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285 Pages·2016·41.48 MB·English

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Copyright© Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences. Published by Elsevier B.V. and Science Press. All rights reserved.
Journal of Environmental Sciences
Volume 44 2016
www.jesc.ac.cn
Highlight article
1 A barrier to metal movement: Synchrotron study of iron plaque on roots of wetland plants
Iris Koch and Michelle M. Nearing
Regular articles
4 Structural and metabolic responses of microbial community to sewage-borne chlorpyrifos in
constructed wetlands
Dan Zhang, Chuan Wang, Liping Zhang, Dong Xu, Biyun Liu, Qiaohong Zhou and Zhenbin Wu
13 An overview of emissions of SO and NO and the long-range transport of oxidized sulfur and
2 x
nitrogen pollutants in East Asia
Yu Qu, Junling An, Youjiang He and Jun Zheng
26 UptakeofradioiodidebyPaenibacillus sp.,Pseudomonas sp.,Burkholderia sp.andRhodococcus sp.
isolated from a boreal nutrient-poor bog
Merja Lusa, Jukka Lehto, Hanna Aromaa, Jenna Knuutinen and Malin Bomberg
38 Development of a field enhanced photocatalytic device for biocide of coliform bacteria
Jeff M. Huber, Krista L. Carlson, Otakuye Conroy-Ben, Mano Misra and Swomitra K. Mohanty
45 Characterisationandseasonalvariationsofparticlesintheatmosphereofrural,urbanandindustrial
areas: Organic compounds
FabriceCazier,PaulGenevray,DorothéeDewaele,HabibaNouali,AnthonyVerdin,FrédéricLedoux,
Adam Hachimi, Lucie Courcot, Sylvain Billet, Saâd Bouhsina, Pirouz Shirali, Guillaume Garçon and
DominiqueCourcot
57 Temporal and spatial changes of microbial community in an industrial effluent receiving area in
Hangzhou Bay
Yan Zhang, Lujun Chen, Renhua Sun, Tianjiao Dai, Jinping Tian, Wei Zheng and Donghui Wen
69 ProfilingkidneymicroRNAsfromjuvenilegrasscarp(Ctenopharyngodonidella)after56daysoforal
exposure to decabromodiphenyl ethane
LianGan,YuanyanXiong,FangDong,YunjiangYu,LijuanZhang,E.Shunmei,LiliuZhou,XiaoxiaLi
andGuochengHu
76 Rapid synthesis of Ti-MCM-41 by microwave-assisted hydrothermal method towards photocatalytic
degradation of oxytetracycline
Hanlin Chen, Yen-Ping Peng, Ku-Fan Chen, Chia-Hsiang Lai and Yung-Chang Lin
88 Environmental application and ecological significance of nano-zero valent iron
Biruck D. Yirsaw, Mallavarapu Megharaj, Zuliang Chen and Ravi Naidu
99 Non-thermal plasma treatment of Radix aconiti wastewater generated by traditional Chinese
medicine processing
Yiyong Wen, Jianping Yi, Shen Zhao, Song Jiang, Yuming Chi and Kefu Liu
109 Do phytotoxic allelochemicals remain in ashes after burning Chrysanthemoides monilifera subsp.
monilifera (boneseed)?
