Effect of Aeration Rates and Filter Media Heights on the Performance of Pollutant Removal in an

13 Pages·2017·2.25 MB·English

water
Article
Effect of Aeration Rates and Filter Media Heights on
the Performance of Pollutant Removal in an Up-Flow
Biological Aerated Filter
JiehuiRen1 ,WenCheng1,TianWan1,*,MinWang1andChengchengZhang2
1 StateKeyLaboratoryofEco-hydraulicinNorthwestAridRegion,Xi’anUniversityofTechnology,
Xi’an710048,China;[email protected](J.R.);[email protected](W.C.);[email protected](M.W.)
2 CollegeofComputing,GeorgiaInstituteofTechnology,Atlanta,GA30301,USA;[email protected]
* Correspondence:[email protected];Tel.:[phone]
(cid:1)(cid:2)(cid:3)(cid:1)(cid:4)(cid:5)(cid:6)(cid:7)(cid:8)(cid:1)
(cid:1)(cid:2)(cid:3)(cid:4)(cid:5)(cid:6)(cid:7)
Received:24August2018;Accepted:7September2018;Published:14September2018
Abstract: Thebiologicalaeratedfilter(BAF)isaneffectivebiologicaltreatmenttechnologywhich
removesthepollutantsinmunicipalwastewatersecondarytreatment. However,westillknowlittle
abouttheinteractionbetweenthepollutantsremovalandmicrobeswithintheBAF.Inthisstudy,
we used an up-flow BAF (UBAF) reactor to investigate the relationships between the pollutants
removal and microbial community structure at different aeration rates and filter media heights.
ThemicrobialcommunityofbiofilmwasanalyzedbyIlluminapyrosequencing. Ourresultsshowed
thattheUBAFachievedabetterremovalefficiencyofchemicaloxygendemand(COD),NH +-N,
4
NO −-N,andtotalphosphorus(TP)atanaerationrateof65L/h. Inaddition,theCODandNH +-N
3 4
removalmainlyoccurredat0–25cmheightoffiltermedia. Themicrobialcommunitystructureinthe
UBAFdemonstratedthattherelativeabundanceofthePlanctomycetesandComamonadaceaeat10cm
heightoffiltermediawere11%and48.1%,respectively,proportionssignificantlyhigherthanthose
underotherstreatments. Finally,thechangesinrelativeabundanceofProteobacteria,Planctomycetes,
andNitrospiraelikelyexplainedthemechanismofnitrogenandphosphorusremoval. Ourresults
showedthatsuitableconditionscouldenhancethemicrobialcommunitystructuretoachieveahigh
pollutantsremovalintheUBAF.
Keywords: aerationrates;filtermedia;microbialcommunity;pollutantsremoval;up-flowbiological
aeratedfilter
1. Introduction
Duringthepasttwodecades,thebiologicalaeratedfilter(BAF)begantosubstitutethetraditional
activatedsludgeprocessforthewastewatertreatmentinmunicipalwastewater[1,2]. TheBAFexhibits
a large number of advantages, such as filtration of solids, organic matter, and nutrient removal,
which occur in the same reactor [1–3]. The BAF systems also enhance the biomass and pollutants
loading capacity, reduce the biomass residence time, improve the environmental shock resistance,
andreducesludgeproduction[1]. TheBAFarerelativelycompact,easytooperate,andmaybemore
efficientincarbonaceousandammoniaremovalthanactivatedsludgesystem[4]. Therefore,theBAF
possessesbroadapplicationprospectsinwastewatertreatment.
TherecentresearchontheBAFfocusesmainlyonoptimizingoftheremovalefficiencyandthe
mechanismofpollutantsremoval[3,5]. Inthatcontext,theaerationusuallyoccupieshalfofthetotal
energy consumption in the biological treatment process [6,7]. Moreover, it plays an essential role
inBAFoperationbycontrollingsupplyofoxygenformicrobialdegradationatlowturbulenceand,
consequently, interferes in the microbial activity [8,9]. Thus, an appropriate aeration rate reduces
the energy costs of the BAF [5,8–10]. Abu Hasan et al. [9] reported that the aeration rate also
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influencedammoniaremovalandeffectivemicrobesintheBAFsystem. Taoetal.[5]optimizedthe
reactorstructureandexploredthechemicaloxygendemand(COD),totalnitrogen(TN),andNH +-N
4
removalatdifferentfiltermediaheightsbysettinganon-aerationzoneatthebottomofthereactor.
Yangetal.[11] studied the pollutants removal using combined UAF–UBAF system, the COD and
NH +-Nremovalreachedto90%and89.0%ofremoval,respectively. However,theyfailedtostudythe
4
mechanismofphosphorusremoval. Removalofbiologicalnitrogenoccursintwostagesoftreatment:
Aerobicnitrificationandanoxicdenitrification[4]. Polyphosphatebacteriaabsorbedthephosphorus
andremovedunderaerobicconditions. Someresearchersalsousedacombinationofprocessesor
methodsincludingFe2+ toeffectivelyremovephosphorus[4,12,13]. IntheBAFsystem,thebiofilm
playsanessentialroleinthemechanismofpollutantsremoval. Toaccessandanalyzethemicrobial
communityinwastewater,previousresearchesappliedseveralmethods,suchas: PolymeraseChain
Reaction,DenaturingGradientGelElectrophoresis(DGGE)[14],Terminal-RestrictionFragmentLength
Polymorphism (T-RFLP) [15], and pyrosequencing of 16S rRNA gene sequences [16]. Nowadays,
manyresearchersanalyzedthefunctionalmicrobesstructureusing454-pyrosequencinganalysis[11,14].
Thefunctionalmicrobialcommunityalsohelpstounderstandthemechanismofpollutantsremoval.
Severalstudiesalreadyinvestigatedthepollutantsremovalefficiencyandoptimizedtheoperation
intheBAF[5,12]. However,fewstudiesexploredtheeffectsofaerationratesandfiltermediaheights
onthepollutantsremovalandmicrobialcommunityintheBAFsystem. Additionally,phosphorus
removalhasalwaysbeenoneofthecriticalrestrictionstotheapplicationofBAF.Suitableaeration
ratesnotonlyachievedhighernitrogenremovalefficiencybutalsosignificantlyincreasingphosphorus
removal. Moreover,intheBAFsystem,thestudyofpollutantsremovalatdifferentfiltermediaheights
becomesincreasinglyimportantforthepracticalapplicationoftheBAF.
Basedonthat,webuiltanup-flowBAF(UBAF)reactortoinvestigatetherelationshipsbetween
thepollutantsremovalandmicrobialcommunitystructureatdifferentaerationratesandfiltermedia
heights. Ourstudyaimed(i)toassesstheeffectofaerationratesonCOD,nitrogen,andphosphorus
removalefficiency;(ii)toexplorethepollutantsremovalmechanismatdifferentfiltermediaheights;
(iii)toanalyzethemicrobialcommunitystructureatdifferentaerationratesandfiltermediaheights
using Illumina pyrosequencing; and (iv) understand the relationship of pollutants removal and
microbialcommunityintheUBAFreactor.
