Screening Of Phosphate Solubilizing Bacteria In Poplar Rhizosphere Soil, Phosphate Solubilizing Characteristics And Growth-promoting Effect Of Rapeseed

Phosphorus deficiency in soil is an severe problem that restricts the development of modern agriculture.Despite the application of phosphate fertilizer can relieve phosphorus deficiency in soil partly,the utilization of phosphate fertilizer is low,and repeated application and abuse of phosphate fertilizer lead soil hardening,soil environmental pollution and other ecosystem problems.Therefore,the research on phosphorus-solubilizing microorganisms?PSB?rise to be a hot area.PSB can dissolve insoluble phosphate in soil and convert it into available phosphorus which can be absorbed and utilized by plants.In this study,the PSB strains were isolated and screened from poplar rhizospheric soil and they were identified through molecular biological methord.The phosphorus-solubilizing characteristics and mechanisms were studied on the high efficiency PSB strains.The growth-promoting effects of the high efficiency PSB strains were studied on canola.The main results were displayed as following.?1?Twenty one strains of inorganic PSB and 14 strains of organic PSBwere isolated and identified.The inorganic PSB strains and organic PSB strains were isolated from the rhizospheric soil of poplar by plate preliminary screening and liquid shake flask culture rescreening on NBRIP and yolk culture media respectively.Through 16S r DNA sequencing and phylogenetic tree analysis,the PSB strains were identified.The inorganic PSB strains were clustered under the genera Pantoea,Ensifer,Cedecea,Rhizobium,Staphylococcus,Cellulosimicrobium,Bacillus,Phyllobacterium.The organic PSB strians were clustered under the genera Bacillus,Delftia,Paenibacillus and Pseudomonas.The two representative strains with high PS activity were identified as Staphylococcus succinus Mp1-Ha3 and Cedecea sp.Mp1-Ha4.?2?The phosphorus-solubilizing activity of 21 inorganic PSB strains and 14 organic PSB strains were determined in NBRIP and Monkina medium,respectively,by molybden um-antimony colorimetric method.The activities of dissolving tricalcium phosphate of 21PSB strains are higher than that of dissolving aluminum phosphate and iron phosphate.Among these inorganic PSB strains,S.succinus Mp1-Ha3,Cedecea sp.Mp1-Ha4,Rhizobium sp.Mp1-Ha7,P.ifriqiyense Mp1-Ha8,Pantoea sp.Mp2-Ha11 showed higher PS activity to tricalcium phosphate,the soluble phosphorus dissolved from tricalcium phosphate by which all beyond 300 mg/L.Furthermore,the PS activities the inorganic PSB strains were negatively correlated with the media p H value and positively correlated with the media titratable acidity.Results of PS Kinetics showed the Mp1-Ha3 and Mp1-Ha4 could secrete organic acids,such as gluconic acid,citric acid,formic acid,pyruvic acid,?-ketog lutaric acid etc.during the PS process.These results elucidated that inorganic PSB strains secrete organic acids to reduce the p H of the environment,thereby dissolving insoluble phosphate and releasing phosphate ions.Fourteen organic PSB strains showed capacity to dissolve lecithin,among which Bacillus sp.Op4-Ha3 and B.cereus Op1-Ha2 have higher PS activities to lecithin,and the dissolved phosphate concentrations in the media were 9.07 mg/L and 2.23 mg/L respectively.?3?In SP medium,K₂HPO₄was added in as exogenous soluble phosphorus source and the levels of exogenous soluble phosphorus were set up as 0,0.5,1.0,5.0,10.0,20.0 and 25.0mmol/L respectively.The effect of exogenous soluble phosphorus on the PS ability of inorganic PSB strain S.succinus Mp1-Ha3 and Cedecea sp.Mp1-Ha4 were determined in the SP media.Along with the increase of the concentration of exogenous phosphorus,the PS ability of the PSB strains Mp1-Ha3 and Mp1-Ha4 gradually decreased,with a negative correlation with the media p H value and a positive correlation with media titratable acidity.The secretion of organic acidsby the PSB strains Mp1-Ha3 and Mp1-Ha4 gradually decreased as the exogenous phosphorus concentrations rised up,and the decrease of gluconic acid secretion was the most obvious.Exogenous soluble phosphorus might inhibit the PS activities of the PSB strains Mp1-Ha3 and Mp1-Ha4 by restraining the secretion of organic acids,especially the gluconic acid.?4?Canola seeds soaked with bacterial suspension of inorganic the PSB strain S.succinus Mp1-Ha3,Cedecea sp.Mp1-Ha4 and the fermentation supernatants germinated earlier and had higher germination rate than that soaked with sterile water.The PSB strains Mp1-Ha3 and Mp1-Ha4 were inoculated on potted canola seedlings respectively,and sterile water was inoculated as control.The seedling height,root length,wet weight and dry weight of the canolo seedlings inoculated with Mp1-Ha3 were 1.26,1.21,1.34 and 1.62 times,respectively,of the control on 60 days after inoculation,correspondingly 1.08,1.33,1.31,1.35 and 1.27 times for Mp1-Ha4.The released soluble phosphorus content of canola rhizosphere soil inoculated with Mp1-Ha3 and Mp1-Ha4 strains were significantly higher than that of control.Both strains could stably colonize in rhizosphere,root,stem and leaf of the canola seedings.These results indicated that,the inorganic PSB strains Mp1-Ha3 and Mp1-Ha4 could promote germination of canola seeds.In addition,the strains could increase soil phosphorus availability in rhizosphere and stablely colonized in rhizosphere and canola tissue,subsequently,which lay the foundation for canola growth promotion.