馬瑞婕,劉永紅*,梁繼東,王 寧,解鳳霞

基于顆粒污泥的短程反硝化USB反應(yīng)器啟動和運行研究

馬瑞婕1,劉永紅1*,梁繼東2,王 寧1,解鳳霞1

(1.西安工程大學(xué)環(huán)境與化學(xué)工程學(xué)院,陜西 西安 710048；2.西安交通大學(xué)能源與動力工程學(xué)院,陜西 西安 710049)

在2個相同的USB反應(yīng)器(R1無載體,R2采用多孔生物填料為載體)中構(gòu)建了短程反硝化工藝,對R1和R2NO3--N→NO2--N轉(zhuǎn)化性能、短程反硝化顆粒污泥物化特性、胞外聚合物(EPS)產(chǎn)生特性以及微生物功能菌群主要特征進行差異分析.結(jié)果表明,反應(yīng)器運行81d,氮負(fù)荷(NLR)為1.2kg/(m3·d)時, NO3--N→NO2--N轉(zhuǎn)化率(NTR)R2(85%)高于R1(80%);載體顆粒污泥(R2)沉降性能優(yōu)于自固定化顆粒污泥(R1)且載體顆粒污泥(R2)更容易截留EPS,PN/PS值R1(1.29)>R2(1.15),污泥體積指數(shù)(SVI)R1(27.07mL/g MLSS)>R2(19.36mL/g MLSS);掃描電鏡發(fā)現(xiàn)R1污泥表面聚集長桿菌,R2污泥表面聚集短桿菌和球菌,與R1相比R2顆粒污泥結(jié)構(gòu)更加規(guī)則密實.微生物高通量測序結(jié)果表明,R2物種豐富度和多樣性高于R1,變形菌門、擬桿菌門和綠彎菌門在短程反硝化系統(tǒng)中占主導(dǎo)地位,R1和R2主要NO2--N積累功能菌屬均為屬(R1-59.18%、R2-46.04%)和屬(R1-6.81%、R2-5.99%).

短程反硝化(PDN)；厭氧氨氧化(Anammox)；亞硝酸鹽積累；PDN顆粒污泥；生物載體

隨著污水中氮素污染物增加,實現(xiàn)污水高效脫氮和達(dá)標(biāo)排放是當(dāng)前污水處理鄰域研究重點之一.厭氧氨氧化(Anammox)工藝因無需外加碳源、污泥產(chǎn)量低、無需供氧,成為一種最經(jīng)濟有效的新型生物脫氮技術(shù)[1].然而,實際污水中Anammox電子受體(NO2--N)的匱乏使其工程應(yīng)用存在局限性[2].短程反硝化(PDN)是將NO3--N還原至NO2--N的不完全生物反硝化過程[3],被認(rèn)為是與Anammox耦合最具前景的NO2--N供給工藝[4].

微生物自固定化形成的顆粒污泥具有良好的沉降性能并能持留較高的生物量,可以滿足高負(fù)荷生物反應(yīng)器的運行需求[5].已有研究[6-7]在SBR反應(yīng)器中以PDN污泥為種泥成功培養(yǎng)出PDN顆粒污泥,實現(xiàn)了較高的NO3--N→NO2--N轉(zhuǎn)化率(NTR).然而污泥自固定化是一個漫長的過程,因此PDN顆粒污泥的快速培養(yǎng)技術(shù)是PDN工藝工程化應(yīng)用的關(guān)鍵技術(shù)之一[8].

相關(guān)研究指出,高上升流速的USB反應(yīng)器有利于顆粒污泥快速培養(yǎng)[9].此外,添加生物載體可加速污泥顆?；约俺至艄δ芫?目前通過添加載體促進厭氧、好氧污泥顆?；难芯枯^多[9-10],但在主流低氮素濃度污水條件下,添加載體促進PDN污泥顆?；难芯枯^少.