Md. Abdullah Yousuf Al Harun, Joshua Johnson and Randall W. Robinson
120 MicrocystinaccumulationandbiochemicalresponsesintheedibleclamCorbiculaleanaP.exposed
to cyanobacterial crude extract
Thanh-Luu Pham, KazuyaShimizu, AyakoKanazawa,Yu Gao,Thanh-Son Daoand MotooUtsumi
CONTENTS
131 Lead toxicity thresholds in 17 Chinese soils based on substrate-induced nitrification assay
Ji Li, Yizong Huang, Ying Hu, Shulan Jin, Qiongli Bao, Fei Wang, Meng Xiang and Huiting Xie
141 Sulfatereducingbacterialcommunityandinsituactivityinmaturefinetailingsanalyzedbyrealtime
qPCR and microsensor
Hong Liu, Shuying Tan, Tong Yu and Yang Liu
148 Rapid degradation of dyes in water by magnetic Fe0/Fe O /graphene composites
3 4
Shan Chong, Guangming Zhang, Huifang Tian and He Zhao
158 Estimating emissions from crop residue open burning in China based on statistics and MODIS fire
products
Jing Li, Yu Bo and Shaodong Xie
171 pH-dependent release characteristics of antimony and arsenic from typical antimony-bearing ores
Xingyun Hu, Xuejun Guo, Mengchang He and Sisi Li
180 Sulfur-based autotrophic denitrification from the micro-polluted water
Weili Zhou, Xu Liu, Xiaojing Dong, Zheng Wang, Ying Yuan, Hui Wang and Shengbing He
189 Thepotentialapplicationofredmudandsoilmixtureasadditivetothesurfacelayerofalandfillcover
system
Éva Ujaczki, Viktória Feigl, Mónika Molnár, Emese Vaszita, Nikolett Uzinger, Attila Erdélyi and
Katalin Gruiz
197 Computersimulationofthecoagulationofsuspendedsolids—TheapplicabilityoftheMüller–Smoluchowski
theory
Regina Wardzynska and Beata Załęska-Chróst
204 Positively charged microporous ceramic membrane for the removal of Titan Yellow through
electrostatic adsorption
Xiuting Cheng, Na Li, Mengfu Zhu, Lili Zhang, Yu Deng and Cheng Deng
213 Distribution and transportation of mercury from glacier to lake in the Qiangyong Glacier Basin,
southern Tibetan Plateau, China
ShiweiSun,ShichangKang,JieHuang,ChengdingLi,JunmingGuo,QianggongZhang,XuejunSun
andLekhendraTripathee
224 Effects of pentachlorophenol on the detoxification system in white-rumped munia (Lonchurastriata)
Peng Jiang, Jianshe Wang, Jinguo Zhang and Jiayin Dai
235 High-performance size exclusion chromatography with a multi-wavelength absorbance detector
study on dissolved organic matter characterisation along a water distribution system
Huiping Huang, Emma Sawade, David Cook, Christopher W.K. Chow, Mary Drikas and Bo Jin
244 Occurrenceandimpactofpolychlorinateddibenzo-p-dioxins/dibenzofuransintheairandsoilaround
a municipal solid waste incinerator
ZhiguangZhou,YueRen,JiazhiChu,NanLi,SenZhen,HuZhao,ShuangFan,HuiZhang,PengjunXu,
LiQi,ShutingLiangandBinZhao
252 The biological effect of metal ions on the granulation of aerobic granular activated sludge
Wen Hao, Yaochen Li, Junping Lv, Lisha Chen and Jianrong Zhu
260 Phosphate recovery from anaerobic digester effluents using CaMg(OH)
4
Xueyu Liu, Zhonghou Xu, Jianfeng Peng, Yonghui Song and Xiaoguang Meng
269 Biochar: A review of its impact on pesticide behavior in soil environments and its potential
applications
Mahdi Safaei Khorram, Qian Zhang, Dunli Lin, Yuan Zheng, Hua Fang and Yunlong Yu
Corrigendum
280 Corrigendum to “Silver nanoparticles tolerant bacteria from sewage environment” [J. Environ. Sci.
(2011) 23 (2) 346–352]
Sudheer Khan, Amitava Mukherjee and Natarajan Chandrasekaran
JOURNAL OF ENVIRONMENTAL SCIENCES 44 (2016) 1–3
Available online at www.sciencedirect.com
ScienceDirect
www.elsevier.com/locate/jes
A barrier to metal movement: Synchrotron study of iron plaque
on roots of wetland plants
⁎
Iris Koch , Michelle M. Nearing
RoyalMilitaryCollegeofCanada,Kingston,Ontario,Canada.E-mail:[email protected]
A R T I C L E I N F O yard. One location had standing water, referred to here as
the “wetter” location, and the other was assumed to be the
Availableonline12May2016 “drier”location.Cu,PbandZnwerepresentatelevatedcon-
centrationsinthesoilandsedimentalongwithhighconcen-
Keywords: trationsofFeandMn.ThetwostudiedplantswerePhragmites
Wetlandplants australis (common reed), obtained from both locations, and
Metalcontamination Typha latifolia (cattail) (Fig. 1), available only from the drier
Urbanbrownfieldsites location.
