Screening,identification And Growth-Promoting Effects Of PGPR From Blueberry Rhizosphere

Blueberry is an important commercial crop and the cultivated area in Shandong Liaoning province is continuously extending.The root of blueberry is undeveloped and the ability of nutrients absorbtion is poor.In addition,the soil pH of blueberry suitable is 4.5-5.5.As a result,the soil condition restricts production of blueberry.PGPR(Plant Growth-Promoting Rhizobacteria)can promote root development,improve soil environment,and contol soil-borne disease.We screened IAA producing bacterium,protease producing bacterium and phosphate solubilizing bacterium and identified the strains.And then,we used the plug seeding and container seeding pot experiments verifying the effects of the selected strains.BIA070 was labeled with GFP to investigate its colonization ability in the blueberry rhizosphere.We used the acid producing ability of microorganisms regulating acid-base property of blueberry soil.The HPLC technology was conducted to analyze the type and content of organic acid in the fermentation liquor.Finally,we tested the survival ability and effects on blueberry soil pH of these acid producing microorganisms by means of soil culture.Thus we can lay the foundation of popularization and application of blueberry microbial fetility.We screened 14 IAA producing bacterium,12 protease producing bacterium and 6 inorganic phosphate solubilizing bacterium.The preliminary plug seeding and container seeding pot experiments confirmed 10 PGPRs which could promote the blueberry growth.And the identifying results showed that BIA032 belongs to Leucobacter aridicollis,BIA044,BIA088,BPR082,BPR088,BPR176 belong to Bacillus aryabhattai,BIA070,BIA141 belong to Enterobacter cloacae,BPR142 belongs to Bacillus amyloliquefaciens,and BIP191 belongs to Pantoea dispersa.In the preliminary pot experiments,BIA032,BIA044,BIA088,BIA141,BPR082,BPR142 and BIP191 improved the plant height and ground diameter of blueberry container seeding.BIA070,BPR088,BPR176 promoted the growth of blueberry plug seeding.In detail,the plant height and the dry weight overground increased significantly compared with CK.The plant height increased 38.92%,61.23%,35.05% respectively,and the dry weight overground increased 64.13%,92.65%,47.58% respectively.The two strains BPR088,BPR176 also increased the ground diameter of the plant significantly.The growth rates were 24.58%,17.47% respectively.The effect of BIA070 on ground diameter was not significant.BPR176 improved the chlorophyll content by 25% compared with CK.BIA070 and BPR088 decreased the MDA content and also increased the peroxidase activity in the leaves.A plug seeding experiment was conducted to make further testing of the selected 10 PGPRs.The results showed that the following 5 strains could promote the growth of blueberry obviously.They could improve the ground diameter significantly,and growth rates were 22.16%,36.22%,21.70%,18.29%,22.06% respectively.The strains also increased the plant height,the leaf number,and the fresh weight and dry weight overground to some extent.The dry weight overground of plant increased 38.89%,35.19%,37.04%,14.18%,29.63% respectively,and the differences were significantly except BPR088.PGPR could also increase the total nitrogen content in the plant.BPR176 produced significant increase compared with CK,and its total nitrogen content increased 23.82%.Moreover,the activities of acid phosphatase,protease,invertase in the soil increased after inoculating the 5 strains.The content of effective nitrogen,rapid available phosphorus,and the organic matter also increased after using the selected strains.The high-throuput sequencing results sugested that Proteobacteria,Acidobacteria and Bacteroidetes are core phyla of bacterial community,and Ascomycota and Basidiomycota are core phyla of fungi community in blueberry rhizosphere soil.BIA070 and BPR176 both changed the bacterial community and fungi community.We labeled BIA070 with GFP,and obtained the labeled strain BIA070(gfp).The gfp gene could keep stable inheritance in BIA070 in the test of plasmid stability.And the pot experiment of BIA070(gfp)showed that the strain could colonize in the blueberry rhizosphere.We isolated 12 acid producing microorganisms from blueberry rhizosphere,11 among them are bacteria,and 1 among them was yeast.The identification results showed that BAC006,BAC011,BAC062 belong to Enterobacter aerogenes,BAC025,BAC031,BAC049,BAC064 belong to E.coli,BAC007 and BAC041 belong to Citrobacter,BAC067 belongs to Enterobacter cloacae,BAC068 belongs to Yokenella regensburgei,and BAC089 belongs to Candida tropicalis.The result of HPLC showed that the types of organic acid which bacterium produced were mainly oxalic acid,acetic acid,and citric acid.The fermentation liquor of BAC006 was mainly oxalic acid and the concentration was 1393.316?g/m L.The fermentation liquors of BAC007,BAC031,BAC067 and BAC068 was mainly acetic acid and the concentration were 9741.763?g/m L,18557.486?g/mL,7731.176?g/mL,9008.221?g/m L respectively.The types of organic acid Candida tropicalis produced were mainly tartaric acid,acetic acid,and succinic acid,and the concentration of succinic acid reached 43055.475?g/m L.We conducted the soil culture experiment and the results showed that BAC068 and BAC089 could survive stablely in the blueberry soil and could decrease the soil pH to some extent.