General crop inspection procedure for potatoes

<i>Ralstonia solanacearum</i>

В статье приводятся результаты анализа сортообразцов картофеля на наличие опасного карантинного объекта для территории Республики Казахстан - вироида веретеновидности клубней картофеля методом обратнотранскриптазной полимеразной цепной реакции с использованием шести различных пар праймеров. В рабочей коллекции сортов и гибридов картофеля Некоммерческого акционерного общества "Казахского агротехнического исследовательского университета имени Сакена Сейфуллина" были выявлены 1 образец в 2021 году и 19 образцов в 2022 гг. казахстанской и зарубежной селекции с фрагментами, обладающимимолекулярной массой, характерной для вироида веретеновидности клубней картофеля.Случаи обнаружения вироида веретеновидности клубней картофеля послужили сигналом для проведения экспресс-анализа фитосанитарного риска для страны. В соответствии со Стандартом ЕОКЗР PM 5/5 (1) «Руководство по анализу фитосанитарного риска: схема поддержки принятия решения для экспресс-анализа фитосанитарного риска» сформировано резюме экспресс-анализа фитосанитарного риска. Результаты анализа подтверждают необходимость изменения статуса карантинного объекта в стране. Сформированные в резюме, результаты исследований, будут переданы в соответствующие органы карантина растений.

Standards to be withdrawn

<i>Diabrotica speciosa</i>

In 1960s, potato spindle tuber was thought to be a viral disease. In 1971, the agent of the disease was characterised as a low-molecular weight infectious ribonucleic acid (RNA), which was named as 'viroid', specifically Potato spindle tuber viroid (PSTVd)). Since then, more than 30 plant diseases in horticultural and ornamental plants have been shown to be caused by different viroids globally. Viroids are single-stranded RNA, covalently closed circular molecule, without any protein coat. They are the smallest known plant pathogen containing RNA genome ranging from 246 nucleotides (Coconut cadang-cadang viroid) to 399 nucleotides (Chrysanthemum chlorotic mottle viroid). Some viroids are located in the plant cell nucleus (pospiviroids) and others in the chloroplast (avsunviroids). With the recognition of pathogenic nature of viroid, specific detection methodologies were developed, which enabled detection of PSTVd in seed-potato tubers prior to their planting in the field, and thus PSTVd was prevented from spreading the disease. As a result, PSTVd was eradicated from Canada in late 1980s. Viroids similar to PSTVd (Pospiviroid) have been discovered and they are detected in symptomless ornamental plants. Although, PSTVd has been eradicated from Canada, there is a strong possibility of viroid introduction from other plants besides potato and tomato and causing PSTVd like diseases.