X-raymicrotomographyand Synchrotron-based techniques were used, which allowed
microfluorescence for minimal sample manipulation to visualize the root
Copper structure and the distribution of metals in the roots of the
Iron plants.Thesamplemanipulationconsistedofsimplycleaning
Lead therootswithdeionizedwater,andthenplacingthesamples
Zinc in holders appropriate for the synchrotron-based method
used.
Onesynchrotron-basedmethodwasX-raycomputedmicro-
tomography (μCMT),carriedoutwithdried rootsamples ina
holding tube, to produce high resolution three dimensional
A wetland with attractive plants hosting birds and other
tomographicimagesoftherootsystems(Fig.2).Thismethod
wildlife is an esthetically pleasing prospect that is gaining
showed high X-ray attenuation in the epidermis (outermost
popularity as a way of stabilizing or remediating metal-
cell layer) of the root tissue. The high X-ray attenuation was
contaminated soils and sediment(Weberand Gagnon, 2014;
attributed to Fe plaque, whose formation on the surface of
Weis and Weis, 2004). In such a wetland it is important
wetlandplantrootsiswellknown(Tripathietal.,2014;Liuetal.,
to understand the processes taking place — for example, if
2010;Panetal.,2014).
stabilization is indeed occurring, or if instead, metals are in
The other synchrotron-based method used in the study
a soluble form that can migrate and contaminate soils and
wasX-raymicrofluorescence(μXRF)atthemicro-meterscale,
sediment elsewhere. Metals in a soluble form may also be
withfreshsamplesthathadbeenfrozenandslicedinoptimal
taken up by the wetland plants, thereby entering the eco-
cuttingtemperature(OCT)compound(acommerciallyavail-
systemandpossiblycontaminatingfoodthatanimalseat.
able formulation of water-soluble glycols and resins) (Feng
A synchrotron study of wetland plants from an urban
etal.,2013;Sakura,2016).μXRFwasusedtomap,orvisualize,
brownfieldsitebyFengetal.(2016)offersinformationabout
the distribution of metals in the epidermis and vascular
a wetland scenario similar to that described above. In this
tissues of the root structures. This analysis confirmed the
study,multiplemetalsinplantsfromwetlandswerestudied
presence of Fe in the epidermis. Pb, Cu, Mn, and Zn were
toseeifthemetalswerebeingsequesteredinsideoroutside
enriched in the same area as the Fe (on the epidermis) in
theplants.Thestudylookedattherootsoftwoplantspecies
Typha and Phragmites from the drier location. This was seen
fromtwolocationsatabrownfieldformerlyusedasarailway
forPb,MnandZninPhragmitesfromthewetterlocationbut
notasmarkedlyforCu.Instead,Cuwasalsoenrichedinsome
⁎ Correspondingauthor. oftheinnerstructuresoftheroot.
http://dx.doi.org/10.1016/j.jes.2016.05.001
1001-0742/©2016TheResearchCenterforEco-EnvironmentalSciences,ChineseAcademyofSciences.PublishedbyElsevierB.V.
2 JOURNAL OF ENVIRONMENTAL SCIENCES 44 (2016) 1–3
TheμXRFresultswereusedtoquantifythemetalsinthe
roots by comparison to standard reference materials (NIST
1832and1833,thinglassfilmonpolycarbonate).Thistypeof
quantificationishighlyuncertainandnotcommonlycarried
out, but no attempt was made to quantify the uncertainty.
The authors found statistically significant differences be-
tween the concentrations in the epidermis and those inside
the plant. Metals were correlated with each other on the
epidermis,butforthemostpartnotinsidetheplant.
Althoughtheauthorsdidnotcompareresultsforthedif-
ferent plant species, or for plants from different wetland
locations(driervs.wetter),anexaminationofthedatareveals
thatMnandPbconcentrationsweresubstantiallylowerinthe
plantepidermisandvasculartissuefromthewetterlocation
comparedwiththesameplantspeciesfromthedrierlocation,
even though the soil/sediment concentrations were similar.
Atthesametime,thedifferencesbetweentheepidermisand
vascular metal concentrations were much less dramatic in
the plant from the wetter location compared with the same
plantfromthedrierlocation.Thustheplaquesequestration
andscavengingofmetalsappearstobeoccurringtoalesser
degree in the wetter conditions compared with drier condi-
tions. This is rather counter-intuitive, considering that Fe
plaqueformationtendstobegreaterinwetterandmorean-
aerobic conditions compared with drier conditions (Tripathi
et al., 2014). Differences in plant metal uptake in different
types of wetlands have been seen in other studies (Zhang
etal.,2011;Hanseletal.,2001).
