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Projects / Programmes source: ARIS

Development of a real-time PCR assay for the detection of fruit, grapevine and hop viroids

Research activity

Code Science Field Subfield
4.03.00  Biotechnical sciences  Plant production   

Code Science Field
B390  Biomedical sciences  Phytotechny, horticulture, crop protection, phytopathology 
Keywords
viroid, real-time PCR, detection, fruit crops, grapevine, hop
Evaluation (rules)
source: COBISS
Researchers (5)
no. Code Name and surname Research area Role Period No. of publicationsNo. of publications
1.  07632  PhD Minka Kovač  Biochemistry and molecular biology  Researcher  2007 - 2009  111 
2.  15489  PhD Irena Mavrič Pleško  Plant production  Head  2007 - 2009  374 
3.  20162  PhD Sebastjan Radišek  Plant production  Researcher  2007 - 2009  606 
4.  22582  PhD Nataša Toplak  Biotechnology  Researcher  2007 - 2009  168 
5.  03853  PhD Mojca Viršček Marn  Plant production  Researcher  2007 - 2009  419 
Organisations (3)
no. Code Research organisation City Registration number No. of publicationsNo. of publications
1.  0401  Agricultural institute of Slovenia  Ljubljana  5055431  20,096 
2.  0416  Slovenian Institute of Hop Research and Brewing  Žalec  5051762000  4,214 
3.  1775  OMEGA svetovanje, inženiring, razvoj in raziskovanje d.o.o. (Slovene)  Ljubljana  5808642  293 
Abstract
Viroids are important plant pathogens which infect different vegetable, ornamental and perennial crops. Some viroids are of big economical importance and the others are important because of possible and known synergistic effects when present in the plant together with other viroids or viruses. The viroids in Slovenia are not well studied. Most of the studies were made on viroids of perennial crops using classical detection methods for viroid detection – hybridisation (dot blot), reverse polyacrilamide gel electrophoresis (R-PAGE) and RT-PCR. Hop stunt viroid (HSVd) and Peach latent mosaic viroid (PLMVd) were found to be present in stone fruits and Hop latent viroid (HLVd) was found in hop. Real-time PCR (qPCR) is a new and powerful tool for detection and identification of plant pathogens and all other pathogens. In frame of the proposed project we will develop qPCR methods for detection of different viroids which infect perennial crops using specific detection by TaqMan chemistry. By development of qPCR assays for different viroids we will be able to detect not only different viroids by running the same test but we will also improve the sensitivity of the detection as well as allowing the quantification. The results will be available in a very short time and by combining different assays on the same plate the costs for the test would be greatly reduced. The results of the research would be of great value also for other researchers and diagnosticians in Slovenia and in the world because no reports about the qPCR detection of viroids except for PSTVd were published until now. Additionally, developed qPCR methods will be available for routine use in sanitary selection and certification programmes.
Significance for science
The available information about viroid detection using RT-qPCR is limited only to detection of Potato spindle tuber viroid - PSTVd. Further RT-qPCR methods for detection of pospiviroids in vegetables and ornamentals were developed in frame of Euphresco project 'Detection and epidemiology of pospiviroids', where Agricultural Institute of Slovenia was involved as a partner. The project ended at the end of 2009. During our project the RT-qPCR methods were developed for detection and identification of Hop stunt viroid (HSVd), Hop latent viroid (HLVd), Grapevine yellow speckle 1 viroid (GYSVd-1), Grapevine yellow speckle 2 viroid (GYSVd-2) and Peach latent mosaic viroid (PLMVd). To our knowledge these are the first RT-qPCR methods for detection of these viroids. The developed methods were already presented at two conferences and the results will be available to researchers and experts after publication in international journals. The experience obtained during the project will be valuable in future when the same techniques will be developed for detection, identification and quantification of other plant pathogens.
Significance for the country
The RT-qPCR methods for detection of viroids, developed in frame of the project and future developments of similar detection methods are of great importance for all their users. They will enable higher sensitivity and reproducibility of results and simultaneous detection of different pathogens due to the universal assay parameters used. The methods will be used in diagnostics of plant pathogens, certification and breeding programes. The costs of pathogen diagnosi will be lower and the results will be available in shorter time. The development of new methods for detection of plant pathogens will contribute to broader knowledge about viroids and to faster and economical viroid detection. They were shown to be more sensitive and could replace the RT-PCR methods for detection of HLVd in hop and HSVd and PLMVd in stone fruits which are used at present. More samples can be analysed using the new methods and they could be used for monitoring and survey of viroids. This will help us to improve the health status of domestic and foreign plant propagation material and higher quality of fruit, grapevine and hop production.
Most important scientific results Annual report 2008, final report, complete report on dLib.si
Most important socioeconomically and culturally relevant results Annual report 2008, final report, complete report on dLib.si
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