A Study Isolation From Soil And Molecular Detection For Sudden Death Syndrome Pathogens Of Soybean

Sudden Death Syndrome of Soybean (SDS) is a relatively new soybean disease, which distributing in three main soybean produce countries, United State, Brazil and Argentina. SDS is caused by 2 Fusarium fungus respectively, Fusarium virguliforme and Fusarium tucumaniae. The loss caused by SDS became more and more severe these years, some even up to 100%. As the soilborn disease, SDS can survive in soil for many years and can be introduced to a new place along with soybean trade. Since China has been importing a big mount of soybean from US, Brazil and Argentina, this study is purposed to establish SDS detection method for quarantine and identification and to apply the method to quarantine the soil sample from ports, in order to prevent the SDS to be introduced into China and to protect soybean industry of China further.In this study, a selective culture medium named PCNB, which can enrich the pathogens of SDS and restrain the other fungi, was used to enumerate the pathogens of SDS from soil.The colony type of SDS pathogens on PCNB was small,round,white,lacked aerial mycelium,and a raised and canary center,and a chalky appearance. In seccession , this colonies continued growing on plates of potato dextrose agar(PDA) by transferred from PCNB. SDS pathogens growed slowly on PDA and produced blue colonies , lacked aerial mycelium ,and exhibited an irregular margin. The colony morphology and pigmentation were the important indexes to identificate SDS pathogens. Under microscope, the spores of SDS pathogens F virguliforme F tucumaniae were very similar to non-SDS Fusarium spp.. They all produced 2-3 septa, falcate macroconidia.An allele-specific PCR-based method was developed to distinguish Fusarium virguliforme, from F. phaseoli and other non-SDS Fusarium spp.. Under the rules of allele-specific primer design, one pair of primers, Fsg-a-1/6 were designed and filtered based on three SNP sites on translation elongation factor 1-α gene, which could distinctively amplify F. virguliforme and produce PCR products of 327 bp. At the same time, to distinguish F. virguliforme and F phaseoli from the other Fusarium spp., one pair of primers, Fu1/2, were designed based on the region of ITS (intergenic...