TheCuinsidetheplantfromthewetterlocationwaseven
higherthanontheepidermis,afindingthatentirelycontra-
dictstheideaofmetalscavengingbyepidermalFeplaqueto
preventinfiltrationbycontaminants.Plantnutritionalneeds
must factor largely into how the Cu is taken up under the
Fig.1–PhotoshowingTyphalatifoliaLatasamplingsitein studyconditions,andindeed,theauthorsconcludethatmetal
LibertyStatePark,NewJersey,USA. transportandaccumulationareelementspecific.
ThispictureiscourtesyofDr.YuQian,MontclairState Overall the study shows that metals are associated with
University. Fe plaque on the outside of the roots of two very common
wetland plants, whichis a good confirmation that stabiliza-
tionbytheplantsisprobablytakingplace.Interestingfuture
studiescouldlookatnon-essentialtoxicelementsadditional
to the Pb examined in the study (e.g., Cd, As, Sb, U, Tl) as
thesemaybeofconcernincontaminatedsitescenarios.For
example, differences in Cd and As uptake by different rice
cultivars were thought to be related to the amount of Fe
plaque formed on the roots: more Fe plaque was associated
withlessCdandAsuptake(Liuetal.,2010;Panetal.,2014).
At the same time, a closer look at the essential elements
thatarecommonatcontaminatedsites–CuandZn–would
be warranted to gain a better understanding of how these
elementsarestabilized;thepresentstudyindicatesthatthere
Fig.2–Three-dimensionalimageofTyphalatifoliaL.root are probably powerful plant controls at work to ensure
withroothairshowingfromSynchrotronX-raycomputed nutritionalneedsaremet.
microtomography(μCMT)measurement.Highattenuation To comprehensively assess wetland stabilization as a
substancesshowinginredareseenintheepidermis. remediation technique, future research scenarios should in-
Althoughthechemicalcompositionofthehigh-attenuation clude the presence and absence of plants, to determine the
substancecannotbeidentifiedusingthecurrentsynchrotron overall ability of the plants to phytostabilize metals in a
μCMTmeasurement,synchrotronX-raymicrofluorescence wetland.However,evenalowphytostabilizationcontribution
(μXRF)measurementhasconfirmedthatFeplaqueis mightbeausefulpartofalowcostremediationmethodfor
concentratedwithinthesesubstances(Fengetal.,2016). brownfields and other contaminated sites that have a com-
Thisfig.iscourtesyofDr.Feng,MontclairStateUniversity. binationofmetalcontaminantspresent.
JOURNAL OF ENVIRONMENTAL SCIENCES 44 (2016) 1–3 3
REFERENCES Pan,W.,Wu,C.,Xue,S.,Hartley,W.,2014.Arsenicdynamicsinthe
rhizosphereanditssequestrationonricerootsasaffectedby
rootoxidation.J.Environ.Sci.26(4),892–899.http://dx.doi.org/
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2013.LeadaccumulationandassociationwithFeonTypha Sakura,2016.Tissue-TekO.C.TcompoundandCryomolds.
latifoliarootfromanurbanbrownfieldsite.Environ.Sci.Pollut. http://www.sakura.eu/Our-products/item/11/Cryotomy/48/
Res.20(6),3743–3750. Tissue-Tek-OCT-Compound-and-Cryomolds(AccessedMay2016).
Feng,H.,Qian,Y.,Gallagher,F.J.,Zhang,W.,Yu,L.,Liu,C.,etal., Tripathi,R.D.,Tripathi,P.,Dwivedi,S.,Kumar,A.,Mishra,A.,
2016.Synchrotronmicro-scalemeasurementofmetal Chauhan,P.S.,etal.,2014.Rolesforrootironplaquein
distributionsinPhragmitesaustralisandTyphalatifoliaroot sequestrationanduptakeofheavymetalsandmetalloidsin
tissuefromanurbanbrownfieldsite.J.Environ.Sci.41, aquaticandwetlandplants.Metallomics6(10),1789–1800.
