First record of a Hop stunt viroid variant on Nagpur mandarin and Mosambi sweet orange trees on rough lemon and Rangpur lime rootstocks

Citrus rootstocks play a very significant role in tree health and productivity. The sub-viral pathogens, viroids pose a major threat to trees and susceptible rootstocks as they bring about slow decline (Ahlawat, 1997). In a survey of citrus orchards in certain parts of Central India, symptoms of bark scaling, bark splitting and leaf yellowing were observed on Nagpur mandarin (C. reticulata) (7 plants) and mosambi (C. sinensis) (10 plants) grafted on rough lemon and Rangpur lime rootstocks (Fig. 1). Following grafting of scions from all these plants onto Etrog Citron (C. medica), an indicator host for Citrus exocortis viroid (CEVd), an unusual symptom of reduced size and yellowing of newly emerging leaves developed. This prompted further investigation.

Electrophoresis under denaturing conditions of total nucleic acid extracted from the grafted Etrog citron plants with symptoms, indicated the presence of viroid-like low molecular weight (LMW) RNA in all the samples. These LMW-RNAs were used in RT-PCR using a set of citrus viroid specific primers (Ramachandran et al., 2003). Amplicons of ~300 bp were obtained in all the samples with primers specific to Hop stunt viroid (HSVd), while no amplification was observed with CEVd specific primers. Cloning and sequencing of these amplicons identified a viroid RNA (Acc. No. AY237168) with 100% sequence similarity to HSVd-Ca905 (Acc. No. AF131250; Reanwarakorn & Semancik, 1999) and HSVd-ycv (Acc. No. AJ490824; Roy & Ramachandran, 2003). This constitutes the first isolation of a HSVd isolate (HSVd-RL) from citrus rootstocks showing bark scaling and splitting symptoms in Central India. This result suggests that HSVd is also a component in producing bark-scaling symptoms in rootstocks, which were previously considered to be due to CEVd-infection alone. This report indicates the need for the testing of citrus plants for HSVd in budwood certification and germplasm evaluation.