Projects / Programmes
Epidemiology and Biodiversity Studies of Plant Pathogens
| Code |
Science |
Field |
Subfield |
| 4.03.05 |
Biotechnical sciences |
Plant production |
Phytomedicine |
| Code |
Science |
Field |
| B390 |
Biomedical sciences |
Phytotechny, horticulture, crop protection, phytopathology |
| Code |
Science |
Field |
| 4.01 |
Agricultural and Veterinary Sciences |
Agriculture, Forestry and Fisheries |
plant pathogens biodiversity, epidemiology,molecular diagnostic methods, new generation sequencing, LAMP, immunoqPCR, CIM, grapevine, potato, PVY, FD, RBDV
Researchers (29)
Organisations (4)
Abstract
Due to the globalization-powered free movement of goods and services, we are exposed to an intensified exchange (import/export) of plants and plant origin products. With them new microorganisms and insects or other vectors fof microbial plant pathogens, are travelling, which, can survive in new warmer areas. New emerging diseases constitute a major threat to agricultural production and forestry, but also present a threat to natural ecosystems and biodiversity. In addition to the emerging of new diseases, which are often difficult to identify, another problem is the genetic variation of already known pathogens. In response to this, the diagnosis of new pathogens or new genetic variants of already known microbes has become an extremely hot topic. New methods for determining the nucleotide sequence (next generation sequencing - NGS) provide new opportunities for pathogens identification.
Viruses occur in the plant as a set of different mutants (quasispecies). After entering a new host, the viral genome can change, or the ratio between the different mutants of the same virus can be altered. Quick mutations arising from replication of viruses give rise to more aggressive isolates that can spread faster and infect a wider range of hosts. Such examples are found in potato with the infection with different Potato virus Y (PVY) virus variants, that threatens the production of potato. Apart from PVY, we will focus on pathogens that infect grapevine. Pathogens of woody plants are problematic as they represent a permanent infection on plants that are grown for many years, and therefore their presence should be detected sooner in order to remove infected plants. Phytoplasma Flavescence Dorée is quarantine pathogen with great economic impact on grapevine production and Raspberry Bushy Dwarf Virus (RBDV) is a newly discovered virus infecting grapevine and its diversity and the importance for the grapevine and original host is still unknown. Determination of genetic traits by NGS and development of new diagnostic methods, such as isothermal amplification (LAMP), high resolution melting analysis (HRM) and immuno real-time PCR (immuno-qPCR), which will be used and developed during the project, will allow rapid detection of plant pathogens. It will enable faster and more reliable discovery of new microbes/pathogens and will contribute to understanding the diversity of quarantine and harmful already identified microbes. The new approach to microbe identification and detection will enable tracking of the route of pathogen introduction and distribution.
Determination of genetic diversity that NGS allows will help to reveal the differences that are important for pathogenicity. In the case of plant viruses one of the bottleneck is a sample preparation in order to make easier the analyzis of the huge amount of data generated by the sequencing, which is non selective. We will introduce a completely original way of preparing samples of virus infected plants for the sequencing of viruses using new generation monolithic chromatographic polymers (CIM).
The involvement of exellent national and international partners, using modern facilities allows us to obtain relevant results, useful both for co-financers (Ministry of Agriculture and environment and the company Omega) and for crop production improvements. The proposed studies belong to the European strategy, which emphasizes the genomics and biotechnology for health, food quality and sustainable development. The project is fully harmonized with the priorities of this public call for proposals – Plant Production.
Significance for science
The introduction of plant pathogens into new environments can decrease the agricultural output of said environemtns and have a negative impact on the biodiversity of local ecosystems. With the help of new methods for the detection and research of plant pathogens on the molecular scale, which we have been developing during this project, we can achieve a more rapid detection of an invasion by novel pathogens. In addition we can study how novel pathogens emerge in the context of changing ecological conditions in different environments. We have also succcessfully used NGS technology in the search for new viruses (viruses in berries). Simultaneously, we have developed a fast and simple workflow for the analysis of NGS results in the context of searching for (new) viruses. In collaboration with other excellent laboratories in this field we have developed framework for the use of this technology for the detection and pursuit of new viruses. The optimisation of protocols for sample preparation with CIM monolithic chromatography before NGS enabled a more efficient assembly of complete viral genomes, and have also gave us a very detailed view into the diversity of the viral populations in the plant. During the project we have established a reliable approach for the study of viral populations (on the case of Potato virus Y) and performed a very exact study of the changes that take place in viral populations during their adaptation to different environments. The results of this part of the project have given us an important view into the mechanisms of viral genome evolution, which broadened our understanding of the processes involved in the emergence of new viral pathogens. The new fast and sensitive diagnostic methods which we have developed and/or validated during the project (LAMP, HRM, immuno-qPCR) allow for a faster and more efficient detection of important plant pathogens and a faster and more exact identification of said pathogens.
Significance for the country
In the context of the project we have carried out relevant and modern studies in the field of plant virology, which facilitated new connections with excellent laboratories in the field as well as our inclusion in international projects related to the topic of the project. We have successfully introduced diagnostic methods for plant viruses based on NGS in Slovenia and developed a procedure for the analysis and discovery of viruses in plant samples, which we are already successfully transporting into the workflow of our diagnostics laboratories. Both the introduction of new generic methods (NGS) as well as the introduction of modern rapid specific methods (LAMP, HRM, immuno-qPCR) for the detection of plant pathogenic microbes allowed for a more efficient and technologically advanced way of working in the field of diagnostics. Using the methods which were introduced during the project we have already discovered new viruses in plants in Slovenia, including previously unknown viruses. We have shared the results and findings of the project to other researchers and the interested public in Slovenia through seminars and lectures (Seminar on plant genomics, lecture in the context of The Symposium of the Slovenian Microbiological Society, public lectures etc.)
Most important scientific results
Annual report
2013,
2014,
2015,
final report
Most important socioeconomically and culturally relevant results
Annual report
2013,
2014,
2015,
final report