因此,本研究在連續(xù)流態(tài)下,分別構(gòu)建有無生物載體的PDN工藝,對比探究PDN工藝啟動及穩(wěn)定運行過程NO2--N積累性能,考察自固定化顆粒污泥和載體顆粒污泥培養(yǎng)過程中污泥物化特性差異,并基于高通量測序技術(shù)分析微生物菌群特征和功能菌屬富集機制,旨在為顆粒污泥技術(shù)促進PDN工藝穩(wěn)定運行及工程化應(yīng)用提供借鑒.

1 材料與方法

1.1 實驗裝置與運行

采用2個相同的有機玻璃材質(zhì)、雙層夾套結(jié)構(gòu)USB反應(yīng)器(R1和R2).反應(yīng)器有效容積為2.6L(內(nèi)徑6cm,高60cm),原水通過蠕動泵連續(xù)進入反應(yīng)器底部.反應(yīng)器設(shè)回流裝置,從下至上平均設(shè)3個取樣口,水浴維持反應(yīng)器溫度為25℃.

本實驗分為2個階段,階段Ⅰ為PDN啟動階段(1～41d),階段Ⅱ為負(fù)荷提升階段(42～81d).R1和R2反應(yīng)器不同階段運行參數(shù)一致,如表1所示.

表1 R1和R2運行參數(shù)

1.2 進水水質(zhì)

進水水質(zhì)采用人工配水,NO3--N和COD由硝酸鈉(NaNO3)和無水乙酸鈉(CH3COONa)按比例配制,礦物質(zhì)組成如下:0.15g/L MgSO4·7H2O, 0.14g/L CaCl2, 0.03g/L KH2PO4, 1.00mL/L微量元素Ⅰ和Ⅱ,微量元素Ⅰ的成分:EDTA·2Na 6.39g/L、FeSO45.00g/L;微量元素Ⅱ的成分:EDTA·2Na 19.11g/L、H3BO40.014g/L、MnCl2·4H2O 0.99g/L、CuSO4·5H2O 0.25g/L、ZnSO4·7H2O 0.43g/L、NiCl2· 6H2O 0.19g/L、CoCl2·6H2O 0.24g/L、NaMoO4·2H2O 0.22g/L[11].采用1mol/L NaOH溶液調(diào)節(jié)進水pH值.

1.3 多孔生物填料及接種污泥

本實驗采用的親水性多孔生物填料(HPBC)是一種具有多孔結(jié)構(gòu)的有機高分子-無機復(fù)合材料,通體呈鱗片狀及通孔狀的親水結(jié)構(gòu),大小為4mm左右,內(nèi)部孔隙大小為5～10μm,比重略大于水[12].該載體結(jié)構(gòu)與顆粒污泥較為相似,有利于微生物在其表面附著生長.

R1和R2接種泥源來源于西安某高校污水站沉淀池污泥,呈黃棕色絮狀.污泥混合液懸浮固體質(zhì)量濃度(MLSS)為11.26g/L,混合液揮發(fā)性懸浮固體質(zhì)量濃度(MLVSS)為4.47g/L.R1接種1.0L污泥;R2接種1.0L污泥并投入30%生物填料.

1.4 分析項目及檢測方法

水樣經(jīng)0.45μm濾紙過濾后,COD采用快速消解分光光度儀[連華5B-3 (C)]測定,COD數(shù)據(jù)為校正值,NO2--N對COD測定理論貢獻(xiàn)為1.14g/g(以COD/NO2--N計)[13];NO2--N和NO3--N分別采用N-(1-萘基) -乙二胺比色法和紫外分光光度法; pH值采用PHS-3C型pH計直接測定法.采用熱提取法[14]分層提取EPS,分別采用考馬斯亮藍(lán)(G-250)法[15]和蒽酮分光光度法[10]測定蛋白質(zhì)(PN)和多糖(PS)含量.

1.5 微生物高通量測序分析

采集2個反應(yīng)器運行第81d底端的少量污泥樣品 (對應(yīng)編號分別為R1和R2),由上海生工公司進行測序,測序方法見文獻(xiàn)[6].

