Analyses Of Biological Characteristics And ITS In Ribosomal DNA Of Synchytrium Puerariae

As a noxious and damaging weed, kudzu causes serious harm to ecology, agriculture and industry in USA. There are many biocontrol factors in China where kudzu originated. Synchytrium pueraria is a parasitic fungus on kudzu and it can bring serious leaf gall symptoms on leaves of kudzu. In the present study, the biological characteristics, developments, inoculation conditions were investigated, the techniques for extracting ribosomal DNA (rDNA) were compared, and the rDNA sequence was analyzed.1. The 'pseudo-rust' disease caused by S. puerariae on Pueraria lobata plants in Guangdong Province was surveyed in March to December, 2006. The results showed that the disease was present from April to December, and it developed quickly during the rainy season, then it became severe in September and October because of abundant rain and strong typhoon, finally it remained serious in December, though spot had a little change and the disease index have decreased with the falling of leaves.2. Biological characteristics of S.puerariae were studied to find out the optimal conditions of sporangium germination. The result showed that sporangium could germinate at 10-35℃; the optimal temperature was 15-20℃with the germination rate of 44.93%; the sporangium germination rate was reduced at temperatures lower than 15°℃and higher than 20℃; the germination rate was only 2.4% at 35℃; the lethal temperature of the sporangium was 46℃. The optimal pH was 6-8 with germination rate of 43.2%. The nutrition hardly affected sporangium germination and the sporangium germinated even in sterile water. The light had little influence on sporangium germination and sporangia germinated under both light and darkness; under continuous light, continuous dark and alternation of light and darkness, the sporangium germination rates were 43.2%, 41.5%, 42.67% correspondently.3. Different methods of inoculation were studied. The result showed that the temperature between 18-20℃was the optimum for inoculation of S. puerariae on Pueraria lobata and the infection rate was about 84% within the temperatures. Relatively longer duration of wetness (48 hours) after inoculation was favorable to fungal infection and disease development.4. Five different DNA extraction methods were tested for genomic DNA extraction from the sori of Pueraria lobata, P. thomsonii and P. montana. The modified technique of Benzyl Chloride was showe the most efficient one to extract genomic DNA. rDNA-ITS were amplified successfully and directly from a single sporangium by PCR. The ITS sequences of the isolates from P. lobata and P. thomsonii were 856 bp and 907 bp respectively and with 92% of sequence similarity; the ITS sequences of two isolates (SPM-1 and SPM-2) from P. montana were 1148bp and 1147 bp, with 54% and 55% sequence similarity, respectively, to the isolates from P. lobata and P. thomsonii. We proposed that the isolates from P. lobata and P. thomsonii were different varieties of the same species and the isolates from P. montana were a different species of Synchytrium; the isolates from P. lobata and P. thomsonii were two closely related species and the isolates from P. montana was a more distantly related species.