Review--Remediation of heavy metal(loid)s contaminated soils _ To mobilize or to immobilize

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Journal of Hazardous Materials 266 (2014) 141–166Contents lists available at ScienceDirectJournal of HazardousMaterialsj o u r n a l h o m e p a g e :w w w.e l s e v i e r.c o m /l o c a t e /j h a z m atReviewRemediation of heavy metal(loid)s contaminated soils –To mobilize or to immobilize?Nanthi Bolan a ,b ,∗,Anitha Kunhikrishnan c ,Ramya Thangarajan a ,b ,Jurate Kumpiene d ,Jinhee Park e ,Tomoyuki Makino f ,Mary Beth Kirkham g ,Kirk Scheckel haCentre for Environmental Risk Assessment and Remediation,University of South Australia,Mawson Lakes,Australia bCooperative Research Centre for Contamination Assessment and Remediation of the Environment,Adelaide,Australia cChemical Safety Division,Department of Agro-Food Safety,National Academy of Agricultural Science,Suwon-si,Gyeonggi-do,Republic of Korea dWaste Science and Technology,Department of Civil,Environmental and Natural Resources Engineering,LuleåUniversity of Technology,LuleåSE-97187,Sweden eCentre for Mined Land Rehabilitation,University of Queensland,St Lucia,Australia fSoil Environmental Division,National Institute for Agro-Environmental Sciences,3-1-3Kannondai,Tsukuba,Ibaraki,Japan gDepartment of Agronomy,2004Throckmorton Plant Sciences Center,Kansas State University,Manhattan,KS,USA hNational Risk Management Research Laboratory,U.S.Environmental Protection Agency,5995Center Hill Avenue,Cincinnati,OH 45224,USAh i g h l i g h t s•Amendments used for remediationof contaminated soils inﬂuence metal(loid)bioavailability.•Case studies demonstrating the application of mobilization and immobilization techniques.•Techniques to monitor the long-term stability of immobilized metal(loid)s.•Environmental implications of mobi-lization and immobilization tech-niques.g r a p h i c a la b s t r a c ta r t i c l ei n f oArticle history:Received 24July 2013Received in revised form 6December 2013Accepted 12December 2013Available online 21 December 2013Keywords:Metals Metalloids Bioavailability MobilitySoil amendmentsa b s t r a c tUnlike organic contaminants,metal(loid)s do not undergo microbial or chemical degradation and persist for a long time after their introduction.Bioavailability of metal(loid)s plays a vital role in the reme-diation of contaminated soils.In this review,the remediation of heavy metal(loid)contaminated soils through manipulating their bioavailability using a range of soil amendments will be presented.Mobi-lizing amendments such as chelating and desorbing agents increase the bioavailability and mobility of metal(loid)s.Immobilizing amendments such of precipitating agents and sorbent materials decrease the bioavailabilty and mobility of metal(loid)s.Mobilizing agents can be used to enhance the removal of heavy metal(loid)s though plant uptake and soil washing.Immobilizing agents can be used to reduce the transfer to metal(loid)s to food chain via plant uptake and leaching to groundwater.One of the major limitations of mobilizing technique is susceptibility to leaching of the mobilized heavy metal(loid)s in the absence of active plant uptake.Similarly,in the case of the immobilization technique the long-term stability of the immobilized heavy metal(loid)s needs to be monitored.© 2013 Elsevier B.V. All rights reserved.∗Corresponding author at:CERAR,Building X,UniSA,University Boulevard,Mawson Lakes,SA 5095,Australia.Tel.:+61883026218;fax:+61883023124.E-mail address:Nanthi.Bolan@.au (N.Bolan).0304-3894/$–see front matter © 2013 Elsevier B.V. All rights reserved./10.1016/j.jhazmat.2013.12.018142N.Bolan et al./Journal of Hazardous Materials266 (2014) 141–166Contents1.Introduction (142)2.Sources of heavy metal(loid)s (143)2.1.Geogenic (143)2.2.Anthropogenic (143)3.Dynamics of heavy metal(loid)s in soils (143)3.1.Sorption/desorption process (143)3.2.Precipitation/dissolution (144)3.3.Oxidation/reduction (144)3.4.Methylation/demethylation (145)4.Approaches and indicators of bioavailability (146)4.1.Approaches to bioavailability (146)4.2.Indicators of bioavailability (146)5.Soil amendments for remediation (146)5.1.Mobilization of soil contaminants (146)5.1.1.Desorbing agents (146)5.1.2.Chelating agents (148)anic amendments (149)5.1.4.Fertilizers (149)5.1.5.Saline waters (150)5.1.6.Microbial mobilization (150)5.2.Immobilization(stabilization)of soil contaminants (150)5.2.1.Phosphate compounds (150)5.2.2.Liming materials (152)anic composts (152)5.2.4.Metal oxides (153)5.2.5.Biochar (153)6.Case studies (154)6.1.Case study1:soil washing for cadmium contaminated paddy soils (154)6.2.Case study2:immobilization of copper and lead in mine spoil (155)6.3.Case study3:immobilization of arsenic in soil contaminated with wood impregnation chemicals (156)6.4.Case study4:immobilization of lead in smelter residue (156)7.Environmental implications of(im)mobilization techniques (157)7.1.Leaching of contaminants (157)7.2.Long-term stability of immobilized metal(loid)s (158)8.Summary and conclusions (159)Acknowledgement (160)References (160)1.IntroductionIndiscriminate waste disposal practices have led to signiﬁcant build up in soils of a wide range of metal(loid)s,such as arsenic(As), cadmium(Cd),chromium(Cr),copper(Cu),mercury(Hg),lead(Pb), selenium(Se),and zinc(Zn).Entry of soil-borne metal(loid)s into the food chain depends on the amount and source of metal(loid) input,the properties of the soil,the rate and magnitude of uptake by plants,and the extent of absorption by animals[1].Health authorities in many parts of the world are becoming increasingly concerned about the effects of heavy metal(loid)s on environmen-tal and human health.Historically,heavy metal(loid)toxicity to human health received attention primarily as a result of series of widespread poisoning.For example,the hundreds of tragic cases of human poisoning of Minamata Bay in Japan(Minamata disease)in the late1950s were believed to have occurred from the ingestion ofﬁsh containing methyl mercuric compounds probably derived through biomethylation of mercuric salts by aquatic organisms[2].More recently high concentrations of heavy metal(loid)s,such as As,Cd,Cu,Pb,and Zn in soils have often been reported in num-ber of countries.For example,signiﬁcant adverse impacts of As on human health have been recorded in Bangladesh,India,and China and it is claimed that millions of people are potentially at risk from As poisoning[3].Similarly,Cd accumulation in the offal of graz-ing animals in New Zealand and Australia made it unsuitable for human consumption and affected access of meat products to over-seas markets[4].Similarly,there have been concerns about urban development of horticultural sites which contained toxic levels of metal(loid)s such as As,Cu,and Pb in soils resulting from excessive use of fungicides and herbicides that are rich in these metal(loid)s [5].Unlike organic contaminants,metal(loid)s do not undergo microbial or chemical degradation,and the total concentration of these metal(loid)s persist for a long time after their introduction in soils[6].With greater public awareness of the implications of contaminated soils on human and animal health there has been increasing interest amongst the scientiﬁc community in the devel-opment of technologies to remediate contaminated sites.This is especially necessary since traditional methods of soil removal and replacement of clean soil is often cost prohibitive.For diffuse dis-tribution of metal(loid)s,remediation options generally include amelioration of soils to minimize the metal(loid)bioavailability. Bioavailability can be minimized through chemical and biological immobilization of metal(loid)s using a range of inorganic com-pounds,such as lime and phosphate(P)compounds,and organic compounds,such as‘exceptional quality’biosolids[7].The more localized metal(loid)contamination found in urban environments is remediated by metal(loid)mobilization processes that include phytoextraction and chemical washing.Removal of metal(loid)s through phytoextraction process and the subsequent recovery of the metal(loid)s or their safe disposal are attracting research and commercial interests[8].However,when it is not possible to remove the metal(loid)s from the contaminated sites by phytoex-traction,other viable options,such as in situ immobilization(e.g., phytostabilization)should be considered as an integral part of risk management(Fig.1).Both phytoextraction and phytostabilizationN.Bolan et al./Journal of Hazardous Materials266 (2014) 141–166143Fig.1.Schematic diagram illustrating the link between(im)mobilization,bioavail-ability and remediation of heavy metal(loid)s.precesses are part of phytoremediation technique employed to manage contaminated soils.An overview of the sources of some of the common heavy metal(loid)s input to soils,their interactions and bioavailability in soils,and the remediation of metal(loid)contaminated soils through manipulating their bioavailability using a range of mobi-lizing and immobilizing soil amendments will be presented in this review.2.Sources of heavy metal(loid)sIn terrestrial ecosystems,the soil is the main repository of chem-ical contaminants.Likewise in aquatic systems,the sediment serves as the ultimate sink for these chemicals.Heavy metal(loid)s reach the soil environment through both pedogenic and anthropogenic processes.Most heavy metal(loid)s occur naturally in soil parent materials,chieﬂy in forms that are not readily bioavailable for plant uptake.Unlike pedogenic inputs,heavy metal(loid)s added through anthropogenic activities typically have a high bioavail-ability[9–11].Anthropogenic activities,primarily associated with industrial processes,manufacturing,the disposal of domestic and industrial waste materials,and the application of P fertilizers are the major source of metal(loid)enrichment in soils[1,12–14].2.1.GeogenicMost of the heavy metal(loid)s occur in nature,the major source of which is weathering of soil parent materials including igneous and sedimentary rocks,and coal.The majority of As is derived from geogenic origin.For example,coal is estimated to release45,000 tons of As annually,while human activities release approximately 50,000tons[15,16].The As content of igneous rocks varies widely (up to100mg/kg);the average content is2–3mg/kg.Sedimentary rocks also vary in their As content,from small amounts in lime-stone and sandstone up to15,000mg/kg in some manganese ores [17].Although the anthropogenic As source is increasingly becom-ing important,the recent episode of extensive As-contamination of ground waters in many countries including Bangladesh,India, China,and Mexico is of geological origin,transported by rivers from sedimentary rocks in the Himalayas over tens of thousands of years [16,18–20].Similarly,seleniferous soils found in the areas of Se-rich rocks, such as black shales,carbonaceous limestones,carbonaceous cherts,mudstones,and seleniferous coal are a major source of Se input to soil.Also,irrigation of Se-rich ground water had shown to contaminate irrigation drain water and surface waters in the San Joaquin Valley,California(United States),and in Punjab,India [21,22].Volcanic and geological activities mobilize natural Hg from deep reservoirs in the earth to the atmosphere.Annual emission of Hg from global mercuriferous belts,the zone along plate tectonic including western North