Location: Sunflower and Plant Biology Research2012 Annual Report
1a. Objectives (from AD-416):
1. Determine genetic variation of S. sclerotiorum from crops grown outside the North Central region using DNA fingerprinting and mycelial compatibility groups. 2. Characterize the virulence of selected isolates on cultivars of soybean, sunflower, canola, dry bean, pea and lentil. 3. Prepare a set of isolates representing the genetic and virulence diversity of S. sclerotiorum in the United States for distribution to researchers on Sclerotinia.
1b. Approach (from AD-416):
Sclerotinia sclerotiorum is one of the most important pathogens of field crops in the North Central USA, causing economic losses in the numerous susceptible crops grown in the northern tier of states. The diseases caused by this pathogen have been very difficult to control. In a project currently funded by the Sclerotinia initiative, 146 isolates of the pathogen from the North Central USA were characterized using mycelia compatibility groups (MCG’s) and DNA fingerprinting. We are currently evaluating the virulence of those isolates on 6 crops grown in the region. We identified 43 MCG’s and the DNA fingerprints of the isolates were strongly associated with MCG’s. This new information on the biology of S. sclerotiorum in the North Central Region has indicated that specific biotypes of the fungus can be widespread within the region and occur on multiple crops. MCG 9, for example is found on sunflower, soybean, dry bean and canola over nine widely separated states. Although we now have good information on the population of the pathogen in the North Central region, we need additional information on the population from the rest of the United States. It is clear from our research that biotypes of the pathogen can be disseminated over large distances. Isolates that occur in areas outside of the North Central area could impact crops and disease management in our region. Knowledge of the genetic variation and virulence of isolates of S. sclerotiorum in the United States is fundamental information needed for understanding the biology of this pathogen. Such information will help us with our overall strategy of using resistance or other controls for this pathogen. It may help explain the mixed results of field screening for resistance that has been observed over the country and the lack of correlation between field and laboratory evaluations. Furthermore, sensitivity of the pathogen to fungicides or other chemicals can be evaluated with greater reliance when there is a population of the pathogen that has been characterized with MCG’s, DNA fingerprinting and virulence traits. The currently funded project which focused on the North Central Region and this new project will establish a collection of isolates that represents the variation of the pathogen in the United States. It will become a benchmark of the pathogen population in the United States. This collection will be made available to researchers working on all aspects of the biology and control of S. sclerotiorum. This research will collect isolates of S. sclerotiorum from various crops grown in areas outside of the North Central region. The mycelial compatibility groups will be identified and microsatellites will be used to characterize the genetic variation. Then, isolates representative of the clones/genetic groups, will be characterized for virulence on six crops (soybean, sunflower, canola, dry bean, pea and lentil) using a standard cut stem inoculating method which measures lesion length as criteria for virulence. A set of isolates representing the genetic and virulence diversity of S. sclerotiorum in the United States will be prepared for distribution to Sclerotinia researchers.
3. Progress Report:
This project was initiated on June 1, 2008, research is ongoing, and the overall objective is to characterize the genetic variation and virulence of S. sclerotiorum on six crops in the North Central Region of the USA. A comprehensive examination of the genetic variation and virulence of Sclerotinia sclerotiorum from various crops across the United States had not previously been examined. In a previously funded project, we studied genetic variation and virulence of this pathogen in the north central United States. This current project is studying the pathogen from other parts of the United States. The objectives of the research were to 1) collect isolates from states outside the north central region and characterize the variation among isolates using mycelial compatibility and microsatellite markers and 2) measure aggressiveness of the isolates on six different susceptible crops. Plant scientists in 35 states were contacted and isolates of S. sclerotiorum from any plant species were requested. Unfortunately, we received only 88 isolates in the form of sclerotia from 15 states. The other states contacted did not have isolates to send. Of those 88 isolates of sclerotia received, only 68 were viable and cultures were established. The isolates were from the following 15 states: AR, AZ, CA, CT, FL, GA, ID, MA, ME, NC, NY, OK, OR, SC and WA. These were collected from 16 plant species and 20 different types of crops: broccoli, butter nut squash, bell pepper, cabbage, canola, carrot, cucumber, cauliflower, collard greens, common bean, gourd, lentil, lettuce, romaine lettuce, parsley, pea, petunia, potato, soybean and tomato. Isolates were evaluated for mycelial compatibility group (MCG) and microsatellite haplotype at twelve loci. There were 49 MCG’s identified within the 68 isolates. The 49 MCG’s were paired with representative isolates of the MCG’s identified from the north central region and 5 of the 49 MCG’s were similar to MCG’s from the north central region (MCG’s 3, 5, 21, and 31).All 68 isolates were genotyped with microsatellite markers. There were 47 unique haplotypes identified among the isolates.Haplotypes corresponded to MCG’s with few exceptions. There were 8 MCG’s with two or more haplotypes (one MCG had 5 haplotypes). Haplotypes within an MCG differed in base pair sizes in 1 to 3 of the microsatellites. Haplotypes from this recent collection will eventually be compared with those from the north central collection. Sixty six of the isolates were evaluated for aggressiveness on soybean and canola and the experiment was repeated. Plants were grown for five weeks in the greenhouse, there were four replications and plants were inoculated using the” cut stem” method. Lesions were measured after three days incubation in a mist chamber. The isolates were also evaluated for aggressiveness on sunflower and dry bean and a repeat of the experiment is in progress. The data from the aggressiveness experiments has not been analyzed. Additional experiments with other crops will be conducted in the future. A collection of isolates from various crops from all regions of the United States has been made and is stored for future researchon the biology of the pathogen. The isolates in this collection have already been used in two separate cooperative research projects on fungicide resistance and mycoviruses in S. sclerotiorum. Genotyping of the collection has been made and aggressiveness of isolates has been tested and recorded. Such information will be available to other researchers who wish to use this information for studies on the biology of the pathogen. This is the first study to obtain a nationwide overview of the genetic variation and aggressiveness of this important pathogen.