )
Antigen-specific binding
Plasmocytes
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Humoral immunity : antibody production
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IgM IgG
B
Secondary response (boost or infection post-vaccination)
Primary response (vaccination /infection )
Plasmocytes
Antigen-specific binding
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Humoral immunity : antiviral activity
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Blocking viral entry and fusion
NDV
By macrophage
Antibody-depended cell-mediated cytotoxicity by NK cells, macrophages and monocytes
HN and F-NDV Antibodies against F or HN induced by
vaccination
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Humoral immunity : detection
Humoral immunity
Jugular vein
Serum
ELISA
Specific IgG – IgM – IgA - Igtot
HI
HN-specific Ig
Seroneutralisation
Neutralizing Ig
IgG or IgY
IgM IgA
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NDV-specific IgG ELISA
Purified whole NDV Saturation
Mouse anti-Chicken IgG - BIOT
NDV-specific IgG
TMB substrat
A450nm
Streptavidine-POD
0,0
0,5
1,0
1,5
2,0
2,5
1 2 3 4 5
Weeks of age
Negative Tracheotropic ND vaccine Enterotropic ND vaccine
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
Rauw F. et al. (2010). Vaccine, 28 : 823-833
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NDV-specific IgM ELISA
NDV-specific IgM
Purified whole NDV
Mouse IgG1 anti-NDV - BIOT
Mouse anti-Chicken IgM Saturation
TMB substrat
A450nm
Streptavidine-POD
0,0
0,5
1,0
1,5
2,0
2,5
1 2 3 4 5
Weeks of age
Negative Tracheotropic ND vaccine Enterotropic ND vaccine
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
Rauw F. et al. (2010). Vaccine, 28 : 823-833
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NDV-specific IgA ELISA
NDV-specific IgA
Purified whole NDV
Mouse IgG1 anti-NDV - BIOT
Mouse anti-Chicken IgA
TMB substrat
A450nm
Streptavidine-POD
Saturation
0,0
0,2
0,4
0,6
0,8
1,0
1,2
1 2 3 4 5
Weeks of age
Negative Tracheotropic ND vaccine Enterotropic ND vaccine
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
Rauw F. et al. (2010). Vaccine, 28 : 823-833
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Immune response in chickens
Chickens respond to vaccination or infection by developing humoral, cellular and mucosal immune responses specific to antigens
o Humoral immunity : • B-cells
• Measured by detection of specific antibodies in sera (HI, seroneutralisation, ELISAs)
o Cell-mediated immunity (CMI) :
• T-cells : T helpers cells (TH cells, CD4 cells ) and cytotoxic T-cells (CTL, CD8 cells)
• Measured by proliferation tests and cytokines production after specific (ex vivo) recall on lymphocytes
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Cell-mediated immunity : fonctions
The major goal of cell-mediated immunity (CMI) is to destroy infected cells or extracellular pathogens
CD4
APC
CD8
o Antigen presenting cells (APCs) :
• Macrophages, B-cells and dendritic cells (DC)
• Able to capture pathogens and present them to T-cells, in the context of the major histocompatibility complex (MHC)
Ag Capture
APC APC
Ag presentation
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Cell-mediated immunity : fonctions
The major goal of cell-mediated immunity (CMI) is to destroy infected cells or extracellular pathogens
Ag Capture
APC APC
CD4
APC
CD8
o Antigen presenting cells (APCs) :
• Macrophages, B-cells and dendritic cells (DC)
• Able to capture pathogens and present them to T-cells, in the context of the major histocompatibility complex (MHC)
o CD4 TH – T helper cells : recognize foreign antigen expressed by the APCs in the context of MHC-II and in response, release cytokines (TH1-TH2)
Ag presentation
CD4
MHC-II on APCs