America,central Europe,and southern China was estimated up to500Mg/year[23,24].2.2.AnthropogenicAnthropogenic activities,primarily associated with indus-trial processes,manufacturing and the disposal of domestic and industrial waste materials are the major source of metal(loid) enrichment in soils.Atmospheric pollution from Pb-based petrol was a major issue in many countries where there was no constraint on the usage of leaded gasoline[25].While biosolids is the major source of metal(loid)inputs in Europe and North America,P fer-tilizers are considered to be the major source of heavy metal(loid) input,especially Cd,in Australia and New Zealand[4,12,26,27].Phosphate compounds contain a range of metal(loid)s[4,12,28]. Cadmium contamination of agricultural soils is of particular con-cern because it reaches the food chain through regular use of Cd-containing P fertilizers.The Cd in most P fertilizers originates mainly from the phosphate rocks(PRs)used for manufacturing P fertilizers.Although many countries have formulated threshold levels for Cd and other heavy metal(loid)accumulation in soils due to the use of municipal biosolids,such limits have not been established from fertilizer use.Large quantities of Cu are used in agriculture,horticulture,and animal industries as a trace element,in many formulations of Cu containing fungicides,such as copper oxychloride and‘Bordeaux’mixture,and as a growth promoter in piggery and poultry units [29,30].Accumulation of Cu in agricultural soils resulting from con-tinuous use of Cu fungicides and biosolids application has been reported in many countries[31–34].One of the major consequences of excessive accumulation of Cu in soils is its toxicity to plants and microbial communities,for instance,formation of bare sterile patches in orchards[35,36].Approximately6400and1600tons of tannery and timber treatment efﬂuents,respectively,are generated annually in New Zealand,and these efﬂuents are considered to be the major sources of Cr contamination into aquatic and terrestrial environments[37]. Chromium is used as Cr(III)in the tannery industry and as Cr(VI) in the timber treatment industry.Chromate is highly toxic and car-cinogenic even when present in very low concentrations in water. Large scale use of Cu–Cr–As(CCA)treated timber as fence post and in vineyards can also result in the release of Cu,Cr,and As to soil environment[38,39].3.Dynamics of heavy metal(loid)s in soilsMetal(loid)ions can be retained in the soil by sorption,pre-cipitation,and complexation reactions,and are removed from soil through plant uptake,leaching,and volatilization(Fig.2).Although most metal(loid)s are not subject to volatilization losses,some metal(loid)s such as As,Hg,and Se tend to form gaseous com-pounds[16,40].The fate of metal(loid)s in the soil environment is dependent on both soil properties and environmental factors.3.1.Sorption/desorption processRetention of charged metal(loid)solute species by charged surfaces of soil components is broadly grouped into speciﬁc and non-speciﬁc retention[41,42].In general terms,non-speciﬁc adsorption is a process in which the charge on the ions balances the charge on the soil particles through electrostatic attraction, whereas speciﬁc adsorption involves chemical bond formation between the ions in the solution and those in the soil surface [43–45].144N.Bolan et al./Journal of Hazardous Materials266 (2014) 141–166Fig.2.The interaction between adsorption reactions of metal(loid)s in soil and their bioavailability.Both soil properties and soil solution composition determine the dynamic equilibrium between metal(loid)s in solution and thesoil solid phase.The concentration of metal(loid)s in soil solution is inﬂuenced by the nature of both organic and inorganic ligand ions,and soil pH through their inﬂuence on metal(loid)sorption processes[46,47].Two reasons have been given for the effect of inorganic anions on the sorption of metal(loid)cations such as Pb and Cd[48–51].Hong et al.[48]indicated that inorganic anions form ion pair complexes with metal(loid)s,thereby reducing their sorption.Naidu et al.[51]indicated that the speciﬁc sorption of lig-and anions is likely to increase the negative charge on soil particles, thereby increasing the sorption of Cd.The effect of pH values>6in lowering free metal(loid)ion activ-ities in soils has been attributed to the increase in pH-dependent surface charge on oxides of Fe,Al,and Mn,chelation by organic matter,or precipitation of metal(loid)hydroxides(e.g.,Pb(OH)3) [52,53].The activity of heavy metal(loid)s(e.g.,Cd and Pb)in solu-tion in naturally acidic soils is found to decrease with increasing pH which is attributed to increasing CEC[54–57].Other chemical interactions that contribute to metal(loid)reten-tion by colloid particles include complexation reaction between metal(loid)s and the inorganic and organic ligand ions.As might be expected,the organic component of soil constituents has a high afﬁnity for heavy metal(loid)cations such as Cu,Cd,and Pb because of the presence of ligands or groups that can form chelates with metal(loid)s[47,58].With increasing pH,the carboxyl,phenolic, alcoholic,and carbonyl functional groups in soil organic matter dissociate,thereby increasing the afﬁnity of ligand ions for these metal(loid)cations.The extent of metal(loid)–organic complex formation,however,varies with a number of factors including tem-perature,steric factors,and concentration.All these interactions are controlled by solution pH and ionic strength,the nature of the metal(loid)species,dominant cation,and inorganic and organic ligands present in the soil solution.3.2.Precipitation/dissolutionPrecipitation appears to be the predominant process in high pH soils and in the presence of anions such as SO42−,CO32−,OH−,and HPO42−,and when the concentration of the heavy metal(loid)ion is high[54,59,60].Precipitation of metal(loid)phosphates/carbonates is considered to be one of the mechanisms for the immobilization of heavy metal(loid)s such as Cu and Pb,especially in substrates containing high concentration of these metal(loid)s.For example, McGowen et al.[61]noticed that P decreased the leaching of Cd,Pb, and Zn.Activity-ratio diagrams indicated that diammonium phos-phate decreased solution metal(loid)concentrations by forming metal(loid)–P precipitates.Similarly,liming typically enhance the retention of metal(loid)s[1].For instance,Bolan and Thiyagarajan [62]found increased retention of Cr(III)with lime-induced increase in pH.As(III) oxidation rate (µM/hr)100200300400500 Bacterialsp.12345678910111213141516Fig.3.Arsenite oxidation potential of various bacterial strains:(1)herminiimonas arsenicoxydans ULPAs1,(2)thermus aquaticus,(3)thermus thermophiles,(4) agrobacterium albertimagni AOL15,(5)agrobacterium tumefaciens-5A,(6)Achro-mobacter sp.GW1,(7)thermus aquaticus AO3C,(8)ochrobactrum tritici SCII24, (9)Achromobacter sp.SY8,(10)Pseudomonas sp.TS44,(11)Alcaligenes sp.RS-19, (12)Marinobacter santoriniensis NKSG1,(13)Pseudomonas arsenicoxydans VC-1,(14) Bosea sp.AR-11,(15)SPB-24,(16)SPB-31[79].Co-precipitation of metal(loid)s especially in the presence of iron oxyhydroxides has also been reported and often such interac-tions lead to signiﬁcant changes in the surface chemical properties of the substrate.Lu et al.[63]conﬁrmed that co-precipitation of Pb(II)with ferric oxyhydroxides occurred at∼pH4and is more efﬁ-cient than adsorption in removing Pb(II)from aqueous solutions at similar sorbate/sorbent ratios and pH.Arsenate(As(V))sorp-tion onto ferrihydrite,and Ni(II)and Cr(III)sorption onto hydrous iron oxides showed that co-precipitation was more efﬁcient pro-cess than sorption for metal(loid)removal from aqueous solutions. Violante et al.[64]noticed that As(V)was desorbed by P from a ferrihydrite on which As(V)was added than from a Fe–As(V)co-precipitate.The potential value of P and liming materials in the remediation of metal(loid)contaminated soils is discussed below.3.3.Oxidation/reductionMetal(loid)s,including As,Cr,Hg,and Se,are most commonly subjected to microbial oxidation/reduction reactions,thereby inﬂuencing their speciation and mobility(Table1).For example, metals(e.g.,Cu and Hg)generally are less soluble in their higher oxidation state,whereas the solubility and mobility of metalloids (e.g.,As)depend on both the oxidation state and the ionic form[77]. The redox reactions are grouped into two categories,assimilatory and dissimilatory[78].In assimilatory reactions,the metal(loid) (e.g.,Se)substrate is involved in the metabolic functioning of the organism by acting as terminal electron acceptor.In contrast,for dissimilatory reactions the metal(loid)substrate has no known role in the metabolic functioning of the species responsible for the reac-tion,and indirectly initiates redox reactions.Arsenic in soils and sediments can be oxidized to As(V)by bac-teria[79,80](Fig.3).Since As(V)is strongly retained by inorganic soil components,microbial oxidation results in the immobilization of As.Under well drained conditions As would present as As(V), whereas under reduced conditions,As(III)dominates in soils,but elemental arsenic[As(0)]and arsine(H2As)can also be present. Oxidation of Cr(III)to Cr(VI)can enhance the mobilization and bioavailability of Cr.It is primarily mediated abiotically through oxidizing agents such as Mn(IV),and to a lesser extent by Fe(III), whereas reduction of Cr(VI)to Cr(III)is mediated through bothN.Bolan et al./Journal of Hazardous Materials 266 (2014) 141–166145Table 1Selected references on the effect of various soil amendments on the redox reactions of metal(loid)s in soils.Metal(loid)sSoil amendmentsObservationsReferencesCr Cattle manureEnhanced the reduction of Cr(VI)to Cr(III).[65,66]Cr Biosolid compost,Manures,spent mushroom Organic amendments increased DOC,which reduced Cr(VI)to Cr(III)in soils.[12]CrComposted cow manureChromate leaching was reduced in soils in the presence of elevated organic matter because of reduction followed by retention on cation exchange sites or precipitation.[67]Cr Zero valent ironEnhanced the reduction of Cr(VI)to Cr(III).[68]Cr Colloidal zerovalent ironA majority of Cr(VI)was reduced at pH 5.[69]Cr Mn oxide salts,birnessite and todorokite The amendments were equally capable of oxidizing Cr(III)to Cr(VI).[70]AsIron oxideAt lower redox,insoluble sulﬁdes can be formed either with iron (AsFeS,arsenopyrite;at strongly reducing conditions)or without iron (As 2S 3,orpiment;at moderately reducing conditions).[71]AsFe(III)and Mn(III)oxidesFe(III)and Mn(III)oxides oxidized As(III)to As(V)through electron transfer reaction.[16]As Goethite [␣-Fe III OOH]Rapidly oxidized As(III)to As(V)[72]Hg Green rusts (mixture of Fe II /Fe III hydroxides)Reduced Hg(II)to Hg(0)in suboxic soils and sediments.[73]Se Green rustIn suboxic conditions,green rust reduced Se(VI)to Se(0).[74,75]SeZero valent ironReduced Se(VI)to insoluble selenide (Se(-II))species.