6Bağışıklık hücrelerinin gelişimi Civcivlerde inkübasyon periyodu: 21 daysBursa’da B hücrelerinin gelişimi:Follikule formasyonu (cortex, medulla)B hücrelerinde gen değişimiB hücrelerinin periferal bölgelere göçüThymus’ta T hücrelerinin gelişimiEmbryonasyonun 9.gününde T hücreleri gelişirT hücreleri çıkımdan sonra perifere göç ederTavuk embriyoları spesifik immun cevabı başlatma yeteneğine sahiptirlerIn ovo aşılama
7bağışıklık Natural or innate immunity Adaptive or acquired immunity CellsPhagocytes, dendritic cells, NK cellsT- and B-lymphocytesHumoral factorsComplement systemCytokinesAntibodiesInterleukinesChemical property of identified structureCarbohydrate, lipidProteinMemoryNoYesProtectionNon-transferableTransferableRapidity of the immune responsePromptFew days or week
9Adaptive or acquired immunity Active immunityspecific immune response against antigensPrimary response (first contact with antigen)IgM IgYImmune memory (B-ly)Secondary response (second contact with antigen)Rapid and longer immune responseIgYPassive immunityMaternal antibody (transmitted from hen to chick via egg yolk)Transfusion of antibodies by sera
10Passive immunity Maternally derived antibodies in offsprings The level of the maternal antibodies can be increased by purposive immunisation of breeder flockProtect the chicks against diseases at early stage of lifeAntibodies decay 2-6 weeks after hatchingThe maternally derived antibodies neutralise the live vaccine viruses → inhibition of active immune response
11Maternal immune transport in the chicken FcRY transports IgY from yolk sac into embrionic blood (West et al., Immunity, 2004)pH dependent IgY binding to the yolk sac membranethe first IgY binding Fc receptor in birds
12Humoral immune response I. Three Ig classes: IgM, IgY, IgAIgM: pentamer, expressed on the surface of B cellsIgY: functionally homologous to mammalian IgGIgA: dimeric or tetrameric forms in mucosal secretionmonomeric molecule in serumcritical role in local immunitySpecific immune responseAntigen → IgMclass switchingmediated by T cells and cytokinesIgY or IgAStrong immune memoryMethods for determination of antibody response: ELISA, SN, IF, agglutination, hemagglutination
15Cell mediated immunity T lympocytes are the principal cells in the cell mediated immunityTwo subsetsCD4+ cells: helper function – central role in humoral and cellular immunityCD8+ cells: cytotoxic functionImportant role in antiviral and antitumor immunity
16Activity of different factors of immune system after viral infection
17Chronic Bursitis after vvIBDV infection Role of the bursa in immune responseGene conversion in B cells → humoral immunityIntact Bursa FabriciusChronic Bursitis after vvIBDV infection
21Enfeksiyon ilk 3 hafta içerisinde immünosüpresyon nedenidir 3-6 haftalar içerisinde hastalığın klınik formu oluşur:Klınik bulgular enfeksiyondan 2-3 gün sonraMortalite enfeksiyondan 5-8 gün sonra
22IBDV infects cells of the cloacal bursa - primarily immature B-cells. CEVA SANTE ANIMALEIBDV infects cells of the cloacal bursa - primarily immature B-cells.Immature B-cells mature into antibody producing cells.If B-cells are destroyed early in life the bird may become immunosuppressed for life.
29Characterisation of the vvIBDV strains PathotypingUp to 100% mortality in SPF chickens50-60% mortality in layers, 25-30% in broilersBreak through higher levels of MDA not possible anymore to protect passively broilersAntigenicityPanel of neutralizing Mabs no (major) antigenic driftHigh cross-neutralization indices measured in SPF embryonnated eggsClassical serotype 1 vaccines still satisfactory on SPF birdsMolecular characterisationSequencing of the variable domain of the VP2 geneAmino acid residues substitutions at positions 222Ala, 256Iso, 294Iso and 299Ser= Genetic fingerprints of vvIBDVRT-PCR of vVP2 followed by sequencing and/or RE Analysis
30Mortality ratesChallenge of 5-week-old Lohmann SPF chickens (n=25-30) with 105 EID50 of IBDV strains of varying virulence in isolated conditions by the oculonasal route.vvIBDV induce higher mortality rates than classical virulent IBDVsConfusion in nomenclatureNeed for standardized challenge model
32Dose effect in IBDV challenge Comparison of a classical virulent IBDV (Cu1wt) and a vvIBDV (HK46):Inoculation of increasing doses 103 107 EID50Classical IBDV: maximum 60% mortalityvvIBDV: up to 100% mortalityExistence of a dose effect in IBDV challenge
33Multiplication ratesTitration of virus in BF of birds dying (or sacrificed) 3-4 days after challengeHigher multiplication rates for more virulent IBDVs1 attenuated IBDV particle 10 particles1 classical virulent IBDV particle 102 to 103 particles1 vvIBDV particle 104 to 105 particles
34PATOGENEZ VIRUS ETKİMESİ GÜÇLÜ VIRUS ZAYIF VIRUS YAŞ (GÜN) LEZYON + MORTALITEGÜÇLÜ VIRUSLEZYONZAYIF VIRUSENFEKSİYON YOK5101520253035404550YAŞ (GÜN)
35WHAT FORM OF GUMBORO DISEASE AT FIELD LEVEL ? WHICH CONTROL STRATEGY ? CEVA SANTE ANIMALEWHAT FORM OF GUMBORO DISEASE AT FIELD LEVEL ?WHICH CONTROL STRATEGY ?
