Passivation Of Pb In Various Contaminated Soil By Phosphate Rock Combined With Phosphate-solubilizing Bacteria

Phosphate rock（PR）can reduce the bioavailability of lead in soil by adsorbing or forming pyromorphite Pb₅（PO₄）₃X（X=Cl^-、OH^-、F^-）,the most stable environmental Pb compounds under a wide range of pH and Eh natural conditions.However,the effectiveness of phosphate rock on Pb immobilization was limited result of its low solubility.Phosphate solubilizing bacteria（PSB）not only promote the dissolution of P from insoluble P compounds,but also dissolve the insoluble P in the soil,which could enhancing the passivation of phosphate rock in the lead soil.Therefore,the objectives of this study are:（i）to explored the dissolving mechanism of 4 strains of PSB which soluting Ca₃（PO₄）₂、FePO₄·4H₂O and AlPO₄,and futher indentify the variation of inorganic P in the lead soil by inoculating 4 strains of PSB;（ii）to examine the remediation of heterogeneous soil by phosphate rock amendment with PSB,and obtained an applicable method to evaluate the bioavailability of heavy metal in soils.The major results of this paper were summarized as follows:（1）The dissolving ability of Ca₃（PO₄）₂ by four strains was shown as:Pseudomonas fluorescens II₈L₄-GFP>Bacillus thuringiensis GL-1>Pseudomonas fluorescens IV₈R₂-GFP>Pantoea ananatis HCR2.The largest amount of phosphorus solubilization was590.60 mg/L,430.17 mg/L,327.71 mg/L and 291.13 mg/L,respectively.The ability of Bacillus thuringiensis GL-1 and Pantoea ananatis HCR2 solubilizing FePO₄·4H₂O were shown as:Pantoea ananatis HCR2>Bacillus thuringiensis GL-1,the maximal amount of phosphorus solubilization was 56.81 mg/L and 37.67 mg/L.Nevertheless,Pseudomonas fluorescens II₈L₄-GFP and Pseudomonas fluorescens IV₈R₂-GFP were almost no effect on FePO₄·4H₂O.The capacity of 4 PSB strains solubilizing AlPO4 was shown as:Pantoea ananatis HCR2>Bacillus thuringiensis GL-1>Pseudomonas fluorescens II₈L₄-GFP>Pseudomonas fluorescens IV₈R₂-GFP,the largest amount of dissolving phosphorus was 119.35 mg/L,106.86 mg/L,94.38 mg/L and 79.01 mg/L,respertively.（2）The pH was sharply decreased,when the 4 PSB was inoculated in the NBRIP growth agar medium with Ca₃（PO₄）₂,FePO₄·4H₂O or AlPO₄ as the only phosphorus sources.After4 days of cultivating,the organic acids were indentified by GC-MS.When Ca₃（PO₄）₂ as only P source,Bacillus thuringiensis GL–1 secreted by pyruvic acid,α-Ketopvalerate acid,citric acid and Succinic acid.Pseudomonas fluorescens II₈L₄-GFP producedα-Ketopvalerate acid,Succinic acid and Propionic acid.Pseudomonas fluorescens IV₈R₂-GFP only secreted oxalic acid.When FePO₄·4H₂O as solo P source,Bacillus thuringiensis GL–1 secreted oxalic acid,pyruvic acid,α-Ketopvalerate acid,citric acid and Succinic acid.Pantoea ananatis HCR2 released glucose acid,pyruvic acid and citric acid.When AlPO₄ as only P source,Bacillus thuringiensis GL–1 secreted glucose acid,α-Ketopvalerate acid and citric acid.Pantoea ananatis HCR2 only produced glucose acid.Pseudomonas fluorescens II₈L₄-GFP released glucose acid,Succinic acid and Malonic acid.Pseudomonas fluorescens IV₈R₂–GFP secreted glucose acid and Succinic acid.Besides,the phosphatase also attributed to P solubilization.It’s detected alkaline phosphatase was positively correlated with Ca₃（PO₄）₂ dissolving while inoculated with Bacillus thuringiensis GL–1and Pseudomonas fluorescens II₈L₄–GFP.（3）Sterile soil cultivation experiment indicated that inoculating Bacillus thuringiensis GL–1,Pantoea ananatis HCR2 and Pseudomonas fluorescens II₈L₄–GFP was significantly decreased the available phosphorus content（14.94%-30.01%）and available Pb concentration in the simulated lead polluted vegetable soil（27.87%-46.54%）.But inoculating Pseudomonas fluorescens IV₈R₂–GFP had no effect on that.4 strains of PSB could reduce the available Pb concentration（9.99%-14.40%）and increased the available phosphorus content in acid muti-metal contaminated soil（except for Bacillus thuringiensis GL-1,significantly reduced available phosphorus content by 4.55%）.In neutral muti-metal contaminated soil,inoculation of Bacillus thuringiensis GL–1,Pseudomonas fluorescens II₈L₄–GFP and Pseudomonas fluorescens IV₈R₂–GFP had no obvious effect on the available phosphorus content,but inoculating Pantoea ananatis HCR2 significantly increased available phosphorus content by 5.43%.（4）By texting the inorganic phosphorus forms in soil,some results were obtained.To reduced the available Pb concentration in simulated lead contaminated vegetable soil,Bacillus thuringiensis GL–1 mainly solubilized Fe-P and Ca-P;Pantoea ananatis HCR2dissolved Al-P and Fe-P;Pseudomonas fluorescens II₈L₄–GFP had obvious effect on all kinds of inorganic P morphology;while Pseudomonas fluorescens IV₈R₂–GFP had no significant effect on various inorganic P in the soil.To precipitated the available Pb in acid muti-metal contaminated soil,4 PSB all significantly dissolved Fe-P,in addition,Pseudomonas fluorescens IV₈R₂–GFP also distinctly dissolved Ca-P.For neutral muti-metal contaminated soil,4 PSB were mainly solubilized Al-P to precipitate the available Pb.（5）Pot experiment reveled that the addition of phosphate rock and inoculating PSB（Bacillus thuringiensis GL–1 and Pantoea ananatis HCR2）not only stimulated the growth and biomass of lettuce（Lactuca sativa L.）,but also reduced its absorption of lead.In the simulated Pb contaminated vegetable soil,adding PR and inoculating PSB could decreased the NH₄NO₃-Pb concentration（8.47%-37.87%）and DGT-Pb content（28.30%-43.90%）,which the effect of inoculating Pantoea ananatis HCR2 was more remarkable.In the simulated Pb contaminated paddy soil,adding PR and inoculating PSB significantly reduced the content of NH₄NO₃-Pb（14.14%-33.37%）,and inoculation of Pantoea ananatis HCR2 was more positive to precipitate Pb（NH₄NO₃-Pb and DGT-Pb content were decreased 29.29%-33.37%and 7.52%-22.56%）.For the neutral muti-metal contaminated soil,the addition of PR and inoculating PSB prominently reduced the content of DGT-Pb（28.09%-38.24%）,It’s still inoculating Pantoea ananatis HCR2 had optimal effect to fixed avaible Pb.（6）Using DGT synchronously examined the Pb concentration and P content.For the neutral muti-metal contaminated soil,the available Pb counld not be extracted by NH₄NO₃.Nevertheless,the DGT technology could accurately express lead concentration in soil,and it had a positive correlation with the lead concentration in plant（P<0.05）.Besides,the P content in the soil which was extracted by DGT was more exact than traditional extraction,that is NaHCO_3.All results recommended that the DGT technology was a feasible way to accurately express the available Pb in soil and the P content which was absorbed by plant.