張 楠 ,何 凱 ,鐘繼承 ,安燕飛 ,袁和忠 ,張 雷 *

藻源性有機質(zhì)沉降對沉積物甲烷釋放促進作用

張 楠1,2,何 凱2,3,鐘繼承2,安燕飛3,袁和忠1,張 雷2*

(1.南京信息工程大學環(huán)境科學與工程學院,江蘇 南京 210044；2.中國科學院南京地理與湖泊研究所,湖泊與環(huán)境國家重點實驗室,江蘇 南京 210008；3.安徽大學資源與環(huán)境工程學院,安徽 合肥 230601)

采集巢湖西北湖灣區(qū)水與沉積物樣品,模擬藻源性有機質(zhì)沉降對 CH4釋放的驅(qū)動作用.培養(yǎng)結(jié)果顯示,加藻組 CH4釋放通量(1.59±0.51)～(98.89±14.30) μmol/(m2· h)顯著高于對照組(0.02±0.016)～(1.37±0.44)mmol/(m2· h)(<0.001).培養(yǎng)過程中,藻源性有機質(zhì)降解迅速,提供了大量產(chǎn)甲烷底物,溶解氧和硫酸根等電子受體減少,創(chuàng)造了有利產(chǎn)甲烷菌的環(huán)境,增加了產(chǎn)甲烷菌數(shù)量,尤其是乙酸營養(yǎng)型產(chǎn)甲烷菌.綜上,藻源性有機質(zhì)的大量輸入,可以增大沉積物產(chǎn)甲烷速率和甲烷釋放通量,其中乙酸營養(yǎng)型產(chǎn)甲烷菌起到重要作用.

釋放通量；產(chǎn)甲烷菌；溫室氣體；二氧化碳；湖泊；水華

甲烷(CH4)年釋放量約737Tg,是僅次于二氧化碳(CO2)的重要溫室氣體,對氣候變暖的貢獻達16%～25%[1-2].目前大氣中CH4濃度已超過1.80× 10-6,是工業(yè)革命以前的近2.6倍[3].湖泊作為CH4的重要自然排放源與CH4代謝微生物的主要棲息地,其CH4年釋放量可達(150.9±73.0)Tg[2].不同湖泊CH4釋放強度差異較大[4],這是產(chǎn)甲烷底物及環(huán)境因子對產(chǎn)甲烷及CH4排放共同影響的結(jié)果[5].底物數(shù)量是限制產(chǎn)甲烷速率的主要因素,有機質(zhì)增加會提高產(chǎn)甲烷產(chǎn)量[6];而不同產(chǎn)甲烷菌所利用底物種類不同,因此產(chǎn)甲烷途徑受限于底物類型[7].湖泊中CH4主要來自嚴格厭氧條件下產(chǎn)甲烷菌對有機質(zhì)的分解過程,除底物外產(chǎn)甲烷過程還會受到氧氣的嚴格限制.另外,湖泊中的硫酸鹽(SO42-),硝酸鹽(NO3-)和金屬氧化還原過程能夠與產(chǎn)甲烷菌群落競爭乙酸,H2等底物從而影響產(chǎn)甲烷[8-10],且SO42-,NO3-和金屬離子的存在可通過厭氧氧化消耗CH4[11-12].

人類活動使得水體富營養(yǎng)化成為全球普遍現(xiàn)象,無論單個大型湖泊的分區(qū)研究結(jié)果[13],還是多個湖泊的對比研究結(jié)果[4],以及不同區(qū)域湖泊的統(tǒng)計分析結(jié)果均表明富營養(yǎng)化可以促進CH4產(chǎn)生與釋放[14];有研究表明這將會導致2100年CH4釋放量升高30%～90%[15].在富營養(yǎng)化湖泊中,大量藍藻繁殖,嚴重堆積后還會形成湖泛,長期監(jiān)測數(shù)據(jù)表明藍藻對CH4釋放有促進作用[16-17],但尚未闡明具體藍藻不同階段,包括生長繁殖階段與衰亡階段,對CH4釋放的影響.懸浮在水體中的藍藻顆粒物衰亡后,最終沉積至沉積物表面,其殘體分解并沉降至沉積物表面,為沉積物帶來大量乙酸[18],甲基硫醚[19]等易降解有機質(zhì),這些有機質(zhì)能促進沉積物CH4的產(chǎn)生[20],且通過改變沉積物中有機質(zhì)種類,使產(chǎn)甲烷菌代謝途徑及優(yōu)勢種變化,影響CH4的產(chǎn)生與釋放[7].所以本研究以藻源性有機質(zhì)的沉降為重點,利用室內(nèi)模擬,探索藻源性有機質(zhì)沉降對CH4釋放的促進作用以及影響機制.

1 材料與方法

1.1 野外采樣

實驗沉積物和水樣采集于巢湖西北湖灣(117.35760°E,31.70499°N).現(xiàn)場利用彼得森采泥器(HL–CN,恒嶺科技,武漢)采集表層沉積物裝于聚乙烯塑料桶中.原水樣用浮游植物網(wǎng)過濾,除去水中的藻類和其他浮游生物,濾后儲存于白色聚乙烯桶中,當天帶回實驗室.同時用浮游植物網(wǎng)采集藍藻生物樣品,濾去多余的水分后收集藍藻樣品帶回實驗室.