Ag
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Cell-mediated immunity : fonctions
The major goal of cell-mediated immunity (CMI) is to destroy infected cells or extracellular pathogens
Ag Capture
APC APC
CD4
APC
CD8
o Antigen presenting cells (APCs) :
• Macrophages, B-cells and dendritic cells (DC)
• Able to capture pathogens and present them to T-cells, in the context of the major histocompatibility complex (MHC)
o CD4 TH – T helper cells : recognize foreign antigen expressed by the APCs in the context of MHC-II and in response, release cytokines (TH1-TH2)
o CD8 CTL – cytotoxic T cells : recognize foreign antigen presented by infected cells in the context of the MHC-I and are able to lyse infected cells that present viral antigens
Ag presentation
CD4
MHC-II on APCs
Ag
CD8
MHC-I on all cells
Ag
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IFN-g, IL-2
Cell-mediated immunity : fonctions
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APC
Il-4, IL-13
IFN-g, IL-2
Ag
APC
CD8
MHC-I
Ag
Infected cells
LYSIS Macrophages
activation
Ag
CD8 CTL– cytotoxic T-cells
Perforins, granzyme
CD4
MHC-II
Ag
IFN-g
Antibody production
Memory TH -cells
Memory CTL cells
Memory B-cells
CD8
MHC-I
Ag
CD4 TH - Helper T-cells
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Cell-mediated immunity : detection
Cell-mediated immunity
Jugular vein
Spleen
Spleen
Peripheral blood
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Cell-mediated immunity : detection
Cytokines production IFN-g
In vivo Specific T-lymphocytes
(CD4 - CD8 T-cells)
Viral proteins
+ Antigen recall activation
Isolation
Lymphocytes
• Spleen
• Blood
Antigens
(pathogens, vaccines)
T-cells
Specific T-lymphocytes (CD4 - CD8 T-cells)
Memory immunity
IFN-g MEASUREMENT
ELISA Enzyme-linked
immunosorbent assay
Ex vivo
Cytokines production Proliferation
IFN-g
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NDV-specific CMI TH1 : ChIFNg production
7/05/2015 26
IFN
-g
0.0
0.5
1.0
1.5
2.0
2.5
3.0
Alsever Heparin Alsever Heparin Alsever Heparin Alsever Heparin Alsever Heparin Alsever Heparin
1 µg/ml 5 µg/ml 10 µg/ml 1 µg/ml 5 µg/ml 10 µg/ml
48h 72h
Anticoagulant, gp-NDV concentration and culture conditions
O.D
.
Negative
Enterotrocpic live ND vaccine
Enterotropic live ND vaccine and challenge
0.0
0.5
1.0
1.5
2.0
2.5
3.0
2 % ChSi 10 % ChSi
PMA/Iono (1 µg/ml)
IFN
-g
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
In the spleen
In the peripheral blood
Rauw F. et al. (2010). Vaccine, 28 : 823-833
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IFN-g, IL-2
Cell-mediated immunity : fonctions
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APC
Il-4, IL-13
Macrophages activation
Ag
CD4
MHC-II
Ag
Antibody production
Memory B-cells
CD4 TH - Helper T-cells
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CMI TH2 : ChIL-4 & ChIL-13
Monoclonal Ab specific to ChIL-4 and to ChIL-13
Polyclonal Ab specific to ChIL-4
and to ChIL-13
rChIL-4 and rChIL-13 produced in E.coli
Bac-ChIL-4 and Bac-ChIL-13 produced in Sf9 insect cells
ELISA Enzyme-linked
immunosorbent assay
ELISPOT Enzyme-linked
immunosorbent spot
pCIneo-ChIL-4 and pCIneo-ChIL-13 produced in
mammalian COS cells
(kindly provided by P.Kaiser)
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Cell-mediated immunity : fonctions
29
Ag
APC
CD8
MHC-I
Ag
Infected cells
LYSIS
CD8 CTL– cytotoxic T-cells
Perforins, granzyme
Memory CTL cells
CD8
MHC-I
Ag
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Immune response in chickens
Chickens respond to vaccination or infection by developing humoral, cellular and mucosal immune responses specific to antigens
o Humoral immunity : • B-cells
• Measured by detection of specific antibodies in sera (HI, seroneutralisation, ELISAs)
