Production and characterization of a monoclonal antibody specific to the M serotype of plum pox potyvirus

A monoclonal antibody to an Albanian isolate of plum pox potyvirus (PPV) was obtained (MAbAL), that specifically recognized strain M of this virus. The specificity of MAbAL, assessed by comparative ELISA on 130 PPV isolates of different geographical origin, 22 of which were also tested by comparative IC-PCR, gave consistent and highly reproducible results. MAbAL seems to be elicited by a stable surface determinant that makes it particularly suitable for successful use under a wide range of conditions. MAbAL is an useful addition to the panel of PPV-specific MAbs available to date.     

Susceptibility to sharka (Plum pox virus) in Prunus mandshurica×P. armeniaca seedlings

Prunus mandshurica [(Maxim.) Koehne] from Central Asia is a species related to apricot and grown in China and Mongolia. This species has been used in apricot breeding as a source of frost resistance. In addition, P. mandshurica has been suggested as the possible origin of some North American apricot cultivars resistant to sharka (Plum pox virus, PPV). The aim of this work was to transmit the resistance to PPV from P. mandshurica to the Spanish apricot cultivar ‘Currot’ by traditional crossing. The resistance to a Dideron PPV isolate of the descendants of P. mandshuricaבCurrot’ and their progenitors was evaluated under controlled conditions in a greenhouse. The results showed the susceptibility of both, the progenitors and the offspring to PPV, as being much higher than in other apricot seedlings. The results showed that the P. mandshurica accession studied is not a good progenitor in breeding apricot for PPV resistance, but just the opposite. The possible role of P. mandshurica as a source of resistance in apricot resistant cultivars is questioned.     

Evaluation of resistance to sharka (Plum pox virus) of several Prunus rootstocks

Sharka (Plum pox virus, PPV) severely affects the production of most Prunus species in the areas affected by the disease. In this study, the resistance of 15 Prunus rootstocks to a Dideron type isolate of PPV was evaluated under controlled conditions in an insect‐proof greenhouse. After four cycles of study,‘GF677’ almond x peach hybrid,‘Myrobolan 29C plum and ‘L2’ cherry did not show any symptoms and were ELISA‐DASI and RT‐PCR negative. These were considered resistant to PPV. The rest of the rootstocks assayed showed symptoms of sharka (confirmed by ELISA‐DASI or RT‐PCR), although the level of susceptibility was different for each rootstock.‘GF305’ peach, ‘Puebla de Soto’ plum and ‘Real Fino’ apricot, were highly susceptible to PPV, showing strong sharka symptoms and being ELISA and RT‐PCR positive. ‘Marianna 2624’ plum,‘AC 9921‐07’ hybrid and ‘CP‐2’ plum showed susceptibility to PPV confirmed by ELISA‐DASI and RT‐PCR positives.‘Nemaguard’ and ‘Nemared’ hybrids, ‘Torinel’ plum and ‘STN2’ hybrid showed an intermediate susceptibility to PPV with slight sharka symptoms and were ELISA‐DASI and RT‐PCR positive. Whereas, ‘Montclar’ peach and ‘Evrica’ hybrid showed moderate resistance to PPV with slight sharka symptoms and were ELISA positive but RT‐PCR negative. The results open new possibilities in the search for different sources of resistance to PPV within Prunus.     

Inheritance of resistance to plum pox potyvirus (PPV) in 'Stella' apricot seedlings

Resistance to sharka (plum pox potyvirus) was studied in 74 seedlings resulting from open pollination of the resistant apricot cultivar ‘Stella’. Each seedling was inoculated at the base by a chip-bud from a diseased GF305 peach tree. To follow tlie spread of virus into the scedlings. a Chip-bud of a healthy GF305 and another from a healthy sensitive Manicot apricot were grafted above the inoculation point in the order described. Six observations of symptoms were made in the leaves of the GF305 used For inoculation, the GF305-control, the Manicot-control and the ‘Stella’ seedlings during four growth cycles. An enzyme-linked immunosorbent assay (ELISA) was applied to the leaves of the ‘Stella’ seedlings. No ‘Stella’ seedling showed symptoms of sharka. Only three seedlings reacted positively to the ELISA. Substantial differences were observed in the speed of virus propagation through the ‘Stella’ seedlings, some of them showing a strong resistance to virus translocation. The possible use of the ‘Stella’ cultivar in apricot breeding programmes to obtain resistant cultivars is discussed.     