2. MaterialsandMethods
2.1. DescriptionoftheUp-FlowBAFSystem
An up-flow BAF reactor is presented in Figure 1. The reactor was designed with a height of
1470mmandadiameterof350mm. TheUBAFreactorincludedasupportzoneof200mm,formed
byanaccumulationofcobblewithadiameterof10–30mmandaheightof200mm. Theupperlayer
ofthesupportzoneconsistedofceramistmaterialwithaneffectivediameterof3–5mmandavoid
ratioof0.43. Theheightoffiltermedialayerwas850mm. Theperforatedplatewasusedtoensurethe
distributionofthewaterandairinsidethereactoratthebottomofthesupportzone. Thetopoffilter
medialayerwassettoawaterzoneof150mm. Sixsamplingportsweredesignedalongthedepth
ofthereactorat100–150mmintervals. Theairwasintroducedintothereactorviaaircompressor,
whichwasinstalledata150mmdepth. Aflowmeter(LZB-4, YinHuan, Changzhou, China)was
usedtodeterminetheaerationrate. Amechanicaldiaphragmpump(GM0090PR9(6)MNN,Nan
fang Pump, Hangzhou, China) was applied to pump the synthetic wastewater into the bottom of
thereactor.
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Figure1.Experimentalschematicoflaboratory-sca leup-flowbiologicalaeratedfilter(UBAF)system.
Figure 1. Experimental schematic of laboratory-scale up-flow biological aerated filter (UBAF) system.
2.2. InoculatedSludgeandSyntheticWastewater
2.2 Inoculated Sludge and Synthetic Wastewater
TheUBAFreactorreceivedsyntheticwastewater. Thesyntheticwastewaterwascomposedoftap
waterThwei tUhBaAmFi xreeadcotofrC reHceivOed (scyanrbthoentisco wuracset)e,wNaHterC. lTahned syKnNthOeti(cn witraosgteewnastoeur rwcea)s, KcomPOpo.s3eHd oOf
6 12 6 4 3 3 4 2
(tpaph owspatheorr wusitsho au rmceix),eadn odf CCa6H2+1,2OM6g (2c+a,rNboan+ ,saonudrcFee),3 +N(Htr4aCcle aenledm KeNntOs3s (onuitrrcoeg).eInn soocuurlactee)d, Ks3lPuOdg4.e3Hw2aOs
c(pohlloecstpehdofrruosm sothuercsee)c, oanndda rCyas2e+,d Mimge2n+, tNataio+,n atnadnk F(eX3+i ’(atnratchei redlewmaesntetws saoteurrcpela).n Itn,oXciu’alna)t.edT hseluadcgtiev awteads
scloulldegcetewd afrsocmul ttihvea steedcoanfdtearryco sleledcitmioennatastifoonll otawnikn g(X: Fi'airns ttlhy,irwd ewcausltteivwaatetedr spluladngt,e Xini'aan6).0 TLhew aactteirvtaatnedk
fseluddwgiet hwCas HcultOiva(t8e0d0 amftger/ Lco),llNecHtioCnl a(s3 5fomllogw/Lin),gK: FNirOstly(8, wmeg /cuLl)t,iKvatPeOd sl(u8dmgge/ inL )a, C60a 2L+ w(2a0tmerg t/anLk),
6 12 6 4 3 3 4
Mfedg 2w+i(t2h5 Cm6Hg1/2OL)6, (N80a0+ m(1g1/0L)m, NgH/L4C),l a(n35d mFeg3/+L)(,1 K.5NmOg3 /(8L )magt/L20), ±K3P3O◦C4 (;8t hmegn/,La),e Craat2e+d (2t0h emmg/wL)i,t hMagi2r+
c(2o5m mprge/sLs)o, rNfao+r (611h0e mvegr/yLd),a ayntdo Foeb3t+a (i1n.5t hmegta/Lrg) eatt o2b0j e±c t3. °FCin; athlley,nt,h aeeraactteivda tthedemsl uwditghe awira scocmulptirveastseodr
ffoorr o6 nhe ewveereyk dwaiyth tod oabiltyairne pthlaec teamrgeentt oobfjtehcet. nFuintrailelyn,t tmhee daicutimva.ted sludge was cultivated for one week
with Dduairliyn rgepbloatchempheanste osf othfes tnaurtt-ruiepnta mndedbiiuomfil.m culture in the UBAF reactor, the ratio of carbon,
nitrogDeunr,ianngd bpohtohs pphhoarsuess soofu srtcaeritn-utphe aenxdp ebriiomfielnmta cluwltaustreew inat ethrew UasB1A00F: 5r:e1a(cCto:2r,0 0thme gr/aLti;oN o:f1 5camrbgo/nL,;
Pni:t3romgegn/, La)n.dIn pohrodseprhtooruans asloyuzrecteh ien ptehref oerxmpearnicmeeonftathl ewUasBtAewFarteear cwtoar,s t1h0e0s:5y:n1 t(hCe:t i2c0w0 amstge/wL;a Nte:r 1in5
tmheg/sLim; Pu: l3a tmiogn/Lo)f. eIxnp oerrdimere tnot aalnpahlyazsee twhea spbearfsoerdmoanntchee ocfo tnhtea UmBinAaFti orenalcetvoer,l sthoef sdyonmtheesttiicc wwaasstteewwaatteerr.
Tinh ethcaer bsoimn,unliattrioogne noaf nedxppheroismpehnotrauls pcohnacseen twraatsio nbsasoefde xopne ritmhee nctaolnwtaamstienwataiotenr wleevreels2 0o0–f 3d00ommegs/tLic,
3w4a–s4t8ewmagte/rL. T(NheH ca+r-bNon: ,3 0n–it4r0ogmeng /aLn;dN pOho−sp-Nho:r4u–s8 cmongc/enLt)r,aatniodns4 –o6f emxgp/erLi,mreensptaelc wtivaestlye.wTatheer twraecree
4 3
e2l0e0m–3e0n0t smcgo/nLc, e3n4t–r4a8ti mong/oLf (CNaH2+4,+-MN:g 320+–,4N0 am+,ga/Ln;d NFOe33−+-Nin: 4e–x8p mergim/Le),n atnaldw 4–a6s tmewg/aLte, rrewspeerceti2v0elmy.g T/hLe,
2tr5amceg e/lLem,1e1n0tsm cgo/nLc,eanntrda1ti.o5nm ogf/ CLa,2r+e, sMpegc2t+i,v Nelay+., Tahnedp FHe3l+e ivne leixnpseyrinmtheenttiaclw waastsetewwaatetrerr awnegreed 2b0e tmwge/eLn,
725a nmdg9/L., 110 mg/L, and 1.5 mg/L, respectively. The pH level in synthetic wastewater ranged between
7 and 9.