[76]abiotic and biotic processes [37].Chromate (Cr(VI))can be reduced to Cr(III)in environments where a ready source of electrons (Fe(II))is available and microbial Cr(VI)reduction occurs in the presence of organic matter as an electron donor [37,81](Fig.4).In living sys-tems,Se tends to be reduced rather than oxidized,and reduction occurs under both aerobic and anaerobic conditions.Dissimilatory Se(IV)reduction to Se(0)by chemical reductants such as sulﬁde or hydroxylamine,or biochemically by glutathione reductase,is the major biological transformation for remediation of Se oxyanions in anoxic sediments [82].Hence,precipitation of Se(0),which has been associated with bacterial dissimilatory Se(VI)reduction,has great environmental signiﬁcance [83,84].In the case of Hg,microor-ganisms,particularly bacteria play a major role in reducing reactive Hg(II)to non-reactive Hg(0),which may be subjected to volatiliza-tion losses.Mercury(II)is reduced to Hg(0)by mercuric reductase,and the dissimilatory reducing bacterium Shewanella oneidensis has been shown to reduce Hg(II)to Hg(0),which requires the presence of electron donors [85].3.4.Methylation/demethylationMethylation is a biological mechanism for the removal of toxic metal(loid)s (e.g.,Hg)by converting them to methyl derivativesM a x i m u m C r r e d u c t i o n (m g /k g )100200300400500600S o i l S o i l 1+B S o i l 1+C M S o i l S o i l 2+B S o i l 2+C M S o i l S o i l 3+B S o i l 3+C M Fig.4.Effect of carbon amendments on maximum Cr reduction (mg/kg)in soils (soil 1,calcic red sandy loam;soil 2,calcic red clay;and soil 3,tannery waste contaminated soil;BC black carbon;CMB chicken manure biochar)[37].that are subsequently removed by volatilization [86].Methy-lated derivatives of As,Hg,and Se can originate from chemical and biological mechanisms and this frequently results in altered volatility,solubility,toxicity,and mobility.Although methyla-tion of metal(loid)s occurs through both chemical (abiotic)and biological processes,biological methylation (biomethylation)is considered to be the dominant process in soils and aquatic envi-ronments.Biomethylation may result in metal(loid)detoxiﬁcation,since methylated derivatives may be excreted readily from cells,and are often volatile and may be less toxic,e.g.,organoarsenicals.Thayer and Brinckman [87]grouped methylation into two categories:trans-methylation and ﬁssion-methylation.Trans-methylation is the transfer of an intact methyl group from one compound (methyl donor)to another (methyl acceptor).Fission-methylation is the ﬁssion of a compound (methyl source),not necessarily containing a methyl group,so as to eliminate a molecule such as formic acid.The ﬁssion molecule is subsequently captured by another compound which is reduced to a methyl group.Microorganisms in soils and sediments act as biologically active methylators [22,88].Organic matter provides the methyl-donor source for both biomethylation and abiotic methylation in soils and sediments.Methylation of Hg is controlled by low molecular weight fractions of fulvic acid in soils [89].Similarly,Lambertsson and Nils-son [90]suggested that organic matter and alternative electron acceptors inﬂuenced methylation of Hg in the sediments.Biomethylation is effective in forming volatile compounds of As such as alkylarsines,which could easily be lost to the atmo-sphere [91,92].Benthic microbes methylate As under both aerobic and anaerobic conditions to produce methylarsines and methyl-arsenic compounds with a generic formula (CH 3)n As(O)(OH)3−n .Monomethylarsenate (MMA)and dimethylarsenite (DMA)are common organoarsenicals in river water.Methylated As species could result from direct excretion by algae or microbes or from degradation of the excreted arsenicals or more complex cellular organoarsenicals [93].Methylation of Hg occurs under both aerobic and anaerobic conditions [94].Regnell and Tunlid [95]showed that the propor-tion of methylated Hg was signiﬁcantly higher in the anaerobic condition than in aerobic systems consisting of undisturbed lake sediment and water.Under anaerobic conditions Hg(II)ions can be biologically methylated to form either monomethyl or dimethyl Hg which are highly toxic and more biologically mobile than the other forms [6].The main methylation mechanism for Hg involves non-enzymatic transfer of methyl groups of methylcobalamin to Hg(II)ions [96].Selenium biomethylation is of interest because it represents a potential mechanism for removing Se from contaminated146N.Bolan et al./Journal of Hazardous Materials266 (2014) 141–166environments as methylated compounds,such as dimethyl selenide(DMSe)are less toxic than dissolved Se oxyanions. Dimethyl selenide can be demethylated in anoxic sediments as well as anaerobically by an obligate methylotroph similar to Methanococcides methylutens in pure culture.An anaerobic demethylation reaction may result in the formation of toxic and reactive H2Se from less toxic DMSe.Aerobic demethylation of DMSe is likely to yield Se(VI),thereby retaining Se in the system.4.Approaches and indicators of bioavailability4.1.Approaches to bioavailabilityThe generic deﬁnition of bioavailability is the potential for living organisms to take up chemicals from food(i.e.,oral)or from the abiotic environment(i.e.,external)to the extent that the chemicals may become involved in the metabolism of the organism.More speciﬁcally,it refers to the biologically available chemical fraction (or pool)that can be taken up by an organism and can react with its metabolic machinery[97];or it refers to the fraction of the total chemical that can interact with a biological target[98].In order to be bioavailable,the contaminants(e.g.,metal(loid)s)have to come in contact with the organism(i.e.,physical accessibility).Moreover, metal(loid)s need to be in a particular form(i.e.,chemical accessi-bility)to be able to enter a plant root.In essence,for a metal(loid) to be bioavailable,it will have to be mobile and transported acrossa membrane and be in an accessible form to the plant.4.2.Indicators of bioavailabilityBioavailability of metal(loid)s in soils can be examined using chemical extraction and bioassay tests that determine a fraction of the metal(loid)s is bioaccessible.Chemical extraction tests include single extraction and sequential fractionation[99–101].Bioassay involves plants,animals,and microorganisms[102,103].A range of chemical extractants including mineral acids(e.g., 1N HCl),salt solutions(e.g.,0.01M CaCl2),buffer solutions(e.g., 1M NH4OAc),and chelating agents[e.g.,diethylene triamine pen-taacetic acid(DTPA)]have been used to predict the bioavailability of metal(loid)s in soils[104–106].Chelating agents,such as ethylene-diamine tetraacetic acid(EDTA–0.05M)and0.05M DTPA,have often been found to be more reliable in predicting the plant avail-ability of metal(loid)s[107,108],since they are more effective in removing soluble metal(loid)–organic complexes that are poten-tially bioavailable.Sequential fractionation schemes are often used to examine the redistribution or partitioning of metal(loid)s in various chemical forms that include soluble,adsorbed(exchangeable),precipitated, organic,and occluded.Although the extraction procedures vary, generally the solubility and bioavailability of metal(loid)s in soils decrease with each successive step of the scheme[99,109].Speciﬁc chemical pools measured by chemical fractionation schemes have been correlated with plant uptake of metal(loid)s and have been successful in predicting the plant availability of metal(loid)s in soils [54,110].The bioavailability of metal(loid)s in soils has recently been examined using physiologically based in vitro chemical fraction-ation schemes that include the Physiologically Based Extraction Test(PBET),Relative Bioaccessibility Leaching Procedure(RBALP), Potentially Bioavailable Sequential Extraction(PBASE),and Gas-trointestinal(GI)Test[111–113].These innovative tests predict the bioavailability of metal(loid)s in soil and sediments when ingested by animals and humans.As in the case of traditional sequen-tial extraction schemes,the ability of these tests to solubilize metal(loid)s increases with each successive extraction step.Despite the recognized nonspeciﬁc(i.e.,operational)nature of chemical extraction methods,their analytical simplicity and rapidity ren-ders them most suitable for routine identiﬁcation of metal(loid) form and estimation of the bioavailability of metal(loid)s under ﬁeld conditions.However,the distribution of a given metal(loid) among the various fractions can only be considered as an estimate at best due to the subjectivity of the steps involved.Measurements of metal(loid)bioavailability and toxicity in soils using soil microorganisms are receiving increasing attention,as microorganisms are more sensitive to heavy-metal(loid)stress than plants or soil macrofauna[114,115].The methods using microﬂora and protozoa have the potential to provide a mea-sure of bioavailability of heavy metal(loid)s in the short-term and even facilitate the measurement of temporal changes.In con-trast,responses by mesofauna(microarthropods)and macrofauna (enchytridae,invertebrates,and earthworms)have cumulative effects.These methods,however,are time consuming and can only provide an overall effect of heavy metal(loid)bioavailability to the species tested.Although molecular techniques are rapidly devel-oped and applied,they are comparatively expensive,and thus the level of information provided by these techniques need to be clearly demonstrated[116,117].5.Soil amendments for remediationA number of amendments are used either to mobilize or immo-bilize heavy metal(loid)s in soils.The basic principle involved in the mobilization technique is to release the metal(loid)s in to soil solution,which is subsequently removed using higher plants.In contrast,in the case of the immobilization technique the metal(loid)concerned is removed from soil solution either through adsorption,complexation,and precipitation reactions,thereby ren-dering the metal(loid)s unavailable for human and plant uptake and leaching to groundwater[12].In this section the potential value of these soil amendments in the(im)mobilization of metal(loid)s in relation to remediation will be discussed.Kirkham[118]reviewed titles of papers cited in theﬁrst30 issues of Current Contents:Agriculture,Biology and Environmental Sciences published in2012and identiﬁed38papers dealing with mobilizing and immobilizing agents.Of the38papers,six dealt with immobilization,29dealt with mobilization,and two dealt with both topics.One paper,by Desaules[119],dealt with meth-ods evaluating how one can tell if a soil is contaminated naturally or anthropogenically,and he pointed out that distinguishing between natural and anthropogenic metal(loid)contents in soils depends upon values for natural content,which range considerably.The fact that three-quarters of the papers reviewed by Kirkham[118]con-sidered mobilization shows that removal of the pollutant is of more concern than stabilization of the contaminant.This reﬂects the pos-sibility that,over time,metal(loid)s once immobilized in the soil will become available for plant uptake[120–123].5.1.Mobilization of soil contaminantsMobilization of contaminants can be achieved through solubi-lization,desorption,chelation,and complexation reactions which result in the redistribution of contaminants from solid phase to solution phase,thereby increasing their bioavailability(Table2).5.1.1.Desorbing agentsAddition of P fertilizers to Pb–As(V)contaminated soils has resulted in an increase in the solubilization and subsequent mobil-ity of certain oxyanions,such as selenite,As(V),and Cr(VI)in soils [152–157].Seaman et al.[158]observed that increasing levels of hydroxyapatite addition to metal(loid)-contaminated sediments resulted in increases in the concentrations of Cr(VI)and As(V)in。

