Programme and abstracts

25 FEBRUARY 2021

9am to 12.30pm GMT

Programme and abstracts

Contents

Programme  2 Sponsors  2 Speakerabstracts  3 Posterabstracts  7 Platformguide  10 co-hostedwith

Programme

Sessionchairs:Professor Fiona Tomley(RoyalVeterinaryCollege,UK) andProfessor Hualan Chen(HarbinVeterinaryResearchInstitute,China)

09.00 – 09.05 Welcome and introduction

Dr Timothy Connelley (IVVN Director, The Roslin Institute, UK)

09.05 – 09.35 New approaches for the development of recombinant glycoconjugate

vaccines for poultry, pigs and ruminants

Professor Brendan Wren (London School of Hygiene & Tropical Medicine, UK)

09.35 – 10.05 Successful control of H7N9 influenza in China

Professor Hualan Chen (Harbin Veterinary Research Institute, China) 10.05 – 10.20 Selectively targeting hemagglutinin antigen to chicken antigen

presenting cell receptor induces faster and stronger immunity against avian influenza

Professor Munir Iqbal (The Pirbright Institute UK) 10.20 – 11.00 Break and poster session in breakout groups

11.00 – 11.20 Aerosol Delivery of oligodeoxynucleotides containing CpG motifs

(CpG-ODN) at hatch against bacterial septicemia in neonatal broiler chickens under field conditions

Professor Susantha Gomis (University of Saskatchewan, Canada)

11.20 – 11.40 Identification of avian influenza viruses subtype H9N2 isolated in 2019

from the backyard and commercial farms in Indonesia

Melina Jonas (Medion, Indonesia)

11.40 – 12.00 IVVN pump-priming grant: Towards edible vaccines for chickens Dr Kate Sutton (Roslin Institute, UK)

12.00 – 12.20 IVVN pump-priming grant: Immunogenicity study of matrix 2

ectodomain proteins displayed on nodavirus-like particles as a universal avian influenza virus vaccine for chickens

Dr Mariatulqabtiah Abdul Razak (Universiti Putra Malaysia, Malaysia) 12.20 – 12.30 Close

Speaker abstracts

New approaches for the development of recombinant glycoconjugate

vaccines for poultry, pigs and ruminants

Professor Brendan Wren

London School of Hygiene & Tropical Medicine, UK

Thislecturewilldescribetheoriginoftheproductionofrecombinantglycoconjugate vaccinesdevelopedthroughthefunctionalcharacterizationofanN-linkedOTasegeneral glycosylationsystemfromtheentericpathogenCampylobacter jejuni.Itwilldescribe howthesebasicstudieswereusedtore-constitutetheglycosylationsysteminE. coli, whichallowedforthefirsttimetheproductionofrecombinantglycoproteinsthrough aprocesstermedProteinGlycanCouplingTechnology(PGCT).Themajorapplication ofPGCTisintheconstructionofaffordablerecombinantglycoconjugatevaccines. Todate,thetechnologyhasbeenusedtoproducenovelrecombinantCampylobacter,

Shigella, Streptococcus pneumoniae, Francisella, Burkholderia pseudomallei, E. coliandMRSA

glycoconjugatevaccines.Thesehavebeenshowntobeprotectiveandsomeareinclinical trials.

IwilldescribetheapplicationsandlimitationsofPCGT fortheconstructionoflow-costrecombinantglycoconjugatevaccinesinthreeareas;(i)designingnovelvaccines againstpathogenswherenocurrentvaccineexists(eg Burkholderia pseudomallei), (ii) improvingexistingglycoconjugatevaccines(egpneumococcalvaccine),and(iii)affordable glycoconjugatevaccinesfortheveterinarymarket(egpoultry,pigsandruminants). FromaOneHealthperspective,low-costglycoconjugatevaccinesforanimalsnot onlyaddstoeconomicprosperitybutcanreduceanimalsufferingandthezoonotic transmissionofdiseasetohumans.Furthermore,itwillreduceusageofantibioticsinthe livestockindustryandthesubsequentspreadofantimicrobialresistance.