EPPO BulletinVolume 52, Issue 3 p. 516-517 EUROPEAN AND MEDITERRANEAN PLANT PROTECTION ORGANIZATIONFree Access Introduction to PM 3 Standards on Phytosanitary procedures PM 3/66 (3), PM 3/77 (2), PM 3/81 (3), PM 3/82 (3), PM 3/94 (1) First published: 20 December 2022 https://doi.org/10.1111/epp.12898AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat 1 APPROVAL EPPO Standards are approved by the EPPO Council. The date of approval appears in each individual Standard. In the terms of Article II of the IPPC, EPPO Standards are Regional Standards for the members of EPPO. 2 REVIEW EPPO Standards are subject to periodic review and amendment. The next review date for this set of EPPO Standards is decided by the EPPO Working Party on Phytosanitary Regulations. 3 AMENDMENT RECORD Amendments will be issued as necessary, numbered and dated. The dates of amendment appear in each individual Standard (as appropriate). 4 SCOPE EPPO Phytosanitary Procedures are intended to be used by NPPOs in their capacity as bodies responsible for the inspection and testing of plants and plant products moving in trade, or for the implementation of surveys against quarantine pests. 5 DEFINITIONS These definitions are extracted from the Glossary of phytosanitary terms (IPPC, 2019): Inspection: Official visual examination of plants, plant products or other regulated articles to determine if pests are present and/or to determine compliance with phytosanitary regulations. Monitoring: An official ongoing process to verify phytosanitary situations. Phytosanitary procedure: Any official method for implementing phytosanitary measures, including the performance of inspections, tests, surveillance or treatments in connection with regulated pests. Survey (of pests): An official procedure conducted over a defined period to determine the presence or absence of pests, or the boundaries or characteristics of a pest population, in an area, place of production or production site. Test: Official examination of plants, plant products or other regulated articles, other than visual, to determine if pests are present, identify pests or determine compliance with specific phytosanitary requirements. 6 OUTLINE OF REQUIREMENTS EPPO Standards on phytosanitary procedures describe the methods to be followed for performing inspections or tests of commodities moving in trade, or surveys against quarantine pests. Methods to be followed for performing treatments of commodities and treatments of crops for containment or eradication of regulated pests are included in series PM10. EPPO Standards on phytosanitary procedures arise from the programmes of activity of the EPPO Panels on Phytosanitary Measures, on Phytosanitary Measures for Potato, and on Phytosanitary Procedures. The full list of phytosanitary procedures Standards (Series PM3) is available at https://www.eppo.int/RESOURCES/eppo_standards/pm3_procedures with direct links to the Standards in the EPPO Bulletin. The new and revised Standards published in the present issue of the EPPO Bulletin are PM 3/66 (3) Guidelines for the management of plant health risks of biowaste of plant origin, PM 3/77 (2) Vegetable plants for planting under protected conditions -Inspection of places of production, PM 3/81 (3) Inspection of consignments for Xylella fastidiosa, PM 3/82 (3) Inspection of places of production for Xylella fastidiosa, PM 3/94 (1) Raising professional operators' awareness of regulated and emerging plant pests. REFERENCES EPPO (2022) EPPO Standard PM 1/2 EPPO A1 and A2 lists of pests recommended for regulation in EPPO countries. In EPPO Standards: General phytosanitary measures. EPPO, Paris (FR). Available at https://www.eppo.int/media/uploaded_images/RESOURCES/eppo_standards/pm1/pm1-002-31-en_A1A2_2022.pdf [accessed on 1 October 2022] or in EPPO Global Database https://gd.eppo.int/standards/PM1/ [accessed on 1 October 2022]. FAO (2008) ISPM No. 31, Methodologies for Sampling of Consignments. IPPC Secretariat. FAO, Rome (IT). Available at https://www.ippc.int/static/media/files/publication/en/2016/11/ISPM_31_2008_Sampling_of_consignments_EN.pdf [accessed 1 November 2020]. IPPC (2019) Glossary of phytosanitary terms. ISPM no. 5. In International Standards for Phytosanitary Measures, pp. 41– 63. IPPC Secretariat, FAO, Rome (IT). Available at https://www.ippc.int/static/media/files/publication/en/2019/06/ISPM_05_2019_En_Glossary_2019-06-26_PostCPM-14-Fixed.pdf [accessed 1 November 2020]. Volume52, Issue3December 2022Pages 516-517 ReferencesRelatedInformation

Synchytrium endobioticum (Schilb.) Perc. is a soil-borne biotrophic fungus causing potato wart disease (PWD) of cultivated potato, one of the most important crops in Poland. S. endobioticum infects epidermal cells of young potato organs, such as eyes, sprouts, young tubers, stolons, stems, leaves, and even flowers, but never roots. S. endobioticum survives in the soil as winter (resting) spores, which germinate, infect the plant, and produce secondary sporangia (summer spores); infection results in galls on the stolons and tubers, in which the pathogen multiplies. Its long persistence in soil and the severe losses it inflicts to potato crops have prompted its inclusion into the A2 quarantine list. The fungus originates from the Andean zones of South America, from where it spread to North America and Europe at the end of the 19th century. S. endobioticum was first reported in the United Kingdom in 1876. This pathogen is quite variable. The first discovery of a new pathotype in Europe occurred in former East Germany in 1941, and there have been 38 S. endobioticum pathotypes identified so far. Pathotypes 1(D1), 2(G1), 6(O1), 8(F1), and 18(T1) are of greatest relevance in Europe. In 2008, the Polish Inspectorate of Plant Health and Seed Inspection (PIORiN) collected soil samples from Mazowieckie Region in Central Poland. Microscopic examinations revealed the presence of viable resting spores of S. endobioticum in a soil sample collected from a crop plantation of ornamentals intended for export. One kilogram of soil contained an average of 300 viable spores. A bioassay (pot tests), recommended by the EPPO standard PM 3/59 (1), showed no wart symptoms because of the very low sporangium density of S. endobioticum. However, concentrating S. endobioticum inoculum by centrifugation and using for a bioassay modified Potocek's tube test (1) allowed us to obtain fresh galls with summer sporangia. The first symptoms of PWD were visible on sprouts of extremely susceptible potato genotypes 7 weeks after inoculation. To identify the pathotype of S. endobioticum, 10 differential potato cultivars (Deodara, Tomensa, Eersteling, Producent, Combi, Saphir, Delcora, Miriam, Karolin, and Ulme) were inoculated with fresh galls of S. endobioticum using the Glynne-Lemmerzahl method (2), according to EPPO standard PM 7/28. Galls were formed on all cultivars except Saphir, Karolin, and Ulme (resistant cultivars). This virulence profile was identical to that of European pathotype 18(T1) of S. endobioticum. This is the first detection in Poland of pathotype 18(T1), which is one of the most virulent pathotypes of this fungus. It should be noted that in 2004, prior to planting, the field was investigated by PIORiN and found to be free of S. endobioticum. The winter sporangia were found on the field with ornamental plants originating from Western Europe, where pathotype 18(T1) is still occurring. S. endobioticum is a classic example of the distribution of plant pathogens by man. Although an infection source has not been determined, the field was probably infested by soil connected with roots of the plantlets. This is an example of alternative ways for S. endobioticum spreading without potato as a main host.

