崔榮陽,劉剛才,胡萬里,付 斌,陳安強(qiáng)*

水位波動(dòng)和氮濃度變化對(duì)氮轉(zhuǎn)化功能基因豐度的影響

崔榮陽1,2,劉剛才1,胡萬里3,付 斌3,陳安強(qiáng)3*

(1.中國科學(xué)院水利部成都山地災(zāi)害與環(huán)境研究所,中國科學(xué)院山地表生過程與生態(tài)調(diào)控重點(diǎn)實(shí)驗(yàn)室,四川 成都 610041；2.中國科學(xué)院大學(xué),北京 100049；3.云南省農(nóng)業(yè)科學(xué)院農(nóng)業(yè)環(huán)境資源研究所,云南 昆明 650201)

為探索淺層地下水氮濃度及水位波動(dòng)對(duì)土壤剖面中氮轉(zhuǎn)化功能基因豐度的影響,以洱海近岸農(nóng)田原狀土壤剖面為對(duì)象,研究了模擬常規(guī)氮濃度的淺層地下水進(jìn)行水位波動(dòng)(SND)和持續(xù)淹水(SNF),以及無氮濃度的淺層地下水位波動(dòng)(0ND)后土壤剖面氮濃度和氮轉(zhuǎn)化功能基因豐度的變化,探討了土壤因子與功能基因豐度的關(guān)系.結(jié)果表明:SNF、SND和0ND處理較試驗(yàn)前土壤剖面中溶解性總氮(TDN)濃度分別降低了44%、21%和30%,NO3?-N濃度分別降低了55%、28%和38%.同時(shí),0ND和SNF處理較SND處理土壤剖面中反硝化功能基因豐度分別降低20%和1%,厭氧氨氧化功能基因豐度則分別增加68%和7%,硝化功能基因豐度分別降低34%和增加23%,土壤含水率(MC)、NH4+-N、NO3?-N和TDN均為功能基因豐度變化的重要驅(qū)動(dòng)因子.土壤剖面持續(xù)淹水會(huì)顯著降低溶解性氮濃度,淺層地下水波動(dòng)及水中氮濃度引起的土壤剖面干濕交替和氮濃度變化是氮轉(zhuǎn)化功能基因豐度變化的主要驅(qū)動(dòng)力.

氮轉(zhuǎn)化功能基因；土壤剖面；干濕交替；淺層地下水位波動(dòng)

土壤氮循環(huán)是生物地球化學(xué)循環(huán)最重要的過程之一,其循環(huán)過程受土壤微生物驅(qū)動(dòng)[1].氮在微生物作用下進(jìn)行著復(fù)雜的轉(zhuǎn)化,保持著土壤中氮素的動(dòng)態(tài)平衡.土壤中氮素形態(tài)轉(zhuǎn)化決定了氮素的植物利用效率[2],影響著氮素向水-氣環(huán)境中的排放量,從而造成了溫室氣體排放、水體富營養(yǎng)化、地下水硝酸鹽污染等環(huán)境問題[3-4].土壤中氮形態(tài)間的轉(zhuǎn)化過程受微生物代謝產(chǎn)生的酶控制,每個(gè)代謝過程中產(chǎn)生的酶均有標(biāo)志性的基因編碼[1],如AOA-和AOB-是參與NH4+-N轉(zhuǎn)化為NO3?-N的關(guān)鍵功能基因[5];NO2?-N轉(zhuǎn)化為N2O的關(guān)鍵編碼基因?yàn)椤⒑蚚1].

