Biocontrol Effect And Mechanism Of A Cotton Endophytic Bacterium Bacillus Cereus Against Both Verticillium And Fusarium Wilts In Gossypium Hirsutum

Verticillium and Fusarium wilts,two of the most important diseases in cotton,pose serious threats to cotton production.Screening effective biocontrol microbial resources has became one of the important ways to against Verticillium and Fusarium wilts.The objective of this study is to screen an efficient antagonistic bacteria against Verticillium and Fusarium wilts,and characterize the biocontrol mechanism of bacteria against Verticillium wilt.The main results are sa following:1.An endophytic bacterium,which can hydrolyze polysaccharides with ?-1,4 linkage was isolated by enrichment medium,which was made of konjac glucomannan as carbon source.The endophytic bacterium was named asBacillus cereus isolate YUPP-10 by the morphology and molecular biology.2.The inhibition rate of YUPP-10 against V.dahliae and F.oxysporum were assessed using the confront culture method.The results showed that YUPP-10 inhibited the colony growth of V.dahliae and F.oxysporum,with the width of the inhibition zone of YUPP-10 was 0.73 and 0.56 cm after 7 days.The inhibition rate of YUPP-10 against V.dahliae and F.oxysporum were assessed using the enclosed chamber test,and the inhibition rate of volatile organic compounds(VOC)of YUPP-10 against V.dahliae was 77.03% after 10 days,but the VOCagainst F.oxysporum was not significant.B.cereus YUPP-10 culture filtrate(CF)suppressed V.dahliae and F.oxysporumspore germination,the inhibition rate range from 17.22% to 71.25% and 27.75% to 51.65%,respectively,and the spore germination in a dose-dependent manner.The inhibition rate on V.dahliae microsclerotia germination and F.oxysporumspore yield by the CF range from 10.69% to 26.62% and 54.49% to 98.41%,respectively,and the microsclerotia germination in a dose-dependent manner.3.Low concentration of CF promoted germination rate,emergence rate and stem length of cotton seeds.The best inhibition rate was 80.60%,when cotton seedings were pro-inoculated with substrate containing YUPP-10 in pot experiment.The inhibition rate was 45.11%,when cotton seedings were pro-inoculated with 1×107 cfu/mL CF in pot experiment.4.YUPP-10 induced disease resistance,including radicle and leaves against V.dahliae.ROS,sedimentation of lignin.The expression level of defense genes in G.hirsutum leaves were detected by real-time quantitative PCR(RT-qPCR).the result showed that some defense genes were induced,such as PAL,POD,PPO,CHI and PR10.5.The encoding gene segments which can hydrolyze polysaccharides with ?-1,4 linkage was isolated from complete genome,construct the expression vector,express and purify its product.The recombinant protein induced defense genes,such as PAL,POD.The pathogenesis-related genesPR1,which is marker genes of the SA-dependent defence pathway,theJAZ1 which ismarker genes of the JA-dependent defence pathway,and theHSR203 J which is considered as marker gene for hypersensitive reaction,were significantly up-regulated in response to recombinant protein.In summary,the cotton endophytic bacterium Bacillus cereus YUPP-10 inhibited the growth of V.dahliae and F.oxysporum,promoted cotton growth andagainst Verticillium and Fusarium wilts.The CF induced immune response of cotton.The CGTase inhibited the growth of V.dahliae and induced immune response of cotton.