Rapid Real-time Viral Load Estimation Technique for chilli leaf curl virus and its validation in different chilli genotypes from Eastern Himalayan Plains.

Abstract

Abstract Chilli leaf curl virus is one of the most devastating virus infecting chilli crops in India. Management of chilli leaf curl disease largely relies on early detection and quantification of the virus. In the present study different Open Reading Frames from chilli leaf curl virus from sub-Himalayan Terai region were cloned, sequenced and submitted in NCBI database with accession number MN851261, MN857412, and MN857413. Comparison of these gene sequences with previously reports revealed that chilli leaf curl virus coochbehar strain exhibits 90–92% similarity with tomato leaf curl joydebpur virus and pepper leaf curl Bangladesh virus. Using these sequence primers were designed from the unique AV2 region of the chilli leaf curl virus coochbehar strain and a SYBR based Rapid Real-time Viral Load estimation (ReViLeR) technique was developed to quantify the virus directly from extracts of infected leaf samples. Seventeen chilli genotypes were evaluated for virus accumulation with ‘ReViLeR’ method after challenge inoculation with chilli leaf curl virus. Traditional landraces like Chuapara, Line boya and White chilli were found to have highest viral titer (36659, 22909 and 25195 viral copies per genomic unit (GU) respectively). On the other hand, from the genotypes like Micro, Pusa Sadabahar, Dalle Khursani no virus was detected. Higher viral load in the susceptible genotypes manifested severe leaf curl symptoms whereas resistant genotypes with no detectable viral load remained healthy. The sensitivity of the newly developed ReViLeR technique was found up to 83% in rapid detection of chilli leaf curl virus.