2.3. OperationofTheUp-FlowBAF
2.3 Operation of The Up-Flow BAF
TheoperationoftheUBAFconsistedofthreephases: (i)Reactorstart-up; (ii)biofilmculture;
and(Tiihi)e soimpeurlaattiioonn oefx ptheer iUmBeAntF. Fcoirnsstliys,tetdh eorfa tphirdeem pehthaoseds:o (fi)a cRtievaactteodr sstlaurdt-guepw; (aisi) ubsieodfildmu rciunlgtutrhee;
panhdas (eisii)o fsirmeaucltaotriosnt aerxt-pueprimanedntb. iFoifirlsmtlyf,o trhme artaiopnid. Tmheethdoodm oefs taicctaivteadteadc tsilvuadtgede wsluasd guese(da wduereikn)gw thaes
dpihlausteesd otof r5e0a0c0tomr gst/aLrt-ounpM anLdSS baionfdilmfe dfotromthateiofinlt. eTrhme eddoimaaebstoicvaete3d0 camctivinattehde sUluBdAgFe r(eaa wctoeer.kT) hweans,
tdhielurteeadc ttoo r5w00a0s mlegt/tLo osnta MndLSfoSr aonnde fdeday t.oL thatee frilotenr, mtheedrieaa acbtoorvoep 3e0r camte dina tthaen UaBeAraFti roenacratoter. oTfh8e0n,L t/hhe
froeracthtorre ewdaasy lse.t Atot esvtaenryd nfoigrh otn,teh deasyu.p Learnteart aonnt, wthaes rderaacitnoerd ofproemrattehde asty satne mae,raantidonan raetqeu oafl v8o0 lLum/he foorf
sthyrneteh edtaicysw. aAstt eewvaerteyr n(Cig:hNt,: Pth=e 1s0u0p:5e:r1n)awtaanst fwedasto dtrhaeinUedB AfrFomre atchteo rs.yNsteexmt,, tahnedr eaanc teoqrubaelg vaonlutomfee eodf
csoynnttihneutiocu wslaysatetwaastmera l(lCw:Nat:Per =fl o1w00o:5f:15) Lw/ahs afnedd gtor atdhue aUllByAreFa rcehaecdtotrh.e Ndeexsti,g tnhfle orweacrtaoter b(1e2gaLn/ hto) afefteedr
acownetienku.oInusaldyd aitt iao ns,mthaell rweaactteorrflwowas ocfo 5n tLin/hu oaunsdly gfreadduwailtlhys ryenatchheetdic twhea sdteewsiagtne rfl(oCw:N r:aPte= (11020 L:5/:h1)) uafntteirl
tah weCeeOkD. I,nN aHd4d+i-tNionr,e mthoev raelaicntotrh eweafsfl ucoennttinacuhoiuevsleyd fsetde awdiyth-s tsaytne.thFeintiac llwy,atshteewexapteerr i(mC:eNn:tPw =a s10s0e:t5u:1p)
tuonstitlu tdhyei CngODth,e NreHm4+o-Nva rleemffiocviaeln icny tohfe peoffllluuetnant taschaitedviefdfe sretenatdaye-rsatatitoen. Friantaelslya,n tdhefi elxtepremrimedeinat hweaigsh stest.
Tuhpe toC sOtuDd,yNinHg4 +th-Ne r,eNmOo3v−a-lN e,ffaicnidenTcPy ionf spaomllpulteasnwtse arte dainffaelryeznetd ateorasttiuodny rathteesp aonldlu ftialtnetrs mreemdoiav ahleiinghthtse.
The COD, NH4+-N, NO3−-N, and TP in samples were analyzed to study the pollutants removal in the
UBAF reactor. The aeration rates were set at 40 L/h to 90 L/h, the waterflow rate was kept at 12 L/h,
and the HRT was kept at 5 h. The reactor operated at room temperature (12 ± 3 °C).
Water2018,10,1244 4of13
UBAFreactor. Theaerationratesweresetat40L/hto90L/h,thewaterflowratewaskeptat12L/h,
andtheHRTwaskeptat5h. Thereactoroperatedatroomtemperature(12±3◦C).
2.4. ChemicalAnalysis
The samples were collected to analyze the pollutants removal at the influent, effluent,
and different filter media height in the UBAF reactor. The DO, pH, and temperature in the water
samples were detected with an HQ d Portable Meter (HQ30d, HACH, New York, NY, USA).
Beforeconductingothermeasurements,thewatersampleswerefilteredthroughqualitativefilterpaper
(Xinxingϕ12.5cm,Hangzhou, China). ThelevelsofNH +-NandNO −-Nweredeterminedusing
4 3
aUVspectrophotometer(DR5000,HACH,NewYork,NY,USA)at420nmforNH +-N,220nmand
4
275nmforNO −-Naccordingtostandardmethods.TPwasmeasuredusinganammoniummolybdate
3
spectrophotometricat700nm. CODwasanalyzedusingthestandardpotassiumdichromatemethod
withaSpeedDigester(DRB200,HACH,NewYork,NY,USA).
2.5. MicrobialCommunityAnalysis
Biofilm samples at the different aeration rates (40 L/h, 65 L/h, 90 L/h) and filter media
heights (10 cm, 40 cm, 70 cm) were collected when the UBAF reactor had a relatively stable
operation. The Power Biofilm™ DNA Isolation kit for biofilm (MOBIO, Carlsbad, CA, USA) was
used to extract the DNA. The extracted steps reference the instructions of manufacturer. Biofilm
samples were firstly weighed out 0.02 g to 0.2 g and place it into a 2 mL collection tube using
a pipette tip so that removing excess liquid. Next, breaking the cell wall and DNA purification
werecarriedoutaccordingtooperationstepsonthekit. NanoDropSpectrophotometer(DC2000,
ThermoScientific,Waltham,MA,USA)andElectrophoresis(DYY-6C,Beijing,China)wereusedto
detecttheextractedDNAconcentrationsandpurity. TheDNAsampleswerestoredat−20◦Cuntil
further analysis. Then, the gene amplification of DNA was performed via the polymerase chain
reaction(PCR)(2720,ABI)usingtheprimers515F(5(cid:48)-GCACCTAAYTGGGYDTAAAGNG-3(cid:48))and907R
(5(cid:48)-TACNVGGGTATCTAATCC-3(cid:48))whichcorrespondedtotheV4domainofbacterial16SrRNAgenes.
Afterthat,theamplifiedDNAwassequencedinIlluminaHiseq2000platformfromShanghaiPersonal
BiotechnologyCo.,Ltd(Shanghai,China).
2.6. StatisticalAnalysis
SPSS 22.0 for Windows (SPSS Inc., Chicago, IL, USA) was used for statistical analyses.
Pearsoncorrelationanalysiswasperformedtodeterminetherelationshipsbetweenthepollutants
removal, aeration rates, and filter media heights. The significance levels used were p < 0.05 * and
p<0.01**.Rpresentedthecorrelationcoefficient.
