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چکیده
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Seven viroids naturally infect the domesticated apple (Malus × domestica Borkh.) and up to now, apple scar skin viroid, apple dimple fruit viroid, and hop stunt viroid have been reported from Iran. To determine which other viroids are present, three pooled samples were subjected to high-throughput sequencing. Each pooled sample contained RNA extracted of leaf samples of ten symptomatic and asymptomatic individual trees collected from different fields from a geographic area. Sequencing performed on an Illumina HiSeq 3000 (2 × 150 nt) by GenWiz (Leipzig, Germany), resulting in ca. 47 million of 150 nt pair-end reads. BlastX analysis showed that apple hammerhead viroid (AHVd) is present. To determine the distribution of AHVd, 137 collected samples from the west and northwest regions of Iran, were tested by RT-PCR using specific primers (Chiumenti et al. 2019). Amplicon with expected size ( ̴ 435 bp) was amplified from nine samples (6.56%) of both regions, indicating the viroid is distributed in Iran. The infected trees exhibited shoot decline and dieback symptoms, loss of apical dominance, radial limb cracking, and small fruits. Although it was not possible to associate any of these symptoms to AHVd-infection because at least one of viruses infecting apple were also detected in the AHVd-infected trees (data not shown). One selected AHVd isolate (HT2) was sequenced confirming AHVd infection. The genome sequence obtained was deposited in NCBI-GenBank under accession numbers PQ273267 had the highest nucleotide sequence identity of 98.05 % (100 % query coverage) to isolate PG_OK15_06 from Canada. AHVd was first reported in apples in China (Zhang et al., 2014) and subsequently it was reported from Africa, Europe, North America, Central and East of Asia, and Oceania (Szostek et al., 2018). To our best knowledge, this is the first report of AHVd infecting apple trees in Iran and Middle East.
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