1.6 NTR計算方法

式中:NO2--Neff和NO2--Ninf分別為USB反應(yīng)器出水和進水NO2--N濃度,mg/L;NO3--Neff和NO3--Ninf分別為USB反應(yīng)器出水和進水NO3--N濃度,mg/L[16].

2 結(jié)果與討論

2.1 PDN系統(tǒng)啟動及運行特性

維持進水NO3--N濃度為50mg/L,R1和R2反應(yīng)器PDN啟動運行特性如圖1所示.

階段Ⅰ-1(1～13d),以COD/NO3--N=3.0啟動R1和R2反應(yīng)器,啟動初期,由于接種原泥中PDN功能菌活性較低,R1出水NO2--N和NO3--N濃度分別為3.00mg/L和23.27mg/L,COD去除率為65%,NTR僅為11.22%; R2出水NO2--N和NO3--N濃度分別為1.40mg/L和7.61mg/L,COD去除率為65%,NTR僅為3.3%.隨著PDN污泥的馴化, R1和R2PDN活性均逐步上升,NTR不斷增加.第7d后,由于系統(tǒng)碳源充足R1和R2全程反硝化菌利用有機物將NO2--N進一步還原為N2,R1和R2NTR均開始降低,R1NTR從54.65%(第7d)下降至24.56%(第13d), R2NTR從58.46%(第7d)下降至5.96%(第13d).

階段Ⅰ-2(14～41d),以COD/NO3--N=2.0運行反應(yīng)器,R1和R2NO2--N積累量從第13d的8.0mg/L和1.8mg/L分別提高到第23d的20.3mg/L和20.9mg/L.表明低COD/NO3--N比使PDN功能菌占據(jù)優(yōu)勢地位,抑制NO2--N還原菌屬增殖[17].第24d進水pH值提高為9,導(dǎo)致NO2--N積累量增加.高pH值會降低有機物的氧化速率, 導(dǎo)致反硝化酶之間競爭電子[18],且高pH值抑制NO2--N還原酶活性,而對NO3--N還原酶活性沒有顯著影響,有利于選擇富集PDN功能菌[19-20].相關(guān)研究認(rèn)為反應(yīng)器出水NTR = 70%是PDN啟動的一個標(biāo)志[16],R1NTR在第41d達(dá)到74.71%, R2NTR在第39d達(dá)到74.50%,說明R1和R2均已成功啟動PDN.

階段Ⅱ-1(42～54d),R1和R2水力停留時間(HRT)由3h降為2h,氮負(fù)荷(NLR)達(dá)到0.6kg/(m3·d), R1和R2平均NTR分別為75.30%、78.57%.階段Ⅱ-2(55～74d),R1和R2HRT降為1h,NLR達(dá)到1.2kg/ (m3·d),R1和R2NO2--N積累量均先下降后上升.階段Ⅱ-3(75～81d),為進一步提高NTR和NO2--N積累量,以COD/NO3--N=3.0運行反應(yīng)器,R1和R2的NTR分別增加為84.46%、89.31%,NO2--N積累量分別為29.3mg/L、29.4mg/L.R1 和R2NTR分別穩(wěn)定維持在80%、85%以上,與R1自固定化顆粒污泥相比,R2載體顆粒污泥在PDN啟動運行性能方面具有優(yōu)勢,R2內(nèi)添加親水性多孔生物載體有利于微生物附著并維持系統(tǒng)穩(wěn)定運行.

2.2 PDN系統(tǒng)污泥特性分析

2.2.1 污泥表觀形態(tài)與微生物相觀察 R1和R2運行過程污泥表觀形態(tài)變化如圖2所示.接種的普通絮狀活性污泥(圖2a,d),經(jīng)過38d的培養(yǎng), R1內(nèi)污泥表現(xiàn)出顆粒特征但小顆粒數(shù)量較多(圖2b);R2內(nèi)絮狀污泥附著在親水性多孔生物載體表面,呈黃棕色顆粒(圖2e).隨著反應(yīng)器運行,顆粒污泥粒徑增大,第81d觀察到R1自固定化顆粒污泥結(jié)構(gòu)較松散(圖2c);R2載體顆粒污泥結(jié)構(gòu)更加規(guī)則密實(圖2f).