Genetic diversity and molecular epidemiology of the T strain of Plum pox virus

Very limited information is available on the origin, diversity and evolution of Plum pox virus (PPV) ‘Turkey’ (T) strain. Phylogenetic analyses based on partial sequences of 421 isolates and complete genome sequences of 57 isolates, representing the geographical distribution of PPV-T in Turkey, revealed the existence of several monophyletic and, in some cases, geographically limited groups within the PPV-T strain (Ankara-Konya1-Kayseri, Ankara-Balkan, Istanbul, Konya2 and Balkan). PPV-T diversity (0.018%) was found to be greater than that of PPV strains D and Rec but lower than that of the M strain when including the newly described and divergent M-Istanbul isolates, suggesting a long evolutionary history for PPV-T. The European part of Turkey in the Balkans, close to Bulgaria where PPV was identified for the first time, appears as a likely centre of origin for PPV-T isolates. The colonization of various parts of Turkey by diverse isolates from that region, followed by secondary local spread, is the most likely scenario for the diffusion of PPV-T in Turkey.     

Resistance of almond cultivars to Plum pox virus (sharka)

Sharka, a disease caused by Plum pox virus (PPV), mainly affects some Prunus species, including apricot, peach and plum, and to a lesser degree, sweet cherry and sour cherry. In almond, different PPV isolates have been transmitted experimentally to the ‘Aï’ cultivar. In this study, the resistance of 10 almond cultivars to a Dideron PPV isolate was evaluated in controlled conditions by grafting the cultivars on to inoculated GF305 peach rootstocks. The results demonstrated a high level of resistance to PPV in all the almond cultivars assayed. They did not show any symptoms and were ELISA and RT‐PCR negative, despite the strong symptoms observed in their GF305 rootstocks. The implications of these results for the dispersion of PPV, and the potential role of almond as a source of resistance to PPV in other Prunus species such as peach, are also discussed.     

Evaluation of resistance of apricot cultivars to a Spanish isolate of plum pox potyvirus (PPV)

The resistance of 12 apricot cultivars to the Dideron type Spanish isolate RB3.30 of plum pox potyvirus was evaluated in controlled conditions in an insect‐proof greenhouse. The results of the evaluation demonstrated the resistance of the cultivars ‘Stella’, ‘Stark Early Orange’, ‘Goldrich’, ‘Harcot’, ‘NJA2’, ‘Pandora’ and ‘Avilara’ to the isolate, and the susceptibility of the cultivars ‘A1453’, ‘Bulida’,‘Currot’, ‘Real Fino’ and ‘Canino’     

Resistance to Plum pox virus in plants expressing cytosolic and nuclear single‐chain antibodies against the viral RNA NIb replicase

The expression of engineered single‐chain variable fragments specific to the NIb RNA replicase of Plum pox virus (PPV) (scFv2A) in transgenic plants was successfully used as a strategy to interfere with viral infection. Different scFv2A fusion proteins were constructed to target those subcellular compartments, such as the cytosol, endoplasmic reticulum (ER) membrane structures and the nucleus, where NIb protein presumably accumulates. Several transgenic lines of Nicotiana benthamiana plants expressing the scFv2A targeted to the cytosol (2A lines), ER (6K2 lines) and nucleus (NLS lines) were obtained. The protective effect of scFv expression was determined by mechanical virus inoculation in five 2A, three 6K2 and four NLS transgenic lines. The strongest resistance was afforded with the 2A‐3 (six non‐infected plants out of 10), 6K2‐1 (17 out of 33) and NLS‐11 (16 out of 19) transgenic lines. The success of this interference with PPV infection opens new possibilities for the control of this RNA virus and could be exploited not only to confer resistance in transgenic plants, but also to elucidate the role of the non‐structural NIb protein in different cell compartments during viral infection.     

Diversity of Plum pox virus isolates in Bosnia and Herzegovina

Sixteen Plum pox virus (PPV) isolates from several stone fruit cultivars, host species, orchards and geographical areas of Bosnia and Herzegovina were selected for typing, using serotype-specific monoclonal antibodies (MAbs) and PCR–RFLP, targeting the 3' terminal region of the coat protein (CP) and P3-6K₁ with restriction enzymes Rsa I and Dde I. Four PPV isolates were identified as PPV-M by serology and PCR; eight isolates were identified as PPV-D based on PCR–RFLP on both genomic regions, but were not recognized by the D-specific MAb4DG5. Four isolates from plum were identified as natural D/M recombinants (PPV-Rec), based on conflicting results of CP and P3-6K₁ typing. To investigate the genetic diversity of Bosnian PPV isolates in more detail, five isolates (three PPV-Rec, one PPV-M and one PPV-D) were partially sequenced in the region spanning the 3' terminal part of the NIb gene and the 5'-terminal part of the CP gene, corresponding to nucleotides 8056–8884. Nucleotide sequence alignment of recombinant isolates showed that they were closely related at the molecular level to previously characterized recombinants from other European countries, and shared the same recombination break point in the 3' terminal part of the NIb gene. This is the first report of naturally infected Prunus trees with PPV-M, PPV-D and PPV-Rec in Bosnia and Herzegovina. The high variability of the Bosnian PPV isolates fits with the presence of this virus in the country over a long period.