Successful control of H7N9 influenza in China

Professor Hualan Chen Harbin Veterinary Research Institute, China

TheH7N9virusesthatemergedinChinain2013werenonpathogenicinchickensbut mutatedtoahighlypathogenicforminearly2017andcausedseverediseaseoutbreaks inchickens.TheH7N9influenzaviruseshavecausedfivewavesofhumaninfection,with almosthalfofthetotalnumberofhumancases(766of1,567)beingreportedinthefifth wave,raisingconcernsthatevenmorehumaninfectionscouldoccurinthesixthwave. InSeptember2017,anH5/H7bivalentinactivatedvaccineforchickenswasintroduced, andtheH7N9virusprevalenceinpoultryaftervaccinationwassuccessfullycontrolled basedonthelarge-scalevirussurveillanceresults.Moreimportantly,sincetheH7N9 vaccinewasappliedinpoultry,onlyfourH7N9humancaseswerereportedinovertwo years(threeH7N9humancaseswerereportedbetweenOctober1,2017andSeptember 30,2018,andonehumancasewasreportedbetweenOctober1,2018andSeptember 30,2019),indicatingthatvaccinationofpoultrysuccessfullyeliminatedhumaninfection withH7N9virus.Thesefactsemphasizethatactivecontrolofanimaldiseaseisextremely importantforzoonosiscontrolandhumanhealthprotection.

Selectively targeting hemagglutinin antigen to chicken antigen presenting

cell receptor induces faster and stronger immunity against avian influenza

Angita Shrestha1,2, Jean-Remy Sadeyen1, Deimante Lukosaityte1, Pengxiang Chang1, Adrian Smith2, Marielle Van Hulten3 and Munir Iqbal1

Affiliations: 1 The Pirbright Institute, UK; 2 Department of Zoology, University of Oxford, UK; 3 Global Poultry R&D Biologicals Boxmeer,

Intervet International BV, MSD Animal Health, Netherlands

WedevelopedaTargetAntigenDeliveryVaccine(TADV)thatselectivelytargetsvaccine antigenstochickenantigenpresentingcell(APCs)andpotentiatesimmunogenicityofthe vaccine.TodemonstratetheeffectivenessoftheTADV,wemodifiedthehaemagglutinin (HA)antigenofH9N2avianinfluenzavirus(AIV)byfusingitwithasinglechainfragment variable(scFv)antibodyspecificforchickenCD83receptorsandproducedarecombinant solubletrimericH9HA-CD83scFvproteinininsectcells.Vaccinationofchickenswith adjuvantH9HA-CD83scFvproteininducedfasterandhigherHAantigen-specific antibodytitrescomparedtountargetedH9HAproteinvaccineorconventionalinactivated virusvaccine.Furthermore,chickensvaccinatedwithtargetedH9HA-CD83scFvvaccine showedreducedH9N2challengevirussheddingcomparedtountargetedH9HA.These resultssuggestthattargetingantigenstoCD83receptorscouldimprovetheefficacyof poultryvaccines.

Aerosol Delivery of oligodeoxynucleotides containing CpG motifs

(CpG-ODN) at hatch against bacterial septicemia in neonatal broiler chickens

under field conditions

Susantha Gomis, Kalhari Bandara Goonewardene and Shelly Popowich Department of Veterinary Pathology, Western College of Veterinary Medicine, University of Saskatchewan, Canada

