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MAD-VIR Microarray Analysis of Minimum 500 Samples) D-JRP4-3.3 (MAD-VIR microarray analysis of minimum 500 samples) st JRP5 - MAD-VIR - ET1 - 1 Call Responsible Partner: SSI GENERAL INFORMATION European Joint Programme Promoting One Health in Europe through joint actions on foodborne full title zoonoses, antimicrobial resistance and emerging microbiological hazards European Joint Programme One Health EJP acronym Funding This project has received funding from the European Union’s Horizon 2020 research and innovation programme under Grant Agreement No 773830. Grant Agreement Grant agreement n° 773830 Start Date 01/01/2018 Duration 60 Months DOCUMENT MANAGEMENT JIP/JRP Deliverable D-JRP4-3.3 (MAD-VIR microarray analysis of minimum 500 samples) Join Integrative/Research JRP5 - MAD-VIR - ET1 - 1st Call Project JIP/JRP Leader Anders FOMSGAARD (SSI) Other contributors Maiken W Rosentierne (SSI) Due month of the deliverable 1st of April 2019 Actual submission month 29th of March 2019 Type R R: Document, report DEC: Websites, patent filings, videos, etc. OTHER Dissemination level CO PU: Public CO: confidential, only for members of the consortium (including the Commission Services) MAD-VIR microarray analysis of minimum 500 samples WP2. Sample collection and analysis Each partner in the project has selected different samples of interest from their biobanks to be tested with the PanVirus microarray in the 1st, 2nd and 3rd round at SSI. A common standardized sample pre- treatment/inactivation protocol was followed when possible. To this date 509 of 671 samples have been tested by the PanVirus microarray. (Table 1 and Table 2). The samples are from wild life (rabbit, wild boar, duck, pheasant, stork, tortoise, hare, wolf, pigeon, springbook, deer, crow, pteropus vampyrus, partridge, eagle, owl, hedgehog and mouse), farm animals (sturgeon, trout, salmon, cat, goat, cow, chicken, horse, turkey, pig, sheep), and ZOO animals (elephant, lemur), aedes, ticks, other insects and human samples (See results in Supplementary Tables 1-5). Table 1: Number of samples analyzed by PanVirus microarray INSTITUTE SSI 1ST ROUND SSI 2ND ROUND SSI 3RD ROUND PIWET 1ST ROUND INIA 1ST ROUND TOTAL PIWET 14/14 25/25 21/21 60/60 INIA 5/5 20/20 3/3 28/28 IZSAM 10/10 20/20 30/30 OIK 16/16 20/20 36/36 ANSES 8/8 20/20 28/28 VRI 3/3 20/20 25/25 48/48 IZSLER 12/12 46/46 58/58 UOS 0 20/20 20/20 APHA 0 17/17 17/17 SSI 43/43 76/76 29/155 148/357 TOTAL 111/111 284/284 54/180 21/21 3/3 473/599 SSI has sent 12 virus positive samples (purified or non-purified inactivated samples) to INIA, PIWET and APHA for ring-testing in 2018 and another 12 samples in 2019 (Table 2). All the samples were Quality Control for Molecular diagnostics (QCMD) samples from different test panels. Table 2: Ring-test of selected samples from different QCMD test panels SAMPLE NO. Virus content Ring-test no.1 Virus SSI PIWET INIA APHA type 1 Zika virus RNA + + + + 2 Chikungunya virus RNA + +1,2 + + 3 Dengue virus type 2, West Nile virus, Yellow fever virus RNA + + + + 4 Measels virus RNA + + + + 5 Varicella Zoster virus DNA + - + - 6 Cytomegalovirus DNA + +3 + - 7 Adenovirus type 1 DNA + + + - 8 Human herpes virus type 1 