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Roenhorst J.W.,National Plant Protection Organization | Boonham N.,UK Environment Agency | Winter S.,Julius Kuhn Institute | Menzel W.,Leibniz Institute DSMZ | van der Vlugt R.A.A.,Plant Research International
EPPO Bulletin | Year: 2013

The availability of characterised reference isolates of plant pathogens is of crucial importance for research and diagnostic laboratories. The Q-bank Plant Viruses and Viroids database contains data and information on plant viruses and viroids, with the unique feature that it is linked to specimens present in publicly available physical collections. The Q-bank database aims to share data and information on the virus and viroid species and their availability between research and diagnostic laboratories. Currently the database focuses on regulated virus species. In future more plant viruses and viroids will be included to provide a comprehensive data and information system. The curators invite virologists worldwide to participate in this international initiative by making their data and isolates available via Q-bank (http://www.q-bank.eu). © 2013 The Authors. Journal compilation © 2013 OEPP/EPPO.


Van de Vossenberg B.T.L.H.,National Plant Protection Organization | Voogd J.G.B.,National Plant Protection Organization | Westenberg M.,National Plant Protection Organization | Karssen G.,National Plant Protection Organization
EPPO Bulletin | Year: 2014

Three different versions of the conventional PCR described by Bulman & Marshall (1997) for the identification of cysts and juveniles of Globodera pallida and G. rostochiensis were compared: the original Bulman & Marshall, Bulman & Marshall as described in EPPO PM 7/40 (2) and an in-house modified version of the Dutch National Plant Protection Organization (NPPO-NL). The versions differ from each other in thermocycler conditions and primer sequences. Two different polymerases (Invitrogen and Roche) were assessed using the different test versions, and performance criteria analytical sensitivity and analytical specificity were determined. Roche-based reaction mixes had the highest amplicon yield and were used for further comparison of the different test versions. The different test versions performed equally well in terms of analytical specificity. No false positive or false negative results were observed. The test version NPPO-NL proved to be the most sensitive test version with a limit of detection of 1 juvenile for both G. pallida and G. rostochiensis. © 2014 OEPP/EPPO.


Ahmed M.,National Plant Protection Organization | Ahmed M.,Ghent University | van de Vossenberg B.T.L.H.,National Plant Protection Organization | Cornelisse C.,National Plant Protection Organization | And 2 more authors.
ZooKeys | Year: 2013

The root-knot nematode Meloidogyne ulmi is synonymised with Meloidogyne mali based on morphological and morphometric similarities, common hosts, as well as biochemical similarities at both protein and DNA levels. M. mali was first described in Japan on Malus prunifolia Borkh.; and M. ulmi in Italy on Ulmus chenmoui W.C. Cheng. Morphological and morphometric studies of their holo- and paratypes revealed important similarities in the major characters as well as some general variability in a few others. Host test also showed that besides the two species being able to parasitize the type hosts of the other, they share some other common hosts. Our study of the esterase and malate dehydrogenase isozyme phenotypes of some M. ulmi populations gave a perfectly comparable result to that already known for M. mali. Finally, phylogenetic studies of their SSU and LSU rDNA sequence data revealed that the two are not distinguishable at DNA level. All these put together, leave strong evidences to support the fact that M. ulmi is not a valid species, but a junior synonym of M. mali. Brief discussion on the biology and life cycle of M. mali is given. An overview of all known hosts and the possible distribution of M. mali in Europe are also presented. © Mohammed Ahmed et al.


van de Vossenberg B.T.L.H.,National Plant Protection Organization | Westenberg M.,National Plant Protection Organization | Bonants P.J.M.,Plant Research International
EPPO Bulletin | Year: 2013