Background The potato spindle tuber viroid (PSTVd) is the smallest of all known potato pathogens. PSTVd is a circular, single-stranded RNA molecule that does not code for proteins. Symptoms caused by PSTVd lead to a significant reduction in tuber yield or death of the plant. PSTVd infection triggers the silencing of host-plant genes and induces disease symptoms in the plant via vd-sRNA. The StTCP23 potato gene which encodes a transcription factor is one of the targets of PSTVd. The aim of the study was to assess the effect of inoculation of potato plants of cv. ‘Colomba’ with the PSTVd viroid NicTr-3 strain on the StTCP23 expression in infected plants.Materials and methods. Potato plants of cv. ‘Colomba’ were inoculated with leaf sap of the tomato cv. ‘Rutgers’ in which the PSTVd viroid NicTr-3 strain was propagated. RT-PCR with specific primers was used to confirm the infection of potato plants with the viroid. Diversity of RNA molecules in the PSTVd population was revealed using RT, cloning, and sequencing of the viroid. The expression of the StTCP23 gene in infected potato plants was assessed by quantitative PCR with the ef1α gene as a reference.Results. Symptoms of infecting ‘Colomba’ plants with the PSTVd viroid strain NicTr-3 were not detected, while the presence of the viroid in potato leaves was confirmed by molecular methods. Ten mutations were found in individual variants of the PSTVd strain NicTr-3, responsible for reducing the viroid’s aggressiveness. An increase in the expression of StTCP23 was shown at 3 time points in ‘Colomba’ plants inoculated with the NicTr-3 strain compared to the control.Conclusion. The mechanism of interaction between PSTVd and the StTCP23 gene in an infected potato plant is not universal and depends both on the potato genotype and viroid strain. Additional studies are required to prove the existence of specific interaction mechanisms between the host-plant and viroid strain genotypes.

Multimeric non-radioactive cRNA probes improve detection of potato spindle tuber viroid (PSTVd)

During a survey conducted in October 2013 in tomato greenhouses in Diano Marina (Imola Province, northwest Italy), in a single greenhouse, unusual disease symptoms were observed in four out 1,400 (~0.3%) of tomato plants cv. Ingrid, grafted on 'Beaufort' rootstock. Symptoms including shortened apical internodes associated with tiny, deformed, and brittle chlorotic leaves, while ripe fruits appeared reduced in size and pale red. Samples of leaves from the four plants were collected and examined using commercial antisera (Bioreba AG, Reinach, Switzerland) by double antibody sandwich (DAS)-ELISA against Tomato spotted wilt virus, Cucumber mosaic virus, Alfalfa mosaic virus, Tomato/Tobacco mosaic viruses, and by indirect plate trapped antigen (PTA)-ELISA against potyviruses (potygroup test). None of the tested viruses were detected in the four leaf samples. In addition, PCR tests for begomoviruses and phytoplasmas were also negative. In a host range study, the original symptoms, consisting mainly of stunting and chlorosis, were reproduced within ~10 days in tomato seedlings (Momor line), mechanically inoculated at two true leaves stage with sap extract obtained from the four symptomatic tomato plants, whereas no symptoms were observed in Chenopodium amaranticolor, C. quinoa, Nicotiana tabacum (cv. Xanthi nc), N. glutinosa, or Phaseolus vulgaris (cv. Borlotto rosso). Total RNAs extracted with the RNeasy Plant Mini Kit (Qiagen, Hilden, Germany) from symptomatic samples were tested in RT-PCR using pospiviroids generic primers PSTVd-32/33 (1), designed to amplify the whole genome of Potato spindle tuber viroid (PSTVd), Tomato apical stunt viroid (TASVd), and Columnea latent viroid (CLVd), and CEVd-FW/RE primers, designed to amplify the whole genome of Citrus exocortis viroid (CEVd) and TASVd (3). Each of the four samples yielded amplicons of the same size (364 bp) with both primer combinations. The identity of the viroid was then determined by sequencing, on both strands, amplicons obtained from the four symptomatic plants at MWG (Ebersberg, Germany). Sequences obtained were identical, showing the highest nucleotide identity (99.7%) with the TASVd isolate Sj1 (AM777161), identified in Germany on Solanum jasminoides. The sequence was deposited in GenBank (Accession No. HG916812) and the field isolate named To1-IT. Two other cases of pospiviroid infection in tomato in Italy have been reported so far and the viroid species detected were PSTVd (2) and CLVd (4), respectively. To our knowledge, this is the first report of TASVd infecting tomato in Italy. The origin of this infection is still unclear, although based on the biological properties and sequence similarity, the To1-IT isolate probably originated from an ornamental species, most likely S. jasminoides, as recently reported for other tomato TASVd isolates, according to their biological and genetic features (5).

