RACING PIGEON SENSITIVITY FOR H5 HPAI
Mieke Steensels, F. Rauw, S. Van Borm , I. Bardos, D.B. Madeira and B. Lambrecht [email protected]
Belgian HPH5N8 Introduction February-March 2017
1. Hobby Holder 4. Black swan + Wild Duck 1 february 21 march Bird species Number With symptoms Mortality Chicken 33 2 6 Turkey 3 0 3 Guinea fowl 3 0 2 Geese 13 0 1 Peacock 7 0 1 Pheasant 1 0 0 Barbary duck 7 0 0 Mallard 7 0 0 5. Bird refuge center 23 march 2. Swan 27 february
3. Wild + tufted duck 27 february Belgian HPH5N8 Re-Appearance June – July 2017
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? A/Mute swan/Hungary/119/2017
63 A/Greylag goose/Hungary/1941/2017 A/Goose/Hungary/64909/2016 A/Goose/Hungary/63743/2016 A/Goose/Hungary/17051/2017 A/duck/Hungary/60441/2016 A/Goose/Hungary/15729/2017 A/Goose/Hungary/59763/2016 A/Goose/Hungary/17261/2017 A/Goose/Hungary/17580/2017 67 A/duck/Hungary/54494/2016 A/Duck/Hungary/54738/2016 A/Harris Hawk/Hungary/120/2017 A/Goose/Hungary/17985/2017 A/Mallard/Hungary/5821/2017 A/Mulard duck/Hungary/62902/2016 A/Mute swan/Hungary/6276/2017 A/Mulard duck/Hungary/60369/2016 A/Mulard duck/Hungary/59163/2016 A/mute swan/Hungary/51049/2016 A/mute swan/Croatia/70/2016 A/shelduck/Italy/17VIR1572-24/2017
62 A/domestic turkey/Hungary/53433/2016 11 A/Turkey/Hungary/53136/2016 A/Mute swan/Hungary/3137/2017 A/goose/Hungary/55128/2016 A/Goose/Hungary/59712/2016
94 A/Duck/Hungary/55764/2016
61 A/chicken/Italy/17VIR1684-2/2017 A/chicken/Italy/17VIR1751-3/2017 Phylogeny 8 A/mute swan/Croatia/78/2016 A/greylag goose/Croatia/33/2017 A/duck/Hungary/55191/2016 A/chicken/Hungary/59048/2016 63 A/Goose/Hungary/982/2017 A/Duck/Hungary/1588/2017 A/mute swan/Croatia/85/2016 3 A/White fronted goose/Hungary/801/2017 A/breeder duck/Croatia/21/2017 29 A/GuineaFowl/Hungary/596/2017 80 A/Rook/Hungary/4975/2017
25 A/mute swan/Croatia/15/2017 A/Mute swan/Hungary/2508/2017 A/Mute swan/Hungary/3139/2017 Europe
3 27 A/Turkey/Hungary/2030/2017
21 A/Mute swan/Hungary/2825/2017
2 A/Mute swan/Hungary/3513/2017 5 41 A/Chicken/Hungary/2496/2017 A/Mute swan/Hungary/1955/2017 2016-2017 17 A/Mute swan/Hungary/3542/2017 A/Mute swan/Hungary/5316/2017 21 A/Harris hawk/Hungary/2762b/2017 37 61 57 A/Common buzzard/Hungary/7061/2017 84 A/Harris hawk/Hungary/2762a/2017
85 A/mute swan/Croatia/30/2017 A/chicken/Croatia/70/2017 A/Duck/Hungary/984/2017 A/turkey/Czech Republic/38-17 5/2017 (H5N8) A/mallard/Hungary/57857/2016 A/Pheasant/Hungary/6553/2017
21 84 A/Pheasant/Hungary/7685/2017 A/Chicken/Hungary/1751/2017 A/chicken/Czech Republic/55-17 1/2017 (H5N8) A/mute swan/Croatia/9/2017 A/mute swan/Czech Republic/54-17 2/2017 (H5N8) A/chicken/Czech Republic/585-17 1/2017 (H5N8) A/mute swan/Poland/108/2016 7 9 A/duck/France/161108h/2016
38 A/chicken/Czech Republic/2644-17 1/2017 (H5N8) A/chicken/Croatia/103/2016 64 79 A/mute swan/Croatia/104/2016 A/Mute swan/Hungary/2193/2017 A/Cormorant/Hungary/6102/2017 Closer to the earlier EU strains 33 A/mallard/Czech Republic/722-17 2/2017 (H5N8) A/goose/Czech Republic/821-17 2/2017 (H5N8) A/mute swan/Czech Republic/722-17 1/2017 (H5N8) 60 A/Indian Runner Duck/Czech Republic/749-17/2017 (H5N8) A/chicken/Poland/114/2016