172–182. Weber,K.P.,Gagnon,V.,2014.Microbiologyintreatment
Hansel,C.M.,Fendorf,S.,Sutton,S.,Newville,M.,2001. wetlands.SustainableSanitationPractice:Outcomesfromthe
CharacterizationofFeplaqueandassociatedmetalsonthe UFZWetlandWorkshopSpecialIssue18(2014),pp.25–30.
rootsofmine-wasteimpactedaquaticplants.Environ.Sci. Weis,J.S.,Weis,P.,2004.Metaluptake,transportandreleaseby
Technol.35(19),3863–3868.http://dx.doi.org/10.1021/ wetlandplants:implicationsforphytoremediationand
es0105459. restoration.Environ.Int.30(5),685–700.http://dx.doi.org/10.
Liu,J.G.,Cao,C.X.,Wong,M.H.,Zhang,Z.J.,Chai,Y.H.,2010. 1016/j.envint.2003.11.002.
Variationsbetweenricecultivarsinironandmanganese Zhang,H.,Cui,B.,Zhang,K.,2011.Heavymetaldistributionof
plaqueonrootsandtherelationwithplantcadmiumuptake. naturalandreclaimedtidalriparianwetlandsinsouthestuary,
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JOURNAL OF ENVIRONMENTAL SCIENCES 44 (2016) 4–12
Available online at www.sciencedirect.com
ScienceDirect
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Structural and metabolic responses of microbial community to
sewage-borne chlorpyrifos in constructed wetlands
Dan Zhang1,2, Chuan Wang1, Liping Zhang1, Dong Xu1, Biyun Liu1,
Qiaohong Zhou1,⁎, Zhenbin Wu1
1.State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan
430072, China. E-mail: [email protected]
2.GraduateUniversityofChineseAcademyofSciences,Beijing100049,China
A R T I C L E I N F O A B S T R A C T
Articlehistory: Long-termuseofchlorpyrifosposesapotentialthreattotheenvironmentthatcannotbe
Received27January2015 ignored, yet little is known about the succession of substrate microbial communities in
Revised8July2015 constructedwetlands(CWs)underchlorpyrifosstress.Sixpilot-scaleCWsystemsreceiving
Accepted20July2015 artificial wastewater containing 1mg/L chlorpyrifos were established to investigate the
Availableonline8February2016 effects of chlorpyrifos and wetland vegetation on the microbial metabolism pattern of
carbon sources and community structure, using BIOLOG and denaturing gradient gel
Keywords: electrophoresis(DGGE)approaches.Basedonoursamples,BIOLOGshowedthatShannon
Constructedwetland diversity(HV)andrichness(S)valuesdistinctlyincreasedafter30dayswhenchlorpyrifos
Chlorpyrifos wasadded. Atthesame time, differencesbetweenthevegetated andthenon-vegetated
BIOLOG systemsdisappeared.DGGEprofilesindicatedthatHVandShadnosignificantdifferences
DGGE among four differenttreatments.Theeffectofchlorpyrifoson themicrobialcommunity
wasmainlyreflectedatthephysiologicallevel.Principalcomponentanalysis(PCA)ofboth
BIOLOG and DGGE showed that added chlorpyrifos made a difference on test results.
Meanwhile,therewasnodifferencebetweenthevegetationandno-vegetationtreatments
after addition ofchlorpyrifosatthe physiological level.Moreover, thevegetation had no
significanteffectonthemicrobialcommunityatthegeneticlevel.Comparisonsweremade
betweenbacteriainthisexperimentandotherknownchlorpyrifos-degradingbacteria.The
potentialchlorpyrifos-degradingabilityofbacteriainsitumaybeconsiderable.
©2016TheResearchCenterforEco-EnvironmentalSciences,ChineseAcademyofSciences.
PublishedbyElsevierB.V.
Introduction non-pointsourcepollution,andincreasedrisktothequality
of aquatic environments (van Dijk and Guicherit, 1999;
Chlorpyrifosisoneofthechlorinatedorganophosphate(OP) Spalding et al., 2003; Leu et al., 2005). Accumulation of
pesticidesthathavebeenusedforpestcontrolinagriculture chlorpyrifos in water bodies could cause potential damage
sincethe1960s(Mayaetal.,2011).Thehalf-lifeofchlorpyrifos (such as carcinogenicity, neurotoxicity, and reproductive
isusually60to120daysinsoil,butitcanrangefrom2weeks anddevelopmental toxicity)tobothaquatic organisms and
toover 1yeardepending on the illuminationintensity, soil humans(Jorgenson,2001;Robles-Mendozaetal.,2009).Thus,
type, temperature and other factors (Anwar et al., 2009). thedemandforcost-effectivemethodstoremovechlorpyr-
Long-term usage of chlorpyrifos has caused agricultural ifosinpesticidewastewaterisincreasing.