淺層地下水位波動(dòng)是造成土壤剖面氮素流失的重要途徑,該流失路徑與土壤微生物調(diào)控的硝化、反硝化、厭氧氨氧化、硝酸鹽異化成銨等氮轉(zhuǎn)化過程密切相關(guān)[6-7].由于水位升降造成土壤剖面環(huán)境變化,改變了土壤微生物群落結(jié)構(gòu)及功能基因豐度,影響著土壤氮素形態(tài)轉(zhuǎn)化,驅(qū)動(dòng)著土壤剖面中氮素的累積和流失[7-8].明確淺層地下水位波動(dòng)下土壤微生物功能基因變化及其主要驅(qū)動(dòng)因子對(duì)于預(yù)測(cè)氮素轉(zhuǎn)化和流失至關(guān)重要.一般來說,水位的周期性升降常發(fā)生在消落帶、濕地、河湖岸帶等區(qū)域,這導(dǎo)致溶解氧、pH值、土壤含水量、溫度、碳源等眾多影響土壤微生物的因子也發(fā)生周期性變化,而水位滯留時(shí)間、流速等同樣影響著土壤微生物及氮轉(zhuǎn)化功能基因豐度變化[9],加劇了這些區(qū)域氮素轉(zhuǎn)化過程及微生物變化的復(fù)雜性.前期研究發(fā)現(xiàn),隨著水位降低和土壤剖面持續(xù)干旱,參與反硝化過程的、、和基因豐度逐漸降低,而AOA-和AOB-基因豐度則逐漸增加,且基因豐度顯著高于[10-12].但較多研究主要關(guān)注表層土壤功能基因豐度變化,而對(duì)于地下水位周期性波動(dòng)引起的土壤剖面干濕交替和底物濃度變化對(duì)氮轉(zhuǎn)化功能基因豐度的影響研究較少.

淺層地下水位波動(dòng)引起的農(nóng)田土壤剖面-地下水界面變化是氮素遷移轉(zhuǎn)化活躍的關(guān)鍵地帶,地下水位波動(dòng)影響著氮素在土壤剖面中的滯留時(shí)間[13]、對(duì)流彌散、吸附解析、有機(jī)氮礦化、硝化和反硝化等過程[14-15],促進(jìn)了水土界面間的氮素交換,使得土壤剖面與淺層地下水之間氮濃度呈顯著正相關(guān)[16-17].淺層地下水位波動(dòng)改變了氮形態(tài)及其濃度在土壤剖面中的空間分布,然而,不同地下水氮濃度及其水位波動(dòng)是否會(huì)造成土壤剖面中氮轉(zhuǎn)化功能基因豐度呈現(xiàn)出差異性變化仍不清楚.本文以洱海湖周農(nóng)田土壤剖面為研究對(duì)象,通過微宇宙試驗(yàn)和PCR技術(shù),研究了高、低氮濃度的淺層地下水,長期淹水與周期升降兩種水位波動(dòng)模式下土壤剖面氮濃度和氮轉(zhuǎn)化功能基因豐度的變化,探究地下水氮濃度和水位波動(dòng)模式對(duì)土壤氮轉(zhuǎn)化功能基因豐度變化的主要驅(qū)動(dòng),以期為認(rèn)識(shí)農(nóng)田土壤剖面-地下水界面氮素生物地球化學(xué)循環(huán)過程提供科學(xué)支撐.

1 材料與方法

1.1 原狀土柱取樣

試驗(yàn)土柱取于洱海西岸湖濱區(qū)大莊蔬菜地(100°12′31″E,25°40′14″N),海拔1966m.氣候類型為低緯亞熱帶高原季風(fēng)氣候,年均降雨量約為1100mm,降雨主要集中在6～11月(占年降雨量85%～90%),年均氣溫為15.7℃.同時(shí),該區(qū)域?qū)儆诘湫图s化露地蔬菜種植區(qū),一年平均種植蔬菜3茬,每茬蔬菜種植的肥料施用量為375kg N/hm2、165kg P2O5/hm2和1200kg/hm2有機(jī)肥料.農(nóng)田土壤類型為水稻土,80年代后期開始種植蔬菜,土壤剖面按發(fā)生層分為4層:耕作層(A層,0～30cm)、犁底層(B層,30～45cm)、潴育層(C層,45～70cm)和潛育層(D層,>70cm),各層土壤特性見表1[7].前期調(diào)查發(fā)現(xiàn),該區(qū)域淺層地下水平均總氮濃度為33.20mg/L[7],雨、旱季降雨差異造成的地下水位波動(dòng)范圍約為101cm[18].

