(cid:2)reation of the virus free nursery plantings of the genus prunus l. by the phytosanitary selection

. Diagnostic of 12 viral pathogens of stone fruit crops, including new to the certification scheme ( Carnation ringspot tombusvirus (CRSV), Myrobalan latent ringspot nepovirus (MLRSV), Petunia asteroid mosaic tombusvirus (PeAMV) , Tobacco ringspot nepovirus (TRSV )) by immune-enzyme analysis was carried out in the Institute for fruit growing in 2020 – 2021. Virus-free clones of varieties and rootstocks forms of stone fruit crops have been identified to create Nuclear stock collection, including: 10 varieties of plums, 3 - cherry plums, 7 - sour cherries, 14 - sweet cherries, 5 - apricots, 4 - peaches, 10 forms of rootstocks of stone crops. Varieties of peach and apricot, as well as rootstocks of these crops, for the creation of Nuclear stock collection were generated in 2021 in Belarus for the first time.


Introduction
Establishment and preservation of healthy repositories in countries with an actively developing fruit growing industry is an important component of the international program of agricultural plant variety improvement. Obviously, the traditional movement of germplasm of perennial vegetatively propagated crops, entails high risks of spreading pathogens with host plants.
Especially, this applies to the pathogens with a long latent period, as well as asymptomatic ones, such as viruses and phytoplasmas, which are in a special risk group. Phytosanitary control of plant diseases from the EPPO A1 and A2 List of pests recommended for regulation as quarantine pests, is the main reason for establishing nuclear stock collections in all countries with high horticulture and developing documents on the maintenance of such collections, their replication, retesting.
The basis and the most interesting part of the Nuclear stock collection in specific horticultural areas are commercial, old local varieties, as well as perspective hybrids and forms. Increased attention is currently paid to the preservation of old local cultivars of plants (landrace), which have acquired their distinctive features as a result of many years of cultivation and selection in a particular area, as well as varieties of their own breeding. These genotypes are only grown in certain areas and require special conservation measures. Before plants are included in the basic collections, they are subjected to a number of tests: In 2018, the total area of seedlings of stone fruit crops in Belarus is 0.89 ha, including: cherry plum, plum, apricot -0.58 ha, cherry, cherry -0.31 ha. Planting material produced less than 150,000 pieces. Almost no Virus free seedlings are produced.
The most common and economically significant viruses of stone fruit crops are Prune dwarf virus (PDV) and Prunus necrotic ringspot ilarvirus (PNRSV), causing yield reduction of up to 100% in some commercial varieties, especially with complex infection [1,2].
In Bulgaria in 2019, infection with PDV of cherries and sweet cherry trees averaged 14.4%. 92.4% of trees were diagnosed with PDV alone, while mixed infection with PNRSV was noted for 7.6%. Only 56.2 % of the trees were visually symptomatic [4].
In Czech Republic, 10 viruses (out of 19 tested) were found in cherry and sweet cherry trees ApMV and LChV-2 were diagnosed for the first time. CGRMV and CNRMV were isolated only from imported plants in collection plantations [5,6].
In recent years, the spread of Cherry detrimental canker has been established in Northern Bavaria. Trees of all cherry cultivars grafted on Prunus avium rootstocks were affected. Petunia asteroid mosaic virus (PAMV) was detected by serological testing in diseased trees [7]. The virus appears on cherry plants as necrosis of shoot tips and necrotic spots on leaf veins and fruits. Earlier, this disease was described in other countries as Cherry detrimental canker. Petunia asteroid mosaic tombusvirus (PAMV) -tombusviruses have isometric particles about 30 nm in diameter, the RNA genome of the virus is singlestranded, 180 protein subunits, and a molecular weight of about 40,000. Cells infected with tombusviruses contain characteristic multivesicular bodies in their cytoplasm [8,9].
In 1987, another virus, Carnation Italian Ringworm Virus (CIRV), was found on cherry plants with similar symptoms. This was the first report of this virus in woody plants. The virus causes the same symptoms as PAMV, but the effect on the tree is less severe. The distribution of PAMV and CIRV virus particles has been found to be highly irregular. Using ELISA, viruses could be reliably detected only in symptomatic plant tissue samples. Therefore, indexing latent PAMV or CIRV infections in cherry trees is currently not possible. Since virus detection correlates significantly with the occurrence of symptoms, PAMV and CIRV are considered to be the causative agent of viral twig necrosis In 1990, Bud blight disease (BBD) was first reported on cherry in Japan. Symptoms of the disease include browning and death of buds, irregular brown spots and chlorotic mottling on leaves, and bark necrosis [11]. Significant yield reduction is the main outcome of the disease. The use of RT-PCR diagnosis made it possible to identify a complex of 5  [12 -14]. A large proportion of cherry trees in Japan has been found to be infected with at least one of these viruses [15]. Prune dwarf virus (PDV) and/or Apple chlorotic leaf spot virus (ACLSV) can also cause Bud blight disease [16].
Studies on the spread of viral pathogens aim to analyze the phytosanitary situation in plantations and to create pathogen-free plants for further replication.
The aim of the research is to create stone fruit cultivars and rootstocks for the creation nuclear stock collection in accordance with EPPO recommendations and "Unified phytosanitary quarantine requirements to quarantinable products and quarantinable items at the customs border and customs territory of the Eurasian Economic Union". ELISA testing of viruses was used to diagnose pathogens [17].

