Overexpression of ghr-miR166b generates resistance against Bemisia tabaci infestation in Gossypium hirsutum plants

In silico identified Gossypium hirsutum ghr-miR166b shows multi-compatible targets in mitochondrial ATP synthase of Bemisia tabaci. Its overexpression in planta has the potential to act as a biopesticide in reducing B. tabaci population, and consequently the spread of whitefly-transmitted plant viruses. Whiteflies (B. tabaci) are hemipterous insects that act as a vector to transmit plant viruses causing enormous losses to the plants. In the present study, G. hirsutum-encoded miRNAs targeting expressed sequence tags (ESTs) of B. tabaci, based on sequence complimentarity and miRNA-target mRNA thermodynamics, were in silico identified. Out of 108 G. hirsutum miRNAs, 55 targeted the protein encoding ESTs. Among them, ghr-miR166b was selected owing to its intrinsic affinity for ATP synthase. Its functional role was validated following expression of ghr-MIR166b (precursor) sequence in G. hirsutum cv. HS6 plants through Agrobacterium-mediated transformation. Total of seven independent transformed (T-0) G. hirsutum lines were obtained. The transcript level of ghr-MIR166b in the transgenic lines was observed to be 2.0- to 17-fold higher as compared to non-transformed plants. Northern-blot analysis of small RNAs isolated from the transgenic plants confirmed the presence of the ghr-miR166b. After feeding on the leaves of transgenic line (HS6-166-30) having highest level of ghr-miR166b expression, B. tabaci population was reduced up to 91% as compared to non-transformed leaves. Further, in the whole plant assay, a maximum of 78% B. tabaci mortality was observed in the same line, while there was an increase in B. tabaci population on the non-transformed plants. Our results revealed that ghr-miR166b supposedly targeting ATP synthase gene of B. tabaci, and subsequently its overexpression in planta has potential to act as biopesticide for reducing B. tabaci population and consequently spread of whitefly transmitted viruses.