表1 土壤剖面理化性質(zhì)

使用直徑30cm、高110cm的PVC管采集100cm深的原狀土柱.首先,挖大約20cm深、直徑略大于30cm的圓形土柱;然后將PVC管底部置于圓形土柱上,用橡皮錘敲擊PVC管頂部,直至挖出的圓形土柱楔入PVC管中,依次重復(fù)上述過程,直到PVC管中土壤剖面達(dá)到100cm;最后在土柱底部放置一塊直徑29.5cm、厚1cm的透水石和孔徑為2mm的尼龍網(wǎng),并用蓋子將PVC管底部和頂部密封,將土柱運(yùn)至實(shí)驗(yàn)室,置于鋼架上靜置1周,使土體逐漸穩(wěn)定.

1.2 試驗(yàn)處理與采樣

微宇宙試驗(yàn)裝置主要由原狀土柱、進(jìn)水口、出水口、供水桶、溶液收集桶和蠕動(dòng)泵構(gòu)成,進(jìn)水口和出水口分別位于原狀土柱底部的蓋子和PVC管頂部的管壁,并通過硅膠管分別與蠕動(dòng)泵和溶液收集桶連接;蠕動(dòng)泵另一端通過硅膠管與供水桶連接.試驗(yàn)設(shè)3個(gè)處理:模擬淺層地下水氮濃度+水位升降處理(SND)、模擬淺層地下水氮濃度+持續(xù)淹水處理(SNF)、無氮添加+水位升降處理(0ND).模擬淺層地下水氮溶液(NH4+-N 0.5mg/L+NO3?-N 30mg/L)由KNO3、(NH4)SO4和蒸餾水配置.

試驗(yàn)開始時(shí),在供水桶中加入配好的模擬淺層地下水溶液,將蠕動(dòng)泵流速調(diào)節(jié)為7mL/min,之后打開蠕動(dòng)泵將溶液通過進(jìn)水口泵入土柱中,在整個(gè)淹水階段,3個(gè)處理的土壤表層均保持薄薄的水層,超過該水層的溶液經(jīng)管壁出水口排至溶液收集桶.整個(gè)試驗(yàn)周期為120d,SNF處理持續(xù)淹水120d,SND和0ND處理分兩次干濕交替,每次干濕交替的試驗(yàn)周期為60d,其中前30d為淹水階段,后30d為落干階段.SND與0ND處理在落干階段停止蠕動(dòng)泵輸送溶液,打開土柱底部入水口,使土柱內(nèi)溶液慢慢滲出直至落干.每隔30d使用直徑2cm的小型土鉆對(duì)A、B、C和D層土壤進(jìn)行取樣(第1次記為F?、第2次記為D?、第3次記為FⅡ、第4次記為DⅡ),一份存儲(chǔ)于4℃冰箱中用于測(cè)定土壤含水率(MC)、NH4+-N、NO3?-N和溶解性總氮(TDN),一份土樣凍干后,存儲(chǔ)于-80℃超低溫冰箱中用于測(cè)定氮轉(zhuǎn)化功能基因豐度.取完土壤剖面樣后,用直徑2cm的PVC管插入取樣留下的洞中,防止土體破壞.

1.3 指標(biāo)測(cè)定

土壤中NH4+-N、NO3?-N采用CaCI2溶液浸提-AA3連續(xù)流動(dòng)分析儀測(cè)定(Bran+Luebbe,德國),TDN采用CaCI2溶液浸提,堿性過硫酸鉀氧化-紫外分光光度法測(cè)定,MC采用烘干法測(cè)定.

