وحید رومی

Turkey is the motherland and first producing country of sweet cherries in the world. It is also one of the largest exporters of sweet cherries and accounts for 20% of the global export market. To date, 44 viruses and three viroids have been described in the 9 main cultivated Prunus species. Seven of these viruses and one viroid have been newly identified in Prunus hosts within the last 5 years thanks to the application of high throughput sequencing technologies (HTS) to detect, identify and quantify known or novel viruses in one step. It has thus been proved to be a sensitive method for detecting putative infectious agents associated with host tissues. Therefore, HTS offers new opportunities for deep characterisation of Prunus viromes. In this study two cherry samples presenting virus-like symptoms but negative for presence of Prune dwarf virus (PDV), Prunus necrotic ringspot virus (PNRSV) and Apple chlorotic leaf spot virus (ACLSV) by DAS-ELISA and RT-PCR were analysed by HTS. Total RNAs were isolated by different protocols including some commercial kits and also homemade protocols with some modifications. The purity and concentration of the total RNA preparations were assessed with a NanoDrop spectrophotometer and the integrity and size distribution of purified RNAs were also evaluated with a Bioanalyzer. The NGS run was performed on an Illumina Nextseq 500 platform yielding a total number of 4.3 and 4.2 million reads (2x150 nt) for sample 1 and 2, respectively. De novo assembly was performed using Geneious (Biomatters Ltd., Auckland, New Zealand) software. The resulting contigs were used for BLAST searches against viral database at NCBI. Mapping of RNAs against the reference genomes of viruses detected in de novo assembly was performed using the Geneious software. The Blastn, Blastx and TBlastx analyses of contigs belong to two different samples gave the same results and confirmed the presence of several viruses in cherries like Cherry rusty mottle-associated vir