Screening Antagonistic Bacteria Against Verticillium Dahliae And Their Application In Cotton Verticilium Wilt Control

Verticillium dahliae Kleb.(Vd)is a kind of devastating phytopathogenic fungus to cause Verticillium wilt(Vw).Vw,also known as cancer of cotton,damages the global cotton product and quality at high cost annually.In the current actual production,the control of cotton Verticillium wilt has four methods.The first method is to use molecular breeding technology to cultivate disease-resistant varieties.The second method is to increase soil permeability through field measures such as deep cultivation and crop rotation to improve plant disease resistance.Thirdly,direct spraying of chemically synthesized pesticides to control diseases.The fourth method is the application of a microbial preparation made of a biocontrol inhibitor.However,due to the lack of high resistant cotton germplasm resources and the poor effects of agricultural actions,chemically synthesized pesticides have become the main control methods in production.In recent decades,chemical pesticides have not only damaged the soil nutrition environment severely,but also have increased the pesticide residues in agricultural products.With the improvement of environmental awareness and the development of biotechnology,biological pesticides have gradually risen since the 1990 s.It is an inevitable trend for crop protection to shift from chemical control to biological control.The main bactericidal factor of biological pesticides is disease-resistant microorganisms.Therefore,the screening of antagonistic microorganisms and the study of resistance mechanisms are the main tasks for the biological control of cotton Vw.The main purpose of this study was to obtain microbial strains resistant to Vw and study the antagonistic mechanism of biocontrol strains.Various samples of soil root microorganisms were collected.The samples were collected from Shanghai,Jiangsu,Zhejiang,Anhui,Heilongjiang,Jilin,Hunan,and Xinjiang Province.And the sources of collected samples were diversified,including cotton?soybean?corn?weeds and so on.In this study,about 8,736 strains of microorganisms were isolated and collected from soil-rooted samples.Inhibition zone assays were performed with Verticillium wilt stains of V07DF2,V08DF1,V15QY1,V08T2,V16DF4 and V991.As a result,148 strains with antagonistic effects to Vd were obtained.The genomic DNA of 15 strains with top resistance was extracted and subjected to genomic sequencing.Based on 16 S ribosomal DNA(rDNA)sequence characterization,7 strains were identified as Enterbacter;4 strains were Pseudomonas;2 strains were Delftia;1 strain was Bacillus;1 strain was Pantoea.In rescreening assay,two strains showed broad-spectrum antagonistic resistance to Vd.One strain was Pseudomonas fluorescens,named XY2F4;the other strain was P.donghuensis,named 22G5.Biocontrol activity planta assays confirmed that: XY2F4 and 22G5 can inhibit the spread of cotton Vw obviously.According ot the comparative genomic analysis of the gene cluster in charge of the secondary metabolites in XY2F4 and 22G5,XY2F4 contains a gene cluster for the biosynthesis of 2,4-diacetylresorcinol(2,4-DAPG);22G5 contains a unique gene cluster for 7-Hydroxtropolone bio-synthesis.Combined with synthetic biology,this research set a corner stone to the construction of a highly efficient industrialized synthetic biological system for Vw.