通過SEM進一步觀察污泥的微觀結(jié)構(gòu),R1和R2運行第81d反應(yīng)器底端污泥SEM圖如圖3所示.R1和R2顆粒污泥培養(yǎng)完成后,R1自固定化顆粒污泥表面桿狀微生物占優(yōu)勢地位(圖3a,b);R2 載體顆粒污泥表面聚集大量短桿菌和球菌(圖3c,d).R1和R2微生物周圍均含有大量的黏性物質(zhì),其可能是細(xì)菌分泌的胞外聚合物(EPS),這些物質(zhì)將細(xì)菌互相黏連在一起,促進PDN污泥顆?；?與自固定化顆粒污泥(R1)相比,載體顆粒污泥(R2)表面微生物排列更加緊密.

圖3 R1和R2第81d 污泥 SEM觀察

2.2.2 污泥濃度與沉降性能分析 由圖4可知, R1和R2的MLSS在運行過程中均先緩慢增長(0～ 40d)后迅速增加(41～80d),第80d,R1和R2的MLSS分別為25.86g/L、41.33g/L.由于接種的污泥活性較低,前期增長緩慢,污泥經(jīng)培養(yǎng)馴化后活性提高,在R2中采用多孔生物填料為載體有助于微生物聚集生長.R1的MLVSS在運行過程中緩慢增長,最大值為7.69g/L.R2的MLVSS初期增長較慢,末期(60～ 80d)迅速增長,最大值達(dá)到15.87g/L.在運行末期, R2的MLSS和MLVSS均大于R1,表明載體顆粒污泥(R2)較自固定化顆粒污泥(R1)更利于生物量的聚集.

R1和R2MLVSS/MLSS比值均先升高后降低趨于穩(wěn)定.在運行末期,R1和R2 MLVSS/MLSS比值分別從最高值53.77%、62.04%下降到29.75%、38.39%.R1和R2污泥中非揮發(fā)性物質(zhì)增加,由于進水中的金屬離子(Ca2+、Mg2+等)在PDN過程形成無機鹽,無機鹽在顆粒污泥內(nèi)部沉淀導(dǎo)致顆粒內(nèi)惰性組分增加[21].已有研究報道,非揮發(fā)性物質(zhì)的積累可促進污泥顆?；痆22].

污泥體積指數(shù)(SVI)能較好地反映出活性污泥的松散程度和凝聚沉降性能,R1和R2運行過程SVI變化如表2所示.

表2 R1和R2 SVI變化

R1和R2SVI小于普通良好的活性污泥SVI (50～120mL/g MLSS之間)[11],表明在R1和R2內(nèi)培養(yǎng)的PDN顆粒污泥均具有良好的沉降性能.在負(fù)荷提升階段(42～81d),R2SVI小于R1SVI,載體顆粒污泥沉降性能優(yōu)于自固定化顆粒污泥.

2.2.3 污泥中EPS含量變化 EPS能夠促進細(xì)胞聚集且維持穩(wěn)定,主要由(PN)和(PS)組成,EPS與顆粒污泥的形成密切相關(guān)[23].分層提取溶解型EPS (S-EPS)、松散附著型EPS(L-EPS)和緊密粘附型EPS(T-EPS),R1和R2顆粒污泥EPS含量變化如圖5所示.

由圖5可知,在階段Ⅰ(1～41d),R2EPS含量高于R1,表明多孔生物載體顆粒污泥更容易截留EPS, EPS有助于微生物聚集生長以及污泥顆?；?使R2污泥顆?；潭容^R1明顯. 在階段Ⅱ(42～81d),由于反應(yīng)器運行后期顆粒污泥成熟導(dǎo)致微生物分泌EPS含量減少且異養(yǎng)菌利用部分EPS作為有機碳源進行內(nèi)源反硝化, R1和R2EPS含量均逐漸降低且R2EPS含量低于R1.第80d,R1和R2EPS含量分別降低至24.10 和17.57mg/gVSS.在整個運行過程中EPS含量T層最高,表明T-EPS對PDN顆粒污泥的穩(wěn)定性具有重要作用,這與大多數(shù)研究結(jié)果一致[24].