Oligodeoxynucleotides(ODN)containingCpGmotifs(CpG-ODN)areeffective immunostimulatoryagentsagainstviralandbacterialdiseases.Wehaverecently demonstratedthatintrapulmonary(IPL)deliveryofCpG-ODNasmicrodropletscan protectneonatalbroilerchickensagainstlethalEscherichiacolisepticemiaunder laboratoryconditions.Theobjectivesofthisstudyweretodevelopacommercial-scale prototypenebulizertodeliverCpG-ODNasmicro-dropletsinneonatalbroilerchickens athatchandtostudytheefficacyofIPLdeliveryunderfieldconditionsinCanada. Neonatalbroilerchicks(n=8,000)receivedCpG-ODNbytheIPLrouteinthemulti-levelcommercial-scalepoultrynebulizer(CSPN)for30minandcontrolbroilerchicks receiveddistilledwater(DW)for30min.Broilerchicksweresampledfromdifferent locationsoftheCSPNfollowingnebulizationandchallengedwithalethaldoseofE. colitostudyefficacyofCpG-ODNbytheIPLdelivery.Broilerchickswereprotectedat asignificantlevelagainstE.colichallengefollowingIPLdeliveryofCpG-ODNinCSPN (P<0.05).ChickstreatedwithCpG-ODNbytheIPLroutehadsignificantlylowerclinical signsandbacterialload(P<0.05).WefoundthatIPLdeliveryofCpG-ODNcaninduce protectiveimmunityasearlyas6hpost-administrationandremaineffectiveuntil day5post-treatment.ResultsofthisstudyconfirmedthatIPLdeliveryofCpG-ODN againstsepticemiainneonatalbroilerswasindustriallyfeasibleandeffectiveunderfield conditions.ThisstudyhasdemonstratedthatCpG-ODNdeliverybyIPLroutecanbea promisingalternativetoantibioticsforinducingprotectiveimmunityinchicksduringthe neonatallife.

Identification of avian influenza viruses subtype H9N2 isolated in 2019

from the backyard and commercial farms in Indonesia

Melina Jonas, Aprilla Sahesty, Theresia Murwijati, Christina Lilis Lestariningsih, Ine Irine, Wahyu Prihartini and Elly Setiawaty

PT Medion Farma Jaya, Indonesia

Lowpathogenicavianinfluenzavirus(LPAIV)subtypeH9N2wasfirstdetectedin Indonesiain2016.Theviruswasspreadoutinvariousprovincesandco-circulatingwith H5N1.Throughout2019,werecordedtwo-hundred-and-seventycaseswithclinical signsfrombothvaccinatedandunvaccinatedflockscollectedfromthebackyardand commercialfarms.MolecularanalysiswithreversetranscriptasePCR(RT-PCR)and sequencingwasconductedtoconfirmthesecases.PositiveAIV-H9N2isolateswere inoculatedtoSPFchickenstoobservethepathologicalsigns.RT-PCRshowedfifty-four AIVH9N2positivesamples,mostlyisolatedfromlayer(46.30%)andbreeder(37.04%) farms.ThephylogenyoftheHAsegmentofAIV-H9N2virusesidentifiedinthisstudy appearedtobeinthesamedistinctclusteras2016-2018isolatesfromtheprevious study.AllIndonesianisolatesshowedhighsimilaritywithsomeVietnamandChina's virusesisolatedfromchickensandduckin2012-2014butformedadifferentcluster withChina'svirusesisolatedbefore2007.Therewerechangesinpathologicalsignsof AIV-H9N2isolatedbeforeandafter2018whentestedinSPFchickens.Haemorrhage occurredlessfrequentinthetracheaandalmostnoneobservedinheartsampleswhile thesesignsweremanifestedprominentlybyvirusesisolatedbefore2018.Swollen kidneyandintestinalhaemorrhagewerenewpathologicalchangescausedbytherecent 2018-2019viruses.WeconcludethattheepidemicofAIV-H9N2continuestoaffect broilers,layers,breedersandnativechickens.AlthoughthenewlyisolatedAIV-H9N2 wereplacedinthesamephylogenyclusterasisolatesfrom2016-2017,theycausedmore severepathologicalsignsonkidneyandintestineandlessobservabletrachealandheart haemorrhage.