75 A/turkey/Italy/17VIR1338-3/2017 66 A/turkey/Italy/17VIR1574-1/2017 29 February-March A/chicken/Poland/79A/2016 14 A/mute swan/Czech Republic/653-17/2017 (H5N8) A/mallard/Czech Republic/2705-17/2017 (H5N8) A/Cygnus olor/Belgium/1567/2017 A/chicken/Poland/85A/2016 A/chicken/Czech Republic/1896-17 1/2017 (H5N8)
59 A/mute swan/Czech Republic/1813-17/2017 (H5N8) 93 A/chicken/Czech Republic/2643-17 1/2017 (H5N8) A/mute swan/Czech Republic/879-17/2017 (H5N8) 29 62 A/turkey/Italy/17VIR973-2/2017
94 A/domestic goose/Poland/33/2016 A/domestic goose/Poland/72/2016 A/turkey/Poland/83/2016 20 A/turkey/Poland/78/2016 A/bronze turkey/Czech Republic/1414-17/2017 (H5N8)
35 A/chicken/Kalmykia/2661/2016 A/wild duck/Tatarstan/3059/2016 A/Common-coot/Egypt/CA285/2016 29 61 A/chicken/Czech Republic/1863-17 1/2017 (H5N8) 43 A/chicken/Poland/002/2017 A/Goose/Hungary/65817/2016 68 92 A/chicken/Czech Republic/206-17 2/2017(H5N8) 40 A/chicken/Czech Republic/206-17 2/2017
38 A/Mute swan/Hungary/6092/2017
89 A/mute swan/Czech Republic/499-17/2017 (H5N8) 95 A/mute swan/Czech Republic/581-17/2017 (H5N8)
94 A/chicken/Belgium/5990/2017
98 A/Brahma chicken/Belgium/6153/2017 A/chicken/Belgium/6178/2017 37 83 A/chicken/Luxembourg/17110447/2017 16 54 A/pochard duck/England/SA12 157809/2016 5 A/gadwall/Kurgan/2442/2016 19 A/chicken/Wales/000023/2016 A/turkey/England/052131/2016 29 June/July Distant from BE 02/03-2017 64 A/turkey/England/003778/2017
13 A/chicken/Italy/17VIR653-12/2017 A/Peregrine falcon/Hungary/4882/2017 A/swan/Italy/17VIR537-2/2017 68 A/Mallard/Hungary/1574b/2017 74 A/Mallard/Hungary/1574a/2017 A/Greylag goose/Hungary/320/2017 22 43 A/duck/Czech Republic/1467-17/2017 (H5N8) 40 A/Goose/Hungary/1030/2017 A/Chicken/Sweden/SVA161122KU0453/SZ0209318/2016
93 A/Chicken/Sweden/SVA161122KU0453/SZ0209321/2016 A/Chicken/Sweden/SVA161122KU0453/SZ0209317/2016 57 A/Chicken/Sweden/SVA161122KU0453/SZ0209316/2016 49 A/chicken/Germany-MV/R8790/2016 86 A/wild duck/Poland/82A/2016 A/chicken/Germany-SH/R8758/2016 A/Common tern/Hungary/8187/2017 A/wild duck/Germany-BW/R8455/2016 26 A/Tufted Duck/Switzerland/V237/2016 38 2 28 A/chicken/Czech Republic/2764-17 1/2017 (H5N8) 99 A/chicken/Czech Republic/2764-17 2/2017 (H5N8) 44 A/tufted duck/Germany/AR8444-L01986/2016
60 A/tufted duck/Germany/AR8459-L01988/2016 A/tufted duck/Germany-SN/R8795/2016 56 59 A/Common Goldeneye/Sweden/SVA161117KU0322/SZ0002165/2016
20 70 A/tufted duck/Germany/AR8444-L01987/2016 A/tufted duck/Germany-SH/R8444/2016 70 41 A/tufted duck/Germany-SH/R8446/2016 A/turkey/Germany-SH/R8595/2016 34 A/herring gull/Poland/84/2016 A/tufted duck/Denmark/17740-1/2016 A/chicken/Voronezh/18/2017 A/chicken/Kalmykia/2643/2016 38 A/long-eared owl/Voronezh/16/2017 A/Ural owl/Voronezh/14/2017 30 79 A/long-eared owl/Voronezh/15/2017 A/chicken/Astrakhan/3131/2016 29 51 A/chicken/Voronezh/19/2017 48 A/chicken/Voronezh/20/2017 A/turkey/Rostov/11/2017 February-March 39 A/goose/Krasnodar/3144/2017
