| Tobacco bacterial wilt is a soil-borne disease caused by Ralstonia solanacearum,which is wide spread and difficult to prevent and control.It is a serious disease that restricts the yield and quality of tobacco leaves over China and around the globe.Biological control,the environmental and sustainable control method,is interested by numerous of researchers in controlling of tobacco bacterial wilt disease these days.There already have registered commercial biological control products made of Paenibacillus polymyxa or Pseudomonas fluorescens.Yet there has no systematical study on the control effects of two biological control agents?BCAs?on tobacco bacterial wilt.This study was conducted with laboratory and field experiments to analyze and compare the control effects of two BCAs,which mainly included the study of inhibition zones,growth of stains,tobacco seeding growth promotion,control efficiency of bacterial wilt,and tobacco plant induced system resistance.The results are as follows.1.Improved plate confrontation and UV spectrophotometry were conducted,the result showed that the diameter of inhibition zones of P.polymyxa and P.fluorescens bacterial colonies were 23.8mm and 23.1 mm,respectively.The log of bacteria suspension[X,log?cfu/mL?]and the diameter of inhibition zones[Y,?mm?]showed a linear relationship,the line of P.polymyxa Y=1.7461X+18.576?R2=0.9458?was above the line of P.fluorescens Y=1.8417X+12.667?R2=0.8514?.Three strains could grow well when pH between 5 and 8.The diameter of inhibition zone of P.polymyxa was in negative correlation to pH,the diameter of inhibition zone was 42.91mm when pH=5.While P.fluorescens had the best inhibition when pH=7,the diameter of inhibition zone was22.00 mm.2.Seed soaking result showed that P.polymyxa had significant promotion on the germination energy and germination rate,while P.fluorescens had significant promotion on the germination energy?P<0.05,similarly hereinafter?.When pouring of BCAs'bacterial suspension alone,P.polymyxa promoted the tobacco seedling height continuously and significantly,it promoted the leaf length and leaf width at the same time,as a result,the shoot dry weight and root dry weight had been significantly promoted by P.polymyxa,while P.fluorescens only significantly promoted the seedling height in late period.When pouring of BCAs'bacterial suspension challenging with R.solanacearum,before the wilting of the tobacco,P.polymyxa and P.fluorescens had promotion on the seedling growth,which showed no significant difference,though.Pot experiment showed that the final control efficiency of P.fluorescens and P.polymyxa were 29.44%and 41.11%,respectively.3.The BCAs were inoculated by pouring into the root,inoculation of BCAs alone and challenge inoculation of BCAs and R.solanacearum were designed as control,respectively.Reference to the onset period of bacterial wilt in chapter 3,the CAT,PAL,POD,PPO and SOD were detected in 5 different times,which were 2d,5d,8d,11d and 14d after the inoculation of BCAs.The results showed that two BCAs promoted five enzymes in different degree.When BCAs were inoculated alone,P.polymyxa promoted the average activity of 5 enzymes for 1.24 times,1.16times,1.29 times,1.36 times and 1.70 times,respectively.P.fluorescens promoted the average activity of 5 enzymes for 1.22 times,1.23 times,1.21 times,1.38 times and 1.58 times,respectively.When inoculating the BCAs challenging with R.solanacearum,P.polymyxa promoted the average activity of 5 enzymes for 1.29 time,1.41 times,1.46 times,1.65 times and 1.44 times,respectively.P.fluorescens promoted the average activity of 5 enzymes for 1.18 times,1.42 times,1.30 times,1.65 times and 1.56 times.4.The populations of cultivated bacteria were counted by diluted plate,the results showed that P.polymyxa and P.fluorescens increased the bacterial population while decreased the fungal populations of the microbiome in tobacco rhizosphere.The Biolog ECO method was used in the study of microbial community functional diversity,the results showed that P.polymyxa and P.fluorescens promoted the soil microbial metabolic activity,P.polymyxa had the best promotion when P.polymyxa inoculated alone,P.fluorescens had the best promotion when inoculating P.fluorescens challenging with R.solanacearum.When inoculating BCAs alone,P.polymyxa significantly promoted the metabolism of carbohydrates,amino acids,carboxylic acids,and amidic acids,P.fluorescens significantly promoted the metabolism of Phenolic compounds.When inoculating BCAs challenging with R.solanacearum,P.polymyxa promoted six carbo sources insignificantly,P.fluorescens significantly promoted the metabolism of amino acids and carboxylic acids.The results of principal component analysis showed that P.polymyxa increased the soil microbial community stability.P.fluorescens increased the soil microbial community stability when inoculated alone,while decreased when challenged with R.solanacearum.Functional diversity index analysis indicated that P.polymyxa and P.fluorescens decreased the soil microbial evenness?McIntosh?,especially P.fluorescens had the significant and stable effects.When BCAs inoculated alone,the species and abundance of soil microorganism raised?Shannon?,while the soil microbial species had been reduced when challenged with R.solanacearum.5.Composition Field experiments were carried out with the combining application of rape seed cake.The result showed that P.polymyxa and P.fluorescens significantly promoted the maximum leaf area and leaf area index of tobacco at rosette stage,vigorous growing stage and budding stage,which were better than the effect of commercial product Tugenben.The control efficiency on tobacco bacterial wilt of P.polymyxa,P.fluorescens and Tugenben were 26.40%,16.41%and 25.64%at 2nd August?when tobacco middle leaves were harvested?,respectively.And the control efficiency on black shank of tobacco were 0.33%,17.01%and 13.40%,respectively.All three of these enhanced the microbial metabolic activity,stability and richness of microbial communities in rhizosphere soil of tobacco,while the evenness of these had been decreased.P.fluorescens significantly enhanced the utilization of carbohydrates and carboxylic acids,P.polymyxa significantly enhanced the utilization of carboxylic compounds and phenolic compounds,and Tugenben significantly enhanced the utilization of Phenolic compounds.The damage ponderance of tobacco bacterial wilt is well-known.By studying the antimicrobial activities,induced systemic resistance,effects of rhizosphere environment and field application,the results showed that both P.polymyxa and P.fluorescens had good control effects on tobacco bacterial wilt,and P.polymyxa showed more advantages.In production practice,we suggested applying P.polymyxa to control tobacco bacterial in soil acidification place. |
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