稱取0.5g凍干土壤樣品,使用E.Z.N.A.?土壤DNA提取試劑盒(Omega Bio-tek, Norcross,美國)進(jìn)行土壤DNA提取,采用1%瓊脂糖凝膠電泳檢測(cè)DNA的提取質(zhì)量,使用NanoDrop2000測(cè)定DNA 濃度和純度.使用實(shí)時(shí)熒光定量PCR檢測(cè)儀(ABI7500,美國)測(cè)定土壤中(AOA-、AOB-)、(、)、、功能基因的豐度.目標(biāo)基因引物、序列和片段大小見表2,定量在20.0μL反應(yīng)體系中進(jìn)行,反應(yīng)體系為:ChamQ SYBR Color qPCR Master Mix(2X)16.4μL、模板DNA 2μL、引物F(5mmol/L)0.8μL、引物R(5mmol/L) 0.8μL.PCR熱循環(huán)條件為:初級(jí)階段3min,然后在95℃/5s、55℃/30s和72℃/1min進(jìn)行40個(gè)循環(huán),擴(kuò)增效率范圍為85%～100%,2399%.

表2 qPCR目的基因擴(kuò)增引物序列

1.4 數(shù)據(jù)處理

每個(gè)階段土壤剖面氮濃度或功能基因豐度為4層土壤的平均值,S?和SⅡ分別為試驗(yàn)前60d和后60d土壤剖面氮濃度或功能基因豐度的平均值,F和D分別為0～30d + 60～90d和30～60d + 90～120d土壤剖面氮濃度或功能基因豐度的平均值.使用SPSS 24.0進(jìn)行正態(tài)分布和顯著差異性(<0.05)檢驗(yàn), Origin 2019b進(jìn)行繪圖,RDA分析和SEM通過R中“vegan”和“l(fā)avaan”包執(zhí)行.

2 結(jié)果與分析

2.1 土壤氮濃度變化

淺層地下水中不同氮濃度及其水位升降引起的干濕交替均會(huì)造成土壤剖面中不同形態(tài)氮濃度變化.隨土壤剖面干濕交替,SND和0ND處理各土層中氮濃度均呈相同變化趨勢(shì),NH4+-N在淹水階段逐漸增加和落干階段逐漸降低,NO3?-N和TDN卻呈相反變化(圖1).隨土壤剖面持續(xù)淹水,SNF處理中各土層NH4+-N濃度呈現(xiàn)出前60d逐漸增加和后60d逐漸降低,NO3?-N和TDN濃度呈現(xiàn)整體性持續(xù)下降.3個(gè)處理中各形態(tài)氮濃度均表現(xiàn)為A層>B層>C層>D層.SND和0ND處理土壤剖面氮濃度在相同階段均呈顯著差異(圖1),與SND處理相比,0ND處理中NH4+-N、NO3?-N和TDN濃度在SI階段分別顯著(<0.05)降低15%、13%和5%,SⅡ階段NO3?-N和TDN濃度顯著(<0.05)降低15%和10%.SNF處理中NO3?-N和TDN濃度在F階段顯著(<0.05)降低9%和增加11%,而在D階段NH4+-N濃度顯著(<0.001)增加81%(<0.001),NO3?-N與TDN濃度則顯著(<0.001)降低55%和50%.相比SND處理,SNF處理的土壤剖面NO3?-N和TDN濃度在整個(gè)試驗(yàn)過程中分別降低37%和29%,0ND處理分別降低14%和7%.

圖1 土壤剖面氮濃度變化

(a)、(c)、(e)為不同處理相同土層的氮濃度變化,(b)、(d)、(f)為不同處理相同階段的氮濃度變化;*表示處理間差異顯著(*<0.05,**<0.01,***<0.001)