Different oligomeric forms of PSTV are detected in nuclei isolated from PSTV-infected potato cells by means of molecular hybridization, using as probes synthetic oligodeoxyribonucleotides with sequence specificity for (+)PSTV and for (-)PSTV. In addition to several species of longer-than-unit-length (-)PSTV molecules, two oligomeric forms of (+)PSTV are detected, which correspond in size to RNA strands of approximately two and three times viroid unit-length. They must be considered as the precursors of the circular and linear (+)PSTV monomers accumulating in the cell nucleus.

<i>Heterodera glycines:</i> procedures for official control

Specific scopeThis Standard describes a diagnostic protocol for adults ofEpitrix cucumeris,Epitrix papa, Epitrix subcrinitaandEpitrix tuberis.Specific approval and amendmentFirst approved in 2011‐09.Revision approved in 2016‐01.

In June 2017, balloon flower (Platycodon grandiflorus) plants with leaf mottling, etiolation, and mosaic symptoms were observed in Kunming, Yunnan province of southwestern China. These symptoms were similar to those induced by tomato spotted wilt virus (TSWV) (Fisher, 2013). Suspected TSWV infected plant samples, as well as thrips (Frankliniella occidentalis) in flowers were collected. Four of six symptomatic plants were positive using TSWV test strips (Adgen Biotechnology). Total RNA was extracted from four TSWV infected plants and 15 individual thrips and analyzed by two steps RT-PCR (PrimeScript™ II 1st Strand cDNA Synthesis Kit and LA Taq, Takara). Three primer sets were used to clone the S-RNA, and five primer pairs to clone the M-RNA as overlapping fragments (Marshall et al., 2017). The expected size fragments corresponding to TSWV S-RNA and M-RNA amplification were obtained from 4/4 leaf and 10/15 thrips samples. Amplified fragments from plants and thrips were cloned using pEASY-T1 Cloning Kit, sequenced and assembled, revealing that the S (GenBank accession No. MF688996) and M (MF688997) genome segments of this isolate shared nucleotide identity of > 95% with an isolate of TSWV from China (JF960235，JF960236; Hu et al., 2011), indicating that the sampled balloon flower plants were infected with TSWV. To our knowledge, this is the first report of TSWV infecting balloon flower in China. Due to the devastating effects of TSWV and the increase of balloon flower cultivation area in southwestern China, it is necessary to implement management strategies to control this disease and avoid the introduction of the virus into new production areas.

Potato spindle tuber viroid (PSTV), a small infectios RNA, has been completely digested with RNase T1 and RNase A, and the resulting oligonucleotides have been sequenced using 5'-terminal 32p-labelling with gamma-32p ATP and T4 polynucleotide kinase, fingerprinting and controlled nuclease P1 digestion. Modified nucleotides have not been detected in 5'-positions of these oligonucleotides. PSTV consists of about 359 nucleotides and contains a remarkable stretch of 18 purines, mainly adenosines; there is no AUG initiation triplet present. The established oligonucleotide sequences preclude a perfect intramolecular base complementarity within the covalently closed viroid circle. Therefore, the rigid, rod-like native secondary structure of PSTV, as seen in the electron microscope, must be based on a defective rather than on a homogeneous RNA helix. The detailed analysis of the bisulfite-catalized modification of cytidine to uridine in PSTV revealed a higher reactivity for the majority of the cytidines than would be expected for a perfect helix. Since only cytidines in single-stranded regions are knonw to be fully reactive, this finding provides additional evidence for defects in the helical secondary structure of PSTV.