重金属积累影响植物和植食性昆虫互作的研究进展

农学学报2021,11(5):42-45Journal of Agriculture0引言随着城市化进程的加速以及自然灾害的破坏，全球可耕地面积呈现减少趋势。

其中工业“三废”的不当处理、农药和化肥的大量使用以及生活污染物的随意排放造成了一部分耕地重金属超标。

土壤中的重金属随着植物根系吸收进入地上部分的其他组织中，从而影响植物正常的生长发育。

动物摄食含有重金属的植物后也会造成重金属在体内的积累，严重时造成重金属中毒。

人类是植物最主要的消费者之一，农业生产中重金属污染已成为影响农产品产量和品质的重要因素。

除此之外，植食性昆虫也是植物的另一大消费者，因此重金属积累也会在一定程度上影响植物和植食性昆虫的互作。

目前，重金属积累对水稻生长发育的影响已经有一些报道，但重金属积累对水稻害虫及其天敌的影响研究相对较少。

本文主要从重金属如何影响植物和植食性昆虫互作的角度，综述了最新的研究进展，以期为水稻害虫的综合防控提供理论指导。

1重金属积累对植物生长发育的影响土壤中的重金属被植物根部吸收后通过各种转运蛋白运输到不同组织，当这些重金属积累过高时会抑制植物地上和地下部分的生长、影响水分和养分的分布、降低光合作用和呼吸作用、促进植物的衰老等[1-2]。

