Arsenic Mobilization Affected By Dissimilatory Iron Reducing Bacteria From High Arsenic Groundwater In Hetao Basin

Arsenic(As)in groundwater is one of the most serious environmental problems worldwidely.More than 140 million people are at the risk of high-As groundwater in the world.In China,high-As groundwater threatens the lives of as many as 19.6 million people.In recent years,many studies have indicated that microorganisms,especially dissimilatory iron reducing bacteria(DIRB),play an important role in As mobilization,transformation and release.At present,the mechanism of DIRB affecting As mobilization remains controversial,and the key factors are not clear.In this study,the study area is located in Hangjinhouqi of Hetao Basin,the typical high-As groundwater area.An integrated approach both geochemical and microbial molecular method was used to investigate the microbial community structure and abundance of typical DIRB in sediments and groundwater.The different effects of various DIRB on iron reduction and As mobilization were analyzed.And the key factors which control As mobilization were studied.The effect of different DIRB EPS on the mobilization and transformation of As were studied.This study provides a theoretical basis for the remediation of high-As groundwater.The main results are as follows:1.In groundwater and sediment,the community structure and abundance of DIRB are different shapped by various environmental factors.The 16S r RNA gene diversity and abundance of two typical DIRB,Shewanella and Geobacter,were analyzed.The results showed that Geobacter argillaceus and Shewanella xiamensis were the dominant population in the high-As groundwater,and G.chapellei and S.glacialipiscicola dominanted in the low-As groundwater.The diversity of the DIRB decreased with the increasing concentration of As.The dominant species of DIRB in sediments were G.bremensis,G.chapellei,S.benthica and S.algae,and the diversity of DIRB increased with increasing depth.The average abundance of Geobacter in groundwater(1.97×10¹-4.30×10⁶ copies/L)was higher than Shewanella(1.07×10¹-4.02×10⁶ copies/L).The mean abundance of Geobacter in shallow(<10 m)sediments(0-7.02×10⁵ copies/kg)was lower than that of Shewanella(6.62×10¹-5.53×10⁵copies/kg),and most of the abundance of the two genera of DIRB was at the same order of magnitudes with that of in deep(>10 m)sediments.Fe(?)/Fe_tot(Total Fe),p H,ORP and depth are the key environmental factors shaping the distribution of typical DIRB in the groundwater.In the samples with high p H value,Geobacter was in higher abundance,and Shewanella predominated in the groundwater samples with higher ORP value.In the sediments,the relative abundance of Geobacter and Shewanella increased with depth.The relative abundances of DIRB have no significant correlation with As(?)and As_tot concentration in the groundwater,which implied that microbial mediated iron reduction might not always cause As release at some high arsenic groundwater.2.With the isolated DIRB strain from high-As groundwater,the effect of iron reduction process on As mobilization and transformation was studied.In this study,eight strains were isolated and numbered as IR-CL1,IR-PC1,IR-R1,IR-K1,IR-K2,IR-C1,IAR-S1 and IR-S2,respectively.According to the 16S r RNA results,these strains belonged to Clostridium,Paraclostridium,Raoultella,Klebsiella,Citrobacter and Shewanella,respectively.All the strains could reduce more than 99%of 20 m M Fe(?)within 5 days,only strain IAR-S1 was found with As reducing capability.Different DIRB showed various effects on As mobilization.Shewanella IAR-S1 and IR-S2,MR-1 and Geobacter IR-G1 strains immobilized As during As-bearing ferrihydrite reduction,Klebsiella IR-K1 strain released As into aqueous phase from ferrihydrite.The X-ray diffraction(XRD),Energy-dispersive mapping(EDS)and scanning electron microscopy(SEM)were used for the mineralogical analysis of final products.The results showed that As-immobilized DIRB generated vivianite and magnetite under the conditions of phosphate and carbonate,and large amount of biological organic carbon were found to participate with the biomineralization.The final p H value was 7.5,and the content of polysaccharide in EPS is high.The As-released DIRB did not generate secondary minerals under both conditions,and biological organic carbon did not participate with the end products.The final p H value of As-released DIRB iron reduction was 6.0,and the