Construction of a multiprobe for the simultaneous detection of viroids infecting citrus trees

Infections with different viroid species are common among cultivated fruit trees and grapevines, and many old-clone citrus varieties contain up to five citrus viroids (CVds) within a single tree. This paper describes the construction of a CVd-Multiprobe consisting of full-length clones of Hop stunt viroid, Citrus exocortis viroid, Citrus bent leaf viroid and CVd-III. The CVd-Multiprobe was tested against RNA transcripts of the four viroids and RNA extracts from plants singly infected with CEVd or HSVd or multiply infected with different CVds. The viroids were effectively diagnosed with the DIG labeled CVd-Multiprobe when tested by Northern hybridization or dot blot analyses. The CVd-Multiprobe does not provide information on the specific viroid resulting in a positive signal. However, this should not be considered as a problem, since most citrus certification programs will discard budwood source trees infected with any of the known CVds.