DNA barcoding protocols for selected EU-regulated arthropods, bacteria, fungi, nematodes and phytoplasmas were developed within the Quarantine organisms Barcoding of Life (QBOL) project financed by 7th framework program of the European Union. DNA barcodes generated with the developed protocols were stored in the Q-bank database. An test performance study (TPS) was set up involving 14 participating laboratories to validate the use of the developed protocols as a diagnostic tool and to identify possible difficulties in the use of the protocols and Q-bank. This paper describes the steps that were used to set up the TPS, to validate the protocols and to identify difficulties. TPS data shows that the developed tests are very robust and produce highly reproducible results. Participants managed to define good consensus sequences which allowed them to correctly identify their samples using Q-bank in 78% of all cases. Q-bank outperformed NCBI and BOLD in terms of diagnostic sensitivity and diagnostic specificity for all organism groups. Using general qualifiers, performance criteria and feedback from TPS participants, difficulties in the set-up of the TPS, the use of the protocols and databases, and the proficiency of participants were identified/evaluated and recommendations for future work were made. The developed DNA barcoding protocols and Q-bank have proven to be useful tools in support of the identification of selected EU-regulated plant pests and pathogens on the desired taxonomical level. © 2013 The Authors. Journal compilation © 2013 OEPP/EPPO.


Roenhorst J.W.,National Plant Protection Organization | Botermans M.,National Plant Protection Organization | Verhoeven J.T.J.,National Plant Protection Organization
EPPO Bulletin | Year: 2013

Test plants are often used for broad screening for plant viruses. Mechanical inoculation of a series of test plants enables generic detection of mechanically transmitted viruses in only 1 assay. Moreover, such an assay is suitable for known as well as unknown viruses and their variants. However, in comparison to serological and molecular methods, quality control in bioassays is almost never addressed. The system of positive and negative controls, blind samples and proficiency tests is applicable, provided that a broader interpretation of positive and negative controls is used. For validation, performance criteria can only be determined for individual viruses. However, results can often be extrapolated. Sensitivity is addressed by dilution and expressed as a relative value. Specificity has to consider the virus species and the plant species to be tested. Selectivity mostly depends on the plant species tested, because some hosts contain components that inhibit transmission. Repeatability and reproducibility, determined for a limited number of samples, appears high, as also substantiated by the authors' experience. This paper details how EPPO Standards on quality control were implemented by the National Plant Protection Organization of the Netherlands (NPPO-NL). This information will be of use for other laboratories that wish to introduce quality control in bioassays for virus testing. © 2013 The Authors. Journal compilation © 2013 OEPP/EPPO.


Alvani S.,Ferdowsi University of Mashhad | Mahdikhani-Moghadam E.,Ferdowsi University of Mashhad | Rouhani H.,Ferdowsi University of Mashhad | Mohammadi A.,Birjand University | Karssen G.,National Plant Protection Organization
Zootaxa | Year: 2016

In order to identify plant-parasitic nematodes (family Tylenchidae Örley 1880) associated with Ziziphus zizyphus in Iran, 360 soil and root samples were collected from South Khorasan province during 2012-2014. Herein, a new species of Basiria and several known members of the family Tylenchidae are reported. B. birjandiensis n. sp. is characterized by short body length (584-748 μm [660.6±72.3]), lip region with flat apex, stylet 11-12 μm (11.3±0.5), excretory pore position varying from isthmus level to the middle of the basal bulb (78-91 μm from the anterior end of the body), post-vulval uterine sac 8-14 μm (10.7±1.9) long, filiform tail (151-181 μm, c= 3.7-4.2, c= 14.3-17.2) and body annuli 0.5-1 μm (0.6±0.1) wide. A checklist of Tylenchidae species from Iran is also presented. © Copyright 2016 Magnolia Press.