比如重金属积累影响水稻的生长发育。

实验研究表明，较高浓度的重金属处理可以抑制水稻种子的萌发基金项目：杭州市农业与社会发展科研项目“重金属积累对水稻害虫及其天敌的影响研究”(20191203B75)。

第一作者简介：黄江南，女，2000年出生，浙江温州人，本科，研究方向：植物保护。

通信地址：310058浙江省杭州市余杭塘路866号浙江大学紫金港校区农生环C 座1004室，E-mail ：547987789@ 。

通讯作者：尉吉乾，男，1984年出生，山东肥城人，高级农艺师，硕士，研究方向：农业生态环境监测。

通信地址：310020浙江省杭州市杭海路768号杭州农业大楼，Tel ：0571-********，E-mail ：wjq5347@ 。

SCI(EI)收录的材料类期刊

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6. Bioremediation of heavy metal-contaminated effluent using optimized activated sludge bacteria

ORIGINAL ARTICLEBioremediation of heavy metal-contaminated efﬂuent using optimized activated sludge bacteriaEbtesam El.Bestawy •Shacker Helmy •Hany Hussien •Mohamed Fahmy •Ranya AmerReceived:14October 2011/Accepted:3December 2012/Published online:16December 2012ÓThe Author(s)2012.This article is published with open access at Abstract Removal of heavy metals from contaminated domestic-industrial efﬂuent using eight resistant indigenous bacteria isolated from acclimatized activated sludge was investigated.Molecular identiﬁcation using 16S rDNA ampliﬁcation revealed that all strains were Gram-negative among which two were resistant to each of copper,cadmium and cobalt while one was resistant to each of chromium and the heavy metal mixture.They were identiﬁed as Entero-bacter sp.(Cu 1),Enterobacter sp.(Cu 2),Stenotrophomonas sp.(Cd 1),Providencia sp.(Cd 2),Chryseobacterium sp.(Co 1),Comamonas sp.(Co 2),Ochrobactrum sp.(Cr)and Delftia sp.(M 1)according to their resistance pattern.Strains Cu 1,Cd 1,Co 2and Cr were able to resist 275mg Cu/l,320mg Cd/l,140mg Co/l and 29mg Cr/l respectively.The four resistant strains were used as a mixture to remove heavy metals (elevated concentrations)and reduce the organic load of wastewater efﬂuent.Results revealed that using the pro-posed activated sludge with the resistant bacterial mixture was more efﬁcient for heavy metal removal compared to the activated sludge alone.It is therefore recommended that the proposed activated sludge system augmented with the acclimatized strains is the best choice to ensure high treatment efﬁciency and performance under metal stresses especially when industrial efﬂuents are involved.Keywords Acclimatization ÁActivated sludge ÁBioremediation ÁDomestic ÁHeavy metals ÁIndustrial wastewaterIntroductionHeavy metals are considered one of the most common and hazardous pollutants in industrial efﬂuents that might cause serious problems to the sewage network pipelines.The deleterious effects of heavy metals on biological processes are complex and generally related to species,solubility and concentration of the metal and the characteristics of the inﬂuent,such as pH,as well as presence and concentration of other cations and/or molecules and suspended solids (Gikas 2008).Metal toxicity results from alterations in the conformational structure of nucleic acids,proteins or by interference with oxidative phosphorylation and osmotic balance (Yaoa et al.2008).The most common mechanisms by which metals are eliminated from wastewater treatment processes depend on precipitation,adsorption to suspended solids during primary sedimentation or adsorption to extra-cellular polymers (Qodah 2006).Use of bio-adsorbentsE.El.Bestawy (&)ÁS.HelmyDepartment of Environmental Studies,Instituteof Graduate Studies and Research,Alexandria University,163Horria Ave.El-Shatby,P.O.Box 832,Alexandria,Egypt e-mail:ebtesamelbestawy@ S.Helmye-mail:shackerh@Present Address:E.El.BestawyDepartment of Life Sciences,Faculty of Science,King Abdul Aziz University,P.O.Box 42805,Jeddah 21551,Kingdom of Saudi ArabiaH.Hussien ÁM.Fahmy ÁR.AmerDepartment of Environmental Biotechnology,City for Scientiﬁc Research and Technology Application,P.O.Box 21934,Alexandria,Egypt e-mail:hhussein66@ M.Fahmye-mail:mohamedfahmy73@R.Amere-mail:ranyaamer@;r.amer@mucsat.sci.egAppl Water Sci (2013)3:181–192DOI 10.1007/s13201-012-0071-0such as bacteria,fungi,algae and some agricultural wastes that emerged as an eco-friendly,effective and low cost material option could offer potential inexpensive alterna-tives to the conventional adsorbents(Valls and Lorenzo 2002).Different species of Aspergillus,Pseudomonas, Sporophyticus,Bacillus,Phanerochaete,etc.,have been reported as efﬁcient chromium and nickel reducers.The response of microorganisms towards toxic heavy metals is very important for reclamation of polluted sites(Conge-evaram et al.2007).Adsorption of heavy metals on the sludge surface is usually attributed to the formation of complexes between metals and carboxyl,hydroxyl and phenolic surface func-tional groups of the extracellular polymeric substances (EPS)produced by many different species of bacteria isolated from activated sludge(Yuncu et al.2006).EPS are metabolic products of bacteria result from the organic matter of the efﬂuent and from microbial lysis or hydro-lysis.They serve as a protective barrier for cells against the harsh external environment and play a crucial role in sequestration and biosorption of metal ions as well as ions which constitute metabolic elements for bacteria.Metal biosorption performance depends on external factors,such as pH,other ions in bulk solutions(which may be in competition),organic material in bulk solution and tem-perature(Comte et al.2007).Identiﬁcation of sludge bacteria that are adapted to the new toxic metal environment provide efﬁcient potential candidates for heavy metal bioremoval from contaminated media.Because of its size(*1,500base pairs),the16S rRNA gene is the most widely used sequence in bacterial identiﬁcation(Weisburg et al.1991;Drancourt et al.2000). Segments from these genes can be easily ampliﬁed by PCR using universal primers and sequenced.Sequence com-parison of16S rRNA has been used as a powerful tool for establishing phylogenetic and evolutionary relationships among organisms(Amer2003).Alexandria is the second largest industrial city in Egypt where more than40%of the industrial activity localized.In Alexandria,industrial efﬂuents are mixed with domestic sewage and discharged into the main sewage network (Hussein1999).The aim of the present study was to isolate and identify heavy metal resistant bacteria from acclimatized activated sludge to be used for remediation of heavy metal contaminated wastewater at their highest expected levels. Materials and methodsSampling and pre-treatmentActivated sludge samples were obtained from the aeration tank of the activated sludge unit at Damanhore Wastewater Treatment Plant located in the North West side of Egypt. Prior to use,sludge was pre-treated by allowing the sludge to settle at room temperature followed by decanting the supernatant and replacement with synthetic wastewater with aeration to maintain dissolved oxygen above4mg/l. After24h,aeration was stopped and the mixture was allowed to settle,then the supernatant was decanted and fresh synthetic wastewater was added to provide the same original volume.The procedure of settling,decanting, addition of new efﬂuent and nutrients was repeated at24-h interval until the desired acclimation time(approximately 1month)had been achieved.Heavy metal determinationFour heavy metals(Cu,Cd,Cr and Co)in the wastewater and sludge were determined using Atomic Adsorption Spectrophotometry(Perkin Elmer)after digestion with microwaves digester following the manual instructions with the addition of nitric acid,hydroﬂuoric acid and hydrogen peroxide for sample digestion(Costley and Wallis2001;Borowski2004).Measurement of dehydrogenase activity(DHA) Dehydrogenase activity was determined using method described by Weddle and Jenkins(1971).Triphenyl tetra-zolium chloride(TCC)and sodium sulfate solutions(5%) were prepared in distilled water.One milliliter of TCC, 10ml liquor or cell suspension and3drops of NaSO3were placed in centrifuge tube and shaken at30°C for1h in dark.This was followed by centrifugation for5min;the color of the supernatant was determined using spectro-photometer at wavelength480nm.Inhibition of the bac-teria and consequently the activated sludge activity were determined according to the following equation: Inhibition%ðÞ¼I0ÀIðÞ=I0½ Â100Where I0represents intensity of the control without treat-ment and I represents intensity of sample after treatment with heavy metal(Weddle and Jenkins1971;Shim et al. 2003).Isolation of heavy metal-tolerant bacteriaTolerant bacterial isolates were collected fromﬁve differ-ent acclimatized activated sludge batch reactors treated by synthetic wastewater(Table1)with elevated levels of Cu, Cd,Co,Cr(prepared as10%stock solutions of CuSO4, CdCl2ÁH2O,K2Cr2O7,CoCl2Á6H2O salts)and metal mix-ture(El.Bestawy et al.2012)as described by O¨zbelge et al. (2005).Tolerant bacteria were isolated from theﬁnal stage of the metal enrichment experiment at all reactors.One-milliliter liquor from each reactor was used to inoc-ulate liquid Luria–Bertani(LB)medium containing the investigated heavy metal(sterilized byﬁltration)in that reactor.Heavy metal concentrations used during bacterial isolation were15,100,40and40mg/l for the individual Cu,Cr,Cd and Co,respectively.While a mixture of Cu, Cr,Cd and Co at2,10,10and10mg/l respectively was used for isolation of strains with multiple accumulation ability.After24h incubation at30°C,100l l of the grown culture was transferred into10-ml fresh LB medium con-taining the same metal concentration and left for another 24h under the same conditions.The highest metal con-centration reduced the activated sludge activity by50% was amended in Petri dish containing20ml sterile LB agar and mixed well.100l l of the grown culture was spread on the surface of the plate and incubated for24h at30°C. Large identical colonies from each plate were picked out and cultured for puriﬁcation and identiﬁcation. Molecular identiﬁcationTotal genomic DNA was extracted from5ml overnight LB culture of the puriﬁed isolates(Sambrook et al.1989).PCR was performed in a light cycler Eppendorf PCR machine. A1,300-bp fragment was obtained by PCR ampliﬁcation of the16S rDNA gene(Ausubel et al.1999)using the primers B341F(50-CCTACGGGA GGCAGC)and1392R(50-ACG GGCGGTG TGTRC-30).The PCR mixture was composed of100ng of genomic DNA,30pmol of each primer, 