This project aims to develop PCR-based detection methods to detect (viable) ArMV and/or TRV in nematodes isolated from soil samples derived from flower bulb, and nursery stock production fields.
Additionally, a serotyping method will be developed for TRV. In the third year of the project the methods will be used to validate current guidelines for reliable sampling of soil.
Arabis mosaic virus (ArMB, nepo virus) and Tobacco rattle virus (TRV, tobra virus) have a broad host range and cause high economical losses in both the flower bulb industry as in the nursery stock industry. Nepo- and tobra viruses are transmitted by Xiphinema species and Paratrhochodorus and Trichodorus species, respectively. Virus-free plants can get diseased when grown on soil contaminated with virus-infected nematodes. The ability to investigate the virus/nematode status of the soil, allows growers to monitor the quality of the soil prior to planting their valuable plant material. For TRV it is an additional value to know the viability and the serotype status of the virus.
The project has started with the construction of a knowledge chart in which aspects that are critical for a successful and reliable result are displayed. The scheme describes the complexity of the method and will be important for knowledge transfer to testing agencies at the end of the project. To obtain viruliferous nematodes and soil samples, a successful model system has been set up for single and multiple nematode virus transmission experiments. Additionally, a PT-project on ArMV epidemiology supplied multiple soil samples of which it was known that ArMV-containing nematodes were present and virus transmission to plants had occurred.
The Naktuinbouw and the Bloembollenkeuringsdienst are currently involved in the project supervision and it was decided that soil sampling and nematode isolation will be carried out according to the Blgg standards. During the first year, a successful DNA-extraction and PCR method was established for the specific detection of Xiphinema nematodes in a population of extracted nematodes. Additionally, a successful RNA-extraction and RT-PCR method was established for the specific detection of ArMV in nematodes. It also appears that ArMV is already PCR-detectable in a few (six) viruliferous nematodes.
For Trichodores identical RNA extraction methods can be used as was developed for ArMV in Xiphinema. So far TRV is not detected in Trichodores nematodes. Partly because the sampled TRV infected Hosta plot contained a very low number of nematodes (2 nematodes per 100 ml soil). The last months of 2009 was worked on the detection of TRV in weeds. TRV could be detected in roots of Stellaria media.
Collaboration with other projects
In October 2009 a 4-year (PT-financed) project will start which aims to optimize an horticultural system to generate virus-free nursery stock plants. Approximately 30 growers will participate in this project and will supply soil samples where TRV and/or ArMV infections have been found. With the initiation of this additional project, this technology-based BO-project will be supplemented in the following years with important agricultural soil samples in order to further optimize and validate the methods.