Occurrence and characterization of Little cherry virus-1 in Serbian sweet cherry and apricot trees

Abstract

Little cherry virus-1 (genus Velaravirus, family Closteroviridae; LChV-1) is considered an important pathogen of cherries worldwide. Together with Little cherry virus-2 (LChV-2) it is associated with little cherry disease. Infection of susceptible cherry cultivars with one or both viruses can lead to severe reduction of fruit quality and loss in yields. It is graft transmissible virus with unknown vector. Over the past twenty years, it has been found that other Prunus species are also hosts of the LChV-1, but mainly without visible symptoms. Since there is no data on the presence of LChV-1 on P. armeniaca and P. domestica in Serbia, except our preliminary results of its presence on P. avium and P. cerasus, the main goal of this study was to analyze the presence of the LChV-1 on these hosts and to characterize Serbian isolates by sequence and phylogenetic analyses. Occurrence of LChV-1 in sweet and sour cherries, apricot and plum was examined over two growing seasons (2017 and 2018). A total of 116 leaf samples were collected from orchards and nurseries of CAC planting materials, out of which 62 were sweet cherry, 16 sour cherry, 21 apricot and 16 plum samples. Samples comprising of 35 varieties and 3 unknown cultivars were collected during the summer season at 16 localities. Total RNAs extracted from the leaves were tested by RT-PCR using LChV-1 specific primers (LCV1U16390/LCV1L16809) targeting an ORF8 fragment. LChV-1 specific PCR product, DNA fragment of 419 bp, was detected in 11 out of 116 tested Prunus samples (9.48%). Nine of the 62 P. avium (14.52%) and two of the 21 P. armeniaca (9.52%) samples were infected with LChV-1. None of the tested P. cerasus and P. domestica was infected. PCR amplicons generated from one selected apricot (LC1) and five sweet cherry isolates (LC2-6), were bidirectionally sequenced (Macrogen, Amsterdam) and characterized. The nucleotide sequences of the 419 bp fragments confirmed the identity of the LChV-1 for all sequenced isolates LC1-6 (with sequences identity between 76.3% and 100%) and revealed a high genetic variability among the Serbian apricot and sweet cherry isolates (in the range of 22.1% to 23.9%). Serbian apricot LC1 isolate grouped together with other LChV-1 apricot isolates from the GenBank and showed 98.1% nt identity with two Czech isolates KX831095 and KX831097. Serbian cherry isolates shared sequence identities in the range from 90.9% to 100%, grouped in two separate groups. Isolates LC2 and LC5 (100% identical) and LC6 grouped together with cherry isolates from Bulgaria MG458879 (shearing 94.5% and 93.6% identity, respectively), sour cherry isolates from Poland HE580104 (98.9% and 97.8% identity) and Japanese isolate originated from P. serrulata MG934545 (98.1% and 97.1% identity). The other two cherry isolates LC3 i LC4 showing 99.2% nt identity were clustered in a separate subgroup within the group with already mentioned cherry isolates. Detection of LChV-1 in apricot and cherry imposes the need for phytosanitary control in the production of planting material. To the best of our knowledge, this is the first report of confirmed detection of LChV-1 on apricot and sweet cherry in Serbia.