2.2 土壤氮轉(zhuǎn)化功能基因豐度變化

淺層地下水中不同氮濃度及其水位升降引起的土壤干濕交替均會(huì)造成土壤剖面中氮轉(zhuǎn)化功能基因豐度變化.隨土壤剖面持續(xù)淹水,SNF處理的各土層中、、和豐度呈整體下降(圖2);而隨干濕交替,SND和0ND處理各土層中豐度表現(xiàn)為在淹水階段下降而落干階段增加,豐度呈整體下降;SND處理B、C、D層中、豐度在淹水階段下降而落干階段增加,0ND的B、D層中豐度也呈相同變化,其它土層中、豐度呈整體下降.3個(gè)處理的土層中功能基因豐度均表現(xiàn)為A層>B層>C層>D層.與SND處理相比(圖2),0ND處理中、、豐度在SI階段分別降低21%、39%和12% (<0.05),豐度增加69%(<0.05);SⅡ階段和豐度降低19%和28%(<0.05),和豐度增加248%和21%(<0.001).水位升降引起的土壤干濕交替同樣改變了氮轉(zhuǎn)化功能基因豐度,與SND處理相比,SNF處理土壤剖面中和豐度在F階段增加49%和53%(<0.05),和豐度降低7%和6%;而、和豐度在D階段分別降低52%、38%和56%(<0.01),豐度增加41%(<0.05).與整個(gè)試驗(yàn)過程中SND處理功能基因豐度相比,SNF和0ND處理土壤剖面厭氧氨氧化功能基因豐度分別增加68%和7%,反硝化功能基因豐度則分別降低20%和1%,硝化功能基因豐度則在SNF處理中降低34%,而0ND處理則增加了23%.

圖2 土壤剖面氮轉(zhuǎn)化功能基因豐度變化

(a)、(c)、(e)、(g)為不同處理相同土層的功能基因豐度變化,(b)、(d)、(f)、(h)為不同處理相同階段的功能基因豐度變化,*表示處理間差異顯著(*<0.05,**<0.01,***<0.001)

3 討論

3.1 淺層地下水中氮濃度和土壤干濕交替對(duì)土壤剖面氮濃度的影響

隨土壤深度增加,土壤中有機(jī)物質(zhì)會(huì)逐漸減少,微生物活性降低[19-20],造成各土層中氮濃度隨剖面深度增加而降低.已有研究表明[6-7,9],水位滯留時(shí)間、地下水中氮濃度均與土壤氮濃度和流失量存在顯著相關(guān)性,這在本研究結(jié)果中也被證明.在淹水階段,各處理土層中NH4+-N逐漸升高,NO3?-N逐漸降低,這歸因于:1)淹水造成土壤剖面形成厭氧環(huán)境,抑制了硝化微生物對(duì)NH4+-N的消耗,促進(jìn)了反硝化微生物活性和加快對(duì)NO3--N的消耗[21];2)厭氧環(huán)境下異化硝酸鹽還原為銨(DNRA)過程變得極為活躍,促進(jìn)了土壤中NO3?-N轉(zhuǎn)化為NH4+-N[9],同時(shí),厭氧環(huán)境也促進(jìn)了土壤有機(jī)氮礦化[22],土壤NH4+-N累積增加而消耗降低導(dǎo)致淹水階段各土層中NH4+-N累積量增加,土層中NO3?-N卻相反.然而,落干階段,由于土壤逐漸由厭氧環(huán)境轉(zhuǎn)變?yōu)楹醚醐h(huán)境,提高了土壤硝化微生物活性,促進(jìn)NH4+-N轉(zhuǎn)化為NO3?-N[23],同時(shí),反硝化及DNRA過程受到抑制[21],導(dǎo)致淹水階段土壤剖面中累積的NH4+-N被消耗而NO3?-N逐漸累積.此外,通過SEM分析也發(fā)現(xiàn)(圖3),MC分別與NH4+-N、NO3?-N和TDN存在直接顯著(<0.05)正效應(yīng),這也證明土壤剖面持續(xù)淹水或干濕交替均顯著影響氮形態(tài)濃度.這些原因造成SNF處理土壤剖面中NH4+-N濃度在整個(gè)試驗(yàn)過程中均高于SND,而NO3?-N濃度則相反.SNF處理持續(xù)淹水60d后,各土層NH4+-N濃度逐漸下降,可能是由于持續(xù)淹水抑制土壤有機(jī)氮礦化[24]和刺激了厭氧氨氧化微生物活性,促進(jìn)NH4+-N轉(zhuǎn)化為N2[25].此外,與SNF處理相比,SND處理的土壤剖面氮濃度在淹水與落干階段波動(dòng)幅度更大,這表明干濕交替加速了土壤剖面氮轉(zhuǎn)化[26-27],主要因?yàn)楦蓾窠惶婕铀偻寥榔拭嬗诤醚?兼氧-厭氧環(huán)境中不斷循環(huán),刺激了好氧或厭氧微生物活性[28],致使土壤剖面氮素不斷轉(zhuǎn)化和相互增加反應(yīng)底物氮濃度.與0ND處理相比,SND處理中土壤剖面各氮形態(tài)濃度顯著較高,一方面原因是地下水中NH4+-N很容易被土壤吸附[29],而NO3?-N雖然不易被土壤吸附,但外源氮大量輸入,激發(fā)厭氧微生物利用外源氮來維持自身的代謝活動(dòng)[30],很大程度上削減了SND處理的土壤剖面氮流失;另一方面是水-土中氮濃度存在較大的濃度差,低氮濃度的地下水與高氮濃度的土壤剖面相互作用,加速了氮從土壤剖面向地下水中釋放,從而0ND處理的土壤剖面氮濃度顯著降低.總體來說,無論持續(xù)淹水或干濕交替,土壤剖面NO3?-N和TDN濃度均呈下降趨勢(shì),這表明地下水位波動(dòng)能夠加速土壤剖面溶解性氮流失.

