Determination of the Dominant Variants of Hop Stunt Viroid in Two Different Cachexia Isolates from North and South of Iran

Citrus plants are hosts of several viroid species, among which, pathogenic variants of Hop Stunt Viroid (HSVd) induce citrus cachexia disease. Stunting, chlorosis, gumming of the bark, stem pitting and decline are symptoms of cachexia in mandarins and their hybrids as susceptible hosts. Based on the pathogenic properties on citrus, HSVd variants are divided in two distinct groups: those that are symptomless on sensitive citrus host species and those that induce cachexia disease. In this study, two cachexia isolates were selected and biological indexing was performed in a controlled temperature greenhouse (40ºC day and 28ºC night) using Etrog citron (Citrus medica) grafted on Rough lemon (C. jhambiri), as a common indicator for citrus viroids. The plants were inoculated with the inocula from a severe symptomatic tree of a newly declining orchard of Jiroft, Kerman province and a mild symptomatic tree from Mazandaran province. Presence of HSVd was confirmed with sPAGE, Hybridization by DIG-labeled probes and RT-PCR using specific primers of HSVd. Primary and secondary structures of the isolates were studied. The consensus sequence of RT-PCR amplicons of the severe isolate (JX430796) presented 97% identity with the reference sequence of a IIb variant of HSVd (AF213501) and an Iranian isolate of the viroid (GQ923783) deposited in the gene bank. The mild isolate (JX430798) presented 100% homology with the HSVd-IIc variant previously reported from Iran (GQ923784). Both isolates were shown to be cachexia inducing according to their sizes, sequences and lack of “non-cachexia expression motif” structures.