⁎ Correspondingauthor.E-mail:[email protected](QiaohongZhou).
http://dx.doi.org/10.1016/j.jes.2015.07.020
1001-0742/©2016TheResearchCenterforEco-EnvironmentalSciences,ChineseAcademyofSciences.PublishedbyElsevierB.V.
JOURNAL OF ENVIRONMENTAL SCIENCES 44 (2016) 4–12 5
Constructed wetlands (CWs) have been proven to be an
effective management practice to reduce aqueousconcentra-
tionsofpesticides(Mooreetal.,2009)andtocontrolpesticide
mitigation(SchulzandPeall,2001;Mooreetal.,2007;Buddetal.,
2011).ThedegradationofpesticidesinCWsmainlytakesplace
by photolytic degradation, substrate sorption, plant uptake,
andmicrobialdegradationprocesses(Zhangetal., 2014). The
efficiencyofdifferentmicrobialdegradationpathwayshasalso
been studied. Numerous studies have concentrated on the
selection of specific pesticide-degrading bacteria in different
pesticide-pollutedsamples(Lakshmietal.,2009;Sasikalaetal.,
2012). Nevertheless, research on the effect of pesticides on
microbial community dynamics should not be neglected.
Vegetated Non-vegetated
Thiscanprovidebackgroundinformationforpesticidetesting
(Engelenetal.,2003).
In addition to microbial degradation, plant uptake is Fig.1–IchnographyofsimulativeverticalflowCWs.
another major method that can decrease chlorpyrifos. Also,
there have been a large number of studies focused on the
interactionof plantsand microbesbeing treatedwithchlor-
pyrifos(Fangetal.,2009;Xieetal.,2010).Therootsofplants CWs.Intheexperimentalstage,chlorpyrifoswasaddedinthe
couldprovideastableenvironmentandsuitableattachment artificial sewage until its concentration reached 1mg/L.
points for rhizosphere microorganisms, and root exudates Chlorpyrifos(purity99.9%)waspurchasedfromSigma-Aldrich
couldalsoprovideenergyandcarbonsourcesforthegrowth Company. The hydraulic loading was 25.48mmd−1. Water
ofmicrobes(Faulwetteretal.,2009). samples were collected on days 4, 7, 17 and 29 after the
In this article, the short-term response (30days) of chlorpyrifoswasadded.Thechlorpyrifosremovalratesondays
microbial community structure was examined before and 4,7,17and29inthevegetatedsystemsamountedto(93.97±
after the addition of chlorpyrifos in both vegetated and 1.62)%, (96.57±1.46)%, (96.14±1.71)% and (95.72±0.64)%, re-
non-vegetated CWs. Both BIOLOG (Garland and Mills, 1991) spectively, and in the non-vegetated systems amounted to
andpolymerasechainreaction-denaturinggradientgelelec- (87.65±3.77)%, (96.12±1.28)%, (94.33±1.03)% and (91.67±
trophoresis(PCR-DGGE)techniques(Muyzer,1999)wereused 2.32)%,respectively.
torevealthebasicfeaturesofthemicrobialcommunity,with
respect to metabolism and genotypic structure, respectively 1.2.Substratesamplecollection
(Bushaw-Newtonetal.,2012).Theaimofthisresearchwasto
exploretheresponseofmicrobesinrhizospheresubstratesof Samples were collected from the rhizosphere substrate, and
constructed wetlands being treated with chlorpyrifos at the eachsubstratesamplewascollectedfrom5uniformlydistrib-
physiologicalandgeneticlevels. uted spots of the surface layer (0–10cm) from one randomly
selected bucket, and then mixed evenly. Substrate samples
weretakenatthebeginningandtheendoftheexperimental
1.Materialsandmethods stage, and stored at −80°C prior to analysis. The samples
collectedatthebeginningandendoftheexperimentalstagein
1.1.Experimentalsystemsandoperatingconditions vegetated systems were designated Pbs and Pas respectively,
whilethoseinnon-vegetatedsystemsweredesignatednon-Pbs
Six sets of vertical flow CWs were made from polyethylene andnon-Pasrespectively.