Towards edible vaccines for chickens

Dr Kate Sutton1, Professor Lonneke Vervelde1, Dr Roger New2 and Professor Damer Blake3

Affiliations: 1 Roslin Institute, University of Edinburgh, UK; 2 Proxima Concepts Ltd, UK; 3 Royal Veterinary College, UK

Inthisprojectwetestedanoveloilformulationasavaccinevehiclethattargetsthe mucosalimmunesystem.Wehaveshowntheabilityoftheoilformulationtobindto intestinalepithelialcellsandinducebothasystemicandmucosalimmuneresponseto modelantigensinchickens.Thisvaccinehasthepotentialtobemixedintofeed,making itsuitableforroutineadministrationtobroilersandbackyardpoultry.Thisoilformulation isofparticularrelevanceinlowandmiddleincomecountrieswherepoultryproduction israpidlyexpandingbuttheyhavelittleornoaccesstocold-chainresourcesrequiredfor currentchickenvaccines.

Immunogenicity study of matrix 2 ectodomain proteins displayed on

nodavirus-like particles as a universal avian influenza virus vaccine for

chickens

Dr Mariatulqabtiah Abdul Razak1, Professor Wen Siang Tan1, Dr Kok Lian Ho1, Professor Abdul Rahman Omar1 and Professor Munir Iqbal2

Affiliations: 1 Universiti Putra Malaysia, Malaysia; 2 The Pirbright Institute, UK

Highlypathogenicavianinfluenza(HPAI)H5N1virusishighlycontagiousamongbirds andconstantlycausesseriousoutbreaksinpartsofAsiaandtheMiddleEast.Failureto eradicateitsspreadandoccurrencecanleadtosevereeconomiclossesandpotential healthrisksinhumans.AllthecommerciallyavailablevaccinesagainstH5N1(inactivated H5N2vaccine,reversegeneticsvaccine,recombinantfowlpoxvirusvaccineandDNA vaccine),whichexpresstheantigenicglycoproteinofavianinfluenza,haemagglutinin, areunabletocompletelystopthevirusfromcirculating.Theinactivatedvaccinesagainst avianinfluenzavirus(AIV)arecommonlydevelopedbasedoncirculatinglowpathogenic avianinfluenza(LPAI)viruses,hencemaynotbeefficaciousagainstothertypesof HPAIorLPAIstrains.Thesevaccinesarestrain-specificandrequirereconstitutionto accommodateanynew,mutatedcirculatingviralstrains.DevelopmentofauniversalAIV vaccinetargetingallAIVstrainsisurgentlyindemand.Ourresearchteamhasshowna promisingprotectiveefficacyofnodavirus-likeparticlesdisplayingthreecopiesofM2e fragmentagainsthumaninfluenzaAvirusinmicemodel.Wehypothesisethatexpression ofthematrix2ectodomain(M2e)proteinsfromavianinfluenzavirusesofavianorigin mayconferauniversalprotectionagainstavianinfluenzavirusesinchickens.

Poster abstracts

1

Surveying Salmonella antigens for use in a bacteriophage-based vaccine

enabling homologous and heterologous protection and

colonization-inhibition effect in poultry

Angela Makumi1, Andrea McWhorter2 and Nicholas Svitek1

Affiliations: 1 International Livestock Research Institute, Kenya; 2 University of Adelaide, Australia

Inthelastdecade,significantincreasesintheproductivityofmodernpoultrystocks havebeenachievedforboththemeatandtheeggproductionsectorsoftheglobal poultryindustry.Regardlessoftheincreaseinproductiontrends,theperformanceof thepoultryindustryisstilllow.Thisismainlyduetolowgrowthrates,highmortality rates,lowreproductiveratesandpoorqualityoftheproduct.Bacterialdiseasessuchas Salmonellosisarerecognizedasanimportantriskfactorinpoultryhealthmanagement. Mostpoultryfarmersareawareoftheriskoftheseinfectionsandtheirsubsequent effectonmortalities,productivity,andprofitability.Consequently,farmersspendmore moneyoncontrolandmanagementofbacterialdiseasesthanonanyotherformof poultrydiseases.Thepastfewdecades,however,havewitnessedasteadyincrease inthenumberanddiversityofantibiotic-resistantbacteria,renderingsomebacterial infectionsvirtuallyuntreatable.Thepervasiveuseofantibioticsforboththerapeutic andnon-therapeuticpurposesisassociatedwiththeoccurrenceofantibiotic-resistant bacteria,duetoselectivepressureinfavourofresistantbacteria.Therecommendationto controlorpreventSalmonellainpoultryflocksincludehygienemeasuresandbiosecurity practiceswhicharegenerallysupportedbyadditionalinterventions,suchastheuse ofantibioticsandinactivatedorattenuatedvaccinesforbreeders,whichcanreduce Salmonellaintestinalcolonization.Thevaccinesthatareusedinthepoultryindustry includewholepathogenvaccinesbothliveandinactivatedwhichwhenusedplayaminor roleincontrollingsalmonellosisinpoultry.Thesevaccinesoffershort-livedprotection againstclinicaldiseaseanddonotelicitacell-mediatedimmuneresponse,whichis indispensablefortheclearanceofSalmonella.Theobjectiveofthestudywillentail movingawayfromwholepathogenvaccinesorsubunitvaccinesintoadoptingstrategies suchasincorporatingthetargetantigensaspartofavirus(bacteriophage)nanoparticle structureinorderproduceamultivalentvaccinesthatwillelicitmucosal,systemic immuneresponsesbutalsocontrolseveralserovarsofSalmonellaenterica.