3.2 水位波動(dòng)對(duì)土壤剖面氮轉(zhuǎn)化功能基因豐度的影響

水位波動(dòng)造成土壤剖面土壤氧化還原環(huán)境、氮濃度和含水率等發(fā)生變化[10-11,31],土壤底物碳氮濃度[32-33]、氧供應(yīng)[34]、土壤理化性質(zhì)[35]等重要因子主要通過影響氮轉(zhuǎn)化功能基因豐度變化,進(jìn)而影響氮的轉(zhuǎn)化過程.通過RDA分析發(fā)現(xiàn)(圖3),土壤NH4+- N、NO3?-N、TDN和MC是土壤氮轉(zhuǎn)化功能基因豐度變化的主要驅(qū)動(dòng)因子,SNF、SND和0ND處理的前兩軸分別解釋了95.5%、98.3%和99.8%的氮功能基因豐度變化.各土層中氮轉(zhuǎn)化功能基因豐度隨剖面深度增加而降低,這歸因于土壤剖面中碳氮濃度和氧擴(kuò)散能力隨土壤深度增加而逐漸降低[36].通常,淹水可增加土壤孔隙中持水量和降低土壤剖面中溶解氧濃度,當(dāng)溶解氧濃度低于2mg/L時(shí)[8],有利于形成反硝化發(fā)生的厭氧環(huán)境,與基因豐度理論應(yīng)該增加.但研究發(fā)現(xiàn),土壤剖面氮濃度整體呈現(xiàn)出的下降趨勢(shì)與和基因豐度變化也一致,水位升降造成的土壤剖面NO3?-N濃度變化才是導(dǎo)致SND與0ND處理的土層中和基因豐度變化的主要原因,SEM分析結(jié)果也表明(圖3),SND與0ND處理中NO3?-N也分別與和呈現(xiàn)出直接的正效應(yīng)(<0.05),這說明NO3?-N作為反硝化過程的反應(yīng)底物,其濃度高低也影響反硝化作用和氮轉(zhuǎn)化功能基因豐度[37-38].相比0ND處理,SND處理的水中較高的NO3?-N濃度為土壤反硝化提供了外源氮,降低土壤剖面中NO3?-N流失,加之土壤孔隙水中NO3?-N濃度也是控制反硝化的關(guān)鍵因素[39],以至于SND處理中與基因豐度在SI和SⅡ階段均顯著高于0ND處理.SNF處理土壤剖面中與基因豐度呈持續(xù)下降趨勢(shì),在F和D階段與SND處理均呈現(xiàn)出顯著差異,且MC對(duì)、均有直接顯著正效應(yīng)(<0.01),這說明SNF處理中MC顯著影響和豐度;而與NO3?-N濃度表現(xiàn)出微弱的負(fù)相關(guān),這與Dandie等[40]研究一致,但并不能否認(rèn)土壤剖面中NO3?-N濃度對(duì)其沒有影響,長期淹水可能導(dǎo)致反硝化微生物所需的碳供應(yīng)不足,抑制了反硝化酶活性.此外,F階段SNF處理中和豐度比SND處理顯著增加49%和53%,而D階段則分別顯著降低38%和56%.這是由于SNF處理處于持續(xù)淹水環(huán)境,更有利于促進(jìn)反硝化微生物生長,但D階段SND處理中累積的NH4+-N轉(zhuǎn)化為NO3?