33 A/wigeon/Italy/17VIR57-3/2017 5 33 43 A/peacock/Belgium/1017/2017 3 A/chicken/Belgium/807/2017 Close to the Italian wild wigeon A/decoy duck/France/161104e/2016 A/turkey/Italy/17VIR5878-3/2017 2 99 A/chicken/Sergiyev Posad/38/2017 35 A/chicken/Sergiyev Posad/39/2017 32 A/chicken/Italy/17VIR7062-1/2017 H5N8
13 A/mute swan/Krasnodar/25/2017 A/turkey/Italy/17VIR576-11/2017 21 14 A/turkey/Italy/17VIR1452-22/2017 15 A/chicken/Italy/17VIR3078/2017 isolate strain and the French decoy A/great crested grebe/Uvs-Nuur Lake/341/2016 A/great crested grebe/Tyva/34/2016
58 A/wild duck/Tyva/35/2016 A/black-headed gull/Tyva/41/2016 97 A/grey heron /Uvs-Nuur Lake/20/2016 A/common tern /Uvs-Nuur Lake/26/2016 A/duck/India/10CA01/2016 A/painted stork/India/10CA03/2016 A/domestic duck/Siberia/50K/2016 99 A/domestic duck/Siberia/103/2016 bird 88 53 A/gadwall/Chany/97/2016 A/green-winged teal/Egypt/871/2016 Eurasia 98 A/green-winged teal/Egypt/877/2016 48 A/turkey/Italy/17VIR538-1/2017 A/chicken/Korea/H903/2017
35 A/mallard duck/Korea/WA137/2017 A/grey heron/W779/2017 93 A/common teal/Korea/W547/2016 2016-2017 A/common teal/Korea/W549/2016
59 A/common teal/Korea/W548/2016 A/common teal/Korea/W550/2016 64 A/common teal/Korea/W555/2017
Qinghai 2016 95 Qinghai 2016
0.0010 Pigeon Sensitivity Study I
DOSE EVALUATION
• 2-5 years old racing pigeon
• Belgian 2017 clade 2.3.4.4. strains, A/Brahma_Chicken/Belgium/6153/2017 (BE_June17) (IVPI 2,98)
6 birds/group
BE_June17 10E4 EID50/bird 10E6 EID50/bird 10E8 EID50/bird Clinical symptoms - - - Mortality 0% 0% 0% Viral Shedding 1 bird: - - 2 dpi, C: 2,71E3 (swabs) 5 dpi, O: 3,33E3 No virus detected (K, LT, B, C, ND ND Organs O, H, Li, S) HI: 0/6 HI: 4/6 HI: 4/6 Serology ELISA: 0/6 ELISA: 5/6 ELISA: 6/6 Pigeon seem poorly sensitive to this clade 2.3.4.4.b strain Passive transmission of humoral immunity
Recovery of eggs, group 10E6 EID50/bird
12 dpi 1x 14 dpi 2x
Transfer of immunity? (IDVET ELISA-NP v2)
12 dpi mix Alb+Yolk +++ 14 dpi 1. Albumen +++ 1. Thick Albumen ++ 1. Yolk ++++ 2. Albumen +++ 2. Yolk +
Minimal protection against early infection? Silent circulation!? Pigeon Sensitivity to HP H5 Data consolidation _ Study II
• 2-5 years old racing pigeon,
• 12 birds/group (6 for clinical follow-up + 6 for viral kinetics)
• 2 HP_H5 strains, 106 EID50/bird
A/Chicken/Belgium/807/2017 (clade 2.3.4.4.b) BE_Febr17 HP_H5N8 (IVPI 2,96)
A/Mynah/Austria-quarantine/13063485-025/13 (clade 2.3.2.1.c) Indo’13 HP_H5N1 (IVPI 2,95)
100% Group II HP H5N1 (Indo’13) HP_H5N1 (clade 2.3.2.1.c) (Indo’13),
80% Group I, HP H5N8_807 (Febr 4 out of 6 birds died
'17) 60%
40% Clinical symptoms %Mortality - Nervous symptoms: trembling/torticollis 20% - Lethargic 0% 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 dpi Neurological symptoms
Lethargy Pigeon Sensitivity to HP H5 Data consolidation
