Abstract Booklet Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL -2- Abstract Booklet Background and program In June 2019, the Dutch National Plant Protection Organization (NPPO-NL), Wageningen University & Research (WUR), and Agriculture and Agri-Food Canada (AAFC) organize a workshop on Synchytrium endobioticum, the causal agent of potato wart disease. The workshop brings together biologists, researchers and policy makers working on all aspects of Synchytrium endobioticum and the interaction with its host potato. Aims of the workshop are: 1. expanding professional networks; 2. sharing knowledge via presentations and posters, and; 3. identifying shared research needs. Workshop attendees are encouraged to present their work, and presentations are grouped in three sessions: Session 1. “Disease occurrence and management” Chair: Dr Nico Horn Director General, European and Mediterranean Plant Protection Organization, France Session 2: Understanding the pathogen Dr Peter Bonants Senior scientist/group leader, Biointeractions and plant health, Wageningen University & Research, the Netherlands Session 3: Host resistance and breeding Prof. Dr Richard Visser Professor of Plant Breeding, Wageningen University & Research, the Netherlands A keynote lecture will be delivered by Prof. Dr Francine Govers, Professor of phytopathology from Wageningen University & Research. -3- Time Activity Wednesday 8:30 - 10:00 Welcome with coffee and registration 26 June 10:00 - 11:00 Opening of the workshop Talks 10:00 Welcome to the Dutch National Plant Protection Organization (Mieke Reyniers, Head National Reference Centre of plant health, Dutch National Plant Protection Organization) 10:15 Welcome and aims of the workshop (Bart van de Vossenberg, Molecular biologist, Dutch National Plant Protection Organization) 10:30 Introduction to EPPO and Euphresco (Nico Horn, Director-General EPPO) 11:00 - 12:00 Session 1: Disease occurrence and management 11:00 Lidia Dimitrova, Bulgarian Food Safety Agency, Bulgaria 11:30 Gerard van Leeuwen, Dutch National Plant Protection Organization, the Netherlands 12:00 - 13:00 Lunch 13:00 - 14:00 Session 1: Disease occurrence and management 13:00 Hale Gunacti, Biological Control Research Institue, Turkey 13:30 Marga van Gent-Pelzer, Wageningen University & Research, the Netherlands 14:00 - 16:00 Session 2: Understanding the pathogen 14:00 Syed Munerul Hoque, National Plant Quarantine Authority, Bangladesh 14:30 Theo van de Lee, Wageningen University & Research, the Netherlands 15:00 Bart van de Vossenberg, Dutch National Plant Protection Organization & Wageningen University & Research, the Netherlands 15:30 Hai Nguyen, Agriculture and Agri-Food Canada, Canada -4- 16:00 - 18:00 Poster presentations and drinks Thursday 8:30 - 9:00 Welcome with coffee 27 June 9:00 - 11:00 Session 3: Host resistance and Breeding See abstract booklet for presenters 9:00 Jaroslaw Przetakiewicz, Plant Breeding and Acclimatization Institute, Poland 9.30 Charlotte Prodhomme, Wageningen University and Research, the Netherlands 10:00 Jarosław Plich, Plant Breeding and Acclimatization Institute, Poland 10.30 Dennis Reckwell, Leibniz Universität Hannover, Germany 11:00 - 12:30 Parallel sessions Workshop/Tour NPPO In subgroups: see registration sheet 12:30 – 13:30 Lunch 13:30 - 16:00 Parallel sessions Workshop/Tour NPPO In subgroups: see registration sheet 16:00 - 17:00 Presentation outcome discussion per group Plenary 19:00 Conference diner (optional) Friday 8:30 - 9:15 welcome with coffee 28 June 9:15 - 10:00 Keynote Lecture 10:00 - 10:30 Project pitches Participants with interesting potato wart projects are invited to give a short description (~5-10 min) 10:30 - 11:30 Discussion on shared statement Plenary 11:30 - 12:00 Closing of the meeting 12:00 – 13:00 Lunch Francine Govers, Professor of Phytopathology, Wageningen University and Research, the Netherlands Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Day Abstract Booklet Oral presentations -5- Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Session 1. Disease occurrence and management Pathotype identification of Synchytrium endobioticum (Schilb.) Perc. in Bulgaria and testing of potato varieties for resistance Lidia Dimitrova (1), Ani Becheva (1), Mariyana Laginova (1) 1. Bulgarian Food Safety Agency, Central Laboratory for Plant Quarantine, Sofia, Bulgaria Potato wart disease (PWD) caused by Synchytrium endobiticum was identified for the first time in Bulgaria in the cool and humid summer of 2004 (region Samokov). In 2012 new outbreaks were reported in the southern part of the country (region Smolyan). The pathogen was detected in tuber samples in small gardens with a long history of potato cropping. The area is mountainous, mainly with light sandy to sandy loam soils, low content of organic matter and pH below 5,5. The continuous growing of potatoes without crop rotation and the use of non certified planting material in courtyards were the main reasons for the spread of the pathogen. A series of laboratory tests for pathotype identification of isolates from both regions were carried out. A set of differential cultivars (EPPO, 2004, 2017) was tested using the Glynne-Lemmerzahl method (IHAR-PIB protocol, Poland). The reaction types of the sprouts were scored after two or three weeks according to the classification scheme established by Langerfeld and Stachewiccz (1994). Based on the results the pathotypes 8(F1), 18(T1) and 38(Nevsehir) were identified. A list of cultivars resistant to the local pathotypes is updated annually, according to Council Directive 69/464/EEC of 8 December 1969 on control of PWD, transposed into Bulgarian legislation through Ordinance № 20 of 2 July 2001. Around 170 varieties were tested for resistance in order to be used in buffer zones surrounding infested fields. Most of the tested cultivars were susceptible. The resistant varieties have no commercial importance. -6- Abstract Booklet Oral presentations Session 1. Disease occurrence and management The role of the National Reference Centre, Wageningen in identification, collection maintenance and disease management of potato wart disease Gerard C.M. van Leeuwen (1), Nico J.B. Mentink (1), Johan P. Meffert (1), Karin H.M. Rosendahl (1) & Patricia C.J. van Rijswick (1) 1. Dutch National Plant Protection Organization, National Reference Centre, section Mycology, Wageningen, the Netherlands The study of potato wart disease in all its aspects has always been a major issue in the Netherlands Plant Protection Organisation. Since its first occurrence in 1915 in the north of the country one has tried to minimise the detrimental effects of this disease. Planting resistant cultivars was ánd is still one of the cornerstones to build management of the disease on. In time, the dominant pathotype 1(D1) has been replaced by new pathotypes, e.g. 2(G1) and 18(T1) in the Netherlands. The focus in the past 10-15 years has laid on pathotypes 1(D1), 2(G1), 