ABSTRACT
The sensitivity of digoxigenin- (dig-) labeled polymerase chain reaction (PCR) was compared with nested PCR and enzyme-linked immunosorbent assay (ELISA) for the detection of the potato ring rot bacterium, Clavibacter michiganensis subsp. sepedonicus, in seed potatoes and stem tissues. The bacterial DNA was extracted from chopped tuber or stem tissue by a modified hot alkaline DNA extraction method. C. michiganensis subsp. sepedonicus specific DNA sequence was amplified by dig-labeled PCR with the primer pair CMSIF1/CMSIR1 previously designed based on the insertion element IS1121 of C. michiganensis subsp. sepedonicus. Dig-labeled PCR products were then blotted on a nylon membrane and the signal was detected by a colorimetric assay using alkaline phosphatase. The new assay procedure has a detection sensitivity close to that of nested PCR and simplicity of the standard ELISA procedure commonly used commercially. The dig-labeled PCR assay was more sensitive than ELISA and can be used to detect C. michiganensis subsp. sepedonicus in symptomless field potato tubers.
ABSTRACT
The potato spindle tuber disease was first observed early in the 20th century in the northeastern United States and shown, in 1971, to be incited by a viroid, potato spindle tuber viroid (PSTVd). No wild-plant PSTVd reservoirs have been identified; thus, the initial source of PSTVd infecting potatoes has remained a mystery. Several variants of a novel viroid, designated Mexican papita viroid (MPVd), have now been isolated from Solanum cardiophyllum Lindl. (papita güera, cimantli) plants growing wild in the Mexican state of Aguascalientes. MPVd's nucleotide sequence is most closely related to those of the tomato planta macho viroid (TPMVd) and PSTVd. From TPMVd, MPVd may be distinguished on the basis of biological properties, such as replication and symptom formation in certain differential hosts. Phylogenetic and ecological data indicate that MPVd and certain viroids now affecting crop plants, such as TPMVd, PSTVd, and possibly others, have a common ancestor. We hypothesize that commercial potatoes grown in the United States have become viroid-infected by chance transfer of MPVd or a similar viroid from endemically infected wild solanaceous plants imported from Mexico as germplasm, conceivably from plants known to have been introduced from Mexico to the United States late in the 19th century in efforts to identify genetic resistance to the potato late blight fungus, Phytophthora infestans.
