Artifical Culture Characters Of Ustilaginoidea Virens And Ultrastructure Of False Smut Balls

Rice false smut is a worldwide fungal disease in rice caused by Ustilaginodea virens (cooke) Tak, which harmed the panicles of rice and produced rice fasle balls. It was all along ignored before 1980s' because of its occasionally happening on the middle- and later-riped rice. Since 1980s', however, with the change of rice cultivars and increasing of fertilization in the paddyfield, rice false smut in China has being more and more serious, which not only influences greatly on the yield and quality of rice, but ustiloxins also cause poisoning to human and animals. Up-to-now, the disease has become one of the major rice diseases in all the major rice production areas in China.It is presumed that chlamydospore may be the major primary infection source of rice false smut and begin to infect the panicles of rice before or after florescence. But it is also supposed, however, that the disease is of systematic infection. Because successful infection rate by artifical inoculation was very low, It is difficult to examine the infection process and growth mode of pathogen in plant. A high effective inoculation method becomes the bottle neck for the research on this disease.This study was focused mainly on the pathogen isolation, artifical culture characters, and the ultrastructure of rice false smut balls to find out the possible infection and growth process of the pathogen in nature, and to establish an artifical inoculation method. Otherwise, the effect of ustiloxins on rice and other saprophytic fungus, Magnaporthe grisea, was also examined in order to explore the biological activity and function of ustiloxins in pathogenesis. The main results gained are as follows:1. Isolation, purification of the pathogen and observation on its artifical culture characters. Ustilaginoidea virens was isolated on XBZ and PSA medium by means of suspended liquid of chlamydospores of false smut balls. The isolatedpathogen fungus was cultured on various media in dark at 26 C to observe itscharacteristics. The media included PSA medium, PS medium, cooked barley and rice grains with glumes, and cooked wheat grains and rice kernels. The culture characters of Ustilaginoidea virens UH17 remarkably changed with serial culture on PSA, but there was no obvious change in strain UH12. After about 20 days, a great amount of chlamydospores were produced. The appearant patterns of chlamydospores were as follows: (1) Chlamydospores dispersed on the whole surface of colonies, and no visible piles of chlamydospores were visibled;(2) There was one to several small piles of chlamydospores on the colony;(3) There was a big ball of chlamydospores on the ceter of the colony. The first two patterns were much common, and the third appeared on occasion. When cultured in PS medium, some isolates never produced chlamydospore, and some could form hyphal pieces floating over the medium at the later stage, on which chlamydospores produced. Under the same condition, the growth of Ustilaginoidea virens in the cooked barley and rice grains with intact glumes was much slower than that in the cooked wheat grains and rice kernels, and more piles of chlamydospores were produced in the later.2. Observation and analysis of SEM and TEM on artifical culture colonies of Ustilaginoidea virens and rice false smut balls. The ultrastructure of rice false smut balls and colonies of Ustilaginoidea virens on different media were conducted by SEM and TEM. In the false smut balls, the outer dense chlamydospores piled around the inner hyphae. There was no crossed interface between the hyphae and endosperm, and there was a large stretch of well developed endosperm in the center part, in which a great amount of starch grains filled. These suggest that the infection of pathogen may occur after the flowering and grouting, and the pathogen may be saprophytic fungus in plant. TEM observation and analysis on fungal tissue of false smut balls found that hyphae were compact and ranged orderly in the interior, and a lot of fat bodies were filled the hyphae;hyphae are sparse and extensed in different directions in the exterior, and fat bodies became faint;in the middle, cell wall of hyphae were thickened locally, in which fat globules were observed. It indicated that Ustilaginoidea virens was distinctly characteristic of saprophytic fungus.3. Effect of ustiloxins on rice and fungus, Magnaporthe grisea. The washingsolution of fresh false smut balls had strong inhibition on the growth of rice coleoptile and radicle, especially on the later, possibly due to the directly exposed to ustiloxins. There was, however, no obvious inhibition on Magnaporthe grisea. And PS filtrate had inhibition on growth of Magnaporthe grisea drastically. These implied that Magnaporthe grisea is not so sensitive to ustiloxins as plants.