Screening Of Biocontrol Actinnomyces Against Sclerotinia Sclerotiorum And Taxonomic Identification Of Micromonospora Parathelypteridis Sp.nov.

Sclerotinia sclerotiorum is caused by the soilborne fungal pathogen Sclerotinia sclerotiorum(Lid.)de Bary.The fungus has a extensive range of hosts.It is known to infect over 400 species of plants and spread all over the world.Due to the resting structure sclerotia produced,it can be remained in soil for a long time.Therefore,the structure makes it is difficult to control.One of the major crops in China is soybean which has a serious losses in yield annually caused by this necrotrophic fungus.Now,fungicides are the major control method for Sclerotinia diseases.According to the chemical agents increased production costs and damaged the environment,it cannot be better to use in practice.Compared with the chemical control method,biocontrol owned more advantages including security,reliable,low-toxin,non-hazardous and so on.So,it is extremely significant to suppress the Sclerotinia sclerotiorum by the biological methods.In this study,choosing five separation medium respectively to isolate and cultivate the endophytic actinobacterium of the health and disease soybean roots which affected with the Sclerotinia sclerotiorum.Observed cultural characteristics and morphology features of the isolated strains and combined the organism which have the same properties.Counted and analysed the total population and the different species of endophytic actinomycetes in roots of soybean plants.Then tablets antibacterial activity against pathogenic fungus S.sclerotiorum in vitro was launched respectively,to test the inhibition activity of the isolated endophytic actinomycetes of the soybean roots and its fermentation crude extract.Incorporated with the field seedling tests,to evaluate the suppressive capacity to the pathogenic fungus,the role in the process of soybean plants growth and the colonization ability in the roots of the host.Later,the obtained bioactive strain which has the highest inhibition rate resistance Sclerotinia sclerotiorum has been described in many aspects,including molecular biology,morphological characteristics,physiological and biochemical characteristics.Provided a new resource in the Sclerotinia stem rot control for the futher study.In addition,a rare actinomycetes of the genus Micromonospora has been identified,enriched the microbes repository.The main results are as follows:(1)With the roots of morbidity soybean plants and the healthy ones that were adjacent to grow,a total of 1574 population were obtained,the numbers of the colony-forming units acquired from the uninfected roots was notable higher than the infected ones.After removing duplicates,70 different actinomycetes of the soybean roots have been obtained.Among them,15 strains onlyisolated from the healthy plants,27 species only isolated from the disease plants,28 organism obtained both in the infected and uninfected roots of soybean plants.The results illuminated that the composition diversity of the disease plants was more richer than the healthy ones.(2)All the 70 screened strains were tested antagonism activity against pathogenic fungi in vitro.The 5 strains have inhibition effect on suppression the fungus.Among them 2 species isolated from both in the infected and uninfected(ZGS 1-10 and ZBCPA 2-13),inhibition rate was from 20.9% to 22.4%;The others(ZBDPA 2-2、ZBAAG 12-4 and ZBAAG 3-15)were all obtained from the disease plants,inhibition rate was from 28.3% to 58.5%.The strain ZBDPA 2-2 has the highest effect on the mycelium growth of the Sclerotinia sclerotiorum,the antibacterial rate was58.5%.Furthermore,ZBDPA 2-2 also showed bioactivity to other 11 tested pathogenic fungus.(3)According to the soybean seedling assay,it was found that the spore concentration of the strain ZBDPA 2-2 achieved 1.0×106 cfu·g-1-1.0×107 cfu·g-1,it was remarkably reduced the disease incidence and disease index of the pathogenic fungus Sclerotinia sclerotiorum.The leaves of soybean tested also revealed the obvious debased of the disease index.Moreover,this concentration has an excellent growth-promoting effect on the total fresh weight,total dry weight,root fresh weight,root dry weight,root length and stem length.And it has a long-term colonization ability in the rhizosphere soil by soybean seedlings,however,it was weakly colonized in the root tissues of soybean plants.(4)The preliminary description of strain ZBDPA 2-2 was proceeded though molecular biology analysis,observation morphology features of this organism,contrasted the physiological and biochemical characteristics.It displayed that it was distinguished from the most similarity species in many aspects and confirmed the strain belong to the genus Streptomyces.(5)In this study,a rare actinomycetes NEAU-JXY5 T was identified by polyphasic approach.The results of 16 S rRNA and gyr B gene analysis shown that strain NEAU-JXY5 T and many type strains of the genus Micromonospora has a highest similarity and formed a stable branch.Compared the morphology features,physiological and biochemical characteristics and chemotaxonmic properties,it conformed to the typical characteristics of the genus Micromonospora,however,differentiated from its closest related species notably.In addition,DNA-DNA hybridization between strain NEAU-JXY5 T and the phylogenetically related species were all below 70%.Illuminated that strain NEAU-JXY5 T represents a novel species of the genus of Micromonospora,for which the name Micromonospora parathelypteridis sp.nov.is proposed.