(PE) buckets with a height of 600mm and a diameter of
250mm, which were equally separated into two groups on 1.3.BIOLOGECOmicroplateexperiment
vegetatedandnon-vegetatedCWsystems.Eachsystemwas
filledwithriversand(1–5mm)withaheightof300mm,and TheBIOLOGECOmicroplatescontained31carbonsourcesof
onlythevegetatedgroupwasplantedwiththreeuniformIris differentgroups: 12 kinds of carbohydrates (CHs), 6 kinds of
pseudacorus. The river sand was passed through a sieve to aminoacids(AAs),5kindsofcarboxylicacids(CAs),4kindsof
removethelargeparticles,andthenwashedwithtapwaterin polymers (PMs), 2 kinds of amines (AMs), and 2 kinds of
ordertoremovethesmallparticles.Therefore,theremaining phenoliccompounds(PCs).A250mLconicalflaskcontaining
riversandwas1–5mmindiameter.TheI.pseudacorususedin 10g substrate and 100mL of sterile saline was shaken for
this experiment was obtained from the cultivation base of 30min (200r/min) at room temperature. The supernatant
vegetation of the institute of hydrobiology. Fig. 1 shows the was diluted 25 times, and 150μL of the diluted bacterial
ichnography of the simulative vertical flow CWs. After an suspension was added to each well of a BIOLOG ECO
acclimatization stage of 15days, the experimental stage microplate.Themicroplatewasincubatedat30°Cindarkness.
lasted for 30days. In the acclimatization stage, the artificial Optical density (OD) was measured with SpectraMax M5 at
sewage, which was composed by 7–8mg/L of total nitrogen 590nm (color and turbidity) and 750nm (turbidity) wave-
(TN),0.2–0.4mg/Loftotalphosphorus(TP),and60–70mg/Lof lengthsevery12hruntilnomoregrowthinODvaluecouldbe
chemical oxygen demand (COD), was discharged into the observed.
6 JOURNAL OF ENVIRONMENTAL SCIENCES 44 (2016) 4–12
1.4.ExtractionofgenomicDNA whereCisthedifferenceinODvalueofeachcarbonsource,R
istheODvalueofthecontrolhole,andnisthetotalnumberof
TheSDSbasedmethodofDNAextraction(Zhouetal.,1996) carbonsourceclasses(n=31inthisresearch).
and E.Z.N.A Gel Extraction Kit (OMEGA, USA) were used to X
obtainthegenomicDNAofallthesamples.ThepreparedDNA H0¼− ðPi(cid:2) logPiÞ
wasstoredat−20°Cbeforethenextstep.
where P (in the BIOLOG experiment) is a probability that
i
1.5.PCRamplification representsthedifferenceofODvaluebetweenamediumpore
and control, namely P ¼ðC−RÞ=∑ðC−RÞ(Dobranic and Zak,
i
The extracted DNA was subjected to PCR to amplify the 1999).ThenaturallogarithmwasusedinthispaperwherePi
bacterialV3regionofthe16SrDNAgene(from1055to1406), (intheDGGEexperiment)standsfortheobservednumberof
withPCRprimerpairGC-338fand518r.GC-338fhada40-bp clones of a given species divided by the total number of
GC clamp CGCCCGCCGCGCGCGGCGGGCGGGGCGGGGGCACG organisms.
GGGGG attached at the 5′-end. PCR reaction system was S=thenumberofreaction(intheBIOLOGexperiment);the
composedof15μL2×TaqPCRMasterMix(ZT201,Zomanbio), observednumberofbands(intheDGGEexperiment).
0.2μL of each primer (100μmol/L) and 1μL of the extracted One-wayanalysisofvariance(one-wayANOVA)wasused
DNA (10–20ng/μL), and then adjusted the volume to 30μL to analyze the data of AWCD, HVand S. The standardized
using sterile deionized water. PCR amplification was per- data of band intensity was processed in PAST3 for principal
formedusingaThermalCycler(Bio-RadT100,USA)underthe componentanalysis(PCA).Principalcomponentanalysiswas
following conditions: 95°C for 5min, followed by 35cycles processed on BIOLOG data as well. The banding patterns of
consistingof95°Cfor1min,55°Cfor1minand72°Cfor1min, DGGE profiles were analyzed using the Quantity One V4.62
with a final extension period at 72°C for 10min. The PCR Software(Bio-Rad).
productsstainingwithGelRedwereverifiedbyrunninga1%
(w/v)agarosegelelectrophoresisin0.5×TBEbuffer.