6(O1) and 18(T1); it is foremost with this series of pathotypes where efforts were done to harmonise methodology and choice of differential cultivars. In conjunction with Germany and Poland an interlaboratory comparison was done on pathotyping in 2009-2011. Later, the Euphresco-project SENDO was initiated by the National Reference Centre (NRC), including partners from nine different European countries (2012-2015). Ultimately, this has lead to a new version of the EPPO Diagnostic Protocol on Synchytrium endobioticum, describing a standardized, validated set of differential cultivars for pathotype identification. Descheduling of infested fields has got much attention in the Netherlands recently. Many of these fields are scheduled now for 20 years, and descheduling according to EPPO Standard PM 3/59 has started. Analysis of soil samples by direct detection (after washing/sieving) ánd bio-assays are executed. Bio-assays are considered as less sensitive, direct examination (detecting resting spores under microscope) is hampered by doubts about the identity of particles seen. Recently, the NRC has started to compare two real-time PCR tests for use in molecular detection of resting spores in soil suspension resulting from the washing/sieving steps. -7- Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Session 1. Disease occurrence and management To be announced Hale Gunacti (1) 1. Biological Control Research Institue, Turkey To be announced -8- Abstract Booklet Oral presentations Session 1. Disease occurrence and management Potato wart disease in the Netherlands: a historic perspective from a molecular view Marga P. E. van Gent-Pelzer (1), Bart T.L.H. van de Vossenberg (1,2), Patricia C.J. van Rijswick (2), and Theo A. J. van der Lee (1) 1. Wageningen UR, Biointeractions and Plant Health & Plant Breeding, Wageningen, The Netherlands 2. Dutch National Plant Protection Organization, Geertjesweg 15, 6706EA Wageningen, The Netherlands To be announced -9- Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Session 2. Understanding the pathogen To be announced Syed M. Hoque (1) 1. Bangladesh Agricultural Research Institute, Gazipur, Bangladesh To be announced - 10 - Abstract Booklet Oral presentations Session 2. Understanding the pathogen The linear mitochondrial genome of the quarantine pest Synchytrium endobioticum; insights in the evolutionary history of an obligate biotroph Bart T. L. H. van de Vossenberg (1,2), Balázs Brankovics (1), Hai D. T. Nguyen (3), Marga P. E. van Gent-Pelzer (1), Donna Smith (4), Kasia Dadej (3), Jarosław Przetakiewicz (5), Jan F. Kreuze (6), Margriet Boerma (7), Gerard C. M. van Leeuwen (2), C. André Lévesque (3) and Theo A. J. van der Lee (2)* * Presenting author 1. Wageningen UR, Biointeractions and Plant Health & Plant Breeding, Wageningen, The Netherlands 2. Dutch National Plant Protection Organization, Geertjesweg 15, 6706EA Wageningen, The Netherlands 3. Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, Canada 4. Canadian Food Inspection Agency, 93 Mount Edward Road, Charlottetown, Canada 5. Plant Breeding and Acclimatization Institute, National Research Institute, Warsaw, Poland 6. International Potato Centre, Avenida La Molina, 1895 Lima, Peru 7. Hilbrands Laboratorium BV, Kampsweg 27, 9418 PD Wijster, Wijster, The Netherlands Chytridiomycota (chytrids) inhabit terrestrial and aquatic environments, and represent a basal lineage in true fungi. Most of the described chytridiomycota are free living saprophytes, but several species are notorious pathogens for plants or amphibians. Synchytrium endobioticum is an obligate biotroph chytrid causing potato wart disease. Quarantine measures have been implemented worldwide to control the disease and to prevent its spread. To determine taxonomical relationships, and to gain insights into the evolutionary history of this plant pathogen we assembled and annotated the mitochondrial genome of S. endobioticum and generated mitochondrial genomes for five additional chytrid species. The mitochondrial genome of S. endobioticum is a linear 72,865 bp molecule with terminal inverted repeats that encodes 14 mitochondrial genes typically found in fungi. Polymorphisms in 30 S. endobioticum isolates shows clustering in four main mitochondrial lineages, and from our data we conclude that the pest was introduced at least three times in Europe. Strains of pathotype 2(G1) and 6(O1) were represented in two mitochondrial lineages, showing that these pathotypes emerged independently. Variations within a strain for polymorphic sites were observed and seem to be consistent in different mitochondrial lineages suggesting that S. endobioticum strains are communities of different genotypes with conserved composition. - 11 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Session 2. Understanding the pathogen Identification of the AvrSen1 gene: towards molecular pathotyping Bart T.L.H. van de Vossenberg (1,2), Charlotte Prodhomme (1), Gert van Arkel (1), Marga P.E. van Gent-Pelzer (1), Marjan Bergervoet (1), Balázs Brankovics (1), Jarosław Przetakiewicz (3), Richard G.F Visser (1), Theo A.J. van der Lee (1), and Jack H. Vossen (1) 1. Wageningen University and Research, Droevendaalsesteeg 1, Plant Science Group, 6708 PB, Wageningen, the Netherlands 2. Dutch National Plant Protection Organization, National Reference Centre, Geertjesweg 15, 6706EA, Wageningen, The Netherlands 3. Plant Breeding and Acclimatization Institute, National Research Institute, Radzików, 05-870 Błonie, Warsaw, Poland Synchytrium endobioticum, the pathogen causing potato wart disease, has a world-wide quarantine status and is included on the HHS and USDA Isolates of S. endobioticum are grouped as pathotypes based on their ability to evade host-resistance in a set of differential potato varieties. So far, thirty-nine pathotypes are acknowledged of which pathotypes 1(D1), 2(G1), 6(O1) and 18(T1) are found most frequently. We hypothesize that differential resistance is caused by recognition of pathotype specific effectors or avirulence (Avr) genes. We set out to identify the effector (AvrSen1) recognized by the potato Sen1 resistance (R) gene product, resulting in pathotype 1(D1) resistance. AvrSen1 is expected to be present in pathotype 1(D1) isolates but absent in others. A comparative genomics approach with fourteen isolates representing six different pathotypes was used to screen S. endobioticum specific secretome. This led to the discovery of an AvrSen1 candidate, because it was present in all pathotype 1(D1) isolates, but absent in others. Expression of this Avrsen1 candidate in potato plants showed a specific hypersensitive defence response in a Sen1 dependant manner. Identification of the AvrSen1 gene represents a first step towards molecular pathotyping based on functional genetic markers. - 12 - Abstract Booklet Oral presentations Session 2. Understanding the pathogen Comparative genome analyses of Synchytrium endobioticum strains from Canadian and European origin representing six different pathotypes reveal pathotype-specific genes and SNPs useful for molecular diagnostics Hai D.T. Nguyen (1), Kasia Dadej (1), Donna S. Smith (2), Wendy Findlay (1) 1. Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, Canada 2. Canadian Food Inspection Agency, 93 Mount Edward Road, Charlottetown, Canada Isolates of Synchytrium endobioticum are grouped into pathotypes, which are defined by their phenotypic virulence or avirulence to a set of potato varieties. Labour intensive and time-consuming bioassays are used in the process of pathotyping isolates. So far, there are no quick molecular diagnostic methods able to discern between various pathotypes. Using the available whole genome data of 14 S. endobioticum strains from Canadian and European origin, representing pathotypes 1(D1), 2(G1), 6(O1), 8(F1), 18(T1) and 38(Nevsehir), three major comparative genomic analyses were performed. Genes in the core genome of the 14 isolates, as well as the genes unique to each pathotype, were determined. The phylogenetic relationships of genes in the core genome were assessed to determine whether certain genes would be good predictors of pathotypes based on grouping patterns. Genome wide SNP analysis was employed to find the union and intersection of pathotype specific sites. Preliminary results show that certain genes and SNPs seem to be unique to a given pathotype although some may be strain-specific. This information will potentially be useful for designing a molecular test for faster pathotyping of isolates compared to the traditional bioassay methods. - 13 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Session 3. Host resistance and breeding Research on the formation of new virulent pathotypes of Synchytrium endobioticum (Schilb.) Perc., the causal agent of potato wart disease Jarosław Przetakiewicz (1) 1. Plant Breeding and Acclimatization Institute - National Research Institute, Department of Plant Pathology, Radzików, Poland. Three different cultivars of potato with varying degrees of resistance to pathotype 1(D1) were used in experience. Irga – extremely resistant, Erika – slightly susceptible and Evora – extremely susceptible to pathotyp1 1(D1) of S. endobioticum. Ten tubers of each cultivars were inoculated with winter sporangia of pathotype 1(D1), using the ring tests. After 2 months of incubation, big and small warts were visible on tubers of Evora and Erica, respectively. No warts were visible on tubers of cv. Irga. The warts were multiplied to check a virulence profile on EPPO differential set using the Glynne-Lemmerzahl method. In both cases the tests showed that the virulence profile was identical to the pathotype 1(D1). Winter sporangia obtained from warts of both cultivars were used to re-inoculate the same varieties as the next generation. After each passage the virulence of each generation were checked. After 2 or 3 passages, there was possible to select virulent pathotype on cv. Erika. The galls obtained from Evora always corresponded to pathotype 1(D1). The virulence profile of isolate selected on cv. Erika was similar to pathotype 6(O1). This increased virulence could be the result of a shift in the heterogeneous 1(D1) population of S. endobioticum due to the selection. - 14 - Abstract Booklet Oral presentations Session 3. Host resistance and breeding Stacking of resistance loci to give full resistance to higher pathotypes of S. endobioticum Charlotte Prodhomme (1), Dong Zhang (1), Herman van Eck (1), Gert van Arkel (1), Marjan Bergervoet (1), Bart T. L. H. van de Vossenberg (1,2), Marga P. E. van Gent-Pelzer (1), Theo A.J. van der Lee (1), Jack H. Vossen (1), and Richard G.F Visser (1), 1. Wageningen University and Research, Droevendaalsesteeg 1, Plant Science Group, 6708 PB, Wageningen, the Netherlands 2. Dutch National Plant Protection Organization, National Reference Centre, Geertjesweg 15, 6706EA, Wageningen, The Netherlands Synchytrium endobioticum, the soil borne parasitic Chytrid responsible for the potato wart disease, is under strict quarantine measures in Europe. Chemical control is not effective against this pathogen which produces spores that can remain viable in the soil for more than 40 years. To contain the multiplication and spread of S. endobioticum, breeding for resistant varieties is required. Breeding programs have been successful in breeding varieties resistant to pathotype 1(D1), but today, few varieties in the breeding germplasm are resistant to the higher pathotypes present in Europe. So far, efforts have been made to identify major resistance genes responsible for pathotype specific resistance, such as Sen1, or for a broader panel of pathotypes, such as Sen2 and Sen3. In this talk, we will emphasize the importance of not using only these major genes in breeding programs and in the field but instead stacking major genes with loci having weaker effects on resistance. In a tetraploid population segregating for pathotypes 2(G1), 6(O1) and 18(T1) resistance, we mapped the Sen3 gene which brings a strong resistance to pathotypes 2(G1) and 6(O1) but a weaker resistance to the pathotype 18(T1) isolate used for the phenotyping. Using a Bulked Segregant Analysis approach, we could identify another locus segregating in the population which, in combination with Sen3, improves the resistance to pathotype 18(T1). More efforts should be made to identify such minor effect loci and to use them in breeding programs. Indeed, the stacking of strong and weak effects resistance loci is necessary to bring a complete resistance, to contain the disease and to hamper the emergence of new pathotypes. - 15 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Session 3. Host resistance and breeding Gene Sen2 – perspective for using in tetraploid potato breeding Jarosław Plich (1), Jarosław Przetakiewicz (2), Beata Tatarowska (1), Bogdan Flis (1), Ewa Zimnoch-Guzowska (1) 1. Plant Breeding and Acclimatization Institute - National Research Institute, Department of Potato Genetics and Parental Lines, Młochów Research Center, 05-831 Młochów, Poland. 