PubMed | Ghent University and National Plant Protection Organization
Type: | Journal: ZooKeys | Year: 2013

The root-knot nematode Meloidogyne ulmi is synonymised with Meloidogyne mali based on morphological and morphometric similarities, common hosts, as well as biochemical similarities at both protein and DNA levels. M. mali was first described in Japan on Malus prunifolia Borkh.; and M. ulmi in Italy on Ulmus chenmoui W.C. Cheng. Morphological and morphometric studies of their holo- and paratypes revealed important similarities in the major characters as well as some general variability in a few others. Host test also showed that besides the two species being able to parasitize the type hosts of the other, they share some other common hosts. Our study of the esterase and malate dehydrogenase isozyme phenotypes of some M. ulmi populations gave a perfectly comparable result to that already known for M. mali. Finally, phylogenetic studies of their SSU and LSU rDNA sequence data revealed that the two are not distinguishable at DNA level. All these put together, leave strong evidences to support the fact that M. ulmi is not a valid species, but a junior synonym of M. mali. Brief discussion on the biology and life cycle of M. mali is given. An overview of all known hosts and the possible distribution of M. mali in Europe are also presented.


PubMed | National Plant Protection Organization, U.S. Department of Agriculture and Colorado State University
Type: Journal Article | Journal: PloS one | Year: 2015

The Old World bollworm, Helicoverpa armigera (Hbner), and the corn earworm, H. zea (Boddie), are two of the most important agricultural pests in the world. Diagnosing these two species is difficult-adults can only be separated with a complex dissection, and larvae cannot be identified to species using morphology, necessitating the use of geographic origin for identification in most instances. With the discovery of H. armigera in the New World, identification of immature Helicoverpa based on origin is no longer possible because H. zea also occurs in all of the geographic regions where H. armigera has been discovered. DNA barcoding and restriction fragment length polymorphism (RFLP) analyses have been reported in publications to distinguish these species, but these methods both require post-PCR processing (i.e., DNA sequencing or restriction digestion) to complete. We report the first real-time PCR assay to distinguish these pests based on two hydrolysis probes that bind to a segment of the internal transcribed spacer region 2 (ITS2) amplified using a single primer pair. One probe targets H. armigera, the second probe targets H. zea, and a third probe that targets a conserved segment of 18S rDNA is used as a control of DNA quality. The assay can be completed in 50 minutes when using isolated DNA and is successfully tested on larvae intercepted at ports of entry and adults captured during domestic surveys. We demonstrate that the assay can be run in triplex with no negative effects on sensitivity, can be run using alternative real-time PCR reagents and instruments, and does not cross react with other New World Heliothinae.


Dullemans A.M.,Wageningen University | Verhoeven J.T.J.,National Plant Protection Organization | Kormelink R.,Wageningen University | van der Vlugt R.A.A.,Wageningen University
Archives of Virology | Year: 2014

The complete genome sequence of chrysanthemum stem necrosis virus (CSNV) was determined using Roche 454 next-generation sequencing. CSNV is a tentative member of the genus Tospovirus within the family Bunyaviridae, whose members are arthropod-borne. This is the first report of the entire RNA genome sequence of a CSNV isolate. The large RNA of CSNV is 8955 nucleotides (nt) in size and contains a single open reading frame of 8625 nt in the antisense arrangement, coding for the putative RNA-dependent RNA polymerase (L protein) of 2874 aa with a predicted Mr of 331 kDa. Two untranslated regions of 397 and 33 nt are present at the 5’ and 3’ termini, respectively. The medium (M) and small (S) RNAs are 4830 and 2947 nt in size, respectively, and show 99 % identity to the corresponding genomic segments of previously partially characterized CSNV genomes. Protein sequences for the precursor of the Gn/Gc proteins, N and NSs, are identical in length in all of the analysed CSNV isolates. © 2014, Springer-Verlag Wien.


News Article | March 7, 2016
Site: phys.org

The polyphagous parthenogenetic root-knot nematodes of the genus Meloidogyne are considered to be the most significant nematode pest in sub-tropical and tropical agriculture. Despite the crucial need for correct diagnosis, identification of these pathogens remains problematic. Scientists of Ghent University (Belgium), the National Plant Protection Organization (the Netherlands) en het International Institute of Tropical Agriculture (Nigeria) refined the identification of the nematodes.

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