Biological and Molecular Characterization of Begomoviruses from Pothwar Region of Pakistan

Abstract

Begomoviruses (family Geminiviridae; genus Begomovirus) are dicot plantsinfecting viruses which are exclusively transmitted by the insect vector whitefly (Bemesia tabaci). These viruses belong to economically important group of viruses which have monopartite and bipartite single stranded DNA genome. The plant diseases induced by begomoviruses are major constraints for successful crop production worldwide. Pothwar region is located in the northern part of Punjab province with an estimated area of 22, 254 km2. Based on the ecological and climatic conditions Pothwar region is adapted for the production of large number of agricultural and horticultural crops. The lack of systematic study on begomoviruses diversity in Pothwar region gave a strong backing to current study for assessing the incidence, distribution and genetic characterization of begomoviruses in the region. A total of 138 plant samples belonging to 17 different plant families were collected from four districts of Pothwar region and capital territory Islamabad. Total genomic DNA was isolated from all plant samples by CTAB method followed by PCR employing universal degenerate primers for begomoviruses detection. Selected positive samples were subjected to Rolling Circle Amplification (RCA) for full genome amplification of DNA-A, DNA-B and satellite genomes which were cloned and sequenced subsequently. The obtained sequences were analyzed using Sequence Demarcation Tool (SDT) and phylogenetic analysis was carried out by Molecular Evolutionary Genetics Analysis (MEGA) 6.0. Plants belonging to 15 families were tested positive for begomoviruses and the overall incidence of begomoviruses in Pothwar region was xxii 77.5 percent while incidence in individual areas was 85.7 percent in Attock, 85 percent (Rawalpindi), 83.3 percent (Islamabad), 81 percent (Chakwal) and 55.5 percent (Jhelum). The begomoviruses were not evenly distributed in Pothwar region with 32 percent in Rawalpindi, 27 percent (Chakwal), 22.4 percent (Attock), 14 percent (Islamabad) and 05 percent (Jhelum). The most significant contribution of the present study is the identification and characterization of new begomovirus specie which was tentatively named as “Euphorbia yellow leaf curl virus” (EYLCV) [Pakistan:Rawalpindi:Euphorbia:2012] [KM978186]. The EYLCV was also isolated and characterized from Hibiscus syriacas and Combretum indicum. Other begomoviruses which were characterized in present study are Mesta yellow vein mosaic virus (MeYVMV) and its association with Cotton leaf curl Multan Betasatellite (CLCuMB) infecting Malvastrum coromandelianum, Rose leaf curl virus (RoLCuV) infecting Duranta repens, Tomato leaf curl New Dehli Virus infecting Solanum lycopersicum and Chilli leaf curl virus and its associated betasatellite infecting Capsicum annum from Pothwar region. The Pothwar region provides an opportunity for helper viruses to develop more complex relationships with satellites DNAs altogether opening a window for the development of unique begomoviruses disease complexes with likely implication of invading the cotton belt that could result in severe economic losses to national economy.