Sensitivity of field tests, serological and molecular techniques for Plum Pox Virus detection in various tissues

Sensitivity of field tests (AgriStrip  and Immunochromato), DAS-ELISA, two step RT-PCR and real-time RT-PCR for <em>Plum pox virus</em> (PPV) detection was tested in various tissues of apricot, peach, plum and damson plum trees infected with isolates belonging to PPV-D, PPV-M or PPV-Rec, the three strains present in Slovenia. Flowers of apricot and plum in full bloom proved to be a very good source for detection of PPV. PPV could be detected with all tested techniques in symptomatic parts of leaves in May and with one exception even in the beginning of August, but it was not detected in asymptomatic leaves using field tests, DAS-ELISA and partly also molecular techniques. PPV was detected only in some of the samples of asymptomatic parts of the leaves with symptoms and of stalks by field tests and DAS-ELISA. Infections were not detected in buds in August using field tests or DAS-ELISA. Field tests are useful for confirmation of the PPV infection in symptomatic leaves, but in tissues without symptoms DAS-ELISA should be combined or replaced by molecular techniques

First report of black raspberry necrosis virus and raspberry leaf mottle virus in Bosnia and Herzegovina

Disease Note on the first report of black raspberry necrosis virus (BRNV) and raspberry leaf mottle virus (RLMV) by high throughput sequencing in raspberry plants in Bosnia and Herzegovina.Peer reviewe

Comparative study of diagnostic methods used for monitoring of common grape vine (Vitis vinifera L.) crown gall (Agrobacterium vitis Ophel & Kerr) in Slovenia

Agrobacterium vitis causes common grape vine (Vitis vinifera L.) crown gall disease that destroyed a lot of Slovenian vineyards more than a decade ago. Eighty isolates of Agrobacterium spp. collected during monitoring in 2006 were identified as A. vitis and A. tumefacies by pehA and multiplex PCR method. Tumor-inducing capacity of these strains was assessed on test plants and with PCR methods for detection of the Ti plasmid responsible for tumor induction. With VCF3/VCR3 primer pair six false negatives and no false positives were detected. The high genetic diversity of pathogenic Agrobacterium spp. strains affects the performance of molecular methods, thus biological test should be performed where results from molecular methods are doubtful

Physiological response of grapevine Vitis vinifera L. to grapevine leafroll associated viruses (GLRaV-1 and GLRaV-1 + GLRaV-3)

Grapevine leafroll disease is one of the most severe viral diseases of grapevine caused by Grapevine leafroll-associated viruses (GLRaVs). Physiological processes were monitored on grapevines with single (GLRaV-1) and mixed (GLRaV-1 and GLRaV -3) viral infection under greenhouse conditions from June to September, in vegetation period 2014. In the mid of the season (July) negative effects of the virus infections on physiological processes were more severe in mixed than in single infection. The net-photosynthesis (Pn) of the leaves infected with GLRaV-1 and GLRaV-3 reached only a half of the Pn in GLRaV-1 infected grapevines. Similar reduction was found for stomatal conductance, transpiration and parameters related to photochemical efficiency (electron transport rate)

Inter-laboratory comparison of four RT-PCR based methods for the generic detection of pospiviroids in tomato leaves and seeds

Four pospiviroid detection methods consisting of a pair of RT-PCR (ANSES 1-2), a pair of real time RT-PCR (Botermans 1-2) and two single RTPCR methods (Luigi and Olivier) were evaluated for their relative accuracy, diagnostic sensitivity, diagnostic specificity, analytical sensitivity and reproducibility through a comparison in eight laboratories. All detection methods were tested on 13 tomato leaf and nine tomato seed samples as well as on 100-fold dilutions of the corresponding RNA extracts. Two different RNA extraction kits and three combinations of RNA extraction and RT-PCR kits were assessed. According to the statistical analysis of the results, ANSES 1-2 method provided the best performance criteria whatever the matrix consisted of, while the relative accuracies of Botermans 1-2 and Olivier methods were not significantly different from ANSES 1-2 for seed and leaf samples respectively. The results also showed that the relative accuracy of the Luigi and Olivier methods significantly increased when primer concentrations were raised to 1 μM, the latter method giving the best relative accuracy of the test in these conditions. No statistical differences were observed between accuracies obtained for the two extraction kits or the three combinations of extraction and RTPCR kits used. ANSES 1-2 method is proposed as international standard for the generic detection of pospiviroids on tomato leaves and seed