Performance of diagnostic tests for the detection and identification of Pseudomonas syringae pv. actinidiae (Psa) from woody samples |
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Authors: | Stefania Loreti Amandine Cunty Nicoletta Pucci Aude Chabirand Emilio Stefani Adela Abelleira Giorgio M. Balestra Deirdre A. Cornish Francesca Gaffuri Davide Giovanardi Richard A. Gottsberger Maria Holeva Aynur Karahan Charikleia D. Karafla Angelo Mazzaglia Robert Taylor Leonor Cruz Maria M. Lopez Joel L. Vanneste Françoise Poliakoff |
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Affiliation: | 1.Consiglio per la Ricerca in agricoltura e l’analisi dell’Economia Agraria-Centro di Difesa e Certificazione - CREA-DC,Roma,Italy;2.Plant Health Laboratory (LSV), French National Agency for Food,Environmental and Occupational Health & Safety (ANSES),Angers,France;3.Unit for Tropical Pests and Diseases, Plant Health Laboratory (LSV), French National Agency for Food,Environmental and Occupational Health & Safety (ANSES),Saint- Pierre,France;4.Department of Life Sciences,Università degli Studi di Modena e Reggio Emilia - UNIMORE,Reggio Emilia,Italy;5.Deputación de Pontevedra, Estación Fitopatolóxica Areeiro,Pontevedra,Spain;6.Department of Science and Technology for Agriculture, Forestry, Nature and Energy,Università ‘La Tuscia’ di Viterbo - DAFNE,Viterbo,Italy;7.The New Zealand Institute for Plant & Food Research Private Bag 3230, Waikato Mail Centre, Hamilton, 3240, New Zealand Physical Address,Hamilton,New Zealand;8.Laboratorio Fitopatologico Regione Lombardia,Vertemate con Minoprio (CO),Italy;9.Institute for Sustainable Plant Production Department for Molecular Diagnostics of Plant Diseases Spargelfeldstr,Austrian Agency for Health and Food Safety (AGES),Wien,Austria;10.Department of Phytopathology, Laboratory of Bacteriology,Benaki Phytopathological Institute (BPI),Attica,Greece;11.Plant Protection Central Research Institute (PPCRI) Gayret Mah,06172 Yenimahalle, Ankara,Turkey;12.Plant Health & Environment Laboratory, Diagnostic and Surveillance Services, Biosecurity New Zealand,Ministry for Primary Industries, (MPI-PHEL),Auckland,New Zealand;13.Instituto Nacional de Investiga??o Agrária e Veterinária (INIAV)UEIS-SAFSV Laboratório de Fitobacteriologia,Oeiras,Portugal;14.Istituto Valenciano de Investigaciones Agrarias,IVIA Centro de Proteccion Vegetal y Biotechnologia Carretera Moncada,Moncada (Valencia),Spain |
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Abstract: | The aim of this study was to characterise the performance of new molecular methods for the detection and identification of Pseudomonas syringae pv. actinidiae (Psa) and to provide validation data in comparison to the assays mentioned in official diagnostic protocols and being currently used. Eleven molecular tests for the Psa detection were compared in an inter-laboratory comparison where each laboratory had to analyse the same panel of samples consisting of thirteen Psa-spiked kiwifruit wood extracts. Laboratories had to perform also isolation from the wood extracts. Data from this interlaboratory test performance study (TPS) was statistically analysed to assess the performance of each method. In order to provide complete validation data, both for detection and identification, this TPS was supplemented by a further study of identification from pure culture of phylogenetically closely related Pseudomonas spp., Psa, and bacterial strains associated with kiwifruit. The results of both these studies showed that simplex-PCRs gave good results, whereas duplex-PCR and real-time PCR were the most reliable tools for detection and identification of Psa. Nested and multiplex-PCR gave false-positive results. The use of the most reliable detection test is suggested for routine analyses, but when Psa-free status needs to be accurately assessed, it is recommended that at least two detection tests are used. This work provides a wide comparison of the available diagnostic methods, giving new information for a possible revision of the official diagnostic protocols (e.g. European and Mediterranean Plant Protection Organization (EPPO) protocol PM7/120 for the detection of Psa). |
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