Niets is zo veranderlijk als een RNA virus. · Animals first exposed or primed with NCP BVDV...
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Blijft BVDV altijd BVDV? Niets is zo veranderlijk als een RNA virus. Frank van der Meer November 30, 2016
Transcript of Niets is zo veranderlijk als een RNA virus. · Animals first exposed or primed with NCP BVDV...
BlijftBVDValtijdBVDV?NietsiszoveranderlijkalseenRNAvirus.
FrankvanderMeerNovember30,2016
ThefirstreportofBVDVintheworld
CanadianJournalofComparativeMedicine1946
Undertheradar
QuotefromDr.JuliaRidpathBovineVeterinarian/May-June2006
Degrote surprise
MutationRates
5Sanjuan,R.,etal,2010J.Virol.Ch.1
Duffyetal.Nat.Rev.Genet.2008 9,267-276.
Quasispecies:Cloudsofgenotypesinapopulation
Immun
esystem
(+vaccination)
Antivirals
Selectiveforcesonviruses
Whyshouldaviruschangeit’sgene,whataretheselectiveforces?
GeneralstructureBVDVvirusgenome
Peterhans,E.etalVeterinaryResearch,41(6),44.2010
DoesBVDVchange?
PhylogenetictreeofPestivirus genus
Bauermann F V et al. J VET Diagn Invest 2013;25:6-15
BVDV3oratypicalBVDV
BVDV-3/Hobi,comingupinafarmnearyou
§ DifficultydetectingusingBVDV1/2tests,%falsenegativeshigh
§ Theclinicalsignsincludedfever(39.4–40.1°C),cough,seromucoid nasaldischarge,leukopenia,andacceleratedpulseandbreath.Mostaffectedanimalsreceivedsupporttreatmentandrecoveredwithin2weeks;however,thedeathof2animalswasreported.Necropsyfindingsincludedseveretracheitis andbronchopneumoniainvolvingtheapicallunglobes.
§ PrevalenceinNorthAmericastill0%
PhylogeneticanalysisandclassificationofBVDVbasedonNpro
Peterhans,E.etalVeterinaryResearch,41(6),44.2010
SubtypesofcattleenteringFeedlotsinKansas
PIPrevalence0.4%
BVDVinCanada
PhylogenetictreeoftheBVDV5'UTRsequencesderivedfromfeedlotcattleinAlbertathatwerediagnosedwithMucosalDiseaseonpathologicalexamination.ThebranchesinthetreerepresentthedifferentstrainsofBVDVvirus.
Month1 Month2-4 Month5-9
Insemination
PersistentinfectedAntibodynegative
Abortion/malformation
NormalanimalAntibodypositive
Abortion/malformation
Gestation
TransientinfectionwithBVDV
EarlyembryonicDeath
Returntoestrus
Pathogenesis
§ Oronasal infection§ Tonsil§ Regionallymphnodes§ Bloodcirculation§ LymphoidtissueoftheGItractandlungs§ Thymus/bonemarrow/spleen§ Allotherorgansdependingonthevirulenceofthevirus
§ Inourexperience:inPI’sallorgansareBVDVpositive
Lymphaticspread
Lymphaticspread
Leadingtoanimmunosuppression
Biosecurity
KyleMizen
UCVMClass2014BVDVtype2a(strain1373)infectionexperiment27steers/5heiferssinglesourceWeight311-690pounds
PublishedinCVJ
Can Vet J 2015;56:1075–1083
Clinicalscoreandweight
Can Vet J 2015;56:1075–1083
Whitebloodcellcount(leukocytes)
Dayspostinfection
Can Vet J 2015;56:1075–1083
DropinallfractionsoftheWBCs
Leucocytes
Granulocytes
Lymphocytes
Monocytes
Can Vet J 2015;56:1075–1083
Virusinbloodandantibodies
2-log
Can Vet J 2015;56:1075–1083
VirusNeutralizingantibody
Dayspostinfection
Can Vet J 2015;56:1075–1083
Nasalexcretionofvirus
Dayspostinfection
Can Vet J 2015;56:1075–1083
OvalbuminantibodyinductionfollowingBVDVtype2infection
0
0.1
0.2
0.3
0.4
0.5
0.6
0.7
0.8
0.9
1
-10 -5 0 5 10 15 20 25 30 35 40
SamplingDay
BVDV2infectedNegativecontrol
ovalbumin
ovalbumin
OpticalDen
sity(OD)
Can Vet J 2015;56:1075–1083
ConsequencesofacuteBVDV2infection
§ BVDVdelaystheimmuneresponse§ Immunosuppression:whitebloodcellsê§ Weightloss§ Diarrhea§ Mortality
§ Persistentinfectioninducedin1-3montholdfetuses
Whatcanyoudoaboutit?Findthemandremovethem
Isitthateasy!
