Heavy Metal Adsorption Behavior And Transcriptome Sequencing Analysis Of Providencia Bacteria

In recent years,the promotion of China's industrialization and improvement of people's living standard level have led soil heavy metal pollution to become more serious.Microbial remediation has become a hot-pot research in soil heavy metal remediation due to its several advantages including no secondary pollution,short growth cycle,low operating cost and good remediation effect.In this study,we isolated a new Mn?II?tolerant bacterial strain,Providencia sp.,from heavy metal contaminated soil around a smelting plant in zhuzhou city,hunan province.The strain has high Mn?II?tolerance and biosorption which were proved by various biochemical and molecular techniques.Several characterization methods such as SEM,EDS,TEM,FT-IR,XRD,XPS and isothermal models were used to investigate the mechanism of absorbing heavy metals by bacterial strains.Biological leaching of the original heavy metal contaminated soil by bacterial liquid was used to prove the ability of the strain to repair heavy metal contaminated soil in practical application.Transcriptome sequencing analysis and real-time quantitative PCR were used to verify the differentially expressed genes and identify the resistance genes.1.Screening and identification of heavy metal tolerant strains.A bacterial strain with high tolerance to Mn?II?was screened from soil and sludge collected from heavy metal pollution area around a smelting plant in zhuzhou,hunan province and the target strain was identified as Providencia sp.LLDRA6.according to the results of physiological and biochemical identification.Its GenBank accession number:MH644827.1.2.Growth curve and characterization of the target strain.The growth curve of the target strain was studied and the optimum pH and the optimum growth temperature were around at 7,35?,separately.The tolerance of the target strain to six heavy metals was studied in detail at the same time.The results showed that the maximum resistances to Cd²⁺,Pb²⁺,Cu²⁺,Cr⁶⁺,Zn²⁺and Mn²⁺were 2,4,4,10,200,500 mM,respectively.Among the six heavy metals,The resistant effect to Mn²⁺was the most evident.The date of SEM-EDS,TEM confirmed that the strain had surface adsorption and intracellular enrichment of Mn²⁺.The date of FT-IR confirmed the movement of the peak for O-H,C-H,N-H,C-N,O-C-O and C-O after adsorption of manganese on the cell surface.XPS and X-ray diffraction further confirmed the LLDRA6 strain was able to oxidize Mn?II?to Mn?III?and Mn?IV?.3.The effects of solution's initial concentration?ranging from 25mM to 200 mM?,pH and temperature on Mn?II?biosorption.According to the pseudo-second-order kinetics and the Langmuir isotherm model,Mn?II?would be mostly adsorbed under optimal conditions,and the maximum biosorption capacity was 291.438 mg/g.The results of adsorption thermodynamic analysis indicated that the adsorption process of the strain was non-spontaneous and endothermic.The results obtained from biological leaching show that Pb,Cr,Cd,Cu,Mn and Zn in soil were removed by 66.3%?49.1%?56%?62.7%?50.7%and 45%,respectively.4.Transcriptome sequencing and analysis of bacterial strains.The results showed that the genome size was 4,34 Mb,and there were 3956genes,while the number of differentially expressed genes were 543,including 257 up-regulated genes and 286 down-regulated genes.Furthermore,GO functional annotation and KEGG biological pathway classification were performed for these differentially expressed genes.Twenty-four significant differential genes were selected from the differential genes for RT-qPCR verification,and the results were consistent with the results of the transcriptome analysis,which confirmed the accuracy of transcriptome sequencing results.