Molecular Analysis of New Crimean Isolates of the Plum Pox Virus
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Molecular Analysis of New Crimean Isolates of the Plum Pox Virus A. A. Shevelevaa, I. V. Mitrofanovab, V. M. Gorinab, and S. N. Chirkova,b,* a
Moscow State University, Moscow, 119234 Russia Botanical Garden, Yalta, 298648 Russia *e-mail: [email protected]
b Nikitsky
Received April 9, 2020; revised May 11, 2020; accepted May 15, 2020
Abstract—Molecular analysis of new isolates of the plum pox virus (PPV) found in the gene pool collections of peach, apricot, and plum from the Nikitsky Botanical Garden (National Science Center, Russian Academy of Sciences), as well as on wild cherry plum trees in the surrounding area, has been carried out. Using the enzyme-linked immunosorbent assay and reverse transcription polymerase chain reaction, 19 new PPV isolates have been detected. Most of them belonged to the strain D; single isolates of the strains Rec and M were also found. The 3'-terminal sequences of genome of the new isolates were sequenced, including the coat protein (CP) gene and adjacent sequences of the NIb gene and the 3'-nontranslated region. Phylogenetic analysis of these sequences revealed the distribution of PPV-D isolates into several clusters or clades. An average divergence rate between sequences of Crimean PPV-D isolates was 2.0%. A relatively high level of genetic variation indicates a long-standing introduction of the ancestors of the current PPV-D isolates to Nikitsky Botanical Garden (NBG). Isolate H23, identified in the apricot collection, belonged to strain M. This is the first detection of PPV-M in Crimea. Isolate AL-Ch (strain Rec) was found on wild cherry plum. In the CP of isolate K87 (strain D), the K59T mutation was detected, which prevents its recognition by PPV-D-specific 4DG5 monoclonal antibodies. In CP of AL-Ch and previously characterized isolates K28 and Kisl-1pl (strain Rec), amino acid substitutions were detected at position 68, which did not affect the recognition of these isolates by AL monoclonal antibodies. Apparently, position 68 is outside the PPV-M/Rec-specific epitope. The results of the work show that PPV most likely entered the NBG with contaminated drupaceous fruit material as a result of several independent introductions from different regions of the world and spread over plantations by aphids. Keywords: stone fruits, gene pool collections, plum pox virus, phylogenetic analysis, epitope, monoclonal antibodies DOI: 10.3103/S0096392520020078
INTRODUCTION Plum pox virus (PPV) causes a disease called sharka in stone fruits of the Prunus genus, which leads to significant losses in the yield of peach, apricot, plum, and other economically significant crops due to mass falling of the fruit, deterioration of their quality, and unsuitability for processing. Infection can inhibit the annual amount growth and shorten the productive life of the plant [1]. PPV belongs to the Potyvirus genus from the Potyviridae family. Virions consist of positive-sense single-stranded RNA molecule of 9800 nucleotides (nt) and approximately 2000 molecules of the membrane protein (CP).
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