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Inter-laboratory evaluation of a duplex RT-PCR method using crude extracts for the simultaneous detection of Prune dwarf virus and Prunus necrotic ringspot virus |
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| Authors: | S Massart Y Brostaux L Barbarossa V César M Cieslinska O Dutrecq F Fonseca R Guillem A Laviña A Olmos S Steyer T Wetzel J Kummert M H Jijakli |
| Institution: | 1. Plant Pathology Unit, Gembloux Agricultural University (FUSAGx), Passage des Déportés, 2, 5030, Gembloux, Belgium 2. Unité de Statistique, Informatique et Mathématique appliquées (SIMa), FUSAGx, Gembloux, Belgium 3. CNR-Istituto di Virologia Vegetale-Sezione di Bari, Via Amendola 165/A, 70126, Bari, Italy 4. Corder ASBL, Unité de Phytopathologie, Croix du Sud, 2/3, 1348, Louvain-La-Neuve, Belgium 5. Virology Laboratory, Research Institute of Pomology and Floriculture, Pomologiczna 18, 96-100, Skierniewice, Poland 6. DNAlis sprl, rue de la Croix-Rouge, 20, 5032, Gembloux, Belgium 7. Molecular Plant Virology Lab, University of Algarve, CDCTPV, Campus de Gambelas, Gambelas, 8005-139, Faro, Portugal 8. Service Régional de la protection des végétaux de la DRAF Aquitaine, 51 rue Kiéser, 33077, Bordeaux, France 9. Deparmento de Protecció Vegetal, Institut de Recerca i Tecnologia Agroalimentaries, Ctra. de Cabrils s/n, 08348, Cabrils, Barcelona, Spain 10. Instituto Valenciano de Investigaciones Agrarias (IVIA), Carretera de Moncada a Náquera km 5, 46113, Moncada, Valencia, Spain 11. Département Lutte biologique et Ressources phytogénétiques, Centre Wallon de Recherches Agronomiques, Rue de Liroux 4, 5030, Gembloux, Belgium 12. Institute for Plant Research, RLP Agroscience, AlPlanta, Breitenweg 71, 67435, Neustadt an der Weinstrasse, Germany |
| Abstract: | The operational capacity of a duplex RT-PCR method for simultaneous detection of Prune dwarf virus (PDV) and Prunus necrotic ringspot virus (PNRSV) has been established by nine European laboratories. A total of 576 samples from Prunus trees with known sanitary status, corresponding to 32 samples in two repetitions for each laboratory, were analysed. The level of sensitivity achieved by the method was 98.3% for PDV and 90.4% for PNRSV. The specificity was 87.4% for PDV and 94.3% for PNRSV. The unilateral 95% confidence intervals were calculated for all these values. Cohen’s Kappa coefficient of repeatability and reproducibility of the technique indicated a strong agreement between data. Likelihood ratios were 7.50 (positive) and 0.02 (negative) for PDV. For PNRSV, the positive likelihood ratio was 15.00 while the negative likelihood ratio was 0.11. In addition, post-test probabilities of infection were calculated to manage the risk associated with the routine use of this method. This allows an accurate test result interpretation to facilitate the integration of this new technique into a certification scheme. |
| Keywords: | Diagnostic Inter-laboratory Plant virus Ring-test |
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