2

Morphological and immunophenotypical evaluations of

maturation-associated chicken dendritic cells by wild type and recombinant fowlpox

virus infections

Sakinah Yusof1, Nor Yasmin Abdul Rahaman1,2, Abdul Rahman Omar1,2 and Abdul Razak Mariatulqabtiah1,3

Affiliations: 1 Institute of Bioscience, Universiti Putra Malaysia, Malaysia; 2 Faculty of Veterinary Medicine, Universiti Putra Malaysia,

Malaysia; 3 Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Malaysia

Fowlpoxisaslow-spreadingcommonviraldiseaseofchickensandturkeys,causedby fowlpoxvirus(FPV).Infectionswillleadtolesionsonthecomb,wattlesandotherskin areas.Althoughmostbirdscansurvivetheinfection,itisconsideredasanimportant viraldiseasetocombatincommercialpoultryfarming,astheinfectionsmayalso resultedinlowerproductionofeggsandreducedbodyweight.Livevaccinesagainst

FPVhavebeenestablishedsince1920s,whiletheirgenomehavebeenmanipulated since1980stobeusedasarecombinantvector.Nonetheless,studiesoninteraction ofchickenbonemarrow-deriveddendriticcells(chBM-DCs)withFPVremainslimited. Therefore,thisstudywasaimedtoevaluatetheresponseofchBM-DCsuponinfections withwildtype(WT)FP9strainofFPV,incomparisontoarecombinantFPVcarrying theH5geneofavianinfluenzavirus(rFPV/H5).First,DCswereisolatedfromchicken femurandculturedfor6daysinthepresenceofrecombinantchickengranulocyte-macrophagecolony-stimulatingfactor(GM-CSF)andrecombinantchickeninterleukine-4 (IL-4).Then,immunofluorescencestainingofFPV-stimulatedBM-DCswasperformed todetectthepresenceofCD86expressionmarkeronthesurface,priortosubsequent immunophenotypingusingAPC-conjugatedanti-chickenCD86andPerCP-conjugated anti-chickenMHCclassII.TheculturedpopulationshowedtypicalmorphologyofchBM-DCswiththeformationofdendrites,whicharethekeymorphologicalcharacteristics thatdefinedasasignofcellmaturation.Theresultalsoshowedthattheinfected chBM-DCsexpressedenhancedlevelsofCD86andMHCclassII.Inconclusion,this studydemonstratesthatchBM-DCsweresusceptibletowardsbothWTFPVandrFPV/ H5infections,andcapableininducingphenotypicmaturationofDCs.Thispreliminary datamayenhancetheunderstandingonFPVandrFPVmechanisms,toimprovefuture developmentofFPV-basedvaccines.