-N,底物濃度增加刺激了反硝化微生物活性,提高了和豐度,這也表明水位升降引起的土壤氮濃度變化對(duì)和豐度起主導(dǎo)作用.然而,土壤干濕交替卻顯著影響各處理中豐度,SEM分析發(fā)現(xiàn),3個(gè)處理中MC對(duì)均有直接的顯著正效應(yīng)(<0.05),并通過調(diào)控TDN和NO3?-N間接影響(<0.05),這表明MC是豐度變化的主要驅(qū)動(dòng)因子.這主要由于AOA和AOB為好氧微生物,干濕交替造成的土壤氧化還原環(huán)境和土壤水分變化更有利于刺激硝化酶活性;另一方面,AOA和AOB對(duì)環(huán)境適應(yīng)偏好并不同,如AOA更能適應(yīng)低氧、酸性、低NH4+-N濃度環(huán)境[41],兩者對(duì)環(huán)境的偏好可能掩蓋了底物氮濃度的重要性.在本研究中,各處理豐度整體均呈現(xiàn)出持續(xù)下降趨勢(shì),這有兩方面原因,一是土壤剖面NH4+-N較培養(yǎng)前增加,這增加了厭氧氨氧化電子供體,但NO3?-N濃度降低導(dǎo)致反硝化底物濃度缺乏,限制了NO2?-N的形成,從而造成厭氧氨氧化電子受體供應(yīng)不足,限制了厭氧氨氧化酶活性[42];二是SND與0ND處理落干階段形成好氧環(huán)境,并不利于厭氧氨氧化過程發(fā)生,通過SEM分析也發(fā)現(xiàn)MC均與呈現(xiàn)顯著正效應(yīng)(<0.05),這在D階段SND處理中豐度顯著低于SNF處理也得以體現(xiàn).綜上,淺層地下水升降及其水中氮濃度分別引起土壤剖面干濕交替和氮濃度變化,兩者共同驅(qū)動(dòng)土壤剖面氮轉(zhuǎn)化功能基因豐度的變化,且淺層地下水中氮濃度影響強(qiáng)度更大,因?yàn)榕cSND處理相比,0ND處理土壤剖面氮轉(zhuǎn)化功能基因豐度變化率遠(yuǎn)高于SNF處理.

圖3 土壤氮轉(zhuǎn)化功能基因豐度與環(huán)境因子的冗余度分析(RDA)和結(jié)構(gòu)方程(SEM)

SEM中黑色和灰色箭頭分別表示正效應(yīng)和負(fù)效應(yīng),實(shí)線和虛線表示路徑系數(shù)的顯著和不顯著,線寬度表示顯著性程度(*<0.05,**<0.01,***<0.001)

4 結(jié)論

4.1 淺層地下水中的氮濃度及其水位波動(dòng)顯著影響土壤剖面中氮濃度,持續(xù)淹水和低氮濃度地下水波動(dòng)將顯著降低土壤剖面溶解性氮濃度.與初始階段土壤剖面中溶解性氮濃度相比,常規(guī)氮濃度地下水波動(dòng)下土壤剖面中NO3?-N和TDN分別下降28%和21%,無氮濃度地下水波動(dòng)下NO3?-N和TDN下降率增加至38%和30%,持續(xù)淹水條件下土壤剖面NO3?-N和TDN下降率高達(dá)55%和44%.