3. ResultsandDiscussion
3.1. PerformanceofUBAFReactor
3.1.1. TheEffectofAerationRatesofCODRemoval
Figure2showsthelevelsofCOD,NH +-N,NO −-N,andTPremovalatvariousaerationrates
4 3
intheUBAFreactor. InFigure2a,theaverageconcentrationofCODininfluentandeffluentwere
257.47±17.99mg/Land23.38 ± 9.34mg/L,respectively. TheeffluentconcentrationofCODwas
50 mg/L based on the China standard limit. The COD removal efficiency did not show apparent
fluctuations (R=0.169, p = 0.464) with the increase in aeration rates (from 40 L/h to 90 L/h).
TheaverageremovalefficienciesofCODataerationratesof40L/h,65L/hand90L/hwere89.49%,
91.86%,and90.61%,respectively. ThesmalldifferenceinCODremovalefficiencycouldbeinterpreted
simplybydifferentdegreeofdenitrificationwhichconsumesorganiccarbon. Theresultwasconsistent
withpreviousresearch[3,5,8].Taoetal.[5]foundthatCODremovalefficiencyreachedto89.38±1.04%
Water2018,10,1244 5of13
inthereactor. Hasanetal.[3,9]showedtheinsignificanteffectofdifferentaerationrates(Q≥3L/h).
Thus,theUBAFreactordemonstratedagoodtreatmentefficiencyonCODremoval.
Water 2018, 10, x FOR PEER REVIEW 5 of 13

(a) COD removal

(b) NH4+-N removal
(c) NO3−-N removal

(d) TP removal
FigureFi2g.uErfef e2c. tEofffeacte roaf taieornatriaotne sraotenst ohne trheem roemvaolveaflfi ecfifeicniecnycoyf ocfh cehmemiciaclalo oxxyyggeenn ddeemmaanndd ((CCOODD)), ,NNHH4+-+-N,
4
NO −N-N, N,aOn3d−-Nto, taanldp thootaslp phhoorsupsh(oTruPs) .(TP).
3
Water2018,10,1244 6of13
3.1.2. TheEffectofAerationRatesofNitrogenRemoval
ThevaluesofNH +-NandNO −-Nrepresentthemaincomponentsofnitrogenandreflectthe
4 3
removalandtransformationofnitrogen. InFigure2b,c,theaerationrateshadanegativecorrelation
withtheNH +-N(R=−0.524,p=0.014)andNO −-N(R=−0.232,p=0.312)removal. Theremoval
4 3
efficienciesofNH +-NandNO −-Nreached55.48%and86.89%,respectively,atloweraerationrate
4 3
(40 L/h). The effluent concentrations of NH +-N and NO −-N were 18.03 mg/L and 1.59 mg/L,
4 3
respectively. However,whenweincreasedtheaerationrateto65L/h,theaverageremovalefficiencies
of NH +-N and NO −-N changed little. At an aeration rate of 90 L/h, the removal efficiencies of
4 3
NH +-NandNO −-Nreducedto46.48%and59.43%,respectively,witheffluentconcentrationsof
4 3
20.71mg/Land3.0mg/Lforthesetwocomponents. NH +-Nremovalpresentedlowervaluesthan
4
thoseobtainedfrompreviousstudies[3,5,9]. Thenitrogenremovalmightberelatedtothebiomass
loss. Becausethehighaerationratecouldresultfromthelowerdensityofmicrobesinthereactor
causedbythesludgerunoffintheBAFsystem. Intheotherhand,themicrobialcommunitystructure
determinedthenitrogenremovalefficiency.
3.1.3. TheEffectofAerationRatesofPhosphorusRemoval
Obviously, the aeration rates had a significantly positive correlation (R = 0.873, p = 2.39 × 10−7)
with TP removal efficiency in Figure 2d. The lower aeration rate removed 20.50% of TP. Under an
aerationrateof65L/h,theTPremovalsignificantlyincreasedandreached49.56±5.45%. Whenthe
aerationratesincreasedto90L/h,TheTPremovalincreased56.74±3.44%andwashigherthanatother
aerationrates. PhosphorusremovalmainlyrelatestothebiomassproductionintheUBAF.Thehigher
aerationratemightmakemorebiomasssplitawayofftheceramicsurfacesanddischargewitheffluent
water.Wangetal.[12]showedthatthetraditionalBAFonlyachievedthephosphorusremovalefficiency
of 9.3%. Thus, TP removal in our study was higher than in previous research [12,13]. In addition,
the TP concentrations in influent maintained at 6.50 ± 0.57 mg/L. The effluent concentrations were
5.61±0.62mg/L,3.22±0.42mg/L,and2.92 ± 0.51mg/Latdifferentaerationrates(from40L/hto
90 L/h), respectively. This TP concentration represented values in effluent higher than the standard
dischargevalue(0.5mg/L),asrequiredbytheEmissionStandardofPollutants,inChina. Phosphorus
hardlyachievesthisdischargestandardsbyonlyrelyingonthetraditionalBAFprocess[12].Theresult
wasconsistentwithpreviousresearch[12,17].Then,toimproveremovalofphosphorus,someresearchers
achievedpartialprecipitationofphosphorusbydosingtheFe2+orFe3+intotheBAF[17].Inthefuture,
we would further analyze the effect of aeration on the biomass suspension characteristics in order to
understandtheremovalmechanismofTP.
3.2. PollutantsRemovalatDifferentFilterMediaHeights
3.2.1. TheCODRemovalatDifferentFilterMediaHeights
Figure3showstheresultsofCODremovalatdifferentfiltermediaheights.TheCODremovalhad
asignificantlypositivecorrelation(R=0.910,p=1.60×10−7)withthefiltermediaheights. TheCOD
removalmainlyoccurredatthebottomoffiltermedialayer. CODremovalefficiencyat0–25cmheight
offiltermediareached81.9±3.60%,73.34±1.46%,and81.22±3.41%ataerationratesof40L/h,
65L/h,and90L/h,respectively. However,at25–85cmheightoffiltermedia,theremovalefficiency
onlyreachedlowervalues,7.59%,18.52%,and9.39%ataerationratesof40L/h,65L/h,and90L/h,
respectively. Thesechangesinremovalefficiencylikelyreflectedthedifferencesintheorganicmatter
concentrationatdifferentheights. Thelayerat25–85cmheightconcentratedlowerlevelsoforganic
matterwhencomparedwiththebottomoffiltermedialayer. Thehighorganicmatterconditionmight
leadtothemicroorganismabundanceincreasing,similartothefindingsinpreviousstudies[11,18].
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Figure3.CODremovalalongwiththeheightofthefiltermedialayer.
Figure 3. COD removal along with the height of the filter media layer.