2.Results
1.6.DGGEexperiment
2.1.BIOLOGanalysisofcultivablebacteria
The DGGE experiment was carried out using a DCode™
Universal Mutation Detection System (Bio-Rad, USA). Using a 2.1.1.Metabolicdiversityandrichnessofmicrobialcommunity
40% polyacrylamide gel at 60°C, PCR-amplified samples (200– The following BIOLOG indices were calculated: AWCD and
300ng)weresubjectedtoelectrophoresisfor7hrat150V.A40% richness (S), which are indicators reflecting carbon source
to60%denaturinggradientwasperformed.Afterelectrophore- utilizationability(ChoiandDobbs,1999;Zhangetal.,2013).In
sis, the gel was stained with SYBR Green I (Molecular Probes, addition, Shannon's diversity (HV), which stands for the
USA)for30minwith1×TAEbuffer.Thenthegelwasexamined diverseutilizationof carbon sourcesbysoil microorganisms
underaGelDoc™XR+ImagingSystem(Bio-Rad,USA). (Shannon, 2001), was calculated. Moreover, S and HVwere
calculatedusingthedatafromthe180hrincubationreadings
1.7.CloningandDNAsequencinganalysis (Table 1). Before the addition of chlorpyrifos, S (BIOLOG)
ranged from 5.25–13.00, and HV(BIOLOG) ranged from 1.43–
The excision and reamplification of the desired bands were 2.20. Beforeadding chlorpyrifos, HV(BIOLOG) and S (BIOLOG)
carried out. The desired bands were excised, resuspended in were both higher in non-vegetated CWs than in vegetated
30μL sterile water and stored at 4°C overnight. The PCR CWs(p<0.05).Whencomparingsamplesaftertheadditionof
amplificationwasperformedasabove,exceptusing338fasthe chlorpyrifostothosebeforechlorpyrifosaddition,S(BIOLOG)
forwardprimer.PCRproductswerepurifiedusingtheAgaroseGel and HV(BIOLOG) in CWs were increased to 18.75–19.83 and
DNA Extraction Kit (TaKaRa) and cloned into pMD-18T vector 2.87, respectively. However, HV(BIOLOG) and S (BIOLOG) had
(TaKaRa).Fivepositiveclonesofeachbandweresequencedby nosignificantdifferencesbetweennon-PasandPas.
WuHantsingkeBioTechCompany.Thesequencesobtainedfrom
thecompanywereanalyzedattheNCBIwiththeBLASTprogram 2.1.2. AWCD and its PCA analysis of microbial BIOLOG-ECO
(http://www.ncbi.nlm.nih.gov/blast). Cluster analysis was per- profiles
formedbyMEGA5.1withtheneighbor-joiningmethod. Theutilizationabilitiesofthesixgroupsofcarbonsourcesby
microbesareshowninFig.2.CHs,AAsandCAswerethemain
1.8.StatisticalanalysisofBIOLOGandDGGE carbonsourcesusedbymicrobesinalltreatments.CHs,AAs,
CAsandAMswereincreasedsignificantlyafteraddingchlor-
Average well color development (AWCD), Shannon–Wiener pyrifos into artificial sewage. However, PCs and AMs had
diversityindex(HV)andrichness(S)werecalculatedtoshow significant difference between non-Pbs and Pbs. The AWCD
thepropertiesofcarbonsourcesandmicrobialcommunity(da values of these two groups could be detected in the non-Pbs
Motaetal.,2005;RogersandTate,2001;Classenetal.,2003). treatment, but decreased to zero in the Pbs treatment. By
Thecomputationformulasofthethreeindexesareasfollows. contrast,therewasnosignificantdifferencebetweennon-Pas
andPasamongthesixgroupsofcarbonsources.
X The first two principal components were sufficient to
AWCD¼ ðC−RÞ=n
explain about 86.99% of the total variance in AWCD data