Detection of ‘ ancestral ’ western lineage of Citrus tristeza virus virulent genotype in declining Arunachal Wakro orange

Eastern Himalayan foothills are known to have optimal agro-climatic conditions for the production of quality citrus fruits including oranges. Among the citrus-growing regions of eastern Himalayas, Wakro in the far eastern state of Arunachal Pradesh is known for its superior quality oranges, popular as the Wakro orange or the Arunachal orange , which has been included in the Geographical Indication Registry by the Government of India. However, during the last few years, Arunachal orange orchards have been severely affected by aphid infestation associated with rapid decline disease, causing catastrophe for the farmers as well as the state economy. Therefore, in 2015, an intensive survey of severely affected orange orchards was carried out to investigate the etiological factors of decline in citrus production. RNA samples extracted from leaf and aphid specimens collected from Wakro orchards were subjected to Citrus tristeza virus (CTV) detection through 3′-UTR specific RT-PCR. Subsequently, ORF1 and CP genetic regions were amplified and clonal-sequencing was performed. Although BLAST search showed close homology of the present sequences with other virulent genotype VT sequences, the detailed phylogenetic analysis demonstrated affinity and clustering of present sequences with VT sequences belonging to the ‘western’ lineage. This finding is considerably distinct from CTV sequences reported from citrus-growing orchards in India and other neighboring countries. Additionally, low diversity of CP gene sequences, recombination patterns, and presence of sequence segments identical to the present ones in other CTV genotypes was also revealed. Our findings suggest that the ancestral nature of present CTV sequences, corroborate well with the proposed origin of CTV in this part of the globe. We here report our finding of the western lineage of CTV virulent genotype VT, which is distinct from CTV molecular epidemiology in other parts of India, and discuss the implications of these findings.