Screening Of Plant Growth-promoting Rhizobacteria Based On Utilization Ability For Rhizosphere Nutrition And Investigation Of Promoting Effects

Excellent microbial strain is the basis of microbial fertilizer,rhizosphere competitiveness is the prerequisite for the performance of plant beneficial roles of the bacterial strains in fertilizer.Screening of plant growth-promoting rhizobacteria must be well adapted to the plant rhizosphere environment.In this study,the root exudates of maize,cucumber and tomato were used as the sole nutrient source of the screening medium respectively,to isolate the fast growing PGPR from the corresponding plant rhizosphere.Their plant growth-promoting characters were determined and further verified by petri dish seedling promoting assay,then the screened strains were evaluated in greenhouse pot experiment and identified by analyzing 16S rDNA.Finally,four Bacillus spp.,JScG,SCmB,XJmG,SDtG were tested for their ability to colonize the rhizosphere of different seedlings,and were optimized for liquid fermentation,the microbial fertilizers based on these strains were developed and their plant-growth promoting effects in pot experiment were investigated.The main results obtained were as follows:1.24 strains were isolated from the first round screening,these strains can grow quickly in the screening medium using root exudates as the sole nutrition source,and having one or more of the following abilities:phosphate solubilization,production of NH3,IAA and siderophores.14 strains were capable of producing ammonia,accounted for 58.3%;20 strains can produce IAA,accounted for 83.3%,JScB strain had the highest production of IAA,reached 73.28 mg L-1;12 strains can produce siderophore,accounted for 50%;15 strains were capable of dissolving organic phosphorus,accounted for 62.5%;All strains had ability of dissolving inorganic phosphorus,but the difference was extensive,ranged from 19.14 to 200.05 mg L-1.12 PGPRs were screened according to the petri dish seedling promoting assay.PGPRs strains that had growth promoting ability for maize were SCmB,SCmD,NMGmA,XJmG;PGPRs strains of SDtC,SDtG,GZtB,GZtE promoted the growth of tomato;PGPRs strains that had growth promoting ability for cucumber were HNcB,HNcH,JScB,JScG.2.The plant growth-promoting effects of 12 PGPRs were eventually determined in pot experiment,inoculation of them to the rhizosphere of corresponding plants(maize,cucumber,tomato respectively).After forty days,the plant height,biomass,root morphology(total root length,root surface area,etc.)and chlorophyll content were significantly higher than control.Through 16S rDNA molecular identification,five isolates were identified as Bacillus genera(Bacillus spp.);three isolates were belong to Ensifer genera(Ensifer spp.);two strains were indentified as Rhizobium genera(Rhizobium spp.);One strain was Ochrobactrum genus(Ochrobactrum sp.);One strain was Microbacterium genus(Microbacterium sp.).In the pot experiment of corn,strain SCmB had the best effect,followed by XJmG;In tomato pot assay,GZtB strain performied the best,followed by SDtG;In cucumber pot experiment,strain HNcB performed the best,followed by JScB and JScG.Four strains of the seven best performed strains were Bacillus spp.,which can produce heat-resistant spores,thus they were ideal strains of microbial fertilizer.We chose these four strains of SCmB,XJmG,SDtG,JScG for further research.3.Biofilm formation and colonization ability for maize,cucumber,tomato of these four Bacillus spp.strains(JScG,SCmB,XJmG,SDtG)were determined.SDtG and JScG had the strongest ability of biofilm formation,followed by SCmB,and XJmG.In the sterile quartz sand colonization experiment,SCmB performed best in maize rhizosphere colonization,reached 9.29×106 CFU g-1 root;In cucumber rhizosphere,SDtG and JScG had the strongest colonization abilities,reached 4.69×106 CFU g-1 root and 4.12×106 CFU g-1 root respectively;XJmG performed the best in tomato rhizosphere,reached 8.07×106 CFU g-1 root.4.Liquid fermentation media of these four Bacillus spp.strains(JScG,SCmB,XJmG,SDtG)were optimized using single factor and orthogonal tests.The optimal liquid fermentation medium of JScG and SCmB was as following:3%molasses,0.6%chemical nitrogen source,0.5%sodium chloride,0.1%calcium chloride,0.05%manganese sulfate;for XJmG:2.5%molasses,0.6%chemical nitrogen source,0.5%sodium chloride,0.1%potassium dihydrogen phosphate,0.1%calcium chloride,0.1%magnesium sulfate,and 0.05%manganese sulfate;for SDtG:3%molasses,0.6%chemical nitrogen source,0.25%sodium chloride.5.According to the growth curves of these four Bacillus spp.strains(JScG,SCmB,XJmG,SDtG)in the liquid fermentation medium,10%(v/w)fermentation cultures at 72 h of these four strains were used to mix with compost to make the microbial fertilizers,withthe spore number reached about 5.0×108 CFU g-1 dry weight,applying 0.5%of these microbial fertilizers to corn and tomato rhizosphere in pot experiment,the plant height,biomass and root morphology(total root length,root surface area,etc.)were significantly higher than control after thirty-five days,SCmB worked best for corn,XJmG performed best for tomato.