o Cell-mediated immunity (CMI) :
• T-cells : T helpers cells (TH cells, CD4 cells ) and cytotoxic T-cells (CTL, CD8 cells)
• Measured by proliferation tests and cytokines production after specific (ex vivo) recall on lymphocytes
o Mucosal immunity :
• Immunity at mucosal surfaces : major route of entry for many infectious agents into the body : local immunity in the head, the digestive, respiratory and reproductive tracts
• Local antibody- and cell-mediated immunity = first line of defense against pathogens
• Poorly investigated in poultry so far
• Difficulties and labor-intensiveness of collecting samples
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Mucosal response induction
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Lumen
Epithelium
Lamina propria
Specific antibody response
Specific cellular
response
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Local immunity in the head
Local immunity in the head
Harderian gland (HG)
Conjunctiva
Tears
Antibody
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NDV-specific lachrymal antibody-mediated immunity
0.0
0.2
0.4
0.6
0.8
1.0
1.2
1.4
1.6
1.8
2.0
2.2
1 week 2 weeks 3 weeks 4 weeks 5 weeks 1 week 2 weeks 3 weeks 4 weeks 5 weeks 1 week 2 weeks 3 weeks 4 weeks 5 weeks
NDV specific IgG NDV specific IgA NDV specific IgM
Time post-vaccination and Ig isotypes
O.D
.
Negative
Tracheotropic live ND vaccine
Enterotropic live ND vaccine
Centrifugation -20°C NDV-specific IgG/M/A ELISA
Rauw F. et al. (2010). Vaccine, 28 : 823-833
Tears
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
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Local immunity in the digestive tract
Local immunity in the digestive tract
Ileum Jejunum
Caecal tonsils (CT)
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Local immunity in the digestive tract
Local immunity in the digestive tract
Duodenum
Antibody CMI
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Local NDV-specific antibody-mediated immunity in the digestive tract
Rauw F. et al. (2010). Vaccine, 28 : 823-833
Ex vivo tissues culture (72h)
Centrifugation -20°C NDV-specific IgG/M/A ELISA
0.0
0.2
0.4
0.6
0.8
1.0
1.2
1.4
1.6
1.8
2.0
1 week 2 weeks 3 weeks 4 weeks 5 weeks 1 week 2 weeks 3 weeks 4 weeks 5 weeks 1 week 2 weeks 3 weeks 4 weeks 5 weeks
NDV specific IgG NDV specific IgA NDV specific IgM
Time post-vaccination and Ig isotypes
O.D
.
Negative
Tracheotropic live ND vaccine
Enterotropic live ND vaccine
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
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Local NDV-specific CMI TH1 (ChIFNg) in the digestive tract
Cuts (5 mm) of duodenal tissues
37 °C
30 min
LPL : Lamina Propria Lymphocytes
0.0
0.5
1.0
1.5
2.0
2.5
3.0
3.5
4 5 6 7 4 5 6 7
1 µg/ml gp-NDV 5 µg/ml gp-NDV
Weeks post-vaccination (challenge at 4 weeks p.v.)
O.D
.
Negative
Enterotropic live ND vaccine
Enterotropic live ND vaccine with challenge
Rauw F. et al. (2010). Veterinary Immunology and Immunopathology, 134 : 249-258
Washes and enzymatic digestion
Viral proteins
+ Antigen recall
activation
IFN-g
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Local immunity in the respiratory tract
Local immunity in the respiratory tract
Trachea
Lung
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Local immunity in the respiratory tract
Local immunity in the respiratory tract
Trachea
Lung
Trachea
Lung
Antibody CMI
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Local NDV-specific antibody-mediated immunity in the respiratory tract (lung)
NDV-specific IgG/M/A ELISA Euthanasia by
i.p injection of Nembutal
0.0
0.2
0.4
0.6
0.8
1.0
1.2
1.4
1.6
1.8
2.0
3 5 3 5 3 5
IgG IgA IgM
Isotypes and weeks post-vaccination
O.D
.