2. Plant Breeding and Acclimatization Institute - National Research Institute, Department of Plant Pathology, Radzików, Poland. Synchytrium endobioticum causes potato wart disease imposing severe losses in potato production and, as a quarantine pathogen in many countries, it results in lost trade markets and land for potato cultivation. Chemical control of the pathogen in field conditions is not possible and therefore cultivation of resistant cultivars is the best solution to restrict its spreading. The main limitation of this method is scarcity of an attractive, market-tailored potato cultivars resistant to most important virulent pathotypes of S. endobioticum. Recently the gene Sen2, which confers potato resistance to common (1(D1)) and at least seven virulent pathotypes of S. endobioticum (2(G1), 6(O1), 8(F1), 18(T1), 2(Ch1), 3(M1) and 39(P1)) was mapped to potato chromosome XI. The gene was identified in diploid potato clone and successfully introduced into tetraploid potato genepool via interploid crosses. Our researches confirmed that gene Sen2 provides the same very high level of resistance against the same broad spectrum of potato wart pathotypes in tetraploid potato clones as it was observed in diploid potato clones. A new DNA markers, suitable for marker assisted selection (MAS) in tetraploid potato clones, were also developed. These results clearly show that gene Sen2 is highly promising and can be used in breeding of a new set of potato cultivars resistant to virulent pathotypes of S. endobioticum. This research was funded by National Science Centre in Poland (grant 2013/11/B/NZ9/01959) and The National Centre for Research and Development (grant Sen2Breed). - 16 - Abstract Booklet Oral presentations Session 3. Host resistance and breeding Development of Diagnostic Tools for Resistance Screening and Pathotype Differentiation Dennis Reckwell (1), Friederike Chilla (1), Annette Bartkiewicz (1), Diro Terefe-Ayana (1), Marian Meyer (1), Kerstin Flath (2), Marcus Linde (1), Thomas Debener (1) 1. Leibniz Universität Hannover, Institute of Plant Genetics, Department of Molecular Plant Breeding, Hannover, Germany 2. JKI, Federal Research Centre for Cultivated Plants, Institute of Plant Protection of Field Crops and Grassland, Kleinmachnow, Germany Resistance against pathotype 1(D1) is included in potato breeding for more than hundred years. The locus for resistance (Sen1) is located on chromosome XI, but the nature of the responsible genes is unknown. To characterise resistance to pathotype 18(T1) a segregating monoparental dihaploid potato population was established to locate the respective locus via resistance screening of recombinant genotypes. The resistance genes were located close to the Sen1 locus on an interval of 777 kbp on the reference genome. Additional genotypes were analysed which offer the potential to locate the genes in an interval of less than 500 kbp. Furthermore, the genome of a highly resistant genotype was sequenced to detect candidate resistance genes. More than 40 pathotypes of Synchytrium endobioticum are known. Pathotypes 1(D1), 2(G1), 6(O1) and 18(T1) are the most important ones in Germany. Genome and transcriptome sequences were used to develop diagnostic markers to distinguish between pathotypes, including SCAR markers and microsatellites. Addition of these markers to biotests is capable to speed up the process of pathogen identification. Effector proteins have a crucial role to overcome the plant defence system. Transcriptome sequences of potato warts were analysed to identify effector candidate genes. Four genes supported the spread of Tobacco Rattle Virus in transformed tobacco leaves, indicating the potential to decrease plant defence. - 17 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Oral presentations Keynote lecture Dissecting the biology and pathology of the late blight pathogen Phytophthora infestans Francine Govers (1) 1. Laboratory of Phytopathology, Wageningen University, Droevendaalsesteeg 1, 6708 PB Wageningen. The Netherlands. francine.govers@wur.nl The plant pathogen Phytophthora infestans is the causal agent of late blight, a devastating disease in potato worldwide. Its first appearance in Europe in the mid-nineteenth century led to the Irish potato famine and to the emergence of plant pathology as a scientific discipline. P. infestans belongs to the oomycetes, a diverse group of organisms comprising pathogenic species infecting plants, animals or microbes, as well as saprophytic species. It has a hemibiotrophic life style and exploits a variable repertoire of effector proteins for manipulating plant defence and facilitating colonization. Comparative genomics revealed features illuminating the success of Phytophthora species as plant pathogens, such as a massive expansion of genes encoding secreted proteins and peculiar gene innovations resulting in proteins with oomycete-specific domain combinations. For successful infection Phytophthora secretes a variety of proteins including a large number of effectors that share the host-cell targeting motif RXLR. Inside host cells these RXLR effectors promote virulence by manipulating the cell machinery via interaction with host targets thereby suppressing host defence. However, in plants carrying matching resistance genes RXLR effectors trigger defence and thus act as avirulence factors. In this presentation I will summarize our current knowledge on the biology and pathology of P. infestans. I will also highlight some aspects of RXLR effectors and the discovery of oomycete specific features that might be instrumental for designing a next generation crop protection agents. - 18 - Abstract Booklet Poster presentations - 19 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Poster presentations Wart Potato Synchytrium endobioticum (Schilb.) Perc. impact on plant host. Avreliya G. Zelya (1), Mykhailo P. Solomiychuk (1), George I. Vorobets (2) 1. Ukrainian Scientific- Research Plant Quarantine Station Institute of Plant Protection National Acadeny of Agrarian Sciences, Ukraine 2. Chernivtsi National University, Ukraine Wart potato is one of the dangerous diseases of potato in Ukraine. The researchers of Ukrainian scientific-research plant quarantine station during 2009-2019 determined changes in biosynthesis of crude protein. The peculiar protein spectrum was received by the way of isoelectric focusing in polycrylamide gel in gradulent pH 3,5-10,0. 59 components were identified in protein spectrum in non-infected by wart potato variety Poliska rozheva. The following results of protein components were recorded on infected potato: 28by common pathotype 1(D1), 30 by 11 (Mizhhirya ), 42 by 13 (Rachiv), 51 by 18 (Yasinya ), 53 by 22 (Bystrytsya ). The extracted protein fractions are differed by quantity and their isoelectric points location. The amino acid content of synthesiszing protein change was recorded during the infecting by wart disease agent. The amount of non-changed amino acide was consisted of 42,3 %, defeated by wart potato ,-35,5% in non-defeated potato variety. The increasing of following components was recorded on 21-st day: glutamic acid from 10,6 to 13,1 %, cysteine and cystine from 1.0 to 2,1%, tyrosine from 5,3 to 6,8%, The decrease of following components content was recorded: lysine from 8.3 to 7.9%, aspartic acid from 8,0 to 7,9%, leucine and isoleucine from 8.6 to 6.9%. There were determined difference during the activity analysis of oxidation-reduction enzyme (peroxidase, catalase). They considered “avarium enzymes”. They are immediately react on any stress on plan: tempearature drop, disease casuative agents impact . It was determined the peroxidase activity 0,62 micromole/ min on infected potato sprout and 0,98 micromole/min on 21 st day after infecting. The analysis of biochemical and