DNA + + + - 9 Parechovirus type 1 RNA + + + + 10 Mumps RNA + + + + 11 Epstein–Barr virus DNA + +1,3 + - 12 Human herpes virus type 2 DNA + + + - Ring-test no.2 13 Human coronavirus OC43 RNA + 14 Human parainfluenza virus 2 RNA + 15 Enterovirus RNA + 16 Human parvovirus B19 DNA + 17 Human Herpesvirus type 5 DNA + 18 Human Herpesvirus type 1 DNA + 19 Zika Virus (French Polynesian) RNA + 20 Human Metapneumovirus A RNA + 21 Human adenovirus type 4 DNA + 22 HHV6 type A, HHV6 type B RNA + 23 Human rhinovirus A90 RNA + 24 Enterovirus 68 RNA + TOTAL 12/ 24 12/24 12/24 1very low signal 2low/strong signal during second testing 3low signal during second testing Supplementary Table 1: 1st round at SSI PCR No Name Species Material Virus present Array result Ct before/after WTA Usutu virus, Africa 2 Usutu RT-PCR 1 ANSES 1 Turdus merula brain Usutu virus lineage 22/18 WNV, lineage 1a, WNV RT-PCR 2 ANSES 2 Caballus equi brain Western Negative no ct/no ct Mediteranean clade WNV RT-PCR 3 ANSES 3 Caballus equi brain or isolate WNV, lineage 2 Negative no ct/no ct 4 Ornithodoros Negative (PanVirus array v2) ANSES 4 maritimus soft isolate Meaban virus Meaban virus (PanVirus array No PCR assay at SSI 5 tick v3) Japanese encephalitis virus, Japanese encephalitis Aedes pseudoscutellaris reovirus, JEV RT-PCR 6 ANSES 5 Tottori isolate isolate virus, genotype 3 Aedes albopictus densovirus, 13/6 Aedes albopictus densovirus 2 Avian rotavirus A, Hum.Rotavirus 7 ANSES 6 Caballus equi brain Avian rotavirus Turkey rotavirus A, RT-PCR Pheasant rotavirus A no ct/no ct VEEV RT-PCR 8 ANSES 7 Cell culture isolate VEEV VEEV 16/4 Eyach virus, Exogenous mouse mammary tumor virus, 9 ANSES 8 Ixodes ricinus brain Eyach virus No PCR assay at SSI Murine leukemia virus, Murine type C retrovirus, Spleen focus forming virus blood (dilution Usutu RT-PCR 10 INIA 1 Partridge 1:10 in BA1 Usutu Virus Avian myelocytomatosis virus no ct/no ct medium) blood (dilution WNV RT-PCR 11 INIA 12 Partridge 1:10 in BA1 WNV Lineage 2 WNV lin 2 25/26 medium) blood (dilution WNV lin 1, WNV RT-PCR 12 INIA 3 Partridge 1:10 in BA1 WNV Lineage 1 Kunjin virus 27/27 medium) WNV lin 1, WNV RT-PCR Kunjin virus, 28/34 Murine leukemia virus, Sample was tested 13 Murine osteosarcoma virus, with (#2) or without brain, macerated Murine type C retrovirus, Spleen (1#) Dnase treatment INIA 59 Mouse WNV Lineage 1 in PBS (1:10) focus forming virus after NA purification in order to verify the presence of 14 WNV lin 1 endogeneous retrovirus WNV lin 2, WNV RT-PCR Murine leukemia virus, 23/24 15 Murine osteosarcoma virus, Sample was tested Murine type C retrovirus, Spleen with (#2) or without focus forming virus (1#) Dnase treatment after NA purification in brain, macerated INIA 63 Mouse WNV Lineage 2 order to verify the in PBS (1:10) presence of endogeneous 16 negative retrovirus. BUT the DNAse treatment also removed the WNV signal ! Alphamesonivirus 1, Cavally virus, supernatant of Ngewotan virus (JKT9982), Usutu RT-PCR 17 ISZAM 4 Culex pipiens insect Usutu virus Houston virus, no ct/no ct homogenate Nam Dinh virus, Hana virus, Dak Nong virus 18 Negative (PanVirus array v2) supernatant of WNV RT-PCR IZSAM 1 Sparrow-hawk brain WNV Lineage 2 WNV lin 2 19 27/14 homogenate (PanVirus array