200l M of dNTPs,1U of Taq polymerase and10l l of10X PCR buffer,the reaction volume was adjusted to100l l in 0.5ml eppendorf tube.The PCR ampliﬁcation conditions were performed by an initial denaturation step at94°C for 10min followed by30denaturation cycles at94°C for 1min,annealing at60°C for1min and an extension at 72°C for1min followed by aﬁnal extension step at72°C for10min.Amplicons of16S rDNA were puriﬁed using PCR puriﬁcation kit(QIAGEN).Each of these puriﬁed products was sequenced by the chain terminator method (ABI3130XL system,DNA technology,Denmark)using the two corresponding PCR primers separately.The resulted DNA sequences were phylogenetically analyzed using the BLAST search program.Multiple sequence alignment and molecular phylogeny were performed using BioEdit software(Hall1999).Screening for heavy metal resistance patternEffect of the individual and mixed tested metals on the growth(measured as optical density,OD at600nm)and dehydrogenase activity of each of the resistant strains was investigated at elevated metal concentrations compared to their controls.An overnight culture of each strain was inoculated in20-ml LB medium supplemented with deﬁ-nite concentrations of the corresponding heavy metals and incubated for24h under the previously mentioned condi-tions.Concentrations of the tested metals(not shown)were calculated according to the IC50which is also included in the screening test.Concentrations of heavy metal mixtures were calculated by dividing the growth IC50of each heavy metal by4and two higher and two lower concentrations were applied for screening bioassay.Treatability studiesThe most efﬁcient strains for metal removal were selected based on the resistance screening bioassays and manipu-lated in activated sludge treatment of mixed domestic-industrial wastewater contaminated with heavy metals and organic matter.The acclimatized sludge was prepared by centrifugation to remove the supernatant which was replaced by wastewater.The resistant strain for each investigated metal was seeded individually in LB-medium and incubated at30°C and200rpm for24h.Cultures were centrifuged and the supernatants were replaced with wastewater and used as inocula for batch reactors for the treatability study with moderate concentrations of heavy metals.Three sets of500-ml reactors were used to compare the removal efﬁciencies of the sludge,bacterial strains and mixture of both.Each set contained four different con-centrations(not shown)of the heavy metals with initial COD of1,400mg/l as following:1.Theﬁrst set consists of200ml of sludge with differentconcentrations of the investigated heavy metals.2.The second set consists of200ml of bacterial cultureonly with different concentrations of the investigated heavy metals.Table1Composition of the concentrated synthetic wastewater Serial Constituents Concentration(g/l)1Tripton122.12NaCl40.73Na2SO4 4.464K2H2PO4 4.465MgCl2Á6H2O0.376CaCl2Á2H2O0.377MnSO40.578H2MoO40.319NaOH0.0810ZnSO40.4611CoSO40.4912CuSO40.7610ml of concentrated synthetic wastewater were added to each 1,000ml of deionized waster3.The third set consists of100ml of sludge and100mlof bacterial culture with different concentrations of the investigated heavy metals.Heavy metal removal per unit biomass was determined using the following equation:M uptake=g¼½MðInf:ÞÀMðEff:Þ =MðMLSS)where M=uptake/g:heavy metal removed per unit mass of biomass,M(Inf.)=the initial heavy metal concentration in the inﬂuent,M(Eff.)theﬁnal metal concentration after treatment, MLSS:Biomass concentration in gramResultsIsolation and molecular identiﬁcation of metal resistant-sludge bacteriaA total of eight strains representing morphologically different bacterial colonies were isolated and puriﬁed. Two showed resistance to copper(Cu1and Cu2),two for cadmium(Cd1and Cd2),two for cobalt(Co1and Co2), one for chromium(Cr)and the heavy metal mixture resistant strain(M1).PCR products of the chromosomal DNA extracts of the resistant isolates were puriﬁed, sequenced and aligned against the16S rDNA sequences of the ribosomal database project http://www.cme.msu. edu/RDP/html/index/html(Maidak et al.1994;Rainey et al.1996).Sequences were deposited in GenBank for obtaining accession numbers.The Phylogenetic relation-ship among the tested isolates and the closely related species(Fig.1)was analyzed using multi-sequence alignment program(BioEdit Sequence Alignment Editor). Accession numbers and similarity of the tested isolates to the related species are presented in Table2.It is well documented that both Gram-positive and Gram-negative bacteria are ubiquitous in nature with highly resistant anionic cell walls.This anionic cell wall canﬁx metal and provide sites for nucleation and growth of minerals, a property that has been the basis for many heavy metal removals using bacterial biomass(Kelly et al.2004). Heavy metal resistance and/or sensitivity of the sludge bacteriaResistance and/or sensitivity of the acclimatized sludge bacteria towards the investigated metals(individuals or mixture)were determined in terms of growth(OD)and dehydrogenase activity to select the most efﬁcient for treatability study.Effect of individual metalsComparing data of the individual effects of the four tested metals on the acclimatized strains(Fig.2)revealed the following points:1.IC50of all the tested metals that induced50%reduction in the growth of all the investigated strains was higher than that induced50%reduction in their DHA.This indicates that tested metal could inhibit the enzyme activity much higher than its inhibition to the growth because dehydrogenase activity is a direct reﬂection of microbial activity(Gong1997;Mathew and Obbard2001)and metal toxicity(Aoyama and Nagumo1997;Kelly and Tate1998;Nweke et al.2006).2.IC50of each metal depends mainly on the microbialspecies as well as acclimatization period,culturing or operation conditions,nutrient status and the presence of other inhibitors.Isolated strains Cu1,Cd1,Co2and Cr had the highest ability to grow individually in the presence of Cu,Cd,Co and Cr with IC50of275,320, 140and29mg/l,respectively.3.Although possessing multiple resistances,most of theacclimated strains exhibited their highest resistance with their corresponding metals.For example Enter-obacter sp.(Cu1and Cu2)showed the highest OD(275 and190,respectively)with copper that declined to190 and46,respectively with Cd followed by more toxicity with Co and Cr.Similarly,Stenotrophomonas sp.(Cd1)and Providencia sp.(Cd2)also showed high resistance to Cd(OD320and260,respectively;DHA 50and15,respectively)compared to their resistance pattern for other metals.4.Ochrobactrum sp.(Cr)isolated from activated sludgeamended with chromium was more sensitive to chromium(OD29)compared to copper and cadmium (OD75and150,respectively).On contrary Ochro-bactrum sp.strain CSCR-3isolated from chromium landﬁlls was able to tolerate and reduce Cr VI to Cr III in cultures containing800mg/l Cr(He et al.2009).This is mainly attributed to factors controlling resis-tance and/or toxicity of metals of which microbial species and the form of the metal are key factors (Gikas and Romanos2006).Results conﬁrmed that gradual increase of the applied metals produced generations with high metal resistance range that can be efﬁciently used in decontamination pur-poses.These results are in agreement and supported by other studies.For example a Cd-resistant Stenotropho-monas sp.showed resistance to Se(Antonioli et al.2007) and Cu(Zaki and Farag2010);a Cd-resistant Providencia sp.showed very high resistance(1,000–1,500mg/l)to Cd,Cr,Ni,Co and Zn(Thacker et al.2006)and Pseudomonas with multi resistance to Cr,Cu,Cd and Ni(Kelly et al. 2004).In addition,Comamonas sp.(Co2)that showed moderate growth and DHA activity(140and82,respec-tively)when cultivated with50mg/l cobalt was found containing plasmid encoding cobalt and nickel resistance (Siunova et al.2009).According to their average DHA(reﬂecting their resis-tance)against the individual metals,acclimatized bacteria can be arranged in the following order from the highly resistant to the highly sensitive Cd1[Co2[Cr[ Cu2[Cu1[M1[Cd2.Therefore,Stenotrophomonas sp. (Cd1)represents the highest DHA activity indicating the lowest toxic effects upon exposure to individual metals.Effect of mixed metalsTheﬁnal stage in the screening process was to investigate each of the acclimatized strain isolated against mixture of the tested heavy metals(Cu,Cd,Co and Cr)based on its IC50concentrations obtained in the previous stage.The concentration of each heavy metal for each strain was calculated by dividing its growth IC50by4,and then two higher and two lower concentrations were used for the screening process(i.e.,ﬁve mixtures with elevated metals concentration).Applying the investigated metals as mix-tures led to completely different responses from the investigated acclimatized strains(Fig.3).Results revealed the followingpoints:1.As with the individual metals,IC50of all testedmetals,those induced50%reduction in the growth,of all the investigated strains were higher than those induced50%reduction in their DHA,indicating that metal inhibits enzyme activity much higher than growth.2.As expected,tolerance of the investigated strains wasreduced dramatically towards the tested metal mixtures compared to their individual application.This was shown by the growth IC50recorded70,81.25,43.75 and12.5mg/l for strains Cu1,Cd2,Co2and Cr, respectively in the metal mixtures compared to275, 320,140and29mg/l,respectively when grown individually in their corresponding metals.3.IC50of DHA of the isolated strains Cu1,Cu2,Cd1,Cd2,Co2,Cr and M against the tested metals(Cu,Cd, Co and Cr)recorded35,20,2and2;37.5,18.75,11.25 and1.5;5,20,7.5and1;5,16.25,1.75and1.75;8,12.5,17.5and2;10,20,3.5and5;andﬁnally18,18,30and6mg/l respectively(Fig.3).4.According to their average DHA against the mixedmetals,acclimatized bacteria can be arranged in the following order from the highly resistant to the highly sensitive M[Cu2[Cu1[Co2[Cr[Cd.Delftia sp.exhibited the highest resistance to the mixture ofthe investigated metals due to the adaptation acquired during the acclimatization process when grown