Negative
Tracheotropic live ND vaccine
Enterotropic live ND vaccine
Rauw F. et al. (2010). Vaccine, 28 : 823-833
Centrifugation 1 min 20 000 x g
10x concentration
Amicon Ultra-centrifugal filter unit
with ultra-100 kDa membrane
Day-old SPF chickens
Occulo/nasal inoculation
1 dose attenuated Tracheotropic or
Enterotropic ND vaccine
Washes
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Local AIV-specific CMI TH1 (ChIFNg) in the respiratory tract (lung)
Cuts (5 mm) of lung
37 °C
2 h
Ficoll HistoPaque
1083 gradient
Erythrocytes
Rauw F. et al. (2011). Veterinary Immunology and Immunopathology, 143 : 27-37
10-day-old SPF chickens
106 EID50 LP H5N2 Oculo/nasal inoculation
Washes and enzymatic digestion
Viral proteins
+
Antigen recall
activation
IFN-g
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Local AIV-specific CMI TH1 (ChIFNg) in the respiratory tract (trachea)
37 °C
2 h
Rauw F. et al. (2011). Veterinary Immunology and Immunopathology, 143 : 27-37
Cuts (5 mm) of trachea
10-day-old SPF chickens
106 EID50 LP H5N2 Oculo/nasal inoculation
Washes and enzymatic digestion
Viral proteins
+
Antigen recall
activation
IFN-g
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Local immunity in the reproductive tract
Local immunity in the reproductive tract
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Local NDV-specific immunity in the reproductive tract
2-week-old 28 weeks
of age
Day-old 11-week-old
Attenuated ND vaccine
17-week-old
Inactivated ND vaccine
32 weeks of age
Local antibody-mediated immunity
Local CMI
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Immune response in chickens
Chickens respond to vaccination or infection by developing humoral, cellular and mucosal immune responses specific to antigens
o Humoral immunity :
• Development of virus specific ELISAs to detect IgG, IgM and IgA specific [Rauw et al., 2009, 2010]
o Cell-mediated immunity (CMI) :
• Tools to measure TH CMI :
• TH1 : ChIFNg production after antigen-specific (ex vivo) recall on lymphocytes :
• In the spleen [Lambrecht et al., 2004, Rauw et al., 2009, 2010]
• In the peripheral blood [Rauw et al., 2010]
• TH2 : ChIL-4 and ChIL-13 production after antigen-specific (ex vivo) recall on lymphocytes : in progress
• Tools to measure CTL : in progress
o Mucosal immunity :
• Tools to measure lachrymal, respiratory and digestive immunity [Rauw et al., 2009, 2010a, 2010b, 2011]
• Reproductive tract : in progress NDV – AIV – IBDV
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Transfert of passive immunity in chickens
o Maternal Derived Antibodies (MDA) or Passive Immunity is the naturally transfer of antibodies from immunized breeder hens to the progeny through the egg
o To protect young chicks against pathogens during a period (1-2 weeks and less than 4 weeks) where their immune system is not fully mature.
How MDA are transferred from the hen to the progeny ?
1. MDA transfer from the hen to the egg
• IgG(Y) are secreted by the chicken ovarian into the developing ova or egg yolk
• IgA and IgM are produced locally in the magnum in the oviduct and transfer to the albumen
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Transfert of passive immunity in chickens
How MDA are transferred from the hen to the progeny ?
1. MDA transfer from the hen to the egg
2. MDA transfer from the egg to the progeny
IgG(Y) :
Are transferred from the egg yolk to the offspring via the embryonnic circulation
Only IgGs are able to cross the intestinal barrier through the embryonic circulation
The transfer starts from 7 ED and reaches its maximum rate at 17-18 ED (3 to 4 days before hatching)
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Transfert of passive immunity in chickens
How MDA are transferred from the hen to the progeny ?
1. MDA transfer from the hen to the egg
2. MDA transfer from the egg to the progeny
IgG(Y) :
Are transferred from the egg yolk to the offspring via the embryonnic circulation
Only IgGs are able to cross the intestinal barrier through the embryonic circulation
The transfer starts from 7 ED and reaches its maximum rate at 17-18 ED (3 to 4 days before hatching)
IgA and IgM in albumen are ingested by chick and protect the intestine.