Serum Earnotch
Heart,lung,liver,kidney,spleen,duodenum,muscle,mes Lnn,lunglnn,urinebladder,gallbladder,bile,feces,urine,ovary/testicle,obex,jejunum,ileum,milkuddertissue,colon,ceacum,skin(=ear),nasalswab,Tonsil,esophagus,serum,whitebloodcells(PBL)
Andthesamefromthefetus(ifpresentandpossible)
EverythingPCRpositiveexceptbileandfeces
BiotypeCPandNCP
Non-cytopathogenic cytopathogenic
http://www.bvd-info.ch
ExampleBVDV:Rearrangementsasgeneticbasisformucosaldisease
Inshort:Thegenomicrearrangementsgeneratingcp viruses:1. DuplicationofBVDVsequences2. Insertionofcellularsequencesot theNS2and
NS3junction
Justtobecomplete…also:1. InframedeletionsaffectingtheNS2-3
polypeptide2. Introductionofpointmutations atvarious
placesinthegenome3. Recombinations betweendifferentBVDV
viruses
Endresult:ProductionoftheNS3proteinnexttoNS2-3polyprotein
Mucosaldisease
MDlesioninthemouthofaPI.
MDcasesinfeedlotanimals
IDEXXHerdchek BVBVAb
InterpretationS/Pvalue<0.2=negative0.2-0.3=suspect³ 0.3=positive
Results:506(67.83%)negative76(10.19%)suspect164(21.98%)positive
BRDproject:October2010
IDEXXHerdchek*IBRgB
Interpretationblocking%:<45%=negative45-55=suspect³ 55=positive
Results:398(53.35%)negative52(6.97%)suspect296(39.68%)positive
746market-derivedmixed-breedbeefcalvesmeanarrivalweight712lbs.
548(73.45%)calvespulledwithin35daysonfeed
CourtesyofBarbaraWolfger andKarinOrsel usedwithpermission
TheBVDVcontainingvaccines:(Jan,2015)
M=modifiedliveK=killed
Doesbiotypematter?WhyCPinavaccine?
AnimalsfirstexposedorprimedwithNCPBVDVbiotypehadsignificantlyhigherantibodytitres
But…alowsecondaryproliferativeresponseagainsttheviruswhateverthebiotypeusedinthesecondaryinoculation.
AnimalsinitiallyinoculatedwiththeCPBVDVbiotype:highTcellreactivityduringthesecondaryexposuredespitethelowerlevelofneutralizingactivityseeninthesecalves.
Thisdistinctionbetweenhumoral andcell-mediatedresponseswouldsuggesttheexistenceofdistinctTh1andTh2 subpopulationsintheregulationofspecificimmuneresponsestoBVDV
Lambot etal.JournalofGeneralVirology(1997),78,1041–1047.