4.2 地下水位波動(dòng)及水中氮濃度引起土壤剖面干濕交替和氮濃度變化,共同驅(qū)動(dòng)著土壤剖面氮轉(zhuǎn)化功能基因豐度變化,且淺層地下水中氮濃度影響強(qiáng)度遠(yuǎn)高于水位波動(dòng).持續(xù)淹水和無氮濃度地下水波動(dòng)條件下土壤剖面厭氧氨氧化功能基因豐度與常規(guī)氮濃度地下水波動(dòng)相比,分別增加7%和68%,反硝化功能基因豐度則分別降低1%和20%,硝化功能基因豐度分別增加23%和降低34%.

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Effects of water table fluctuations and nitrogen concentration variations on the abundances of nitrogen-transforming functional genes in soil profiles.

CUI Rong-yang1,2, LIU Gang-cai1, HU Wan-li3, FU Bin3, CHEN An-qiang3*

(1.Key Laboratory of Mountain Surface Processes and Ecological Regulation, Chinese Academy of Sciences, Institute of Mountain Hazards and Environment, Chinese Academy of Sciences and Ministry of Water Conservancy, Chengdu 610041, China；2.University of Chinese Academy of Science, Beijing 100049, China；3.Agricultural Environment and Resources Institute, Yunnan Academy of Agricultural Science, Kunming 650201, China)., 2022,42(11)：5378～5386

To explore the effects of nitrogen concentration in shallow groundwater and its water table fluctuations on the abundance of soil nitrogen-transforming functional genes, taking the undisturbed soil profile from cropland around Erhai as the object, changes in nitrogen concentrations and abundance of nitrogen-transforming functional genes in soil profiles under shallow groundwater table fluctuations (SND) and continuous flooding (SNF) with conventional nitrogen concentration, and shallow groundwater table fluctuations (0ND) without nitrogen were studied. The relationship between soil environmental factors and abundance of functional genes was discussed. The results indicated that, compared with the nitrogen concentrations in soil profile before the microcosmic experiment, the total dissolved nitrogen (TDN) concentrations in SNF, SND and 0ND decreased by 44%, 21% and 30%, and NO3?-N concentrations decreased by 55%, 28% and 38%, respectively. Meanwhile, compared with the abundance of nitrogen-transforming functional genes in soil profile in SND, the denitrification function gene abundances in 0ND and SNF decreased by 20% and 1%, while the anammox function gene abundances increased by 68% and 7%, and the nitrification function gene abundances decreased by 34% and increased by 23%, respectively. Changes in functional gene abundances were mainly driven by soil moisture content (MC), NH4+-N, NO3?-N and TDN. In conclusion, continuous flooding in soil profiles would significantly reduce dissolved nitrogen concentrations, and changes in alternation of drying-flooding and nitrogen concentrations in soil profile caused by the nitrogen concentrations in shallow groundwater and its water table fluctuations were the main drivers for changes in the abundance of nitrogen-transforming functional genes.

nitrogen-transforming functional gene；soil profile；alternation of drying-flooding；shallow groundwater table fluctuation

X172;X523

A

1000-6923(2022)11-5378-09

崔榮陽(1993-),男,云南昆明人,中國科學(xué)院、水利部成都山地災(zāi)害與環(huán)境研究所博士研究生,主要從事土壤氮素遷移轉(zhuǎn)化及其環(huán)境效應(yīng)研究.發(fā)表論文13篇.

2022-04-15

國家自然科學(xué)基金資助項(xiàng)目(41977319,42067052);云南省科技人才與平臺(tái)計(jì)劃項(xiàng)目(202205AM070002);云南省財(cái)政廳專項(xiàng)(530000221100000648476)

* 責(zé)任作者, 研究員, chaq163@163.com