Table1showsthedistributionofdissolvedoxygen(DO)concentrationsatdifferentfiltermedia
Table 1 shows the distribution of dissolved oxygen (DO) concentrations at different filter media
heights. DOconcentrationsininfluentandeffluentweresignificantlyhigherthoseatthefiltermedia
heights. DO concentrations in influent and effluent were significantly higher those at the filter media
layer, and distributed in the range of 8.0–9.21 mg/L. In addition, as the aeration rates increased,
layer, and distributed in the range of 8.0–9.21 mg/L. In addition, as the aeration rates increased, the
theDOconcentrationofthefiltermedialayerpresentedadecreasingthenincreasingtrend. TheDO
DO concentration of the filter media layer presented a decreasing then increasing trend. The DO
concentrationwasdeterminedbyoxygendiffusionfrombubblestowaterandoxygenconsumptionby
concentration was determined by oxygen diffusion from bubbles to water and oxygen consumption
microbesintheBAF.ThelowerDOconcentrationsatanaerationrateof65L/hcouldbeinterpretedby
by microbes in the BAF. The lower DO concentrations at an aeration rate of 65 L/h could be
thehigherabundanceandactivityofaerobicmicrobesunderthiscondition. Thus,theCODremovalat
interpreted by the higher abundance and activity of aerobic microbes under this condition. Thus, the
anaerationrateof65L/hwasdifferedfromtheaerationratesof40L/hand90L/h. Accordingtothe
COD removal at an aeration rate of 65 L/h was differed from the aeration rates of 40 L/h and 90 L/h.
resultsinSection3.1,wefoundthattheaerationratescouldachieveahigherpollutantsremoval.
According to the results in Section 3.1, we found that the aeration rates could achieve a higher
pollutants removal.
Table1.Thedistributionofdissolvedoxygen(DO)concentrationsatdifferentaerationratesandfilter
mediaheights(Unit:mg/L).
Table 1. The distribution of dissolved oxygen (DO) concentrations at different aeration rates and filter
mediaA heeriagthiotsn (RUantiet:s mg/ILn)fl. uent 10cm 25cm 40cm 55cm 70cm Effluent
40L/h 8.59 1.96 2.42 1.81 1.70 2.76 8.60
Aeration Rates Influent 10 cm 25 cm 40 cm 55 cm 70 cm Effluent
65L/h 8.67 0.95 1.35 1.13 0.90 1.71 8.56
40 L/h 8.59 1.96 2.42 1.81 1.70 2.76 8.60
90L/h 9.03 1.63 2.76 2.91 3.85 4.95 8.99
65 L/h 8.67 0.95 1.35 1.13 0.90 1.71 8.56
90 L/h 9.03 1.63 2.76 2.91 3.85 4.95 8.99
3.2.2. TheNitrogenRemovalatDifferentFilterMediaHeights
3.2.2.F Tighue rNei4trsohgoewn sRethmeorveaslu altts DoifffNerHen4t+ F-Niltearn Md eNdOia3 −H-eNigrhetms oval at different filter media heights.
NH +-Nremovalhadasignificantlypositivecorrelation(R=0.933,p=1.73×10−8)withthefilter
4
mediFaighueirgeh 4t sshinowFisg tuhree r4easu.lAtst o0f– N25Hc4m+-Nh eaingdh tNoOffi3−-ltNer rmemeodviaa,l tahte dfiiflfteerrernetm fioltveer dmmedoirae hNeiHgh+ts-N. NtHha4+n-
4
N removal had a significantly positive correlation (R = 0.933, p = 1.73 × 10−8) with the filter media
those observed under other filters. At 25–85 cm height of filter media, when filter media heights
hinecirgehatsse dinb Fyig1u5rcem 4,at.h Aetr a0t–e2o5f cNmH he+i-gNhtr eomf ofivltaelrr memeadiinae, dthceo nfislttearn trethmaotvweedr em5o.0r2e% N,H4.41+5-N% ,tahnadn 3th.5o6s%e
4
observed under other filters. At 25–85 cm height of filter media, when filter media heights increased
at different aeration rates, respectively. The results were mainly related to the supply of DO and
bthye 1d5i sctrmib, utthieo nraotfeo orgf aNnHic4m+-Nat treer.mHovoawl erveemr,atihneedN Ocon−s-tNanrte mthoatv awlesrheo w5.e0d2%a, s4ig.1n5i%fic, aanntldy 3n.e5g6a%ti vaet
3
different aeration rates, respectively. The results were mainly related to the supply of DO and the
correlation(R=−0.817,p=3.52×10−5)withthefiltermediaheightsanddifferedfromtheNH +-N
4
dreimstroivbaultiionnF iogfu roer4gba.nTich emraetmteorv. aHleofwficeiveenrc, ytohfe NNOO−3−--NN rreeamchoevdal9 4s.h85ow±e3d. 9a5 %siagtnfiiflitcearnmtleyd inaehgeaitgivhet
3
coofr0r–e1la0ticomn. (ARt =1 0−–02.851c7m, ph e=i g3h.5t2o ×f fi1l0te−5r) mweitdhi at,hNe Ofilt−er- Nmreedmiao vhaeligehffitsc iaenndcy ddififderneodt fcrhoamn gteh.eN NOH−4+--NN
3 3
rreemmoovvaall tinen Fdigtourdee 4cbre. aTsheea rte2m5–o8v5acl meffhiceiiegnhctyo offfi NlteOr3m−-Ned rieaa.cThheedN 94O.8−5 -±N 3.r9e5m%o vata lfioltcecru mrreeddiaas hreeisguhltt
3
of 0–10 cm. At 10–25cm height of filter media, NO3−-N removal efficiency did not change. NO3−-N
ofthebiologicaldenitrificationprocessintheUBAF.ThelowDOconcentrations(lessthan0.5mg/L)
removal tend to decrease at 25–85 cm height of filter media. The NO3−-N removal occurred as result
inducedtothedenitrificationprocess[4]. Table1alsoshowedthatDOconcentrationsatthebottomof
of the biological denitrification process in the UBAF. The low DO concentrations (less than 0.5 mg/L)
filtermediaremainedlowerthanthatobservedunderotherlayers. Therefore,theDOdistribution
induced to the denitrification process [4]. Table 1 also showed that DO concentrations at the bottom
likelyexplainedtheNO −-Nremoval. Inaddition,aerationratedidnotaffecttheremovalofnitrogen
3
of filter media remained lower than that observed under other layers. Therefore, the DO distribution
atdifferentfiltermediaheightssignificantly.Theaerationratesof40L/hand65L/hremovedNH +-N
4
likely explained the NO3−-N removal. In addition, aeration rate did not affect the removal of nitrogen
at different filter media heights significantly. The aeration rates of 40 L/h and 65 L/h removed NH4+-
N and NO3−-N more efficiently than the aeration rate of 90 L/h. Previous researches with BAF system
also reported similar results of NH4+-N and NO3−-N removal [19].
Water2018,10,1244 8of13
Waatenr d20N18O, 10,− x- FNORm PoErEeRe RffiEVciIeEnWtl y thantheaerationrateof90L/h. PreviousresearcheswithBAF8 soyf s1t3e m
3
alsoreportedsimilarresultsofNH +-NandNO −-Nremoval[19].