Results and discussion
Allocation of initial plants of promising cultivars and rootstocks for further testing was carried out in the departments of biotechnology, nursery and fruit crops breeding of the Institute for fruit growing. Visually healthy plants with typical cultivars features, without symptoms of viral and virus-like diseases were selected based on the results of plantation inspection in the spring period. The selected plants in previous years were characterized by good indicators of winter hardiness, general condition of plants, productivity, resistance to fungal diseases.
During the pomological evaluation of cultivars, the following features were taken into account: crown (shape, density); shoot (length, thickness, growth character, pubescence, length of internodes); leaf blade (size, shape, base, tip, serration, pubescence); fruit (size, shape, surface, skin color, color and consistency of pulp). During pomological characterization of the rootstock, the following characters were taken into account: mother bush (shape, growth strength); shoot (bark color, thickness, curvature, pubescence); bud (base, size, bud position relative to shoot); leaf blade (apex, base, leaf position relative to shoot, serration); petiole length; leaf stalk shape; anthocyanin color of shoot tip; internodes length.
Pomological   presumably affected by PRMV showed an excess of optical density for samples isolated only from lignified shoots. Samples from leaves and annual shoots are reliably free of the virus. Due to the ambiguity of the results, only clones with an optical density of about to negative control were distinguished as Nuclear stock. For Nuclear stock collection have been determined virus free clones from the cultivars: (Donetsk bely, Iskra, Alex, Zolotoy Jubilee) and rootstock for peach -Vesennee Plamya.
Virus-free graft material is stored on rootstocks (1660 pcs.) for laying a new rotation of the repository of stone fruit crops.

Conclusion
Diagnosis of viral pathogens (PeAMV, CRSV, MLRSV, PRV, ToRSV) new to the certification scheme in Belarus was carried out for the first time. Reliable infection with viruses of initial plants of stone fruit cultivars and rootstocks was not established by ELISA method.
Virus-free clones of cultivars and rootstock of stone fruit crops have been identified to create Nuclear stock collection in accordance with EPPO recommendations and "Unified phytosanitary quarantine requirements to quarantinable products and quarantinable items at the customs border and customs territory of the Eurasian Economic Union", including: - -cultivars peaches (Donetsk bely, Iskra, Alex, Zolotoy Jubilee) and rootstock Vesennee Plamya, free from ACLSV, ApMV, PPV, PNRSV, PRMV, PDV, TomRSV, TRSV, CRSV, PeAMV, MLRV.