Complex mixtures of viroids identified in the two main citrus growing areas of Iran

The citrus industry in Iran is based on graft propagation of local and imported cultivars on a limited number of rootstocks such as sour orange, Mexican lime and Bakraee, a local natural hybrid. Infection of the available cultivars with graft-transmissible agents, viroids in particular, may have deleterious effects on the productivity of citrus and, more importantly, limits the choice of rootstocks. Rootstocks other than sour orange and Mexican lime will be needed as a consequence of the introduction and spread of Citrus tristeza virus (CTV) in the country. In the present study, two cv. Sanguinello sweet orange and two cv. Duncan grapefruit trees were selected on the basis of the bark scaling symptoms observed on the trifoliate orange and Troyer citrange rootstocks. Analysis by sPAGE and Northern blot hybridization of nucleic acid preparations of Etrog citrons graft-inoculated with these sources revealed the presence of several viroids: Citrus exocortis viroid (CEVd) and Hop stunt viroid (HSVd), the causal agents of exocortis and cachexia, respectively, as well as Citrus bent leaf viroid (CBLVd), Citrus dwarfing viroid (CDVd) and Citrus bark cracking viroid (CBCVd). These viroids were sequenced and compared with the corresponding reference sequences. To extend these results, samples from different citrus cultivars were collected in the two main citrus growing regions and analyzed for viroids. Of the 49 samples tested, 22 were viroid-positive, witnessing the occurrence of cultivars infected with CEVd, HSVd, CBLVd, CDVd, or CBCVd as mixed infections in the surveyed regions. One sample was also co-infected with the newly described Citrus viroid V (CVd-V)