此外,在階段Ⅰ(1～41d),R2EPS 中PN和PS含量均高于R1,研究表明,PN具有較高黏結(jié)強度且有助于微生物維持自身生長和代謝活性[25].粘性和親水性的 PS能夠促使生物群體通過橋聯(lián)作用形成交叉的網(wǎng)狀結(jié)構(gòu),從而促進污泥形成顆粒狀結(jié)構(gòu)[26]. PN/PS一般作為評價污泥沉降性能的重要指標(biāo),相關(guān)研究報道顆粒污泥PN/PS在4.0以下時具有良好的沉降性能,過高的比例會導(dǎo)致沉降性能差[27].本研究第80d R1和R2顆粒污泥的PN/PS分別為1.29、1.15,再次表明載體顆粒污泥(R2)沉降性能優(yōu)于自固定化顆粒污泥(R1).

2.3 PDN系統(tǒng)微生物群落多樣性分析

2.3.1 物種豐富性和多樣性 R1和R2運行第81d污泥樣品豐富性和多樣性指數(shù)如表3所示.R1和R2污泥樣品中共產(chǎn)生46518和47643個有效序列,OTUs分別為415和425.R1和R2的覆蓋率估計值均在0.99以上,表明測序結(jié)果能夠全面分析微生物群落結(jié)構(gòu).ACE和Chao值代表物種豐富度, Shannon和Simpson值估算樣品中物種的多樣性.由表1可知R2的物種豐富度和多樣性比R1高,這與SEM觀察到的結(jié)果一致.

表3 R1和R2污泥豐富度和多樣性

2.3.2 門水平微生物群落結(jié)構(gòu) R1和R2運行第81d污泥樣品門水平微生物群落結(jié)構(gòu)如圖6(a)所示.按門水平相對豐度高低排序,R1主要菌門分別為變形菌門(Proteobacteria)(84.00%)、綠彎菌門(Chloroflexi) (2.36%)、擬桿菌門(Bacteroidetes) (2.14%)和衣原體門(Chlamydiae)(1.80%);R2依次為變形菌門(Proteobacteria)(73.34%)、擬桿菌門(Bacteroidetes) (5.49%)、綠彎菌門(Chloroflexi) (3.50%)和浮霉菌門(Planctomycetes)(2.58%).

變形菌門在氮素循環(huán)和有機物利用過程中發(fā)揮重要作用,研究表明變形菌門在PDN系統(tǒng)中占比很高[28].R1和R2的絕對優(yōu)勢菌門是變形菌門,高豐度的變形菌門是高效積累NO2--N的保證[29].擬桿菌門具有降解有機物和促進污泥顆?；墓δ?綠彎菌門可以提供絮體形成的絲狀支架并具有降解碳水化合物的功能[6-7],R2擬桿菌門和綠彎菌門占比均高于R1.

2.3.3 屬水平微生物群落結(jié)構(gòu) R1和R2運行第81d污泥樣品屬水平微生物群落結(jié)構(gòu)如圖6(b)所示.按屬水平相對豐度高低排序,R1主要菌屬分別為(59.18%)、(6.81%)、(2.68)和(1.58);R2主要菌屬分別為(46.04%)、(5.99%)、(2.13%)、(1.69%).

屬于變形菌門的屬和屬均是兩個反應(yīng)器的優(yōu)勢菌屬.屬在污水處理系統(tǒng)發(fā)揮高效的反硝化性能,張星星等[16]研究發(fā)現(xiàn)在PDN-SBR反應(yīng)器中屬豐度達(dá)40.6%,推測屬在PDN系統(tǒng)具有重要的作用.屬是廣泛報道的 PDN反應(yīng)器中實現(xiàn)NO2--N積累的功能菌屬[30].本研究培養(yǎng)馴化的自固定化顆粒污泥和載體顆粒污泥均成功富集PDN功能菌屬,R1和R2 反應(yīng)器具有良好的NO2--N積累性能.