v3) Ngewotan virus (JKT9982), Nam Dinh virus, Houston virus, Hana virus, 20 Dak Nong virus, Alphamesonivirus 1, Cavally virus (PanVirus array v2) Ngewotan virus (JKT9982), Nam Dinh virus, Houston virus, supernatant of Hana virus, WNV RT-PCR 21 IZSAM 2 Culex pipiens insect WNV Lineage 2 Dak Nong virus, 26/no ct homogenate Alphamesonivirus 1, Cavally virus (PanVirus array v2) Ngewotan virus JKT9982, Negev virus, Nam Dinh virus, Houston virus, 22 Dianke virus, Cavally virus, Alphamesonivirus 1, WNV lin 2 (PanVirus array v3) Supernatant of Negative (PanVirus array v2) WNV RT-PCR 23 IZSAM 3 Crow brain WNV Lineage 2 32/no ct homogenate WNV lin 2 (PanVirus array v3) Supernatant of Usutu RT-PCR 24 IZSAM 5 Blackbird brain Usutu virus Usutu virus 22/not analyzed homogenate Bluetongue virus, Ovine enzootic nasal tumour Bluetongue virus, virus, 25 IZSAM 6 Sheep Whole blood No PCR assay at SSI serotype 3 Jaagsiekte sheep retrovirus, Enzootic nasal tumour virus of goats Supernatant of 26 Bluetongue virus, Negative IZSAM 7 Sheep spleen No PCR assay at SSI serotype 4 27 homogenate Negative supernatant of Bluetongue virus, Culicoides Bluetongue virus, 28 IZSAM 8 insect Caribou feces associated No PCR assay at SSI imicola serotype 4 homogenate gemycircularvirus Pteropine Orthoreovirus, Melaka orthoreovirus, Pteropus Supernatant of Pteropine 29 IZSAM 9 Pulau reovirus, No PCR assay at SSI vampyrus tissue culture Orthoreovirus Kampar orthoreovirus, Cangyuan orthoreovirus RVF RT-PCR 30 IZSAM10 Springbook Serum Rift valley fever virus Rift valley fever virus 30/16 Hum parvovirus PCR Canine parvovirus, IZSLER no ct/no ct 31 CAT Intestinal content PARVOVIRUS Feline astrovirus 2, 10 Hum astrovirus RT-PCR Feline leukemia virus 26/19 Turkey rotavirus A (all segments), Turkey astrovirus, Turkey astrovirus 3, Turkey astrovirus 2, Hum astrovirus RT-PCR IZSLER Turkey astrovirus 1, No ct/no ct 32 TURKEY intestinal content ASTRO + 13 Pheasant rotavirus A (5 Hum rotavirus RT-PCR segments), No ct/no ct Avastovirus 1, Turkey avisivirus, Avian rotavirus (6 segments), Gyrovirus GyV7 Pseudocowpox virus, orf virus, IZSLER Skin proliferative PARAPOX+++ Parapox PCR 33 BOVINE Bovine papular stomatitis virus, 15 lesions PAPILLOMA +++ 16/20 Bovine papillomavirus 2, Bovine papillomavirus 13 No PCR assay at SSI IZSLER European brown hare syndrome 34 HARE Liver LAGOVIRUS (EBHSV) 16 (1#) virus Sample was tested with (#2) or without (1#) Dnase treatment after NA purification in IZSLER European brown hare syndrome 35 HARE Liver LAGOVIRUS (EBHSV) order to verify the 16 (2#) virus presence of endogeneous retrovirus IZSLER Hum adeno PCR 36 PIGEON Intestinal content ADENOVIRUS Pigon adenovirus 1 21 No ct/no ct Y73 sarcoma virus, Turkey parvovirus, Rous sarcoma virus, Infectious bursal disease virus, Fowl adenovirus D, Fowl adenovirus 9, Fowl adenovirus 3, BIRNAVIRUS Fowl adenovirus 2, IZSLER 37 CHICKEN pool of organs (INFECTIOUS BURSAL Fowl adenovirus 11, 29 (2#) DISEASE VIRUS) chicken parvovirus, Caribou feces associated gemycircularvirus, No IBDV PCR assay at avian Sarcoma virus, SSI hum ADV PCR Aviam myelocytomatosis virus, no ct/no ct Avian leukosis, Avian leukemia virus, Sample was tested
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