polysaccharide of generated EPS was less than that of As-immobilized DIRB.The results suggest that p H values,EPS,especially polysaccharides,may be an important factor affecting the formation of secondary minerals.3.The various factors affecting the As mobilization were studied,including p H values,geochemical characteristics,As reducing ability and the species of As-bearing minerals.The HEPES(4-hydroxyethyl piperazine ethylsulfonic acid)solution and sodium citrate buffer solution were used to adjust the p H of the reaction system.The results showed that the amount of As release was decreased in the weakly alkaline environment,while the amount of As release was promoted in the weakly acidic environment.The slight variation of p H value(6.0-7.5)did not change the mobilization trend of As in the iron reduction process by DIRB.Phosphate,carbonate and sulfate were added into the microbial incubation system to simulate the in situ different geochemical types,and the results showed that the order of effect on As release was phosphate>sulfate>carbonate.The contribution of sulfate and carbonate on As release is much greater than phosphate in our study area.The difference of geochemical types could only affect the amount of As mobilization.The As-reducing bacteria AR-C1 and two DIRB strains(IR-S2 and IR-K1)were selected for co-culture to study the As mobilization.It was found that the As-reducing ability of DIRB or the addition of As-reducing bacteria could only affect the amount of As release and the time efficiency of As immobilized.Compared with ferrihydrite,iron reduction and As release ability of two DIRB strains(IAR-S1 and IR-K1)decreased significantly when the two strains were incubated with the synthesized As-bearing goethite and hematite.Moreover,IAR-S1 could not completely sequester As into the solid phase.IAR-S1 formed secondary minerals of magnetite and vivianite by reducing goethite and hematite,respectively.No secondary mineral was found to be formed with strain IR-K1.In conclusion,p H value,geochemical types,As reduction capacity and the species of As-bearing mineral type could only affect the amount of As mobilization,but not the mobilization trend of As.These conditions were not the key factors affecting the As immobilization or release during iron reduction with DIRB.The differences of EPS components in various DIRB and the effects of EPS on As mobilization were studied,and the molecular mechanism of EPS metabolism was analyzed.Centrifugation,ultrasonic and thermal extraction were used to extract different EPS components of DIRB.The results showed that the amount of EPS,especially the polysaccharide content,produced by Shewanella IAR-S1 and Geobacter IR-G1 was higher than that of Klebsiella IR-K1.With additional 10?M As(?),the extra polysaccharide produced by strains IAR-S1 and IR-G1 were much higher than that of strain IR-K1.The As mobilization and iron reduction capability of DIRB was decreased by EPS removing.The results of incubation with additional different components of EPS(S-EPS,LB-EPS,TB-EPS)showed that LB-EPS promoted iron reduction and S-EPS promoted As immobilization.The addtional S-EPS of strain IAR-S1 can promote the immobilization of As with strain IR-K1,and the concentration of As in liquid phase is reduced to 4.26?M.Adding S-EPS of strain IR-K1 could inhibit the immobilization of As by strain IAR-S1,and the concentration of As in the liquid phase was stabilized at 4.13?M,which was not completely immobilized.These results indicated that S-EPS of DIRB was the key factor affecting As mobilization.The main different components of S-EPS were succinic and mannan.The addition of succinic promoted the release of As with strain IAR-S1 and reduced the formation of vivianite.The mannan promotes the immobilization of As and the formation of the secondary mineral by strain IR-K1.The results indicated that succinic acid and mannan were the key factors controlling As mobilization.The KEGG database was used to analyze the metabolic pathway of different species DIRB.The results showed that Klebsiella had a complete succinic metabolic pathway,while Shewanella and Geobacter lacked the key succinate dehydrogenase gene.Shewanella and Geobacter have a complete mannan metabolic pathway,and Klebsiella lacked the key phosphorylase gene in this pathway.These results indicated that the molecular mechanisms of EPS generation in DIRB were different,which directly controlled As mobilization.