3

Generation of a recombinant chickenized monoclonal antibody against

neuraminidase of H9N2 avian influenza virus

Fei Wang1, Zhimin Wan1,2, Hongxia Shao1,2,3, Kun Qian1,2,3, Jianqiang Ye1,2,3 and Aijian Qin1,2,3

Affiliations: 1 Ministry of Education Key Lab for Avian Preventive Medicine, Yangzhou University, China; 2 Jiangsu Co-Innovation Center for

Prevention and Control of Important Animal Infectious Diseases and Zoonoses, China; 3 Jiangsu Key Lab of Zoonosis, China.

H9N2avianinfluenzaviruses(AIVs)havebeenprevalentinChinasincethefirstreport inGuangdongprovincein1992.Itisverydifficulttocontrolandcausesgreateconomic lossesinpoultryindustry.Basedonourpreviouslyreportedmonoclonalantibody(mAb) 1G8againsttheneuraminidase(NA)ofH9N2withsignificantNAinhibitoryactivity,the geneoffragmentofvariableregionofthemonoclonalantibodywassequenced.Then thegenewasclonedandfusedwiththegeneofconstantregionofchickenIgY.The chickenizedmAb(rCmAb)wasexpressedinCOS-1cellsandabletosecrettosupernatant. ThesignificantNAinhibitoryactivityandneutralizingabilitywereconfirmedinenzyme-linkedlectinassay(ELLA)andmicro-neutralization(MN)assay.Itcouldbeverygood passiveimmunizationforH9N2inthefuture.

4

Evaluating the effectiveness of vaccines in commercial poultry against

Newcastle disease viruses of genotype VI and VII

Muhammad Zubair Shabbir1, Sameera Akhtar1, Yi Tang2, Tahir Yaqub1, Arfan Ahmad1, Ghulam Mustafa1, Muhammad Azhar Alam1, Diwakar Santhakumar3, Venugopal Nair4 and Muhammad Munir3

Affiliations: 1 University of Veterinary and Animal Sciences, Pakistan; 2 The Pennsylvania State University, PA, USA; 3 Department of

Biomedical and Life Sciences, Lancaster University, UK; 4 The Pirbright Institute, UK

Newcastlediseaseviruscauseseconomicallydevastatingdiseaseinavianspecies

aroundtheworld.NewcastlediseaseisenzooticinPakistanandrecurrentoutbreaksare frequentinmultipleavianspeciesevenaftercontinuousandextensiveuseofvaccines. AnumberofAPMV-1andpigeonparamyxovirusserotype1(PPMV-1)strainshavebeen

isolatedandgeneticallycharacterizedinrecentyears.However,theimpactofrecently characterizedwildbird-originAPMVsindomesticpoultry,andthepotencyofroutinely usedvaccinesagainstthesenovelandgeneticallydiversevirusesremainedelusive.Here, weappliednext-generationsequencingforunbiasedcompletegenomecharacterization ofAPMV-1andPPMV-1strainsisolatedfromclinicallydiseasedpeacocks(Pavocristatus) andpigeons(Columbalivia),respectively.Globalphylodynamicsandevolutionaryanalysis demonstratesPigeon/MZS-UVAS-Pak/2014isclusteredintolineage4(orgenotype VI)andPeacock/MZS-UVAS-Pak/2014intolineage5(orgenotypeVII).Thegenomes ofbothisolatesencodedforpolybasicresiduesatthefusionproteincleavagemotif (112RRQKR↓F117)alongwithanumberofimportantsubstitutionsinthesurface glycoproteinscomparedwiththevaccinestrains.Clinico-pathologicalandvaccine-driven immunologicalinvestigationsindomesticatedchickensindicatethattheseisolatescan potentiallytransmitbetweentestedavianspecies,cancausesystemicinfections,and caninduceantibodiesthatareunabletopreventvirusshedding.Anoticeableantigenic variationbetweenvaccinestrainandwild-originatedstrain(R-value=0.26)wasalso observedwhileperformingcross-HIassay.Collectively,thedatafromthesegenomic andbiologicalassessmentshighlightthepotentialofwildbirdsintransmittingAPMVsto domesticatedchickens.Thestudyalsodemonstratesthatthecurrentvaccineregimens areincapableofprovidingcompleteprotectionagainstwildbird-originAPMVsand PPMVs.

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