The Research Of Oidium Heveae Indoor Artificial Inoculation Of New Host-Arabidopsis Thaliana Mutants Eds1

Oidium heveae BA Steinmann(OH) is the pathogenic fungus of powdery mildew on rubber tree. Powdery mildew is the most impotant disease of rubber tree, which results in abnormal leaf fall and significant ruduce of the rubber latex. It has been widely accepted so far that OH is the obligate parasitic fungus and can only complete itself life history by infecting the tender organs of rubber tree. Therefore, the materials needed for OH research could only inconveniently be abtained from the leaves in the canopy of the tall rubber tree. On the other hand, great effort on the genetic transformation of rubber tree has not been succeeded and the research on molecular biology of interaction between rubber tree-OH has been unable to carry out.Searching out an easy operation host of OH is the bottleneck to solve the problem.We roport that OH is able to complete its disease cycle by infecting Arabidopsis thaliana mutant edsl as follows.1. The conidia were respectively inoculated on the leaves of Arabidopsis thaliana wild type Col-0, A. thaliana mutant edsl and Srl-4D, rubber tree clone RY7-33-97. The conidium germination and appressorium formation of OH were observed with optical microscope. The results of observation showed that the conidia were able to germinate and differentiate into the appressoria on the leaves of A.thaliana tested. However, the conidium germination rate and the number of appressoria formed were significantly different among the 4 plants. The RY7-33-97 ranked the first, edsl the second, Col-0 the third and Srl-4D the least. These results meant that the conidium germination and appressorium formation of OH had not yet encountered the resistant system of the plant tested.2. Systematic observation on the infection and development of OH after appressorium formation on A. thaliana were observed with optical microscope. The results showed that the haustoria began to appear on the leaves of A. thaliana mutant edll at 24h after OH inoculated,the meshy mycelia began to appear at 5d,lot of conidiophores began to appear at 12d,white color powder that similar to the symptom of powdery mildew began to appear at 6d. As the extension of time after inoculation,the symptoms on A. thaliana mutant edsl were more and more typical as those on rubber tree. At 37d after inoculation, the leaves of A. thaliana mutant edsl appeared in malformation and white color powder distributed all over the face of theleaves. By examination under the microscope, the white color powder was found to be the conidia with the same morphology as the ones of OH,and the morphology of haustoria were also very similar to the one of OH.However, only very few mycelia and conidiophores were observed on the leaves of Col-0 and Srl-4D after inoculated with OH, and no any haustoria and conidia were observed all the time. These observation results meant that A. thaliana mutant edll had lost, while the Col-0 and Srl-4D had not, its resistace to OH.3 . The white color powder incurred by OH were harvested from the leaves of A.thaliana mutant edll and inoculated on to the leaves of pot plantlets of RY7-33-97.The symptom of powdery mildew began to appear at 5d after inoculation, and typical symptom appeared at 12d.By examination the microscope, the fungi from lesions found to be the mycelia, conidiophores and conidia with morphology similar to the ones of OH. These results confirmed that OH growing on edsl was able successfully to infect rubber tree .4. PCR were carried out by using ITS5/IST6 of rDNA-ITS of OH as primers and total DNA abstracted from the white color powder on lesions of edsl and powdery mildew on rubber tree as templates. The results showed that the amplification patterns were the same between the 2 DNA templates. PCR aimed at the pathogenic-related genes 12732, 18993 and 9493 of OH as the amplified targets showed that the sequence similarity of the 3 genes was more then 99% between the 2 DNA templates.These results also confirmed that the fungi from edsl incurring the powdery mildew on rubber tree was OH.In summary, OH was able to infect A. thaliana mutant edsl and took advantage of this plant as non-host to complete its life history. The discovery and confirmation A.thaliana mutant edsl as the non-host of OH can greatly help setting up the convenient technique for large scale reproduction of OH, and also help peeping the interaction between rubber tree-OH.