Isolation,Identification And Control Effects Of Biocontrol Bacteria Against Phytophthora Sojae

Soybean Phytophthora root rot caused by Phytophthora sojae is an important and destructive disease on soybean worldwide.Biological control is an effective and safe way for control of the disease at present.In this dissertation,the isolation,screening,identification and the biocntrol effects of the biocotrol agents of the disease were conducted systematically,in order to provide new biocntrol resource and experimental evidences for the control of soybean blight.The main results are as follows.1.Isolation,identification of biocontrol bacteria and determination of their physiological and biochemical characteristicsFour strains of antagonistic bacteria with inhibitory effects on P.sojae were isolated from the soybean field soil of Dayangdian,Shushan District,Hefei City,Anhui Province,and the soil of the greenhouse of Anhui Agricultural University.Two of them had strong antagonistic effects on P.sojae.First of all,colony morphology and biological and biochemical properties of biocontrol bacteria were observed.Biocontrol bacterium YG-2 was Gram negative and aerobic,and the colony is round,colorless,translucent,smooth.The colony edge is smooth.The experiment of glucose oxidation fermentation is an oxidation type.It can not liquefy gelatin,decompose hydrogen peroxide,hydrolyze starch,and can not hydrolyze cellulose.It has salt resistance.The OD₆₀₀00 value of LB culture containing 1%,2% and 7% NaCl has little change.It can grow in the environment of pH 5-10,and the optimum pH is 7,which can grow at 10-45? and the optimum temperature is 37?.Enough to use a variety of substances as a carbon and nitrogen source.By 16 SrDNA sequence alignment,the homology with serial number CP019667.1 was 99%.Accordingly,the biocontrol bacterium YG-2 was initially identified as Burkholderia cenocepacia.The biocontrol bacterium YG-4 is close to Bacillus,Gram-negative,aerobic bacteria,Oxidation-type glucose oxidation and fermentation experiments,can make gelatin liquefaction,hydrogen peroxide can be decomposed,contact enzyme reaction is rapid,phenylalanine solution Ammonia positive,can make the starch hydrolyzed,the reaction is negative,the oxidase test is positive,can not use malonate oxidase and citrate,nitrate reduction reaction is negative,VP assay test is negative,can not hydrolyze fiber Suspension,without salt tolerance,grew slowly in LB broth containing 7% NaCl.It can be grown in an environment with a pH of 5-10,with an optimum pH of 7,which can be grown at 10-45?and an optimum temperature of 37?.Biocontrol bacteria YG-4 can take advantage of a variety of carbon and nitrogen sources.By 16 SrDNA sequence alignment,the sequence number:JQ229807.1 has a homology of 99%.Accordingly,the biocontrol bacteria YG-4 was initially identified as Bacillus amyloliquefaciens.2.Tests of inhibitory rates of YG-2 and YG-4 against P.sojae and determination of antibacterial spectrum of YG-4In the course of the experiment,there was some difference in the inhibition rates of P.sojae strains in different areas with bio-control bacteria.Therefore,the strains YG-2 and YG-4 were used against 8 strains of P.sojae P6497,XX2,and GZ6 in different regions.GZ37,SX8,30,LB5-7,GY18-1 were measured for inhibition rate.Biocontrol bacteria YG-2 and YG-4 have different inhibition rates on P.sojae from different regions,and the same biocontrol bacteria have different inhibition rates on P.sojae from different regions.The highest inhibition rates of biocontrol bacteria YG-2against P.sojae SX8 reached 78.31%?at the 6th day of pupal culture?,and the inhibition rate to GZ37 was only 16.88%?at day 6?,and the inhibition rate to P6497was 59.52%.The inhibition rate for XX2 was 62.35%.Since the YG-2 antibacterial spectrum was not broad enough and the inhibition rates against other pathogens were not high,the inhibition rates against 13 pathonic fungi were measured using the biocontrol bacterium YG-4.The tested fungi included Alternaria alternata,Trichothecium roseum,Fusarium oxysporum,F.avenaceum,Rhizoctonia cerealis,F.graminearum,Sclerotinia sclerotiorum,Phytophthora infestans,A.longipes,A.solani,Colletotrichum gloeosporioides.The inhibitory effects of the biocontrol bacterium YG-4 on different pathogenic fungi were very different.YG-4 has the highest inhibition rate against S.sclerotiorum,which is 79.63%,followed by the inhibition rate against R.cerealis.The inhibition rate was74.07%.The inhibition rates agaist A.alternata,T.roseum,F.avenaceum,A.solani,Bipolaris maydis and A.longipes varies from 37.04%to 55.56%.The biocontrol bacterium YG-4 had weaker inhibitory effects on F.oxysporum.It had almost no inhibitory effect on F.graminearum,P.infestans and C.gloeosporioides.The inhibition rates were below 10%.3.Control experiment of biocontrol bacteria YG-2 and YG-4 on soybean blightThe leaf protective effect of YG-2 fermentation solution on soybean blight was78.57%,and that of YG-4 fermentation solution on soybean blight was 60.71%.The control effect of the biocontrol bacterium strain YG-2 fermentation broth on soybean blight in potted plants reached 60.67%,and the control effect of the biocontrol bacteria strain YG-4 fermentation broth on soybean blight in potted plants reached 60.00%.The metabolites of Burkholderia YG-2 and Bacillus YG-4 could reduce the pathogenic effect of P.sojae on soybean leaves,and the control effect of YG-2 metabolite is stronger.Since the metabolites are filtered through the bacterial filter,it is suggested that the control effect of the biocontrol bacteria YG-2 and YG-4 is not due to the competition of the viable bacteria between the strains,and it may be caused by some product in the metabolites.The future experiments are necessary to explore the biocontrol mechanism of the biocontrol bacteria YG-2 and YG-4 against P.sojae and other plant pathogenic fungi.