The Effect Of Plant Growth-promoting Endophytic Bacterial Strain S61 On The Quality Formation Of Astragali Radix And Associated Mechanism

Background: A growing body of evidences has shown that plant growthpromoting rhizosphere bacteria(PGPRs)play as important members of the host symbiotic system by producing HCN,iron carriers,antibiotics and nutrients,blocking ethylene production,inducing systemic resistance,and increasing phosphate dissolution and nitrogen fixation.In brief,PGPRs are widely involved in processes such as plant growth and defense responses.However,the beneficial roles of PGPRs is still in infancy in medicinal plants as well as underlying mechanism,which is urgent to carry out associated research to explore their application in green cultivation of medicinal plants.Objective: Endophytic bacterial strain S61 has been isolated from fresh roots of Astragalus mongholicus in Hunyuan,Shanxi and can produce plant growth hormone indole acetic acid(IAA).Here,the strain was adapted to explore its potential role in the quality formation of Astragali Radix under normal water supply and drought stress conditions.In our view,this was the first report on the beneficial roles and associated mechanism of endophytic bacteria in the quality formation of Astragali Radix.Methods:(1)Extraction and quantification of extracellular polysaccharides: overnight liquid culture of the strain S61 has been used and the extraction was performed by alcohol precipitation,and the quantification was performed by phenol-sulfuric acid method.(2)The strain growth and IAA production has been characterized by measuring OD600 and OD530 in the liquid cultures under variable stressed conditions(i.e.,salinity,drought and acidility)as well as extreme stressed conditions.(3)Experiment designment and sample collection: the seeds were firstly surface-disinfected,then incubated with the strain S61 and germinated under light conditions,finally transplanted into the pots after the true leaves expanded.When the seedlings had grown for six and five weeks,they were treated with 20% PEG6000 and hexanal for 4 and 2 weeks,respectively;after the experiment,the root and shoot parts including mature leaves were collected from individual seedlings,weighted then frozen with liquid nitrogen,and finally stored at-80?.Pretreatment with phenidone had been performed to understand whether the mediation of hexanal was involved in the lipoxygenase signaling.(4)Chemistry analysis: the spectrophotometric method had been performed to determine the contents of chlorophylls and lipid peroxidation products;HPLC and UPLC-MS had been performed to determine the contents of calycosin-7-glucoside,ononin and astragaloside IV,respectively.Major results:(1)Endophytic strain S61 can produce extracellular polysaccharide and be more robust in the production of IAA under drought stressed conditions than the others.(2)Under drought stressed condition,the seedling growth and root accumulation of the two isoflavone compounds were significantly improved by the incubation of the endophytic strain S61 while the strain S61 exhibited no growth-promoting effect under normal water supplying condition,which might be related to the higher level of the lipid peroxidation in the shoot part of A.mongholicus.(3)Under normal water supply condition,the incubation of the endophytic strain S61 synergistically promoted the seedling growth of A.mongholicus as well as the root accumulation of the two isoflavone compounds.The synergistic effect also included the inhibition the transformation from chlorophyll a to chlorophyll b and decreased lipid peroxidative damage of the aerial part of A.mongholicus caused by n-hexanal and the strain alone.Phenidone pretreatment further showed that the lipoxygenase signaling pathway could inhibit the biosynthesis and accumulation of astragalosides,calycosin-7-glucoside and formononetin in control and treated seedling roots with n-hexanal.And the lipoxygenase signaling was not associated with the synergistic promotion of the combination of the strain S61 and n-hexanal.Conclusion: The endophytic strain S61 of A.mongholicus imporved the host plant growth and the root accumulations of calycosin-7-glucoside and ononin under drought and normal water supply conditions which combined with hexanal treatment.The promotion under drought stress might be related to the increased level of lipid peroxidation in the aerial parts of A.mongholicus seedlings while the synergistic promotion by the combination of the strain and n-hexanal was regardless of the lipoxygenase signaling under normal water supply condition.