in the presence of heavy metal mixture.These results are consistent with those of Jackson et al. (2009)who reported a multiple heavy metal resistance of Delftia sp.when used within a consortium for bioremedi-ation of River Plankenburg water in South Africa con-taminated with aluminum,nickel,copper and manganese due to the presence of highly incidence plasmids(Zolg-harnein et al.2007).Treatability bioassayThree parallel strategies were used to evaluate the reme-diation capability of the proposed system towards heavy metal-and organic matter-contaminated wastewater. Treatment technologies included the proposed activated sludge unit amended with the acclimatized bacterial strains (ASBS),the plain activated sludge unit(AS)and the acclimatized bacterial cultures(BS)alone represented by Cu1,Cd1,Co2and Cr.The performance of the three pro-cedures was evaluated to determine the most efﬁcient application.Also a comparison was made between the removal efﬁciency of the contaminated metals andtheCFig.1continuedorganic matter load (as COD).Four different concentra-tions of each heavy metal were investigated in the three planned treatment procedures and the removal was deter-mined after 24h exposure.Treatment of wastewater efﬂuent under individual metal stressesWastewater collected from the Western Wastewater Treatment Plant (WWTP),Alexandria was subjected to 24h treatment using ASBS,AS and BS technologies after the addition of four elevated concentrations of individual heavy metals (20–200mg/l for each of Cu,Cd and Co and 20–300mg/l for Cr).Results achieved (Figs.4and 5)can be summarized asfollows:Table 2Resistant isolates’accession numbers and their similarity to the related species Isolate Accession no.Similar Strain Similarity (%)Cd 1FJ205387Stenotrophomonas sp.99Cd 2FJ205388Providencia sp.98Cu 1FJ205389Enterobacter sp.98Cu 2FJ205390Enterobacter sp.99Co 1FJ205391Chryseobacterium sp.98Co 2FJ205392Comamonas sp.97Cr FJ205394Ochrobactrum sp.98M 1FJ205393Delftia sp.9950100150200250300350Fig.2IC50for the growth (OD)and dehydrogenase activity (DHA)of the acclimatized strains after 24-h exposure to individual heavy metals:Cu copper,Cd cadmium,Co cobalt,Cr chromiumFig.3IC50for the growth (OD)and dehydrogenase activity (DHA)of the acclimatized strains after 24h exposure to the heavy metals in mixture:Cu copper,Cd cadmium,Co cobalt,Cr chromium1.As a general rule,removal of all the investigated metals was stimulated with increasing metal levels in the three investigated procedures reaching maximum efﬁciency at the highest applied concentration while COD removal decreased.Moreover,the removal of the investigated metals by the proposed technologies recorded the following order Co [Cd [Cr [Cu.2.The highest Cu removal achieved (at 200mg/l)by the ASBS,BS and AS technologies recorded 15.84,22.29and 31.37Cu/g biomass,respectively.Therefore,the magnitude of increasing Cu removal efﬁciency (RE %)is AS [BS [ASBS.On the contrary,COD removal under the highest Cu stress recorded 58.10,57.82and 38.16%using the ASBS,BS and AS technologies,respectively.Therefore,the magnitude of increasing COD RE %is ASBS [BS [AS.3.The proposed technologies ASBS,BS and AS could achieve Cd removal (at 200mg Cd/l)of 53.17,69.36and 56.25mg Cd/g sludge biomass,respectively.Therefore,Cd removal magnitude power order is BS [AS [ASBS technology.While COD removalwas recorded as 23.12,16.64and 50.99%using the ASBS,BS and AS technology,respectively.Therefore,the magnitude of increasing COD RE %is AS [ASBS [BS.4.Co removal achieved (at 200mg/l)by the ASBS,BS and AS technologies recorded 60.93,70.72and 67.12Co/g biomass,respectively indicating that Co removal efﬁciency by the three techniques is in the following order BS [AS [ASBS technology.COD removal under Co stress recorded 64.44,48.44and 55.16%using ASBS,BS and AS technology,respec-tively indicating magnitude of increasing COD RE %as ASBS [AS [BS.5.Cr removal (at 300mg/l)recorded as 55.17,47.89and 44.04mg Cr/g using ASBS,BS and AS technology,respectively indicating an order of ASBS [BS [AS technology in Cr removal efﬁciency.COD removal under the highest Cr stress recorded 67.51,10.51and 51.05%using the ASBS,BS and AS technology,respectively.Therefore,the magnitude of increasing COD RE %is ASBS [AS [BS.C DFig.4Comparison among the three bioremediation technologies AS,ASBS and BS in the removal efﬁciency of a Cu,b Cd,c Co and d Cr after 24-h exposureTreatability bioassays of the contaminated wastewater in the presence of the four tested metals applied as individuals achieved the following points:1.Native activated sludge (AS technology)exhibited the maximum efﬁciency in the removal of Cu.2.For Cd and Co removal,BS technology where acclimatized bacterial cultures were used,exhibited the highest efﬁciency followed by AS technology and ASBS technology with no signiﬁcant variations.3.Activated sludge amended with the acclimatized bacteria (ASBS technology)performed the highest efﬁciency for removing Cr.4.Activated sludge amended with the acclimatized bacteria (ASBS technology)performed almost the highest efﬁciency for removing COD from wastewater under the stress of all the investigated metals even under their highest levels.Metal biosorption by microbial biomass is a well-known advanced biotechnological tool used for efﬁcient removal of contaminant metals.Optimization tools of metalbiosorption included many factors among which pH is the most important one (Comte et al.2007)as well as the C/N ratio of activated sludge (Yuncu et al.2006)and temper-ature (Zouboulis et al.2004).Acclimatization is a very important process for acquiring new advanced features of microorganisms that can be efﬁciently applied in metal biosorption.During the present study,using strains previ-ously acclimated by exposing microbial biomass to metal concentration gradients (El.Bestawy et al.2012)proved high efﬁciency in the enhancement of activated sludge performance for metals and organic matter removal even at high metal stresses.For example,Enterobacter sp.(Cu 1)acclimated to survive under high levels of Cu stress (200mg/l)showed high efﬁciency for Cu (31.37Cu/g biomass)and organic matter (58.10%)removal compared to 2.93mg Cu/g achieved as the maximum removal by plain activated sludge (Principia et al.2006).Similar results were obtained by manipulating metal binding capacity of Nocardia amarae cells to optimize the overall Ni,Cu and Cd binding capacity of activated sludge (Kim et al.2002).As in the present study the pure cultureofC DFig.5Comparison among the three bioremediation technologies AS,ASBS and BS in the removal efﬁciency of COD in the presence of elevated levels of a Cu,b Cd,c Co and d Cr after 24-h exposureNocardia exhibited signiﬁcantly higher metal sorption capacity than the activated sludge biomass attributed to the fact that the Nocardia cells growing at stationary phase have substantially more speciﬁc surface area than that of activated sludge.The metal sorption capacity of activated sludge increased proportionally with the amount of Nocardia cells present in the mixed liquor.Sirianuntapi-boon and Ungkaprasatcha(2007)also reported efﬁcient removal of Pb and Ni as well as organic matter by com-paring acclimatized and un-acclimatized biosuldge.Reductions in the COD removal with increasing metal concentrations during the present study are attributed mainly to the toxic effects and inhibition of the biodegra-dative microbes in the proposed treatment systems induced by the investigated metals at their elevated levels(Jefferson et al.2001;Gikas2007).Treatment of wastewater efﬂuent under mixed heavy metal stressesTreatability study was also performed with mixtures of the four investigated metals with elevated concentrations after 24-h exposure.Three technologies were applied here included BS,where mixture(strains Cu1,Cd1,Co2and Cr) of the acclimatized bacteria was used,ASBS,where acti-vated sludge supplied with the acclimatized bacteria was used and plain activated sludge(AS).Heavy metal removalsAs general trends for all the investigated metals and the three proposed treatment technologies,increasing metal concentrations from5to75mg/l enhanced their removal which was in the following order Cu\Cr\Cd\Co. The highest achieved RE(s)%of Cu,Cd,Co and Cr recorded were6.61,16.58,27.58and19.83mg/g sludge, respectively using ASBS technology(Fig.6a);4.39,22.89, 30.38and26.52mg/g biomass,respectively using BStechnology(Fig.6b)andﬁnally7.09,15.02,21.08and 18.58mg/g biomass,respectively using AS technology (Fig.6c).The efﬁciency of using activated sludge under the stress of microbial inhibiting agents(i.e.heavy metals)is one of the issues with lots of environmental concern.Many factors affect the efﬁciency of activated sludge systems used for metal removal from contaminated media.pH considered as the most critical factor for microbial metal biosorption (Ozdemir et al.2003;Sheng-lian et al.2006;Congeevaram et al.2007)which is mainly attributed to organism-speciﬁc physiology(Congeevaram et al.2007).Other factors such as initial metal concentrations,activated sludge dosage, temperature,contact time(treatment duration),presence of other ions and organic materials in the bulk solutions (which may be in competition)could affect the efﬁciency of activated sludge for heavy metal removal.(Sheng-lian et al.2006;Comte et al.2007;Pamukoglu and Kargi 2007).Bieszkiewicz and Hoszowski(1978)reported that acti-vated sludge could tolerate up to0.8,1.15and20mg/l only of Cu,Cr III and Cr(VI),respectively after which the activated sludge experienced inferior puriﬁcation and reduced intensity of respiration of its microorganisms.In another study,removal efﬁciency%of Cu,Cd and Cr using activated sludge recorded only80,62and46%, respectively with initial concentrations of0.8,20.9and 0.63mg/l of the same metals(Petrasek and Kugelman 1983).Comparison with the present work indicated remarkable enhancement of the metal tolerance acquired 051015202530355101520253035510152025303502550750255075 0255075 ACBFig.6Comparison among the removal efﬁciency of the investigated metals mixture of Cu,Cd,Co and Cr using the proposed a ASBS, b BS and c AS technology for24-h exposure。

