Screening Of Plant Growth-promoting Rhizobacteria And Analysis Of Growth-promoting And Biocontrol Mechanism Of Its Volatile Organic Compounds

With the development of agriculture,the excessive use of chemical pesticides and fertilizers increased production of crops,and caused great damage to the environment and human health at the same time.It has become the most important factor influencing the world's agricultural economic and social development.Thus,there is an urgent need for more effective and safe products.In recent years,microbial agents attracted much attention for its characteristics of being friendly to environment,disease prevention and yield increase.Plant growth promoting rhizobacteria(PGPR)colonizing the plant rhizosphere has many beneficial effects on crop growth and productivity.PGPR promotes the plant growth directly via phosphate solubilization,nitrogen fixation,phytohormone,siderophores and 1-aminocyclopropane-1-carboxylate deaminase production,volatile organic compound(VOC)production,biofilm formation,or protects plants via systemic acquired resistance or induced systemic resistence,acting as bioelicitors,rhizoremediation,tolerating stress,etc.VOC produced by PGPR have getted widespread attention,but of which there was no complete understanding of plant growth promoting and disease biocontrol mechanisms.Thus,the present study further clarified the mechanism of how to regulate plant growth and biocontrol diseases by VOC.Of 838 bacterial strains isolated from rhizosphere soil,107 strains were selected for amplified ribosomal DNA restriction analysis(ARDRA),based on their activities of nitrogen fixation,decomposition of organic phosphorus,solubilization of inorganic phosphorus,and production of IAA and siderophore in vitro.Of these 107 strains,20 potential microorganisms from different clusters were choosed to assessment for their plant growth promotion in greenhouse.And four strains 3 YB11,YB3 3,Y1 and JC01 were selected by their growth-promoting efficiency on different crops.3YB11 had best growth-promoting efficiency on water spinach and cotton by 35.87%,19.76%respectively.YB33 had best growth-promoting efficiency on rice by 13.97%.Y1 had best growth-promoting efficiency on wheat by 18.35%.JC01 had best growth-promoting efficiency on corn by 13.52%.Further study showed that JC01 and JC03 could produce VOC,They were identified as Bacillus subtilis by 16S rRNA sequence analysis.By using the bipartite Petri dishes(I-plates)system,of which A seedlings were transferred to one side and JC01 or JC03 was inoculated on another side,we found that they could promote A.thaliana well.The IAA signaling pathway key gene ctd1,ABA signaling pathway key gene SlABF4 and ETH signaling pathway key gene ERF2 were tested by Real-time PCR after treatment of JC01 and JC03.And the result showed that the expression of gen ctd1 was up-regulated by nearly 18,6 fold respectively,and that of ERF2 had no difference with the control.In addition,SlABF4 expression of JC01 treatment was dow-regulated,but that have no effect for JC03 treatment.To further discuss the mechanism of volatiles enhancing plant growth,JC01 and JC03 were tested against a series of defective A.thaliana mutants.While exposed to JC015 the strigolactones-insensitive mutant CCD7 and the IAA receptor mutant ARF1 did not exhibit any growth promotion,suggesting necessary roles for the Strigolactones(SL)and IAA signaling pathways in growth promotion of volatiles.And that of JC03 may be mainly dependent on IAA,SL signaling pathways,rather than primarily dependent on ethylene(ETH),abscisic acid(ABA),gibberellins(GA)pathways.10 specific chemical volatiles emitted from JC01 and JC03 were identified by employing Headspace-SPME/GC-MS.To evaluate the contribution of volatiles to A.thaliana seedling,we selected the pure compounds,inculding 3-Hydroxytetrahydrofuran,2-Heptanone,2-Decanone,2-Nonanone,2-Ethylhexanol,Tridecane instead of strains in I-plates system test.And we spotted different concentrations of the compounds to find the most effective dose.Plant growth promotion(PGP)efifect was highest,when 1?g/?L 3-Hydroxytetrahydrofuran,10ng/?L 2-Heptanone,0.1?g/?L 2-Decanone,10ng/?L 2-Nonanone,1?g/?L 2-Ethylhexanol,or 10ng/1?L Tridecane was applied respectively.While application of higher concentration than the best one,it would inhibite seedlings growth,even kill them.In addition,there had been lots of studies suggested that some compounds we have identified could have biocontrol efficiency to plant diseases.2-ethyl-1-hexanol could inhibit mycelial growth or sclerotia formation.2-nonanone and 2-heptanone had antibacterial,antifungal activities and killing activitys of nematodes as well as flies.2-Decanone has antibacterial effect.3-hydroxy-2-butanone(acetoin)and n-tridecane could protects plants by induced systemic resistance.So,these suggesting their potential role of JC01,JC03 in biocontrol of diseases.