37OIE I. B. D. V. Classification N OIE I.B.D.V. Classification N. Eterradossi - AFSSA Lab - Ploufragan FranceCAPSIDE ANTIGENSCharacterisation by Monoclonal Antibodies (Mab)8 Mab put in contact with the virus = bind or not.R.N.A. :Characterisation by sequencing: Sequence of Nucleotids and deduced a.a.
38Specific Antigen-Capture ELISA OIE I.B.D.V. Classification N. Eterradossi - AFSSA Lab - Ploufragan FranceSpecific Antigen-Capture ELISARevelationMabspecificFixation or notCapture of the virusPolyclonal anti -IBDV
39Classical IBD virus : All Mabs bindings positive OIE I.B.D.V. Classification N. Eterradossi - AFSSA Lab - Ploufragan FranceClassical IBD virus : All Mabs bindings positiveAg1Ag3Ag4Ag5Ag6Ag9Ag7Ag8VP2 Protein
40vv IBD virus : Bindings to Mab 3 and 4 are negative OIE I.B.D.V. Classification N. Eterradossi - AFSSA Lab - Ploufragan Francevv IBD virus : Bindings to Mab 3 and 4 are negative--Ag1Ag5Ag6Ag9Ag7Ag8VP2 Protein
41CEVA’S APPROACH OF GUMBORO CONTROL CEVA SANTE ANIMALECEVA’S APPROACH OF GUMBORO CONTROL
43CEVAC IBD L Antigenic typing CEVA’S APPROACH OF GUMBORO CONTROLCEVAC IBD L Antigenic typingOrigin Mab 1 Mab 3 Mab4 Mab5 Mab6 Mab7 Mab8 Mab9F52/70 (Classic) 66% 91% 78% 75% 110% 114% 96% 77%(vvIBD) 61% 1% 5% 68% 118% 113% 80% 53%Cevac IBD L 64% 89% 74% 23% 94% 93% 91% 91%CEVAC IBD L derived from classical IBDV strainand NOT from vvIBDVAFSSA 2001
44Molecular typing of IBDV strains PCR and sequencingSegment BVP 1VP2VP4VP3VP5Segment A709Hypervariable region11461. PCR reactionP1P258712292. PCR reactionP2.3P5.37211176Nucleotide numbering according to IBDV-STC strain (Kibenge et al, J. Gen. Virol (71) ).
45Differentiation of IBDV strains CEVA’S APPROACH OF GUMBORO CONTROLDifferentiation of IBDV strains(VP2 gene 414bp BseD I RFLP)CEVAOKYM52/70STCVarELIBDVD6950 bp DNA ladder2512Gp82FSP3IBD B.WSVIBD B.WSVD142FSD69AEG145050040030020010050400Jakabszállás350300250200150250100150502512vvvaccineclassicalvarvery virulentsubc
46Phylogenetic study of IBDV strains CEVA’S APPROACH OF GUMBORO CONTROLMOH94/94H"MOH97/97H"Csengele/97H"PECS/97H"Phylogenetic study of IBDV strainsOKYMT/95JDV86/86NvvOKYM/91JKABA/97H"UK661/86GB(based on the hypervariable region ofthe VP2 gene, 414bp)CS89/89GBFS/97H"AEG1/97H"Toszeg/97H"Diosjeno/97H"849VB/87BvaccIBDL(2512)/80US"STC/67USclassicalP1II/75H"P1/75H"52/70GBPBG98/76GBGP82/75H"D78"vaccTAD"Cu1/76DLIBDV/75H"P2/73DVarE/85USUS varU28/88USVarA/85US3212/88USserotype IGLS/87USP3/77H"P6/78H"subcP7/78H"P9/81H"P10/81H"serotype IIP11/81H"00273/73AusOH/82US1918161412108642
47CEVA’S APPROACH OF GUMBORO CONTROL CEVA SANTE ANIMALECEVA’S APPROACH OF GUMBORO CONTROL
48Aşı türleri Canlı attenue Inaktive Mild Intermedier Intermedier plus („hot”)İlk ve devamı aşılamalar içinInaktiveSadece canlı aşılama devamınaYüksek titre ve uzun süreli korunma
49Conventional IBD Vaccines Mild IBD Vaccine - Easily eliminated by low levels of maternal antibody.Intermediate Vaccine - relatively safe to use in ovo or at hatch. Eliminated by low to moderate levels of maternal immunity.Intermediate Plus Vaccine - not safe to give in ovo unless modified in some manner. Eliminated by high level maternal immunity.