4 3
Water 2018, 10, x FOR PEER REVIEW 8 of 13

(a) NH4+-N removal (b) NO3--N removal
Figu(are) N4.H N4+i-tNro rgeemn orveaml oval along with the height of the f(iblt)e Nr mO3e--dNia r leamyoevr.a l
FiFgiugruere4 .4N. Nitirtoroggeennr reemmoovvaall aalloonngg wwiitthh tthhee hheeiigghhtt ooff tthhee fifiltleter rmmedediai alalyaeyre. r.
3.2.3 The Phosphorus Removal at Different Filter Media Heights
3.23.3.2..T3 hTeheP hPohsopsphhoorurussR Reemmoovvaalla att DDiiffffeerreenntt FFiilltteerr MMeeddiiaa HHeeigighhtsts
Figure 5 shows the TP removal at different filter media heights. The TP removal had a poor
correlatFioigFnui g(rRue r5=e 0s5h. 2so3hw1o,ws pst h= t eh0e.T3 PT57Pr) e rwmemiothov vatalhlea a tftid ldtiefiffrfe emrreeenndtti fiafi llhtteeerri gmmhetesdd. iiTaa hhheee iaiggmhhtotssu. .nTTth heo efT TTPP Pr errememmovooavvla alhl aohdsac dail lpaaotepodor or
as ctohrecr oefrillrateetlirao tmnioa(nRt e(Rr=i a0=l . 20in.32c13r,1ep,a p=s e=0d .0,3 .w3557h7)e)w rweiatihtsh, t tthhheee firfeilltmteerro mvmaeeldd eiifaaf ihhcieeeiignghchyttss .a. TtT hhheieg aham maoeouruantnti toonof f TrTPatP eresrm ermeoavocavhla eoldso chsiliclgailhtleaedtre d
vaalusetashs. e tThfihele tfe irlrteemsru amltte arrteeialrailatieln dicn rtceorae tsahesede d,bw,i wohmheeraerseasas ls,o,t sthhse.e Trreehmme oolovvwaall eeaffeffiriccaiiteeinoncncyy r aaattt ehh irgieghmh aoaeverareatdito ioonnn rlayrta e1tse0 rs.e9ra8ec ah±c eh9d.e5 hd3i%ghhi goehrf er
TPv raelvumaelosu.veTasl.h Tienhr eee sfrfuelulstuerlnett lr.a etTleahdteet dore ttdohu etchbteii oobnmio oamsf saTslPos slrsoe.smTs.h oTevhaleol wloocwaceu arraertreiadotn iaotrn a 2tr5ea–tre4e 5mr ecmomvoe vadnedod n o5lny5l–y180 51.90 c8.9m±8 h±9 e9.i5.g53h3%%tso oofffT P
filtreerm TmoPv eradelmiain.o vWeaffllh iuenne en fttfh.lueT ehaneet.rr aeTtdhiouen crt eirdoauntecot ifionTncP roerfa eTsmePdo r vetamol oo6v5cac lLu o/rhrcec, duTraPret d2re 5am–t 4o255v–cam4l5 s acignmnd iaf5ni5cd–a 8n555tl–yc8m 5i mchmpeir ghohevitegsdho tfsb fiuolftt er
redmuecfdielidtae .ra Wtm 2he5ed–ni4a0t. h cWemah eehrnea ittgihohent aoreafr tfaeitliitnoencrr meraaetsede diian.tc oUre6na5dsLeed/r hato,nT 6aP5e rrLaemt/hioo, nvT arPal tsreieg monfoi fi9v0caa lL ns/tihlgy,n TiimfPic parrneomtvlyoe dvimablpu crtoornvetedidnu ucbeeuddt at
to 2in5c–rr4eed0auscceme. dTh haeeti g 2rhe5mt–4oo0fv ficamllt ee hrffemiicgiehednt icoayf. fUeislntseedrn emtriaeadlnliyaa .re eUrmantadioiennre ardan tu eaneocrfhat9ai0nongL e/rdaht ae,t To 5Pf5 9-r80e5 mL c/omhv, aThlPeci ogrenhmtti oonfvu faeilld tceotron mtiinnecdureieada. se.
ThTeh hetoigr iehnmecsrotev araselem.e Toffihvceai ler enemfcfyiocveiesasnl eecnfyft iircaeilealnycchryee dmes 5ase6inn.7et4ida ±lul y3n .rc4eh4ma%na.gi nTeehddea suten5 rc5eh-sa8un5lgtcsem dr eahvte e5iag5l-he8td5o ctfhmfia lhtt eethirgemh hte iodgfih afie.lrtTe ahre merahetidigoihan.e st
ratree mpTrohovem ahloiegtfehfidec siTte Pnrec mryeomrevaoacvl heaeflf,d iwc5ie6hn.i7cc4yh ± rseua3gc.4gh4ee%dst .e5Td6.h 7teh4s a±et 3rte.h4s4eu% lhts.i gTrhehv eaeseear lraeetdsiouthnlta srt artetheve emhaliieggdhh etth rraaeted truhaecti eho ntighrheae tmre aipcerrrooamtbioioantle d
rate promoted TP removal, which suggested that the high aeration rate might reduce the microbial
bioTmParsesm loovsas.l ,Twhheircehfosrueg, gweset efdurtthhaetrt hinevheisgthigaaeterdat itohne radtievemrsigithyt arenddu ceevethnenemssi croofb itahleb imomicraosbsialol ss.
biomass loss. Therefore, we further investigated the diversity and evenness of the microbial
comThmeruenfoitrye., w efurtherinvestigatedthediversityandevennessofthemicrobialcommunity.
community.

FiFgiugruere5 .5T. TPPr eremmoovvaalla alloonngg wwiitthh tthhee hheeiigghhtt ooff tthhee ffiiltleterr mmeeddiai alalyaeyre. r.
Figure 5. TP removal along with the height of the filter media layer.
3.3. DiversityandEvennessoftheMicrobialCommunity
3.3. Diversity and Evenness of the Microbial Community
3.3. Diversity and Evenness of the Microbial Community
ThediversityofthemicrobesforeachsamplearelistedinTable2. Librarieswithsampleswere
The diversity of the microbes for each sample are listed in Table 2. Libraries with samples were
comThpeo dseivdeorsfit4y3 3of( 4th0eL m/ihc)r,o4b3e0s f(o6r5 eLa/chh )s,a4m14pl(e9 a0rLe /lihst)e,d4 1in3 T(1a0blcem 2)., L4i7b6ra(r4ie0sc wmi)t,ha snadm4p5le8s (w70ercem ).