3 結(jié)論

3.1 在25℃, pH=9.0和COD/NO3--N為2～3條件下, R1和R2均成功啟動PDN.NLR達(dá)到1.2kg/ (m3·d),R1和R2NTR分別高于80%、85%,添加多孔生物填料有助于提高 NO2--N積累性能.

3.2 以絮狀普通活性污泥為接種物連續(xù)運行USB反應(yīng)器81d均成功培養(yǎng)PDN自固定化顆粒污泥和載體顆粒污泥.添加親水性多孔生物填料促進了污泥顆?；M程,與PDN自固定化顆粒污泥(R1)相比, PDN載體顆粒污泥(R2)結(jié)構(gòu)規(guī)則緊密、沉降性能優(yōu)且污泥濃度高.

3.3 R1和R2的主要菌門均是變形菌門、擬桿菌門和綠彎菌門,兩個反應(yīng)器均成功富集PDN功能菌屬屬和屬,保證了系統(tǒng)高效穩(wěn)定積累NO2--N.R2載體顆粒污泥的物種豐富度和多樣性高于R1自固定化顆粒污泥,添加親水性多孔生物填料有利于微生物聚集生長.

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Study on start-up and operation of USB reactor for partial denitrification based on granular sludge.

MA Rui-jie1, LIU Yong-hong1*, LIANG Ji-dong2, WANG Ning1, XIE Feng-xia1

(1.School of Environment and Chemical Engineering, Xi′an Polytechnic University, Xi′an 710048, China；2.School of Energy and Power Engineering, Xi′an Jiaotong University, Xi′an 710049, China)., 2022,42(5)：2129～2135

In this study, the partial denitrification process was constructed in two identical Up-flow Sludge Bed reactors (R1 without the carrier, R2 with porous biological carrier). The transformation performance of nitrate-to-nitrite, the physicochemical characteristics of partial denitrification granular sludge, the production characteristics of extracellular polymeric substances (EPS), and the dominant characteristics of functional bacteria generawere compared and analyzed. The NO3--N→NO2--N transformation ratio(NTR) in R2 (85%) was higher than that of R1(80%) when the reactors ran for 81 days and the nitrogen loading rates (NLR) was 1.2kg/(m3·d). The sedimentation performance of carrier granular sludge (R2) was better than that of self-immobilized granular sludge (R1), and the carrier granular sludge in R2 was easier to capture EPS. The PN/PS value of R1 (1.29) was greater than that of R2 (1.15), and the sludge volume index (SVI) of R1(27.07mL/g MLSS) was also greater than that of R2 (19.36mL/g MLSS). Under scanning electron microscopy, it was found that Long bacillus gathered on the surface of R1 sludge, while short-bacillus and coccus were observed on the surface of R2 sludge. In comparison, the structure of granular sludge in R2 was more regular and dense than that in R1. Microbial high throughput sequencing showed that the species richness and diversity of R2 were higher than that of R1.,, andwere dominant in the partial denitrification system. The main NO2--N accumulating genera in both R1 and R2 were(R1 accounted for 59.18% and R2 accounted for 46.04%) and(R1 accounted for 6.81% and R2 accounted for 5.99%).

partial denitrification(PDN)；anaerobic ammonia oxidation(anammox)；nitrite accumulation；PDN granular sludge；biological carrier

X703

A

1000-6923(2022)05-2129-07

馬瑞婕(1996-),女,山西大同人,西安工程大學(xué)碩士研究生,主要從事污水處理技術(shù)研究.

2021-10-15

國家自然科學(xué)基金青年科學(xué)基金資助項目(22008188);陜西省重點研發(fā)計劃項目(2018ZDXM-SF-023,2019GY-154,2020SF-413);西安工程大學(xué)2021年度研究生創(chuàng)新基金資助項目(chx2021023)

* 責(zé)任作者, 教授, liuyhxa@hotmail.com