51Aşı ve enfeksiyon sonu lezyon gelişimi (4 hft.SBF) IDBV strainsDays post infection1234581421mild-intermedier●● Rintermedier plusclassical●*very virulentvaccine strain [* clinical signs, death** 4-week old SPF birdsR: regeneration
52Immunity induced by strains of different virulence ** 4-week old SPF birds
53Immunity induced by strains of different virulence - CONCLUSIONS - Strains of higher virulence cause earlier bursa damage and earlier serological reponseOnly strains that casue bursal damage induce immune responseThe tissue-culture adopted „mild” vaccine strain does not induced immune response in 4-week old SPF chicks (probably due to lack of receptors)
54The influence of maternal antibody level on the replication of classical IBDV strain positive bursaInfection at 11 days agenegative bursa
55Infection at 11 days of age The influence of maternal antibody level on the replication of vv IBDV strainpositive bursaInfection at 11 days of agenegative bursa
56Vaccination at 15 days of age The influence of maternal antibody level on the replication of intermedier plus IBDVpositive bursaVaccination at 15 days of agenegative bursa
57Vaccination at 18 days of age The influence of maternal antibody level on the replication of intermedier IBDV strainpositive bursaVaccination at 18 days of agenegative bursa
58IBD: The concept of constant protection CEVA SANTE ANIMALEIBD: The concept of constant protectionAGE AT VACCINATION VARIES ACCORDING TO VACCINE TYPE and EPIDEMIOLOGICAL SITUATIONAntibody titrePassive immunityVaccine takelevelIntermediate + V.Protection levelv.v. IBDVVaccine takelevelIntermediate V.Protection levelSubclinical IBDVD1AgeAge at vaccination
59Vaccination programme versus epidemiology CEVA SANTE ANIMALEVaccination programme versus epidemiologyFarmNo IBDscl. IBDvv IBDNeighborhoodMildIntermediateIntermediate plus
60Farm No IBD scl. IBD vv IBD Cevac Bursa L Cevac Gumbo L Cevac IBD L CEVA SANTE ANIMALEFarmNo IBDscl. IBDvv IBDNeighborhoodCevac Bursa LCevac Gumbo LCevac IBD L
61CEVA’S APPROACH OF GUMBORO CONTROL CEVA SANTE ANIMALECEVA’S APPROACH OF GUMBORO CONTROLFarmNo IBDscl. IBDvv IBDNeighborhoodCevac Transmune IBD
62CEVA’S APPROACH OF GUMBORO CONTROL Is it immunosuppressive ? CEVAC IBD L field use:Is it immunosuppressive ?
63CEVA’S APPROACH OF GUMBORO CONTROL Economical Interest of Using an Intermediate Plus Vaccine in a Farm Contaminated with vvIBDVIntermediate plusvaccinationNet income (FF/m2)105- 5- 10- 15- 20O Benefitlevel(After Y. Gardin, 1992)round 1round 2round 3round 4
64IMMUNOSUPPRESSION IN CHICKENS CEVAC IBD L field useTURKEY
65IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENS
66IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENS
67IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENSPI = livability x average slaughter BW / FCR> 240: very good: good: medium: poor
68IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENSIntermediate Plus IBD live vaccines use in Europe: CEESA figures.CEESA means European Centre for Animal Health (Centre Europeen de la Sante Animale, in French).
69IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENSIntermediate Plus IBD live vaccines use in Europe: CEESA figures.It includes 12 countries:Austria,Belgium,Denmark,France,Germany,Italy,the Netherlands,Portugal,the Republic of Ireland,Spain,Switzerland,the United Kingdom.
70IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENS
71IMMUNOSUPPRESSION IN CHICKENS CEVA SANTE ANIMALEIMMUNOSUPPRESSION IN CHICKENS
72IMMUNOSUPPRESSION IN CHICKENS CEVAC IBD L field useBRAZIL
73IMMUNOSUPPRESSION IN CHICKENS 90,000 broiler chickensVaccination with CEVAC IBD L
78IMMUNOSUPPRESSION IN CHICKENS COMMENTSIncreased performances thanks to the optimum control of vvIBD in Brazil by Cevac IBD L.Other countries constantly report the same benefit.