biophysical indexes of potato plant’s infecting processes is continued. - 20 - Abstract Booklet Poster presentations Choice of potato varieties resistance to Synchytrium endobioticum. George V. Zelya (1), Tetyana M. Oliynyk (2), Volodymyr M. Gunchak (1) 1. Ukrainian Scientific Research Plant Quarantine Station Institute of Plant Protection National Academy of Agrarian Sciences, Chernivtsi, Ukraine 2. Institute for potato research, National Academy of Agrarian Sciences, Ukraine Ukrainian Scientific- Research Plant Quarantine Station Institute of Plant Protection National Academy of Agrarian Sciences provides researches by evaluation and selecting new potato resistant varieties to wart of Ukrainian and foreign breeding for more than 80 years. The resistance test have been conducted by the old methods for evaluation and selection. The testing is conducted as per standard EPPO PM 7/28 after the providing training by Dutch National Plant Protection Organization in 2013 (Wageningen, the Netherlands), by Spieckermann and Glynne-Limmerzahl methods in laboratory test. Field trials on naturally infected soils were conducted in the area of Chernivtsi. Zakarpattia (11- Maydan, Mizhhirya district, 13- Rakhiv, 11- Yasinia) and Ivano- Frankivsk region (22- Bystrets, Verkhovyna district). 1677 samples of potato breeding material were received from 6 scientific- research institutes and breeding centres of Ukraine. 3700 samples of potato breeding material were extracted from 4677 samples during 2011-2018. They were resistant to common pathotype (D1) S. endobioticum in previouis testing and 130 samples were approved in state testing. Among all samples tested, 75 (49%) were identified as resistant to 11- Mizhhirya pathotype,5 (33,9%) samples were resistant to 13 Rakhiv pathotype, 55 (35,9%) samples were resistant to 8-Yasinya, 74 (48,3%) were resistant to 22-Bystrets pathotype. The tests revealed that the national breeding program targeted on resistance against S. endobioticum, common pathotype (D1), 11-Mizhhirya, 22-Bystrets, was the most effective, but it was least effective against 13, 18 pathotypes of wart potato. The already registered potato varieties Bozhedar, Solokha, Khortytsa, Bazys, Santarka which were found to have a multiple resistance to all pathotypes were recommended for use in the breeding process as sources of resistance and also for the eradication programs in sources of wart potato in Ukraine. - 21 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Poster presentations Potato varietal resistance cannot always reduce the impacts caused by the introduction of Synchytrium endobioticum (Schilb.) Perc. into a new area - the case of Greece Irene Vloutoglou (1); Konstantinos B. Simoglou (2); Helias Eleftheriadis (2); Christos Kritikos (1); Dimitrios Tsirogiannis (1); Ioanna Sarigkoli (2); Nikolaos Nikolaidis (2); Christos Arampatzis (3) and Dimitra Gkilpathi (4) 1. Benaki Phytopathological Institute, Department of Phytopathology, Kifissia, Attica, Greece. 2. Region of Eastern Macedonia & Thrace, Regional Unit of Drama, Department of Quality and Phytosanitary Control, Drama, Greece. 3. Hellenic Ministry of Rural Development and Food, Department of Phytosanitary Control, Athens, Greece. 4. Hellenic Ministry of Rural Development and Food, Department of Biocide Products, Athens, Greece Potato wart caused by the quarantine soil-borne fungus Synchytrium endobioticum (Schilb.) Perc. was first reported from Greece in one of the main ware potato-producing areas (Kato Nevrokopi, Northeastern Greece), during the 2011 official surveys. The pathotype present in the contaminated fields was identified as pathotype 18(T1). Since then, strict phytosanitary measures, including the demarcation of a safety zone (c. 200 ha) around the contaminated fields to prevent further spread of the pathogen, have been implemented in the area, in compliance with the EU and National Plant Health legislation. In addition, official testing under field and controlled environmental conditions was performed to identify potato varieties resistant to pathotype 18(T1) to be used in the safety zone. Fifty European potato varieties of various end-uses (table varieties, processing industry/starch varieties), nine of which were reported by other EU Member States and/or potato breeding companies as resistant to pathotype 18(T1), were screened for resistance in a naturally contaminated field over four consecutive years. Varieties showing field resistance were further evaluated in pot tests using artificially contaminated substrate. Results showed that, only two starch varieties constantly exhibited field resistance to pathotype 18(T1) and furthermore provided adequate protection against secondary infections by the pathogen, in line with Council Directive 69/464/EEC. Nevertheless, in the absence of potato starch manufacturers in the country, the farmers are very reluctant to grow these two varieties in the safety zone. As a result, the impacts caused by the introduction of S. endobioticum in Greece remain very high.for use in the breeding process as sources of resistance and also for the eradication programs in sources of wart potato in Ukraine. - 22 - Abstract Booklet Poster presentations The occurence and testing method of Synchytrium endobiticum in the Czech Republic Iveta Svobodová (1); Vladimír Gaar (2) 1. Central Institute of Supervising and Testing in Agriculture, Division of Diagnostics, Department Diagnostic Laboratory Olomouc, Czech Republic 2. Central Institute of Supervising and Testing in Agriculture, Division of Diagnostics, Department Diagnostic Laboratory Prague, Czech Republic Synchytrium endobioticum, the agent of potato wart disease, is an A2 pest for the EPPO region. The occurence of potato wart disease Synchytrium endobioticum in the territory of the present Czech Republic has been known since 1915. Many pathotypes (races) of S. endobioticum are known to occur, in the Czech Republic 19 pathotypes of Synchytrium endobioticum have been identified. However, the last finding of Synchytrium endobioticum in the Czech Republic on the field was in 2010. - 23 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Poster presentations Occurrence of potato wart in Georgia Sopio I. Ghoghoberidze (1); Tsisana M. Tsetskhladze (1); Ketino T. Sikharulidze (1) 1. Institute of Phytopathology and Biodiversity, Batumi Shota Rustaveli State University, the Autonomous Republic of Adjara, 6010, Batumi, Georgia Potato wart is an important disease in Georgia causing yield loss of potato crops along with other diseases. The disease was first found in the private plots of Khulo municipality in 2009. Nowadays more infested foci were detected in two separately located western (Khulo region) and northern (Zemo Svaneti region, Mestia district) areas. Among them, 17 infested foci located at an altitude of about 183 meters above sea level were found in Khulo region, and 2 infested foci located at an altitude of about 2100 