GENOME CHARACTERIZATION AND DEVELOPMENT OF DIAGNOSTICS OF CITRUS YELLOW VEIN CLEARING VIRUS (CYVCV) ASSOCIATED WITH CITRUS IN INDIA

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Date
2017
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Citrus yellow vein clearing virus (CYVCV) belongs to genus Mandarivirus, produces conspicuous yellow vein clearing symptoms on Etrog citron and was first reported in 1997 in India. Our study indicated that it is not confined to Etrog citron and produces chlorotic irregular ring spots on other citrus species such as kinnow mandarin and sweet orange and mimic the symptom of Indian citrus ringspot virus (ICRSV). The virus was initially identified by the RT-PCR using degenerate primer designed from RdRp gene of the alphaflexiviruses. The complete genome sequence of four isolates of CYVCV (ECAI;_RMGI; PALI and KPMI) were determined following overlapping primers strategy and sequences of 5’ & 3’ ends was obtained by RACE. Genome of all the Indian isolates were of 7531 nucleotide long excluding the 3’poly (A) tail and comprises of six open reading frames. The phylogenetic analysis indicates that ICRSV is the most closely related virus and shares 72% overall sequencing identity at nucleotides level, while the CP gene shared identity of 73.8 to 74.2% nucleotide level and 80.7 to 81.3% amino acid level with the ICRSV K1 isolate. Based on molecular and serological features both the viruses are distinct but with regard of genome organization, virion particles and host range they shares similarity. Genetic distance among the Mandarivirus was 0.025+ 0.009 in Indian CYVCV isolates. The recombination analysis revealed stable nature of the genome and found some traces of recombination events. To gain insight into biodiversity and molecular variability of virus, surveys were conducted during 2012 to 2014 of citrus orchards in various geography of India. Forty isolates were examined and diverse symptoms, viz. irregular ring spots, yellowing of veins and vein lets, water soaking of veins, leaf distortion and mottling, necrosis, conspicuous ringspots, chlorotic spots and mixed infection governed mosaic like patterns on different citrus cultivars were observed. Some of these symptoms may be governed by mixed infection of viruses. Further, to determine molecular variability, coat protein was targeted in RT-PCR and from fifteen isolates the CP sequences of CYVCV were retrieved and submitted to NCBI database. The pair wise analysis of the coat protein sequences showed 96 to 99% homology with the CYVCV-ECAI isolate and 73% sequence identity with the ICRSV isolates. In phylogenetic tree all isolates of the CYVCV clustered together on
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