Potato spindle tuber viroid eradicated in Western Australia
Potato spindle tuber viroid (PSTVd) was detected for the f irst time in an isolated hydroponic greenhouse tomato (Lycopersicon esculenturn) erop in the great southern region of Western Australia (WA). An urgent eradication plan was activated because of the highly contagious nature of the pathogen. The tomato erop on the infected property was destroyed and the greenhouses and surrounding area were decontaminated. Surveys of 15 other hydroponic tomato crops did not detect any other PSTVd infected tomato plants in WA.
Analysis of Sequence Polymorphism and Population Structure of Tomato chlorotic dwarf viroid and Potato spindle tuber viroid in Viroid-Infected Tomato Plants
The sequence polymorphism and population structure of Tomato chlorotic dwarf viroid (TCDVd) (isolate Trust) and Potato tuber spindle viroid (PSTVd) (isolate FN) in tomato plants were investigated. Of the 9 and 35 TCDVd clones sequenced from 2 different TCDVd-infected plants, 2 and 4 sequence variants were identified, respectively, leading to a total of 4 sequence variants of 360 nucleotides in length. Variant I was identical to AF162131, the first TCDVd sequence to be reported, and the rest exhibited 1 to 3 nucleotide differences, all in the TR domain, from AF162131/variant I. Of the 33 and 29 PSTVd clones sequenced from 2 different PSTVd-infected plants, 8 and 9 sequence variants were found, respectively, leading to a total of 15 variants ranging in length from 356 to 359 nucleotides. The variant I was identical to EF044303, a PSTVd reported in Russia. The rest exhibited 1 to 11 nucleotide differences scattering in all five domains from EF044303/variant I. The results demonstrated for the first time that TCDVd, like many other viroids including PSTVd, exists in host plants as a collective group comprised of various sequence variants. However, in comparison to PSTVd, TCDVd is less polymorphic in tomato plants as fewer variants and lower haplotype/nucleotide diversities were observed.
Improved RNA Extraction from Woody Plants for the Detection of Viral Pathogens by Reverse Transcription-Polymerase Chain Reaction.
An efficient procedure for the extraction of high-quality RNA from woody plants without the use of phenol, organic solvents, or alcohol precipitation is described. The method employs commercially available spin-column matrices and mitigates the inhibitory effects of plant polysaccharides and polyphenolic compounds commonly observed on subsequent polymerase chain reaction amplification when conventional extraction methods are applied to woody plant species. The method described has been successfully used in the development of highly sensitive reverse transcription-polymerase chain reaction (RT-PCR) techniques for the detection of a number of viruses in their woody hosts. The viruses detected included apple stem grooving capillovirus (ASGV), apple stem pitting virus, Prunus necrotic ringspot ilarvirus (PNRSV), grapevine fanleaf and Arabis mosaic nepoviruses, and grapevine leafroll-associated closterovirus type 3. The method described was equally effective for the extraction of viral RNA from either budwood, leaves, or flower blossoms as determined by the equivalent RT-PCR detection of ASGV and PNRSV from these tissues. Detection of viral RNA in samples of total plant RNA prepared using this method was found to be as sensitive as was previously described for the immunocapture RT-PCR technique.