Related Experiment Video
Updated: May 21, 2026
Bioassay-Guided Identification of Natural Products for Biocontrol by Thin Layer Chromatography-Direct Bioautography
Published on: July 26, 2024
Construction of a bio-test for infecting red clover plants with Sclerotinia trifoliorum
T Vleugels1, J Baert, E Van Bockstaele
1Institute for Agricultural and Fisheries Research (ILVO), Plant Sciences Unit, Caritasstraat 21, BE-9090 Melle, Belgium. tim.vleugels@ilvo.vlaanderen.be
Abstract:
Sclerotinia trifoliorum causes clover cancer in red clover crops. Clover cancer is difficult to control and completely resistant red clover varieties are not available. Breeding for resistant red clover varieties is being slowed down because little is known about the diversity of European S. trifoliorum populations and because of the lack of bio-tests that are useable in breeding programs. The first objective of this research was to develop a reliable high-throughput bio-test, useable in breeding programs. The second objective was to optimise another bio-test, based on isolated leaves, for more precise studies. First, we optimised a method for ascospore production of S. trifoliorum. Once produced, the ascospores were used to evaluate the effects of climate conditions, ascospore concentration and plant age on the high-throughput bio-test. For the bio-test on isolated leaves, the effects of infection method, incubation conditions, incubation period, ascospore concentration, leaf growth stage and mechanical damage were evaluated. In the high-throughput bio-test, disease levels rose with increasing ascospore concentration up to 20,000 spores/ml. The plant age had a small, yet significant effect on the disease level. For the isolated leaf bio-test, the most effective and most repeatable infection method was spraying of an ascospore suspension. Disease levels continued to increase with rising concentrations and incubation time did not interact with plant susceptibility levels. The youngest completely opened leaf yielded the most repeatable results. Both bio-tests were shown to be correlated and could be valuable instruments for breeding programs and for studying plant-pathogen interactions.
Insights
Researchers developed two reliable bio-tests to assess resistance to clover cancer, caused by Sclerotinia trifoliorum. These methods will aid breeding programs in developing resistant red clover varieties.
Area of Science:
- Plant Pathology
- Agricultural Science
- Genetics
Background:
- Sclerotinia trifoliorum infection causes significant crop losses in red clover.
- Current red clover varieties lack complete resistance to clover cancer.
- Developing resistant varieties is hindered by limited knowledge of pathogen diversity and lack of effective bio-assays.
Purpose of the Study:
- To develop a reliable high-throughput bio-test for Sclerotinia trifoliorum suitable for plant breeding programs.
- To optimize an isolated leaf bio-test for precise studies on clover cancer development.
- To improve breeding strategies for red clover resistance.
Main Methods:
- Optimized ascospore production for Sclerotinia trifoliorum.
- Evaluated climate conditions, ascospore concentration, and plant age in a high-throughput bio-test.
- Assessed infection method, incubation, concentration, leaf stage, and damage in an isolated leaf bio-test.
Main Results:
- High-throughput bio-test showed increased disease with ascospore concentration up to 20,000 spores/ml; plant age had a minor effect.
- Spraying ascospore suspension on the youngest fully opened leaves provided the most repeatable results in the isolated leaf bio-test.
- Both bio-tests demonstrated correlation and effectiveness.
Conclusions:
- Developed and validated two distinct bio-tests for Sclerotinia trifoliorum.
- These bio-tests are valuable tools for accelerating red clover breeding programs.
- The optimized methods will facilitate further research into plant-pathogen interactions and resistance mechanisms.