LED3A土柱淋洗Pb与Zn污染砂土的效果

２０１９年２月
第３２卷㊀第２期
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环㊀ ቤተ መጻሕፍቲ ባይዱ㊀科㊀学㊀研㊀究
Ｖｏｌ.３２ꎬＮｏ.２Ｆｅｂ.ꎬ２０１９
ＬＥＤ３Ａ土柱淋洗Ｐｂ与Ｚｎ污染砂土的效果
乔洪涛１ ꎬ 赵保卫２ ꎬ 于雄胜１
１. 忻州师范学院化学系ꎬ 山西忻州㊀０３４０００２. 兰州交通大学环境与市政工程学院ꎬ 甘肃兰州㊀７３００７０
第２期
乔洪涛等:ＬＥＤ３Ａ土柱淋洗Ｐｂ与Ｚｎ污染砂土的效果
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㊀㊀土壤重金属污染因其具有隐蔽性㊁滞后性㊁积累性㊁不可逆转性和难治理性已成为环境科学领域的研究热点
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能同时去除污染土壤中的重金属和有机物 [２０￣２２] . 该类表面活性剂水溶性好ꎬ 对哺乳动物和水生生物无毒ꎻＯＥＣＤ试验结果表明ꎬ 该类表面活性剂易于生物降解ꎬ环境风险低 [２１] ꎻ 现已广泛应用于洗涤㊁矿物浮选㊁相转移催化㊁乳液聚合等领域 [２２] . 但是ꎬ到目前为止采用该类表面活性剂强化土壤重金属污染的原位淋洗修复的研究却鲜见报道. 因此ꎬ 该研究拟选用螯合型表面活性剂ＬＥＤ３Ａ( Ｎ￣十二酰基乙二胺三乙酸用土柱淋洗试验ꎬ 探究不同流速条件下ＬＥＤ３Ａ对Ｐｂ㊁Ｚｎ污染砂土的淋洗作用ꎬ 同时通过优化的ＢＣＲ供理论参考依据. 法ꎬ 研究土柱中重金属的迁移转化规律ꎬ 以期为ＬＥＤ３Ａ在化学淋洗修复重金属污染土壤中的应用提钠盐) 作为淋洗试剂ꎬＰｂ㊁Ｚｎ为重金属污染物代表ꎬ采
２.ＳｃｈｏｏｌｏｆＥｎｖｉｒｏｎｍｅｎｔａｌａｎｄＭｕｎｉｃｉｐａｌＥｎｇｉｎｅｅｒｉｎｇꎬ ＬａｎｚｈｏｕＪｉａｏｔｏｎｇＵｎｉｖｅｒｓｉｔｙꎬ Ｌａｎｚｈｏｕ７３００７０ꎬ Ｃｈｉｎａ

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Review--Remediation of heavy metal(loid)s contaminated soils _ To mobilize or to immobilize

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重金属积累影响植物和植食性昆虫互作的研究进展

SCI(EI)收录的材料类期刊

6. Bioremediation of heavy metal-contaminated effluent using optimized activated sludge bacteria

LED3A土柱淋洗Pb与Zn污染砂土的效果

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