composed of 433 (40 L/h), 430 (65 L/h), 414 (90 L/h), 413 (10 cm), 476 (40 cm), and 458 (70 cm). The
coTmhpeohseigdh oerf C43h3a o(410a Lnd/hA), C4E30i n(6d5e xLi/nhd),i c4a1t4e s(9a0r iLc/hh)m, i4c1r3o b(1ia0l ccmom),m 47u6n i(t4y0[ 2cm0].),I nanadd d4i5t8io (n7,0t hcemS)i. mTphseo n
higher Chao1 and ACE index indicates a rich microbial community [20]. In addition, the Simpson
higahnedarn SdCh hSaahnoan1no nnaoninnd id nAedxCeixEn ifnionfrodmremxed eidnb dboiotchtahtr eircsich han nereiscsssh a anmnddic eervvoeebnniannlee csssos mooffm mmuiicncrriotobybi ai[a2l l0cco].om mInmm auudnnditiyitt yi[o2[n12],1 . t]Lh.aeLr agSreigmr eipnrsdinoendx ex
anvda Slvuhaealsuniennsod niinc iadntidecdeaxthe iding fhhoeirrgmheveerde n benvoeetshns nroeifcshms nioecfrs osm bainiacdlroc eobvmiaelmn ncueonsmist myo.fuT mnhiietcytr.o otTbaihlaenl uctomomtbalme rnsuunomfitoybp e[er2sr1 a]to. ifLo naorapgleetrara xitoniondnoeamxl ic
vaulunetisats xio(nOndoTimcUaistce) uden sithtismi g(haOteTerdU esb)vy eesntthnimeesCasth edaoof b 1ym etihscterio mCbhaiaatloo r c1wo emestrmiemu9an1to0itry( 4w. 0eTLrhe/ e9h 1)t0,o 9t(a460l2 Ln(6/uh5m)L, b9/e6hr2)s ,( 6ao5nf Ldo/h8p9)e,5 raa(n9tdi0o 8nL9a/5lh ),
taxon(o90m iLc/ hu)n, iatsn d(O tThUe sA) CesEt imesatitmeda tboyr wtheer eC h1a34o0 1.0 e1s t(i4m0 aLt/ohr) ,w 1e3r4e1 9.4190 ((6450 LL//hh)),, a9n62d (16257 L7./4h3) , (a9n0 dL 8/h9)5,
(90 Li/nhd)i,c aatnindg ththea tA tCheE aeesrtaitmioant orra twe eorfe 6 51 3L4/0h.0 e1x h(4ib0i tLed/h )m, o1r3e4 1th.4e9 ri(c6h5n eLs/sh )o, f amndic r1o2b7ia7l. 4c3o m(9m0 uLn/ihty),.
indicating that the aeration rate of 65 L/h exhibited more the richness of microbial community.
Water2018,10,1244 9of13
and the ACE estimator were 1340.01 (40 L/h), 1341.49 (65 L/h), and 1277.43 (90 L/h), indicating
thattheaerationrateof65L/hexhibitedmoretherichnessofmicrobialcommunity. Accordingto
theShannonindex, themicrobialcommunitypresenteditshighestevennessatanaerationrateof
65L/h. Theresultverifiedtherelevantinferencesofnitrogenandphosphorusremovalthatwere
relatedtothebiomasslossinhighaerationrate. Accordingtopreviousresults, wefoundthatthe
richnessofmicrobialcommunityatanaerationrateof65L/hmightcontrolremovalefficiencyof
pollutants. Inaddition,diversityandevennessofmicrobialcommunityatdifferentfiltermediaheights
alsodifferedsignificantlydifferentatdifferentaerationrates. Ourfindingsalsoshowedthatat70cm
heightoffiltermediathemicrobialdiversityreacheditshighestevenness,likelyaconsequenceofthe
higherdissolvedoxygenatthesurfaceoffiltermedia.
Table2.Therichnessanddiversityof16SrRNAsequencesatdifferentconditions.
Sample OTUs Simpson Chao1 ACE Shannon
40L/h 433 0.9819 910 1340.01 6.64
65L/h 430 0.9775 962 1341.49 7.21
90L/h 414 0.9580 895 1277.43 6.53
10cm 413 0.9530 943 1362.56 6.72
40cm 476 0.9646 825 1166.10 6.85
70cm 458 0.9709 1032 1417.76 7.17
3.4. ComparativeAnalysisofMicrobialCommunity
3.4.1. TheEffectofAerationRatesonMicrobialCommunity
Figure6showstherelativeabundanceofmicrobialcommunityatthreetaxonomiclevels(phylum,
class, and genus) at different aeration rates. Proteobacteria corresponded to the most abundant
with abundance ranging between 42.2–50.4%, followed by Actinobacteria (14.5–20.5%), Firmicutes
(12.7–17.6%),Bacteroidetes(10.0–15.3%),andPlanctomycetes(3.3–4.4%)inFigure6a. Theaerationrates
hadaneffectinthemicrobialcommunitystructure,indicatingamicrobialcommunitystructuremore
resistanttochanges. Ourstudyoperatedfor15daysatthesameaerationrate,whichmightexplain
thesimilardistributionofmicroorganisms. TherelativeabundanceoftheProteobacteriaandFirmicutes
increased following the changes in the aeration rates, likely compensated by the reduction of the
Planctomycetesrelativeabundance. PreviousresearchfoundthatspeciesofthePlanctomycetesperform
theanammoxactivity,whichcouldconvertdirectlyNH +-NintoN [22]. However,thesebacteria
4 2
weremoresensitivetothechangingofdissolvedoxygen. Underanaerationrateof65L/h,therelative
abundance of Nitrospirae was 3.9%, higher abundance for this group. The presence of Nitrospirae
indicatedtheexistenceofnitriteoxidizingbacteria(NOB)astheycanoxidizenitritetonitrate[23].
TheProteobacteriarepresentsadominantphylum,whichcanremovephosphorusandconvert
nitrite to nitrate in the wastewater biological treatment [5,16]. Therefore, we performed a further
analysis of the Proteobacteria group. Figure 6b shows the five classes of Proteobacteria at different
aerationratesandfiltermediaheights. Betaproteobacteriapredominatedatdifferentaerationratesand
filtermediaheights. TheresultswereconsistentwithpreviousresearchperformedbyWangetal.[21]
that reported Betaproteobacteria as the main supplier of protons and electrons during the growth
and metabolism of phase. The aeration rates produced no significant effects on the microbial
communitystructure.
Water2018,10,1244 10of13
Water 2018, 10, x FOR PEER REVIEW 10 of 13

(a) Phylum level (b) Five classes of Proteobacteria
FigurFeig6u.rMe 6ic. rMobiciraolbciaolm commumnuitnyityco cmomppoosistiitoionn aatt tthhee pphhyylluumm aannd dclcalsass: s(a:)( aP)hyPlhuymlu lemvelle avte dliafftedreinffte rent
aeration rates; (b) five classes of Proteobacteria at different aeration rates.
aerationrates;(b)fiveclassesofProteobacteriaatdifferentaerationrates.