TlymphocytesareinducedwhenvaccinationtakesplaceIFOMA
§ Pentavalent MLVvaccinewas§ EvaluatedforstimulationofCD4,CD8,andotherTcell
responsestoBVDVtypes1and2,BHV1,BRSV,andPI3§ Calveswithmaternalantibody§ Thevaccinedidnotstimulateanantibodyresponse§ Did stimulateallthreeTcelltypestorespondtoBVDV
typesIandIIinvitro§ Protectedthecalvesfromchallengewithvirulent
BVDVtype2.§ NoevidenceofantibodyorTcellresponsestoBHV-1,
BRSV,orPI-3incalveswithmaternalantibody.InductionofTlymphocytesspecificforBVDvirusincalveswithmaternalantibody.ViralImmunol,17(1):13-23,2004
Biologicals Volume31,Issue2,June2003,Pages123–125
BVDVMLVvaccinatedcalveswithmaternalantibodies
PlattetalVaccine27(2009)4508–4519
AsingledoseofacommercialMLVvaccinecontainingBVDVtype1(Singerstrain),BVDVtype2(Bolin296strain),BHV-1(Coloradostrain),BRSV(proprietarystrain)andPI-3(proprietarystrain)withproprietaryadjuvant(Express5™,Boehringer IngelheimVetmedica,Inc.,St.Joseph,MO)wasadministeredsubcutaneously(2 mL).
4calvespercontrolgroup,8calvespervaccinatedgroup
1V/C=1-2weeksold;2V/C=4-5weeksand3V/C=7-8weeks
EffectofBVDVvaccinationonpregnancyrisk
Theriogenology83(2015)360–365
VaccinationwithBVDVvaccinedoesNOT
changepregnancyrates
EffectofBVDVvaccinationon…
Theriogenology 83(2015)360–365
…abortion …fetalprotection
Inmoredetail:Fetalprotection
Theriogenology83(2015)360–365
10PI’sanalyzed
PI 1
PI 2
PI 3
PI 4
NADL (M31182)
PI 8
Oregon (AF091605)
PI 5
Osloss (M96687)
PI 6
PI 7
PI 9
PI 10
100
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100
100
0.05
Farm 1
Farm 2
Farm 3
Farm 4
Farm 5
PI 1
PI 2
PI 3
PI 4
NADL (M31182)
PI 8
Oregon (AF091605)
PI 5
Osloss (M96687)
PI 6
PI 7
PI 9
PI 10
100
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100
100
0.05
Farm 1
Farm 2
Farm 3
Farm 4
Farm 5
BVDV1B
BVDV1A
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PI 1 97
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8198
0.002
O7 O8
O4 O1
O2
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PI 3
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PI 5
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PI 7
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PI 8
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PI 4
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Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2 88
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PI 9
Cluster 9.1
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PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
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PI 8
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PI 4
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Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
88
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PI 9
Cluster 9.1
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PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
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PI 8
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PI 4
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Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
88
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PI 9
Cluster 9.1
86
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PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
DiversityofBVDVinPI’sDow et al. PLOS ONE | DOI:10.1371/journal.pone.0131972 July 1, 2015
BVDVclustersbyTissue
E1-E2 NS5B
Green=colon,blue=MLN,purple=obex,orange=serum,teal=ileum,red=tonsil
Dow et al. PLOS ONE | DOI:10.1371/journal.pone.0131972 July 1, 2015
ClusteringisbyTissueisSignificant
E1-E2 NS5B
Dow et al. PLOS ONE | DOI:10.1371/journal.pone.0131972 July 1, 2015
Dow et al. PLOS ONE | DOI:10.1371/journal.pone.0131972 July 1, 2015
86
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PI 2
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72
79
96
94
89
0.002
PI 3
89 0.002
PI 5
75PI 6
72
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71
0.002
PI 7
77
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0.002
PI 8
0.002
PI 4
72
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0.002
Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2 88
75
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0.002
PI 9
Cluster 9.1
86
0.002
PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
86
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88
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0.002
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95
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0.002
O7 O8
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O2
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PI 2
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PI 3
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PI 5
75PI 6
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0.002
PI 7
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0.002
PI 8
0.002
PI 4
