Screening,Identification And Herbicidal Active Study Of Streptomyces Sp.NEAU-240

There are about 200 species of farmland weeds in China,of which 30 species of weeds reduce the annual output of farmland crops by 12.3-16.5%.With the development of the rural economy,about 50% of the main farmland crops are applied with herbicides.Chemical weeding make the labor and material resources liberated.At the same time,the continued use of large amounts of herbicides can cause residual phytotoxicity,trigger weed transfer problems and the emergence of weed-resistant plants in crops,as well as soil pollution,water quality degradation,harm to humans and other organisms.Therefore,microbial herbicides have gradually entered people's sights,the use of microorganisms or their metabolites to control weeds has become a major trend under the concept of green agriculture and assured agricultural products.In this study,six isolation media were used to isolate and purify actinomycetes from the rhizosphere soil and leaves of dogberry.Firstly,the spores and the fermentation broth of the actinomycetes were used for germination experiment test to obtain the strain of high herbicidal activity.Then the active strains were subjected to a plate and pot experiment to verify the herbicidal activity and carry out a safety analysis of the herbicidal ability.The herbicidal active compound of the active actinomycetes was isolated and extracted,the herbicidal activity mechanism of the compound was preliminarily explored.Finally,the active actinomycetes was identified at the molecular level,morphological culture characteristics,physiological and biochemical characteristics.Provide data materials for the study of actinomycetes in herbicidal activity and provide strain resources to develop new microbial herbicides.The main results of this study are as follows:(1)214 strains of actinomycetes were isolated from the rhizosphere soil and leaves of dogberry.(2)20 strains inhibited the roots and shoots of wheat were obtained from 214 actinomycetes,three strains with good inhibition effect(TDZ44,TDZ51,NEAU-240)were obtained using the liquid fermentation broth of 20 actinomycetes for screening,among them strain NEAU-240 works best.Therefore,the next step is to study strain NEAU-240.(3)The result of herbicidal activity plate experiment showed that the fermentation supernatant liquid diluted 100 times of strain NEAU-240 had the root inhibition rate was higher than 90% on Echinochloa crusgalli(L)Beauv,Amaranthus retroflexus L.and Setaria viridis L.,the inhibition rate of shoot was higher than 30%;dilution 10 times can significantly inhibited the germination of weeds seeds.(4)The result of herbicidal pot experiment showed that the fermentation supernatant liquid diluted 100 times of strain NEAU-240 has the fresh weight inhibition rate of aerial parts of E.crusgalli(L)Beauv and A.retroflexus L.higher than 32%,the fresh weight inhibition rate of underground part was higher than 15%.Dilution 10 times,the fresh weight inhibition rate on the A.retroflexus L.and E.crusgalli(L)Beauv was 78.4% and 45.3%,the fresh weight inhibition rate in the underground part was higher than 20%.Therefore,it was verified that the strain NEAU-240 has good herbicidal activity,which could be further studied.(5)Crop safety experiments showed that the fermentation supernatant broth of strain NEAU-240 had almost no effect on caraway,less impact on crops such as wheat,corn and cucumber,greater harm to tomatoes and rapeseed.The results showed that the active substance of the strain NEAU-240 was harmful to some crops.Therefore,it was suitable for selective weeding.(6)Preliminary study on the herbicidal mechanism of the compound 2-[2-(3,5-Dimethyl-2-oxo-cyclohexyl)-6-oxo-tetrahydro-pyran-4yl]-acetamide was carried out,the results showed that the compound inhibited the activity of ?-amylase,simultaneously,the activity of acetyl-lactate synthase(ALS)was inhibited,which inhibited the germination of seeds and the growth of weeds.(7)The antibacterial activity of strain NEAU-240 was investigated.Its spore antibacterial activity experiment showed that the strain has antibacterial activity against a variety of phytopathogenic fungi such as Verticillium dahlia and Fusarium oxysporum.At the same time,the herbicidal active compound 2-[2-(3,5-Dimethyl-2-oxo-cyclohexyl)-6-oxo-tetrahydro-pyran-4yl]-acetamide was tested for its antibacterial activity and found to inhibit the growth of mycelia such as Alternaria mali Roberts,Pyricularia oryzae and Rhizoctonia solani Kuhn.The antifungal activity of the compound was determined for the first time,which provided theoretical basis for further research and development of the compound.(8)Through the analysis of 16 S rRNA gene sequence and phylogenetic tree,the strain NEAU-240 was established on the same evolutionary branch with the closest strains Streptomyces setonii NRRL ISP-5322T(99.65%)and Streptomyces anulatus NRRL B-2000T(99.65%),and combined with the analysis of physiological and biochemical characteristics and morphological culture characteristics,the strain NEAU-240 was identified as the genus Streptomyces.(9)At the same time,16 S rRNA sequencing was performed on the species-specific actinomycetes of 214 strains,6 rare genera of bacteria were found,including Kribbella,Cellulomonas,Agromyces,Actinoallomurus,Nocardia and Micromonospora.The strain NEAUTCZ24 T assigned to the genus Cellulomonas was a potential taxon,which was identified as a novel species of the genus Cellulomonas by polyphasic taxonomic analysis,for which the name Cellulomonas rhizosphaerae sp.nov.is proposed.