meters above sea level were detected in Mestia district, where the abiotic factors are respectively different. To identify the pathotypes, the wart material collected from the private plot of Didajara village, Khulo district (the western part of Georgia) was sent to the Netherlands, and the laboratory, pot and field tests were also conducted in the Institute of Phytopathology and Biodiversity according to the EPPO standards. The results obtained from both laboratories showed the presence of the pathotype 38 (Nevsehir) in the western part of Georgia. By the end of 2018, the wart tissue, collected in the northen part of Georgia (Mestia district) was again sent to the Netherlands, and as well as the pot and lab (using Glynne–Lemmerzahl method) tests for the same wart tissue is being conducted in Georgia. - 24 - Abstract Booklet Poster presentations Preliminary Screening of Potato Cultivars for Resistance to Potato Wart in Georgia Zoya V. Sikharulidze (1); Sopio I. Ghoghoberidze (1); Tsisana M. Tsetskhladze (1) 1. Institute of Phytopathology and Biodiversity, Batumi Shota Rustaveli State University, 6010, Batumi, Georgia The average yield of potato in Georgia is low due to many reasons. Quarantine disease potato wart, caused by fungal pathogen Synchytrium endobioticum, is an important constraint. The pathogen is widely distributed in the world including Georgia where it was first found in 2009 in Khulo district. The most effective way to control the disease spread is the cultivation of varieties resistant to the pathotypes present in the infested fields. The introduced potato cultivars were assessed for the reaction to potato wart using pot and field tests. When tubers formed, wart symptoms were evaluated in the pot test by the Spieckermann scale and in the fields conducted in Khulo villages (Skvana, Uchkho) according to the scale specified in the EPPO Diagnostic Protocol. The majority of cultivars: Marfona, Pekaro, Panamera, Saturna, Spunta, Annalena, Nandina, Glorietta, Bernadette, Estela, Skrab, Uladar, Briz, Lileya, Europrima, Sylvana, Finca, Laura, Figaro, Impala, Jelly, Alwara, Artemis, Milva and Sante showed susceptible reaction and cultivars Arnova, Sofia, Arizona, Fabula, Caruso, Omega, Arinda were resistant in the pot test to pathogen isolate originating from Khulo, Didadjara village. The main part of cultivars showed the susceptibility in both field trials. Cultivars Catania, Leandra, Cardinia and cultivars Sylvana, Agria, Marabel were resistant in the Uchkho and Skvana trials, respectively. As a result of preliminary screening, some potato cultivars showed the different reactions to potato wart in pot and field tests; therefore, further testing of these cultivars is needed to obtain more reliable results. - 25 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Poster presentations The linear mitochondrial genome of the quarantine pest Synchytrium endobioticum; insights in the evolutionary history of an obligate biotroph Bart T. L. H. van de Vossenberg (1,2), Balázs Brankovics (1), Hai D. T. Nguyen (3), Marga P. E. van Gent-Pelzer (1), Donna Smith (4), Kasia Dadej (3), Jarosław Przetakiewicz (5), Jan F. Kreuze (6), Margriet Boerma (7), Gerard C. M. van Leeuwen (2), C. André Lévesque (3) and Theo A. J. van der Lee (2) 1. Wageningen UR, Biointeractions and Plant Health & Plant Breeding, Wageningen, The Netherlands 2. Dutch National Plant Protection Organization, Geertjesweg 15, 6706EA Wageningen, The Netherlands 3. Agriculture and Agri-Food Canada, 960 Carling Avenue, Ottawa, Canada 4. Canadian Food Inspection Agency, 93 Mount Edward Road, Charlottetown, Canada 5. Plant Breeding and Acclimatization Institute, National Research Institute, Warsaw, Poland 6. International Potato Centre, Avenida La Molina, 1895 Lima, Peru 7. Hilbrands Laboratorium BV, Kampsweg 27, 9418 PD Wijster, Wijster, The Netherlands Chytridiomycota (chytrids) inhabit terrestrial and aquatic environments, and represent a basal lineage in true fungi. Most of the described chytridiomycota are free living saprophytes, but several species are notorious pathogens for plants or amphibians. Synchytrium endobioticum is an obligate biotroph chytrid causing potato wart disease. Quarantine measures have been implemented worldwide to control the disease and to prevent its spread. To determine taxonomical relationships, and to gain insights into the evolutionary history of this plant pathogen we assembled and annotated the mitochondrial genome of S. endobioticum and generated mitochondrial genomes for five additional chytrid species. The mitochondrial genome of S. endobioticum is a linear 72,865 bp molecule with terminal inverted repeats that encodes 14 mitochondrial genes typically found in fungi. Polymorphisms in 30 S. endobioticum isolates shows clustering in four main mitochondrial lineages, and from our data we conclude that the pest was introduced at least three times in Europe. Strains of pathotype 2(G1) and 6(O1) were represented in two mitochondrial lineages, showing that these pathotypes emerged independently. Variations within a strain for polymorphic sites were observed and seem to be consistent in different mitochondrial lineages suggesting that S. endobioticum strains are communities of different genotypes with conserved composition. - 26 - Abstract Booklet Poster presentations Sensitive detection of Synchytrium endobioticum in soil samples using zonal centrifuge Kurt Heungens (1) 1. ILVO, Plant Unit, Merelbeke, Belgium Detection of Synchytrium endobioticum in soil samples is relevant in surveys of potentially infected fields. Several methods have been described, including extraction of the winter spores from soil using a zonal centrifuge, filtering, DNA extraction and qPCR as described by Wander et al. (2007) and van Gent-Peltzer et al. (2010). This method is very sensitive, as it combines high efficiency extraction from 100 g soil samples with sensitive qPCR assays. We validated this method and included optimization steps relating to incubation time in the separation liquid, qPCR assay, and use of cloned DNA target for the standard curve. Using these optimized methods we conducted recovery experiments from soil samples spiked with decreasing numbers of resting spores. Even down to 2 spores per 100 g of soil, recovery was successful in all replicates. Especially at lower concentrations, the recovery rate was above 70%. - 27 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL - 28 - Abstract Booklet Parallel sessions - 29 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL The workshop brings together biologists, researchers and policy makers working on all aspects of Synchytrium endobioticum and the interaction with its host potato. Apart from knowledge exchange through presentations, a tour of the NPPO facilities is organized, and research needs and experiences are discussed in six subgroups of approximately 10 persons. During registration you were assigned to a discussion