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MDA interference with vaccination
0
10
20
30
40
50
60
70
80
90
100
0 10 20 30 40
% p
rote
ctio
n
Time post vaccination (days)
Efficiency of attenuated ND vaccine administered to day-old chicks
Control (SPF)
Control (NDV MDA)
Vaccinated (SPF)
Vaccinated (NDV MDA)
Bennejean et al., 1978, Avian Pathology, 15-27
SPF chickens
Conventional chickens
Early NDV vaccination
Rauw F. et al. (2009). Vaccine, 27 : 3631-3642
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To study the impact of MDA
o Development of an artificial model mimicking natural passive immunity in SPF chicken by injection of a NDV- and/or AIV-specific polyserum :
• Production of NDV- and AIV-specific polyserum :
7/05/2015 50
Attenuated vaccine (106 EID50) by oculo-nasal route
Day-old 4-week-old
Inactivated vaccine
2-week-old
NDV-specific polyserum
Serum HI titer = 12 log2
3 weeks
Lardinois. et al. (2014). Avian Pathology, 143 : 118-124
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To study the impact of MDA
o Development of an artificial model mimicking natural passive immunity in SPF chicken by injection of a NDV- and/or AIV-specific polyserum :
• Production of NDV- and AIV-specific polyserum :
• Model of artificial passive immunity induction :
7/05/2015 51
Attenuated vaccine (106 EID50) by oculo-nasal route
Day-old 4-week-old
Inactivated vaccine
2-week-old
NDV-specific polyserum
Serum HI titer = 12 log2
3 weeks
AIV-specific polyserum
6-week-old
Inactivated purified virus
with CFA
8-week-old
Inactivated purified virus
with IFA
11-week-old
HPAIV (106 EID50) by oculo-nasal
route
Serum HI titer = 8 log2
10 days
200 µl in egg yolk at 14 ED
200 µl at day-old by im route
Lardinois. et al. (2014). Avian Pathology, 143 : 118-124
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Research on NDV specific immunity
WHY ?
o By improving immunological parameters and further characterizing the chicken immune response elicited by current ND vaccines, a correlation with clinical protection and shedding reduction could be done
o Good parameters of protection could be established
o Challenge experiments could be reduced
To improve the efficacy of ND vaccination programs
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Live vector vaccines
o Definition :
• A live vector vaccine is a vaccine that uses an attenuated virus (vector) to transport pieces of the virus (pathogen) in order to stimulate an immune response against this virus
o Development and production :
• To determine the protective gene to introduce in the virus vector
• To insert the protective gene into the genome of the vector (herpesvirus, poxvirus, adenovirus…)
• To produce the recombinant vector = live vector vaccine
NDV – AIV – IBDV
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ND Live vector vaccine : rHVT-F
NDV F
gene
Homologous HVT
sequence
Pec promoter
SV40 polyA
Homologous HVT sequence
HVT genome
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Live vector vaccines
o Definition :
• A live vector vaccine is a vaccine that uses an attenuated virus (vector) to transport pieces of the virus (pathogen) in order to stimulate an immune response against this virus
o Development and production :
• To determine the protective gene to introduce in the virus vector
• To insert the protective gene into the genome of the vector (herpesvirus, poxvirus, adenovirus…)
• To produce the recombinant vector = live vector vaccine
o Validation :
• Being intracellular, live vector vaccine should be able to prime a humoral response, to induce a strong cellular and mucosal immune responses against the inserted gene and strong B & T-cell memory immunity
NDV – AIV – IBDV
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1 2 IL-4, IL13
Immunity induced by live vector vaccine
7/05/2015 56
B cells
First in vivo replication of live vector vaccine by
targeted cells
Priming: Viral antigen capture and
presentation by APCs - Th cells activation - Cytokine production - CTL and B activation
IL-2, IFN-g
Effective responses: - Ab production - CTL activity