SEROCONVERSION BE_FEBR ’17 Indo_’13 CLADE 2.3.4.4.b CLADE 2.3.2.1.c ELISA-NP 90 % (9/10) 100 % (7/7)
HI 70 % (7/10) 86 % (6/7) homologue VIRAL EXCRETION CLS TRS 8 CLADE 2.3.4.4.b
7 O > C: # + quantity
6 10/12 12/12 3/7 5 3/12 4/12 1/9 2/10 8/12 4 1/8 1/7 2/12 6/9 CLADE 2.3.2.1.c 3 1/8
log10(EID50/ml) 2 2/12 O ~ C: # + quantity 1
0 2 dpi 5 dpi 7 dpi 9 dpi 2 dpi 5 dpi 7 dpi 9 dpi GR I: HP H5N8 Be_Febr GR II: HP H5N1 Indo’13 SUMMARY STUDY I + II
Day of death or viral excretion Viral kinetics seroconv euthanis oral cloacal Organs ersion ation 2 5 7 9 2 5 7 9 S O B H K C TL Br L E HI 1 PIGEON SENSITIVITY 2 3 4
Clade 2.3.2.1.c > Clade 2.3.4.4.b 5 (10Exp8 EID50/ml) H5N8_June'17 6 1 5 dpi - - 2 5 dpi - - 3 7 dpi +/- - CLADE 2.3.4.4.b 4 7 dpi + - 5 9 pdi 6 9 pdi 2 related viruses, with different viral excretion 7 8 9 and kinetics patterns 10
11 (10Exp6 EID50/ml) Prolonged circulation = reduced pathogenesis? H5N8_February '17 12 1 5 dpi 2 5 dpi 3 Ϯ, 7 dpi 4 5 Does this reduced pathogenesis also apply for 6 7 Ϯ, 7 dpi wild birds? 8 Ϯ, 9 dpi 9 10
11 Ϯ, 8 dpi (10Exp6 EID50/ml) H5N1_Indon'13 12
dpi HI
Liver
ELISA
Heart Brain
Ovary Bowel
Spleen
Kidney
Caecum Trachea+Lung position: relative to first start Alternative orf SEGMENT codon nt 807 nt Brahma aa 807 aa Brahma aa 807 aa Brahma 1 PB2_227 C T PB2_76 T I 1 PB2_268 A T PB2_90 M L 1 PB2_643 A G PB2_215 T A 1 PB2_746 G A PB2_249 G E 2 related viruses, with 1 PB2_899 A G PB2_300 Q R 1 PB2_945 G A PB2_315 M I different pathogenicity 1 PB2_1199 T C PB2_400 C A 1 PB2_1676 C A PB2_559 T N 2 PB1_155 A G PB1_52 K R pattern in pigeon 2 PB1_294 A C PB1-F2_29 H P 2 PB1_314 A C PB1_105 N T PB1-F2_36 T P 2 PB1_632 A G PB1_211 K R 2 PB1_1165 C A PB1_389 L I 2 PB1_1183 C A PB1_395 L M Molecular marker? 2 PB1_1363 A G PB1_455 N D 2 PB1_1735 C A PB1_579 L M NGS_full genome 3 PA_556 A G PA_186 S G PA-X_186 S G 3 PA_611 G A PA-X_203 N K 3 PA_871 A T PA_291 S C 97 mutations detected 3 PA_1201 G A PA_401 G R 3 PA_1348 A G PA_450 I V 37 non-silent 3 PA_1673 C T PA_558 S L 4 HA643 T G HA215 S A 4 HA715 C H(A in sanger) HA239 R S Signification? 5 NP_929 C G NP_310 T S 5 NP_1348 G A NP_450 G S 6 NA406 G A NA136 A T 6 NA560 G A NA187 R K 6 NA793 A G NA265 T A 6 NA907 A G NA303 I V NA1402- NA468- 6 GATAA TTATC DK LS 1406 469 7 M_459 G T M1_153 Q H M2 no differences 8 NS_49 Y T NS1_17 H/Y Y 8 NS_245 C T NS1_82 A V 8 NS_264 C T NS1_88 R C 8 NS_610 A G NS1_204 R G 8 NS_651 G A NS2_64 S N FUTURE PERSPECTIVES
• Identification of molecular markers pigeon pathogenicity
• Interspecies Transmission? ON infection + 6 sentinel SPF WL layers HP H5 1 dpi
A/Chicken/Belgium/807/2017, Clade 2.3.4.4b, Febr ’17 A/Brahma_Chicken/Belgium/6153/2017, Clade 2.3.4.4.b, June ‘17
A/Mynah/Austria-quarantaine/13063485-025/13, Clade 2.3.2.1.c THANK YOU
Thierry van den Berg
Bénédicte Lambrecht Dr D. B. Madeira Steven Van Borm Dr I. Bardos Fabienne Rauw Virginie Roupie
Alexander Ausloos Marc Boschmans Siam Mahibullah Catherine Rasseneur