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0.002
Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
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PI 9
Cluster 9.1
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PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
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0.002
PI 1 97
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PI 3
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PI 5
75PI 6
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PI 7
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PI 8
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PI 4
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0.002
Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
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0.002
PI 9
Cluster 9.1
86
0.002
PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
DiversityofBVDVinPI’s
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0.002
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0.002
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O4 O1
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PI 2
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72
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94
89
0.002
PI 3
89 0.002
PI 5
75PI 6
72
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0.002
PI 7
77
7180
0.002
PI 8
0.002
PI 4
72
84
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0.002
Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
88
75
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0.002
PI 9
Cluster 9.1
86
0.002
PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
86
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0.002
PI 1 97
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0.002
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PI 2
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72
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96
94
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0.002
PI 3
89 0.002
PI 5
75PI 6
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71
0.002
PI 7
77
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0.002
PI 8
0.002
PI 4
72
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710.002
Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
88
75
84
0.002
PI 9
Cluster 9.1
86
0.002
PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
86
90
71
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88
78
99
0.002
PI 1 97
95
8198
0.002
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O2
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94
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94
89
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PI 3
89 0.002
PI 5
75PI 6
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PI 7
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0.002
PI 8
0.002
PI 4
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0.002
Cluster 1.1
Cluster 2.1
Cluster 8.1
Cluster 8.2
88
75
84
0.002
PI 9
Cluster 9.1
86
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PI 10
Colon Ileum Milk MLN Obex Serum Tonsil
FourmontholdfetusLactatingdairycow(mother)
Virusesthroughageneticfunnel
Barriersintheplacenta?
Practice:BVDVstrainsWesternCanada
Chernick,Godson,vanderMeer:Infection,GeneticsandEvolution28(2014)367–374
WhendidBVDV1acametoWesternCanada?
Chernick,Godson,vanderMeer:Infection,GeneticsandEvolution28(2014)367–374
BVDVPhylogeny(E2)
50.0
V051_E2
V050_E2
V012_E2
V010_E2
V015_E2
V045_E2
V026_E2
V070_E2
V079_E2
V040_E2
V020_E2
V059_E2V099_E2
V083_E2
V027_E2
V085_E2
V057_E2
V089_E2
V042_E2
V036_E2
V073_E2
V081_E2
V088_E2
V033_E2
V060_E2
V014_E2
V054_E2
V023_E2
V076_E2
V002_E2V024_E2
V046_E2
V065_E2
V008_E2
V028_E2
V022_E2
V005_E2
V029_E2
V019_E2
V047_E2
V001_E2
V100_E2
V097_E2
V009_E2
V052_E2V039_E2
V075_E2
V044_E2
V035_E2
V086_E2
V062_E2
V098_E2
V037_E2
V095_E2
V034_E2
V031_E2
V043_E2
V056_E2
V077_E2
V017_E2
V082_E2
V067_E2
V048_E2
V092_E2
V021_E2
V025_E2
V041_E2
V074_E2
V013_E2
V006_E2
V091_E2
V063_E2
V080_E2V049_E2
V078_E2
V087_E2
V007_E2
V016_E2
V011_E2 1a
1b
2
~100years
~200years
~500years
50Chernick etal.Submitted
>95%BVDVreads<1%BVDVreads
Nextgenerationsequencing
RTwithrandomprimers RTwithBVDVspecificprimers
Nextgenerationsequencingapproach
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ChartTitle
PI199 PI204 PI206 PI209
PI269 PI269F PI422 PI581
PI730 PI730F 50per.Mov.Avg.(PI199) 50per.Mov.Avg.(PI204)
50per.Mov.Avg.(PI206) 50per.Mov.Avg.(PI209) 50per.Mov.Avg.(PI269) 50per.Mov.Avg.(PI269F)
50per.Mov.Avg.(PI422) 50per.Mov.Avg.(PI581) 50per.Mov.Avg.(PI730) 50per.Mov.Avg.(PI730F)
DiversityofBVDV1aonaminoacidlevel
Chernick etal.Submitted
Contemporarystrainsvs.referencestrains
E2
InfluenzaH7N7crossingthespeciesbarrier
Chicken
Turkey Bataille A,vanderMeerF,Stegeman A,KochG(2011)PLoS Pathog 7(6):e1002094.
AutomatedImageProcessing
Eschbaumer,M.,etal.2014J.Virol.Methods
Questions?