Induction of memory immunity
Vector vaccine
IL-4
IL-13
1
1
2
Viral antigen
2
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1 2 IL-4, IL13
Immunity induced by live vector vaccine
7/05/2015 57
First in vivo replication of live vector vaccine by
targeted cells
Priming : Viral antigen capture and
presentation by APCs - TH cells activation - Cytokine production - CTL activation - B cells activation
B cells
IL-2, IFN-g
Effective responses: - Ab production - CTL activity
Induction of memory immunity
Vector vaccine
IL-4
IL-13
1
1
2
Viral antigen
2
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1 2 IL-4, IL13
Immunity induced by live vector vaccine
7/05/2015 58
First in vivo replication of live vector vaccine by
targeted cells
Priming : Viral antigen capture and
presentation by APCs - TH cells activation - Cytokine production - CTL activation - B cells activation
IL-2, IFN-g
Effective responses : - Ab production - CTL activity
Induction of memory immunity
Vector vaccine
IL-4
IL-13
1
1
2
Viral antigen
2
B cells
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Immunity induced by rHVT-ND vaccine
Day-old SPF chickens
One dose of rHVT-ND Subcutaneous inoculation
Every 2-3 days or every week
6-10 weeks
Serum
Humoral immunity
HI and ELISA-IgG/M/A tests
CMI
Spleen
ChIFNg production
Local antibody-mediated immunity
Duodenum
ELISA-IgG/M/A tests
Lung
HVTIMMUNOVAC-ND-1,2,3,4,5,6,7,8
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NDV-specific IgG ELISA
*
*
* * *
* * *
Day-old SPF chickens
1 dose of rHVT-ND Subcutaneous inoculation
*
* * * *
* *
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NDV-specific IgM ELISA
*
* *
*
* * *
*
*
Day-old SPF chickens
1 dose of rHVT-ND Subcutaneous inoculation
*
* *
*
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NDV-specific IgA ELISA
*
* *
* * * * *
* * *
* *
*
*
Day-old SPF chickens
1 dose of rHVT-ND Subcutaneous inoculation
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ND vaccination schedule on conventional layer chickens
7/05/2015 63
Oculo-nasale inoculation
Day-old conventionnal layer chickens
Live ND vaccine 1 dose
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ND vaccination schedules on conventional layer chickens
7/05/2015 64
+ Oculo-nasale
inoculation
Day-old conventionnal layer chickens
Live ND vaccine 1 dose
rHVT-F vaccine 1 dose
Subcutaneous inoculation
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• protection : • 90% to 100 % (challenge at
5 weeks) • virus shedding
• OP swabs: 60% to 50% • Cl. swabs: 80% to 40%
ND vaccination schedules on conventional layer chickens
rHVT-ND/Live ND vaccines
Live ND vaccine
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ND vaccination schedules on conventional layer chickens
7/05/2015 66
Oculo-nasale inoculation
Day-old conventionnal layer chickens
Live ND vaccine 1 dose
rHVT-F vaccine 1 dose
Inactivated ND vaccine 1 dose
Subcutaneous inoculation
Subcutaneous inoculation
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ND vaccination schedules on conventional layer chickens
7/05/2015 67
+ Oculo-nasale
inoculation
Day-old conventionnal layer chickens
Live ND vaccine 1 dose
rHVT-F vaccine 1 dose
Inactivated ND vaccine 1 dose
Subcutaneous inoculation
Subcutaneous inoculation
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ND vaccination schedules on conventional layer chickens
7/05/2015 68
+ Oculo-nasale
inoculation
Day-old conventionnal layer chickens
Live ND vaccine 1 dose
rHVT-F vaccine 1 dose
Inactivated ND vaccine 1 dose
Subcutaneous inoculation
Subcutaneous inoculation
+
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• protection : • 90% to 100 % (challenge at
5 weeks) • virus shedding
• OP swabs: 60% to 50% • Cl. swabs: 80% to 40%
ND vaccination schedules on conventional layer chickens
• protection : • 90% to 100 % (challenge at
10 weeks) • virus shedding :
• OP swabs: 90% to 40% • Cl. swabs: 20% to 0%
rHVT-ND/Live ND vaccines
Live ND vaccine
rHVT-ND/Live ND vaccines Inactivated ND/Live ND
vaccines
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Thanks !