79IMMUNOSUPPRESSION IN CHICKENS COMMENTSRate of usage of Cevac IBD L in the top 4 Brazilian broiler integrations in 20041SADIA100%2PERDIGAO3FRANGOSUL0%*4SEARA* Use intermediate vaccine only.
80IMMUNOSUPPRESSION IN CHICKENS 2005: Sadia (2nd largest Brazilian integration): Cevac IBD L is used in 100% of its capacity, i.e. 45 million broilers per month.From its introduction (2000) in Brazil, Cevac IBD L is now used at a rate of 140 million doses per month:Four Brazilian broiler chickens produced out of 10are vaccinated usingCevac IBD L.
81IMMUNOSUPPRESSION IN CHICKENS CEVAC IBD L Immunodepression study
82IMMUNOSUPPRESSION IN CHICKENS Immunosuppression study according to the European Pharmacopoeia requirementsCeva-Phylaxia study11-day old SPF birds.Vaccination by eye drop (Day 0) with Cevac IBD LControl50 birds per group.Four days later (D4, ie at maximum IBD vaccine replication in the bursa), ND vaccination by eye drop with Hitchner B1 strain (Cevac Uni L).ND challenge (Herts 33 strain, 106 EID50 per bird, by im route): day 18 (14 days after ND vaccination).ND challenge assessment: day 32.
84CONCLUSIONS Cevac IBD L is safe upon the immunological functions. Cevac IBD L is the right tool to prevent vvIBD.Cevac IBD L is the right way to come back to performances and profits.
85CEVA’S APPROACH OF GUMBORO CONTROL CEVA SANTE ANIMALECEVA’S APPROACH OF GUMBORO CONTROLAnd tomorrow…CEVAC TRANSMUNE IBD
86Main issues around the protection against IBD: CEVA SANTE ANIMALEIBD: INNOVATIVE VACCINATION STRATEGYMain issues around the protection against IBD:Differences in level of MDAVariability in quality of vaccine applicationDrinking water administrationTo adapt to the epidemiological context
88Cevac Transmune IBD CEVA’s answer to address these issues: CEVA SANTE ANIMALEIBD: INNOVATIVE VACCINATION STRATEGYCEVA’s answer to address these issues:Cevac Transmune IBDWinterfield 2512 strain of IBDVSpecific AntibodiesLive, immune-complex vaccineInjectedIn-ovo, orSub-cutaneous route at day-old
89IBD: INNOVATIVE VACCINATION STRATEGY BackgroundAbility of the chick / the embryo to develop an immune response.1985, Sharma: First in ovo Gumboro vaccination but MDA or safety problems:Mild vaccines: safe for Embryos but susceptible to MDAIntermediate and intermediate + vaccines: Efficacious in front of MDA but unsafe for SPF Embryos.
91Virus + Antiserum IBD: INNOVATIVE VACCINATION STRATEGY The IMMUNE-COMPLEX concept ( )It will CONTROL and PROTECT the vaccine virus:Virus + Antiserum
92IBD: INNOVATIVE VACCINATION STRATEGY The IMMUNE-COMPLEX concept ( )MECHANISM ??
93IBD: INNOVATIVE VACCINATION STRATEGY 1 / Antiserum neutralises the vaccine virus embryo and vaccine are protected2 / Progressively, the MDA are metabolised, as well as antiserum Release of vaccine virus is synchronised with decrease of MDA3 / When vaccine virus is stronger than antiserum + MDA action, it starts to replicate and to Vaccinate !!!
94IBD: INNOVATIVE VACCINATION STRATEGY Antibody titreVACCINATIONAge (days)Not protected chickenProtected chicken- 3Maternally Derived AntibodiesProtection on individual basis.Bird by bird !!!Active Immunity
96Competitor vaccine dw 12 days CEVAC TRANSMUNE IBDProtection of the Bursa of Fabricius after vvIBD challenge at 3,4,5 weeks of age (histology)Age at challenge345Cevac Transmune IBD SQYESCompetitor vaccine dw 12 daysNO
98IBD: INNOVATIVE VACCINATION STRATEGY Advantages of Cevac Transmune IBDVaccination as soon as possibleMass vaccination – quick and correctSingle vaccination – no follow-upsNo need for date determinationNo effect of variable MDA levelsWinterfield 2512 safety and efficacyCompatible with Marek vaccines, …
99IBD: INNOVATIVE VACCINATION STRATEGY THANK YOUFOR YOUR ATTENTION !