Submitted to: Rice Technical Working Group Meeting Proceedings
Publication Type: Proceedings
Publication Acceptance Date: December 15, 2005
Publication Date: N/A
Technical Abstract: Rice wild relatives (Oryza spp.) are an important source of novel pest resistance genes, as well as tolerance to abiotic stresses and yield enhancing traits. Rice sheath blight caused by Rhizoctonia solani Kühn and leaf blast, Magnaporthe grisea (T.T. Herbert) Yaegashi & Udagawa, are major fungal diseases of cultivated rice (O. sativa L.). Resistance to blast and sheath blight has been reported in Oryza spp. accessions. Most of the Oryza spp. accessions have low seed set, poor germination, and shatter seed. Thus, it is advisable to grow the Oryza spp. accessions in the greenhouse. In order to determine the resistance of these accessions to U.S. isolates of M. grisea and R. solani it is necessary to adapt the procedures used for evaluating blast and sheath blight resistance for use with the Oryza spp. accessions and their progenies. The objective of this presentation is to describe the procedures presently being used to screen the Oryza spp. and their progenies for blast and sheath blight resistance. To ensure the M. griesa isolate being used for inoculation was virulent, the culture was started from a stock culture of the isolate stored at -20°C by placing the dried culture on an oatmeal agar medium or an agar medium containing alfalfa pellets and rice hulls. After the initial culture grew, it was sub-cultured to obtain the spores needed for inoculation and if needed, subcultures were started that could be stored for future use. Seeds of the Oryza spp. accessions or progenies to be inoculated were dehulled, surface sterilized and placed on an orchid medium in a lighted incubator set at 27°C and 12 hr light/12 hr dark for approximately 10 days. After the seedlings were approximately 12 cm tall, 4-6 plants were transferred to 6.5 cm square pots containing a soil mixture (1 part soil: 2 parts RediEarth). When the seedlings were at the 3 to 4 leaf stage, they were drought stressed in preparation for inoculation. M. griesa spores were placed in a xanthene gum mixture and diluted to 100,000 to 200,000 spores per ml. The spore solution was applied to the leaves of the drought stressed seedlings with an airbrush and then, the seedlings were placed in a controlled environment (dew) chamber for 24 hr at 24°C. When the seedlings were removed from the chamber, they were watered with a 20-20-20 liquid fertilizer solution and rated approximately one wk later for blast symptoms on a 0=no disease to 9=dead rating scale. To evaluate the Oryza spp. accessions for sheath blight resistance, this growth chamber/greenhouse method was developed because the accessions and their initial progenies cannot be grown in the field due to seed shattering and the limited availability of seed. Also, many of the accessions are tall which often gives false ratings for sheath blight resistance. To start new cultures, sclerotia of selected R. solani were placed on potato dextrose agar medium. Approximately 2.0 cm pieces of toothpick were infiltrated with potato dextrose broth and placed on potato dextrose agar medium. Next, R. solani sclerotia were placed on the medium with the toothpicks, so the R. solani mycelia would infiltrate the toothpick pieces. After 7 to 10 days, Oryza plants at the heading stage (R3-R6) were inoculated with R. solani by placing the aforementioned toothpick piece into the leaf collar of the second, third and fourth leaves of each tiller. Plants were covered with a plastic bag to increase humidity and placed in a growth chamber set at 28°C, 100% humidity and 14 hr light/10 hr dark for 7 to 10 days. Each pot containing 2 to 4 plants was given an overall sheath blight rating on a 0=no disease to 9=dead rating scale. To determine the infected area, the total leaf length of the leaf blade (collar to tip) and the leaf sheath (collar to soil surface) was measured first. Next, the disease infection was measured on the leaf blade (collar to end of the infected area) and leaf sheath (collar to distal point of the infected area). These measurements were compared to those of rice cultivars evaluated under field conditions. If the infected plants recover and produce ratoon tillers, this method can be used on ratoon tillers, especially when there is limited seed is available. Results obtained from inoculating and rating Oryza spp. accessions using both of these methods will be presented with U.S. rice cultivars included for comparison. In conclusion, methods currently used by rice pathologists for evaluating O. sativa lines need to be adapted for evaluation of Oryza spp. accessions and their crossed progenies which have low seed set and a varied plant type.