2003 2007 2012
CELOVAC project MUCOVAC project HVTIMMUNOVAC project
2015
12 years
CEVA-CERVA collaboration
Vilmos Palya
Yannick Gardin
Scientific promoters
Technical staff Eva Ngabirano
Sophie Lemaire
Martine Gonze
Fiona Ingrao (PhD)
PUBLICATIONS : • Rauw F. et al. (2007). Journal of Interferon and Cytokines, 27 : 11-118. • Rauw F. et al. (2009). Vaccine, 27 : 3631-3642. • De Vriese et al. (2010). Avian diseases, 54, 246-252. • Rauw F. et al. (2010). Veterinary Immunology and Immunopathology, 134 : 249-258. • Rauw F. et al. (2010). Vaccine, 28 : 823-833. • Rauw F. et al. (2011). Vaccine, 29 ; 2590-2600. • Rauw F. et al. (2011). Veterinary Immunology and Immunopathology, 143 : 27-37 • Rauw F. et al. (2012). Avian Diseases, 56: 913-922 • Rauw F. et al, (2014). Avian Pathology, 43:26-36 • Rauw F. et al. (in press). Avian Pathology
Bénédicte Lambrecht
Thierry van den Berg
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Publications
o Rauw F., Gardin Y., Palya V., Gonze M., Lemaire S., van Borm S., van den Berg T. & Lambrecht B. (2009). « Humoral, cell-mediated and mucosal immunity induced by oculo-nasal vaccination of one-day-old SPF and conventional layer chicks with live Newcastle Disease vaccines ». Vaccine, 27 : 3631-3642.
o Rauw F., Gardin Y., Palya V., Anbari S., Gonze M., Lemaire S., van den Berg T. & Lambrecht B. (2010). « The positive adjuvant effect of chitosan on antigen-specific cell-mediated immunity after chickens vaccination with live Newcastle disease vaccine ». Veterinary Immunology and Immunopathology, 134, 249-258.
o Rauw F., Gardin Y., Palya V., Anbari S., Lemaire S., Boschmans M., van den Berg T. & Lambrecht B. (2010). « Improved vaccination against Newcastle disease by an in ovo recombinant HVT-ND combined with an adjuvanted live vaccine at day-old ». Vaccine, 28 : 823-833.
o Rauw F., Anbari S., van den Berg T. & Lambrecht B. (2011). « Measurement of systemic and local respiratory cell-mediated immunity after influenza infection in chickens ». Veterinary Immunology and Immunopathology, 143, 27-37.
o Rauw F., Palya V., Van Borm S., Welby S., Tatar-Kis T., Gardin Y., Moore Dorsey K., Aly M.M., Hassan M.K., Soliman, M.A., Lambrecht B. & van den Berg T. (2011). « Further evidence of antigenic drift and protective efficacy afforded by a recombinant HVT-H5 vaccine against challenge with two antigenically divergent Egyptian clade 2.2.1. HPAI H5N1 strains ». Vaccine, 29, 2590-2600.
o Rauw F., Palya V., Gardin Y., Tatar-Kis T., Moore Dorsey K., Lambrecht B. & van den Berg T. (2012). « Efficacy of rHVT-AI vector vaccine in broilers with passive immunity against challenge with two antigenically divergent egyptian clade 2.2.1 HPAI H5N1 strains ». Avian Diseases, 56, 913-922.
o Rauw F., Gardin Y., Palya V., van den Berg T. & Lambrecht B. (2014). « Combination of attenuated vaccine at day-old with rHVT is more protective against NDV challenge than combination with inactivated vaccine ». Avian Pathology, 43 (1), 26-36.
o Rauw F., Van Borm S., Welby S., Ngabirano E., Gardin Y., Palya V. & Lambrecht B. (in press). « Quantification of rHVT-F genome load in feather follicles by specific real-time qPCR as indicator of NDV-specific humoral immunity induced by day-old vaccination in SPF chickens ». Avian Pathology,.