بررسی فیلوژنیکی جدایه های ویروس سوختگی سیاه برگ چغندرقند ایران و دنیا بر اساس ناحیه 3'UTR

Beet black scorch virus (BBSV) is a positive-strand RNA virus belonging to the genus Betanecrovirus in the family Tombusviridae and it is one of the new sugar beet soil-borne viruses in Iran. BBSV genome consists of a 3644 nucleotides single-stranded RNA without a 5′ cap structure nor 3′ polyA tail. But at the end of 3′UTR is a structure that is involved in translating and replicating the virus. The virus has been identified from most sugar beet growing areas in Iran, especially Khorasan province. In this study, sugar beet root samples were collected from different sugar beet cultivation areas of Khorasan, and then Total RNA was extracted using RNeasy Plant Mini Kit. The virus was detected in samples of Mashhad, Fariman, Chenaran, Jolgeh Rokh, Shirvan, and Jovein sugarbeet farms using reverse transcription-polymerase chain reaction. cDNA synthesis of virus isolates was achieved using the Easy cDNA Synthesis Kit. The Taq DNA Polymerase Master Mix Red kit and specific forward and reverse primers were used to amplify 315 nucleotide fragments by a polymerase chain reaction. Amplification fragments of the isolates were cloned into the pTG19-T plasmid vector. The recombinant plasmids were then transfected into Escherichia coli strain DH5α. The 3'UTR of six different isolates was sequenced following cloning and then submitted to NCBI. To study the genetic diversity, the sequences obtained in this study were aligned with isolates registered in NCBI from Iran and the world after sequencing using Geneious Prime, 2019 software. Phylogenetic tree and grouping of these nucleotide sequences Phylogenetic relationships of isolated Iranian and non-Iranian isolates were implemented using MEGA-X software. The percentage similarity matrix of the sequences was designed using Sequence Demarcation Tool software and the nucleotide similarity percentage was calculated with Geneious Prime, 2019 software. Phylogenetic analysis based on nucleotide sequences of the 3'UTR region showed that the entire world isolates could be classified into two main groups II and I and each one was divided into two subgroups. Isolates of this study were grouped with 23 isolates from Iran in subgroup IA and Four isolates from Khorasan province were placed in the IB subgroup. Four Iranian BBSV isolates from the West were grouped in the subgroup IIA. All Chinese isolates and the European isolates together with the USA isolate clustered in subgroup IIB. The results of comparing the average nucleotide identity of 3'UTR isolates in this study showed that they are identical to Iranian isolates in the range from 97.75% to 97.98%. Nevertheless, they had the least identity with Kermanshah isolates (92.52%). These results show that there is genetic diversity in this region of the virus genome in Iranian isolates.