Effect Of Bio-matrix Pot Addition On Phytoremediation Of Iron Tailing Sands And The Microbial Communities

Tailing sand is a kind of granular material with high compactness and small particle size produced in smelting or beneficiation process.It is easy to flow and leak,which often leads to dam break accident of tailing pond.Vegetation restoration is an effective bioremediation method for dam prevention.In this paper,the effects of phytoremediation on tailings sand with bio-matrix pot combined Pennisetum giganteum and Medicago sativa were studied.The change of plant biomass,nutrient of tailings sand,microbial communities were performed;The effective growth-promoting bacteria screened from rhizosphere of P.giganteum,the growth-promoting effect on maize and P.giganteum was explored after inoculation of effective growth-promoting bacteria.Firstly,the effect of bio-matrix pot on the growth of P.giganteum,nutrient content and microbial characteristics of the tailings was investigated.The result showed that,compared with control,the shoot dry weight and root dry weight of P.giganteum increased by 1.34times and 1.24 times respectively,at 360 d after adding bio-matrix pot.The contents of total nitrogen,total phosphorus and total potassium increased by 11.21%,6.57%and 10.91%,respectively.Organic matter,alkali-hydrolyzable nitrogen,available phosphous and available potassium increased by 27.21%,5.50%,98.33%and 58.85%,respectively.The numbers of bacteria,fungi and actinomycetes increased by 28.78%,16.46%and 32.27%,respectively.The activities of urease,phosphatase and catalase in iron tailing increased significantly.BOXA1R-PCR analysis of the bacteria isolated from the rhizosphere showed that,74 strains got at the 90 day’s sample formed 11 genetic groups at 48%similar level;the 100strains got at the 180 day’s sample were divided into 12 genetic groups at 46%similar level;the 128 strains obtained from the 360 day’s sample formed 11 genetic groups at 45%similar level.Phylogenetic analysis of 16S r RNA gene sequence of representative strains showed that these strains were distributed in 4,4 and 11 phylogenetic clades,indicating that bacterial population diversity was increasing during restoration.The bacterial communities of tailings sand on the 90^th,180^th and 360^th days were analyzed by Miseq sequencing.At the phylum level,Proteobacteria was the most abundant,accounting for 25.73%-51.37%;and followed by Acidobacteria,Chloroflexi and Actinobacteria.After adding bio-matrix pot,the abundance of Chloroflexi,Euryarchaeota,Gemmatimonadetes,Latescibacteria and Planctomycetes increased obviously at the 360^thday.At the genus level,the abundances of Geobacter,Sideroxydans and Geoalkalibacter were significantly increased,which were 3.93%,3.98%and 0.98%,respectively.The KEGG results showed that,addition of bio-matrix pot,the abundance of glycosyltransferase,glycosylhydrolyase,nitrogenase gene and denitrifying genes involved in carbon metabolism in rhizosphere tailing sand increased obviously at 360 d.Then,this paper discussed the effect of bioremediation of tailings by using bio-matrix pot combined with M.sativa and P.giganteum.The results showed that the shoot dry weight and root dry weight of P.giganteum was 1.60 times and 1.71 times of that of M.sativa,respectively,and the total potassium content was 1.53 times of that of M.sativa.However,the contents of total nitrogen and total phosphorus in M.sativa were 4.54 and 1.78 times higher than those in P.giganteum.Planting M.sativa and P.giganteum effectively increased the content of organic matter,alkali-hydrolyzed nitrogen,and available potassium in the rhizosphere tailing sand.The highest organic matter content was 6.09 g·kg^-1 in the rhizosphere tailing sand of cultivating M.sativa.The contents of alkali-hydrolyzable nitrogen and available potassium in rhizosphere tailing sand were 24 mg·kg^-1 and 49.35mg·kg^-1,respectively;Cultivating M.sativa and P.giganteum increased the nitrogen fixation ability of the rhizospheric tailings.Miseq sequencing platform was used to analyze the bacterial communities of tailings samples from each treatment on 360^th day.Proteobacteria,Actinobacteria,Firmicutes,Acidobacteria,Chloroflexi and Patescibacteria were the dominant phylum;Of them,Proteobacteria occupied the most abundant,with the abundance range from 40.36%to52.21%,and the samples of M.sativa rhizosphere tailing ore were the highest.There were significant differences in the dominant flora in the rhizosphere tailing sands of different plants.Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium was the dominant bacteria in the M.sativa rhizosphere tailing（12.16%）,while Bacillus was the most abundant in the rhizosphere tailing samples of P.giganteum（12.23%）.Metagenome Seq analysis showed that,at the genus level,Allorhizobium-Neorhizobium-Pararhizobium-Rhizobium,Mesorhizobium,Novosphingobium,Brevundimonas,Nocardioides,Saccharimonadales and Ilumatobacter were significantly enriched in the rhizosphere tailing sand of M.sativa;However,Bacillus,Pseudomonas,Haliangium,Ramlibacter and Iamia were significantly enriched in the rhizosphere tailings of P.giganteum.Analysis of fungal communities in tailings by Miseq analysis showed that,Ascomycota,Mucoromycota,and Basidiomycota（abundance less than 1%）were the dominant at phylum level.Fusarium,Humicola,Rhizopus,Ascobolus and Staphylotrichum were the dominant（abundance>5%）at the genus level.Furthermore,this paper studied the growth promoting effect of rhizosphere bacteria on maize and P.giganteum to prove the growth promoting effect during restoration.Three effective growth-promoting bacteria were screened from 65 strains isolated from the rhizosphere of P.giganteum,which were Acinetobacter Soli S18166,Bacillus Flexus S18232,Pseudomonas laurylsulfativorans S18259.Growth promotion test showed that the maximum leaf area of maize seedlings increased 43.49 cm²and 46.18 cm² after inoculation of S18232 and S18259.The highest peroxidase（POD）and superoxide dismutase（SOD）activity were 4554 mg·g^-1·h^-1 and 49.92 mg·g^-1·h^-1 in maize seedlings inoculated with S18259.Field experiment showed that the inoculation effect of S18232 was the best,with the maximum yield of 295.8 g per plant,the highest soluble sugar content and protein content,27.52 mg·kg-1 and 7.82%,respectively.Compared with the control,the chlorophyll activity,root activity and dry weight increased by 13%,30%and 41.53%after inoculation with S18232.q PCR results showed that the copies of 16S r RNA and nif H genes in tailings increased by 1.25 and 1.11 times after inoculation with S18232,reaching a significant difference level（p<0.05）.This paper suggested that P.giganteum and M.sativa have good restoration ability to tailings,the addition of bio-matrix pot can significantly promote plant growth,M.sativa and P.giganteum can significantly improve the physical and chemical properties and microbial diversity of tailings.The growth promotion test showed that more than 83%of the rhizosphere bacteria of P.giganteum had the promotion ability,and Bacillus Flexus S18232had a good growth promotion effect,indicating that the dominant bacteria enriched in the rhizosphere tailing sand played a positive role in the restoration process of the tailing sand.