3.4.2.3T.4h.2e. EThffee cEtffoefctF oilft FeirltMer eMdeiadiHa eHigeihgthstso onnM Miiccrroobbiiaall CCoommmmuunnitiyt y
Figure 7 shows the microbial community composition at the phylum, class, and genus level at
Figure7showsthemicrobialcommunitycompositionatthephylum,class,andgenuslevelat
different filter media heights. The Proteobacteria represents a dominant phylum. The differences of
differentfiltermediaheights. TheProteobacteriarepresentsadominantphylum. Thedifferencesof
microbial community distribution at different filter media heights were more obvious than at
microbialcommunitydistributionatdifferentfiltermediaheightsweremoreobviousthanatdifferent
different aeration rates. The relative abundance of Proteobacteria (49.6%), Firmicutes (23.1%), and
aerationrates. TherelativeabundanceofProteobacteria(49.6%),Firmicutes(23.1%),andPlanctomycetes
Planctomycetes (11.0%) peaked at 10 cm height of filter media. Moreover, in the COD test, we observed
(11.0%th)apt ethaek oedrgaatni1c0 mcamttehre riegmhtoovaflfi olctecurrmreedd miaa.iMnlyo raet o0v–2e5r, cimn thheeigChOt oDf ftieltsetr, mweedoiab. sTehrev eodrgtahnaict tmhaettoerrg anic
mattedrisrterimbuotvioanl oimccpurorrveedd mthea ignrloywatth 0a–n2d5 mcmetahbeoilgishmt ooff mfilitceroromrgeadniiasm. Ts.h Aecocorgrdainnigc tmo athttee sredctiisotnr ibofu tion
impro3v.2e.2d, twhee fgoruonwdt thhaatn nditmroegteanb omliasimnlyo rfemmiocvroedo ragta tnhies mbost.toAmc coof rfdilitnerg mtoedthiae. STehcet iroensu3lt.s2 .s2h,owweefdo tuhnadt that
nitrogPelanncmtoaminyclyeterse mcoouvlde dbea trethspeobnositbtolem foorf tfihlet enritmroegdenia .loTshs eanreds uenlthsasnhcoe wtheed ntihtraotgPenla nrecmtoomvyacl.e tTeshec ould
microbial community is the primary responsible for the COD and nitrogen removal mechanism. The
beresponsibleforthenitrogenlossandenhancethenitrogenremoval. Themicrobialcommunityisthe
Bacteroidetes and Chloroflexi abundance peaked at 40 cm height of filter media. In addition, the relative
primaryresponsiblefortheCODandnitrogenremovalmechanism. TheBacteroidetesandChloroflexi
abundance of Nitrospirae (4.7%) peaked at 70 cm height of filter media. The result explained why
abundancepeakedat40cmheightoffiltermedia. Inaddition,therelativeabundanceofNitrospirae
(4.7%r)epmeoavkaeld efaftic7ie0nccmy ohfe NigOh3t−-oNf fishltoewremde ad idae.cTrehaesirnegs utrletnedx palsa tihnee dfilwtehr ymreedmiao hveailgehftfis ciniecnrecayseodf N(seOe −-N
Section 3.2.2). 3
showedadecreasingtrendasthefiltermediaheightsincreased(seeSection3.2.2).
Figure 7b shows the five classes of Proteobacteria at different filter media heights.
FBiegtauprreot7ebobsahctoerwias ptrheedofimveincaltaesds east doifffPerroetneto fbialtcetre rmiaedatiad hifefiegrhetns.t Tfihlete rresmueltds iwaehreei cgohntssi.sBteentta pwriotthe othbaec teria
predormesuinlta toef ddiaftfedreifnfte raeenrattfiiolnte rramtese.d Tiaheh reeilgahtitvse. Tabhuenrdeasnuclet sowf tehree Acolpnhsaipsrtoetneotbwactiethriat hpeearkeesdu latto 1f0d cimffe rent
aeratihoenighrat toefs .filTtehre mreedlaiati.v Tehaeb ruenladtiavne caeboufndthaneceA lopfh aAplprohtaeporbotaecotbearcitaerpiae adkeecrdeaasted1 0asc mtheh ediigsshotlvoefd filter
medioax.yTghene irneclraetaivseeda (binu nTdabalne c1e), owfhAiclhp hsaupgrgoetsetoebda ctthearti athdee dcirsesaoslveedda osxtyhgeend cisosnoclevnetrdatoioxny gperonminotcerde ased
bacterial growth. Figure 7c–e shows the relative abundance of five classes of Betaproteobacteria at
(in Table 1), which suggested that the dissolved oxygen concentration promoted bacterial growth.
different filter media heights. The relative abundance of Comamonadaceae and Dechloromonas at 10 cm,
Figure7c–eshowstherelativeabundanceoffiveclassesofBetaproteobacteriaatdifferentfiltermedia
40 cm, and 70 cm heights of filter media responded to 48.93%, 33.73%, and 39.02% of the total
heights. TherelativeabundanceofComamonadaceaeandDechloromonasat10cm,40cm,and70cm
microbial abundance, respectively. These bacterial groups represented the dominant denitrifying
heightsoffiltermediarespondedto48.93%, 33.73%, and39.02%ofthetotalmicrobialabundance,
bacteria found in our study [24]. The results explained why nitrogen presented a high removal at the
respectively. These bacterial groups represented the dominant denitrifying bacteria found in our
bottom of filter media. In addition, the relative abundance of Zooglea at different filter media height
studyw[e2r4e ]2.7.T9h0%e,r 3e6s.u6l9t%s,e axnpdla 3in7.e2d8%w, hreyspneicttrivoegleyn. Tphree sbeanctteerdia amhigighht bree mmoorvea lseantsitthivee btoo ttthoem hiogfh filter
medidai.ssIonlvaeddd oitxiyogne,nt. hNeortaeblalyti,v theea rbeulantidvae nacbeunodfaZnoceo golfe aDeacthdloirfofmeroennast afit ldteifrfemreendt ifailthere imgehdtiaw heeriegh2t7s. 90%,
36.69%we,raen 0d.8367%.2, 86%.5,1%re,s panedct i4v.5e3ly%.,T fhroemb a1c0t ecrmia tmo i7g0h ctmbe, rmesopreectsievnelsyi.t iTvheet obathcteerhiaig whedreis snooltv oendlyo xay gen.
Notabdleyn,ittrhiefyrinelga tbiuvte aalbsou nphdoasnpcheaotef-aDccecuhmlourloamtinong amsiactrodoirffgearneinsmtfis.l tTehrem reesduilat hise sigtrhicttslyw reerlaete0d.8 6to% t,h6e. 51%,
distribution of dissolved oxygen and pollutants. Our result showed that the microbial community
and 4.53%, from 10 cm to 70 cm, respectively. The bacteria were not only a denitrifying but also
structure explained the removal mechanism of COD, nitrogen, and phosphorus. However, it remains
phosphate-accumulatingmicroorganisms. Theresultisstrictlyrelatedtothedistributionofdissolved
unclear which specific microorganisms played a role in the nitrogen and phosphorus removal.
oxygen and pollutants. Our result showed that the microbial community structure explained the
Therefore, further studies are necessary to explore the specific functional microbes of nitrogen and
removalmechanismofCOD,nitrogen,andphosphorus. However,itremainsunclearwhichspecific
phosphorus in the UBAF system.
microorganismsplayedaroleinthenitrogenandphosphorusremoval. Therefore,furtherstudiesare

necessarytoexplorethespecificfunctionalmicrobesofnitrogenandphosphorusintheUBAFsystem.