group. On Thursday 27 June we will be active with three sessions. A. B. C. A tour to the facilities of the NVWA demonstrating equipment and procedures with emphasis on spore extraction from soil suspensions, molecular methods and the wart disease compost collecti on. Workshop part 1: Discussion sessions on questions are grouped under three topics (Disease oc currence and management, Pathogen biology, and Host resistance/Breeding), which will serve as starting point for the discussion on needs in potato wart research and management. Discussion groups are invited to comment on the questions below which serve to start the discussion. Subgroups can put emphasis on a specific topic, or may address other relevant topics. Workshop part 2: groups are asked to summarize the items discussed in a PowerPoint presentation. Groups will be equipped with laptop computers and a template PowerPoint presentation. Each group selects a person to plenary present (max. 4 slides, 10 min) the outcomes of the subgroup. Activities for the different subgroups during the parallel workshop and tour sessions 11:00 – 12:30 13:30 – 14:45 14:45 – 16:00 Group 1 Tour Workshop 1 Workshop 2 Group 2 Tour Workshop 1 Workshop 2 Group 3 Workshop 1 Tour Workshop 2 Group 4 Workshop 1 Tour Workshop 2 Group 5 Workshop 1 Workshop 2 Tour Group 6 Workshop 1 Workshop 2 Tour - 30 - Abstract Booklet Questions for the discussion Discussion groups are invited to comment on the questions below which serve to start the discussion. Subgroups can put emphasis on a specific topic, or may address other relevant topics. 1. Disease occurrence and management Do you think problems with potato wart disease are decreasing/increasing or remain similar in your country? Do you think all infections are noted? Do you have indications of emerging/new pathotypes that “break” resistance? Do you think potato varieties offering partial resistance could be used in management strategies? Do you have indications of specific factors that might influence epidemics? Do you have indications of long lasting infestation in particular regions. If so do you have indications of particular conditions that support the presence of S. endobioticum in these regions? Do you have indications alternative hosts may play a role in the epidemics? 2. Pathogen biology Have you implemented pathotyping methods, do you pathotype all isolates, and which differential set and bioassay is used? Do you think the currently used protocol for bioassays are sufficiently harmonized? Do you think that there is a need of a standardized set of pathogen inoculum to determine potato resistance for different pathotypes? Do you think the current used differential set is sufficient to reliably determine the pathotype identity? Would molecular characterization of isolates be something that would be of use? If yes”, for which characteristics would this be possible/which would be of interest to you? 3. Host resistance Do you think your breeding material is sufficiently characterized for wart resistance? What are the main bottlenecks for testing wart resistance? Do you know of any mischaracterizations in of genotypes in public databases? Is the use of molecular markers required to obtain/characterize resistant material? Is the stacking of R genes required? How important is wart resistance for new cultivars? Other topic related to potato wart disease - 31 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL - 32 - Abstract Booklet Participants - 33 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Country Name Position Institute/Organization E-mailadres Austria Sonja Axmann Senior Expert Austrian Agency for Health and Food sonja.axmann@ages.at Safety (AGES) Bangladesh Syed Munerul Quarantine Patho- National Plant Quarantine Authority, Hoque logist Department of Agricultural Extension, syedhq74@gmail.com Bangladesh Belgium Bulgaria Kurt Heungens Ani Becheva Senior researcher Research Institute for Agriculture, Fis- mycology heries and Food (ILVO) Head of depart- Bulgarian Food Safety Agency a.besheva@bfsa.bg ment Canada Lidia Dimitrova Chief expert Bulgarian Food Safety Agency clkr_samokov@mail.bg Hai Nguyen Research Scientist Agriculture and Agri-Food Canada (AAFC) hai.nguyen.1984@gmail. com Czech Re- Iveta Svobodová Mycologist public Denmark Finland Central institute for supervising and tes- iveta.svobodova@ukzuz. ting in agriculture cz Mette Vestergård Researcher Aarhus University mvestergard@agro.au.dk Ea Riis Sundmark Breeder Danespo eri@danespo.com Mikko Lehtonen Project Researcher Finnish Food Authority mikko.lehtonen@ruokavirasto.fi France Nico Horn Director-General European and Mediterranean Plant Pro- nico.horn@eppo.int tection Organization (EPPO) Veronique Wilson French Agency for Food, Environmental veronique.wilson@anses. and Occupational Health & Safety (ANSES) fr Georgia Germany Zoya Sikharulidze Head of Depart- Institute of Phytopathology and biodiver- zsikharulidze@ymail.com ment, senior sity, Batumi Shota Rustaveli State Univer- scientist sity Dorothee Kaem- Head of the work Bayerische Landesanstalt für Landwirt- Dorothee.Kaemmerer@lfl. merer group ‘quarantine schaft bayern.de labaratory head Bayerische Landesanstalt für Landwirt- Peter.Buettner@LfL. mycology schaft bayern.de Phytopathologist Böhm-Nordkartoffel Agrarproduktion mgelszat@boehm-kartof- GmbH & Co. OHG fel.de measures potatoes’ Peter Büttner Karsten Buhr Rafal Zgadzaj Bettina Beerbaum Friederike Chilla Project Coordi- Böhm-Nordkartoffel Agrarproduktion nator GmbH & Co. OHG desk officer Federal Ministry of Food and Agriculture bettina.beerbaum@bmel. (BMEL) bund.de Julius Kühn-Institut (JKI) friederike.chilla@juli- Wissenschaftliche Mitarbeiterin us-kuehn.de - 34 - Abstract Booklet Country Name Position Institute/Organization E-mailadres Germany Kerstin Flath Senior Scientist Julius Kühn-Institut (JKI) kerstin.flath@julius-kue- (continued) hn.de Stephan Koenig Scientist - Labora- Julius Kühn-Institut (JKI) tory Diagnostics Karl-Heinz Pastrik Special consultant stephan.koenig@julius-kuehn.de Landwirtschaftskammer Niedersachsen karl-heinz.pastruk@ lwk-niedersachsen.de Dennis Reckwell Postdoc Leibniz Universität Hannover reckwell@genetik. uni-hannover.de Marcus Linde Senior Scientist Leibniz Universität Hannover linde@genetik.uni-hannover.de Thomas Debener Professor Leibniz University Hannover debener@genetik. uni-hannover.de Joachim Weinert Sachgebietsleiter Pflanzenschutzamt Mykologie Greece Irene Vloutoglou Ireland Andy Bourke Head of the Labo- joachim.weinert@lwk-niedersachsen.de Benaki Phytopathological Institute i.vloutoglou@bpi.gr ratory of Mycology Latvia Kristine Paruma Laboratory Analyst Department of Agriculture, Food and the Expert mycologist Andy.Bourke@agriculture. Marine, Ireland gov.ie State Plant Protection Service of Latvia kristine.paruma@vaad. gov.lv Poland Jarosław Plich Assistant professor Plant Breeding and Acclimatization Insti- j.plich@ihar.edu.pl tute - National Research Institute (IHAR) Jaroslaw Przeta- Senior Scientist kiewicz Russia Yulia Tsvetkova agronomist Plant Breeding and Acclimatization Insti- j.przetakiewicz@ihar.edu. tute - National Research Institute (IHAR) pl All-Russian Plant Quarantine Center ( yutska@mail.ru FGBU “VNIIKR”) Sweden Sofia Windstam Plant Health Swedish Board of Agriculture Officer Johanna Boberg Pest risk analyst sofia.windstam@jordbruksverket.se Swedish University of Agricultural Scien- Johanna.Boberg@slu.se ces Switzerland Stéphanie Schürch scientific collabo- Agroscope stephanie.schuerch@ Plant Breeder Agrico Research a.cucerdean@agrico.nl Plant Breeder Agrico Research j.brommer@agrico.nl Phytopathology Averis Seeds B.V. hilde.room@avebe.com Breeding Specialist Averis Seeds B.V. jasper.tammes@avebe. rator the Nether- Alina Cristina lands agroscope.admin.ch Cucerdean Gerrit Jan Brommer Hilde Room Technologist Jasper Tammes com - 35 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL Country Name the Nether- Nico Rookmaker Position Institute/Organization E-mailadres Breeder Averis Seeds B.V. nico.rookmaker@avebe. lands (continued) com Bart van de Vos- Molecular Biolo- Dutch National Plant Protection Organiza- b.t.l.h.vandevossenberg@ senberg gist tion (NPPO-NL, part of NVWA) nvwa.nl Dirk Jan van der Dutch National Plant Protection Organiza- d.j.vandergaag@nvwa.nl Gaag tion (NPPO-NL, part of NVWA) Gabriela M. Ferrari Supervision plant Dutch National Plant Protection Organiza- g.m.ferraricalcena@nvwa. Cálcena health tion (NPPO-NL, part of NVWA) Gerard van Leeu- senior phytopa- Dutch National Plant Protection Organiza- g.c.m.vanleeuwen@nvwa. wen thologist tion (NPPO-NL, part of NVWA) Jos van Meggelen Senior Advisor Dutch National Plant Protection Organiza- j.c.vanmeggelen@nvwa.nl International Coo- tion (NPPO-NL, part of NVWA) nl nl peration Mieke Reyniers Ineke van Holst Head National Dutch National Plant Protection Organiza- m.m.reyniers@nvwa.nl Reference Centre tion (NPPO-NL, part of NVWA) analyst molecular HLB B.V. i.vanholst@hlbbv.nl biology margriet Boerma projectleider HLB B.V. m.boerma@hlbbv.nl Doretta Boomsma Program leader HZPC Research B.V. doretta.boomsma@hzpc. Plant Pathology & nl Cell Biology Paul Schaap Charlotte Prod- Solynta Ph.D. Student paul.schaap@solynta.com Wageningen University & Research (WUR) charlotte.prodhomme@ homme wur.nl Dong Zhang Ph.D. Student Wageningen University & Research (WUR) zhang.dong@wur.nl Jack Vossen Senior Scientist Wageningen University & Research (WUR) jack.vossen@wur.nl Marga van Research techni- Wageningen University & Research (WUR) marga.vangent@wur.nl Gent-Pelzer cian Peter Bonants Teamleader Wageningen University & Research (WUR) peter.bonants@wur.nl Richard GF Visser Professor and Wageningen University & Research (WUR) richard.visser@wur.nl Head of Department Theo van der Lee Senior Scientist Wageningen University & Research (WUR) theo.vanderlee@wur.nl Turkey Hale Gunacti Senior Scientist Biological Control Research Institue hale.gunacti@tarim.gov.tr Ukraine Avreliya Zelya Chief of Labora- Institute of Plant Protection - National avrelia.zelya@gmail.com tory Academy of Agrarian Sciences of Ukraine Scientific resear- Institute of Plant Protection - National georgetrex7777777@ cher Academy of Agrarian Sciences of Ukraine gmail.com George zelya - 36 - Abstract Booklet Country Name Position Institute/Organization E-mailadres United Alexandra Sch- Senior Plant Pa- Science and Advice for Scottish Agricultu- alexandra.schlenzig@sasa. Kingdom lenzig thologist re (SASA) gov.scot Matthew Everatt Plant Health Policy Department for Environment, Food and United States of John Bienapfl Molecular Biolo- Matthew.Everatt@defra. Rural Affairs gov.uk USDA APHIS PPQ S&T john.c.bienapfl@usda.gov gist America - 37 - Synchytrium endobioticum workshop - 26 to 28 June 2019, NPPO-NL - 38 - Abstract Booklet Biorisk management at the National Reference Centre Plant Health Hans Derks (Biorisk officer) National Plant Protection Organization, National Reference Centre, 6700 HC, Wageningen, the Netherlands, Email: j.h.j.derks@nvwa.nl Introduction. To enable safe handling of plant pests and diseases, the National Reference Centre (NRC) of the National Plant Protection Organization (NPPO) has quarantine facilities and procedures that minimize the risk of escape of harmful organisms. The facilities consist of both laboratories and greenhouses of different containment levels to allow working with organisms with different risk profiles. Together these physical containment and procedures form the base of Biorisk management, including both biosafety and biosecurity. Physical containment Biorisk Plant Classes Based on the biology of the harmful organisms the facilities allow working at appropriate safety levels, based on different Biorisk Plant Classes for laboratories (BPL) and greenhouses (BPG). Tables 1 show the characteristics of these different classes: I, II and III, from low to high, respectively. Table 1 Characteristics of Biorisk Plant Classes Biorisk Plant Risk for Class escape BPL-I, Low risk BPG-I BPL-II, Medium BPG-II risk BPL-III, High risk BPG-III Spread by / Survival Infrastructure Characteristics no spread: - closed transport and storage of harmful organisms - decontamination of waste water before disposal - closed windows (insect screens for greenhouses) - no permanent administration working place - authorised access - protective clothing –> white lab coat - disinfection laboratory - disinfection tools - disinfection hands on leave - treatment biological waste spread by: - contact - water - soil - survival forms like cysts - biosafety cabinet (laboratories) - windows air-tight (e.g. sealed) - functional arranged laboratories/greenhouses - see level I - protective clothing –> blue-labelled lab coat plus gloves - paper work separated from lab work spread by: - air - vegetative reproduction - spores - negative pressure - Hepa filtration of exhausted air - double door with vestibules in between - see level II - protective clothing –> red-labelled lab coat plus gloves and shoe covers - paper not released unless treated as biological waste Figure 1 Facilities minimising the risk of escape Figure 2 Facilities ensure containment of laboratory BPL-III level Procedures The safe working procedures, embedded in the Biorisk Management System are an integral part of the ISO 17025 accreditation of the NRC. The desired practices are described in procedures and related documents and include the following topics: - Organisation of biosafety and biosecurity - Supervision by Biorisk officer - Training of staff and external workers - Authorisation for access to containment areas - Rules of conduct - Maintenance of essential biosafety equipment - Release of equipment for external service - Transport and storage of biohazard materials and waste - Emergency procedures, including: - informing the NPPO, risk analysis - decontamination and corrective measures - Change of containment level - 39 -