1.2 培養(yǎng)裝置

用12根有螺口的有機玻璃管(內(nèi)徑8.4cm,高度30cm)建立培養(yǎng)裝置.將采集的沉積物樣品過30目篩以去除大顆粒物,并用攪拌器使沉積物樣品攪拌均勻,將沉積物樣品注入有機玻璃管中裝填至20cm處,之后緩緩注入原位水樣至柱高30cm.裝樣完成后,將培養(yǎng)裝置放入恒溫水浴培養(yǎng)箱中,20℃下穩(wěn)定一周.將12個培養(yǎng)裝置隨機分為兩組,每組6個.一組命名為加藻組,每根柱子加入5g 凍干藍藻和少量干沉積物顆粒,用玻璃棒緩慢攪拌,使干藻絮凝沉降;添加量根據(jù)藍藻大量聚集時的生物量與含水率確定[23-24].另一組作為對照組,不添加藍藻.淺水湖泊水底通常氧氣條件良好,為模擬水體覆氧過程,避免沉積物-水界面隨培養(yǎng)時間的延長而出現(xiàn)缺氧,每天用小型充氣泵對各柱樣上覆水曝氣10min,空氣流量為0.5L/min;同時為避免曝氣對水體監(jiān)測的影響,曝氣安排在每天21:00進行.

1.3 分析方法

1.3.1 培養(yǎng)裝置的取樣 干藻加入微型實驗裝置穩(wěn)定1d后,開始持續(xù)19d的培養(yǎng)實驗,并在培養(yǎng)過程監(jiān)測上覆水,間隙水和沉積物化學指標變化,在培養(yǎng)結(jié)束時分析各組沉積物產(chǎn)甲烷菌變化.每個實驗組選擇3根柱樣監(jiān)測上覆水的溶解氧(DO),CH4和CO2溶存濃度及通量,根據(jù)實驗進程每2或3d監(jiān)測1次,監(jiān)測均在上午進行.另外3根柱樣,分別在實驗第5,12,19d用來監(jiān)測間隙水中SO42-,CH4溶存濃度及沉積物中葉綠素(Chl)含量.

1.3.2 沉積物和上覆水理化性質(zhì)分析 用便攜式溶氧測定儀(JPB–607A,雷磁,上海)測定水面下方5cm 處DO含量.間隙水SO42-用微型Peeper獲取間隙水[25],首先將組裝好的Peeper板放置于超純水中用高純氮氣曝氣3h,隨后將其插入培養(yǎng)裝置的沉積物中,平衡2d后取出,取其中的間隙水置于酶標板中,用硫酸鋇沉淀法測定間隙水中SO42-濃度[26].將沉積物柱樣每2cm分為一層,每層沉積物取一定量樣品先冷凍干燥,然后研磨粉碎過100目篩,用90%丙酮在4℃條件下萃取其中Chl,用分光光度法分析其中的Chl含量[26],用過硫酸鉀氧化法對沉積物樣品進行消解,再分別用紫外分光光度法和鉬酸銨分光光度法測定總氮(TN),總磷(TP)含量[27],采用重鉻酸鉀法測定沉積物中有機質(zhì)(OM)的含量[27].

1.3.3 CH4及CO2溶存濃度及通量測量 上覆水CH4和CO2溶存濃度監(jiān)測:首先在水面下方5cm處取15mL水樣注入預置有2g氯化鉀,預先抽真空并注入15mL高純氮氣的30mL頂空瓶中,每個位置取兩個平行測定水樣中溶存的CH4與CO2,將所取樣品放置于恒溫培養(yǎng)箱中靜置12h,使氣相和液相達到平衡后測定CH4與CO2,所有樣品在24h內(nèi)分析完畢.對于培養(yǎng)裝置水–氣界面通量的觀測,先用裝有三通閥的有機玻璃瓶蓋密封瓶口,再用注射器通過三通閥抽取10mL氣體注入50mL 鋁箔采樣袋中,每間隔30min取樣一次,累積取樣5次.在前述沉積物分層中,沉積物中的CH4含量通過用頂端截斷的5mL注射器獲取每2cm一層的沉積物樣品,裝入預先裝有50mL 2.5% NaOH 溶液的100mL血清瓶中,立即用丁基橡膠塞密封,再利用高純氮氣通過注射針吹掃殘留空氣2min,每層沉積物做3個平行樣品,倒置保存在恒溫培養(yǎng)箱內(nèi)待測CH4.氣袋,上覆水及沉積物中CH4含量均用配有火焰離子檢測器的氣相色譜儀(GC–FID,7890B,Agilent)測定.

總CO2釋放當量通過CH4和CO2的CO2當量之和表示.先將CH4和CO2釋放通量以單位時間釋放的質(zhì)量表示,再乘對應(yīng)的全球變暖潛能值(GWP),得到二者相應(yīng)的CO2當量.其中CO2的GWP為1, CH4的GWP為28[28].最后,用所計算出來的CO2和CH4的CO2當量相加,即為培養(yǎng)實驗的總CO2釋放當量.

工序任務(wù)對應(yīng)的物料節(jié)點等同于其父節(jié)點(即某個部件任務(wù))對應(yīng)的部件節(jié)點，用t.Parent表示t的父節(jié)點，有

1.3.4 分子生物學分析 從對照組,加藻組各取3份表層(0～2cm)沉積物樣品,放入凍干機凍干,用Omega公司的DNA提取試劑盒(Omega,cat:M5635–02)提取總DNA,嚴格按照試劑盒說明書操作.將提取的DNA進行基因的PCR擴增,引物序列運用mcrA–F(GGTGGTGTMGGDTTCA-CMCARTA)和mcrA–R(CGTTCATBGCGTAGTTV- GGRTAGT).測定樣品時,標準品與未知樣品同時進行PCR循環(huán),根據(jù)未知樣品的閾值循環(huán)值(C),結(jié)合標準曲線求得未知樣品cDNA的起始拷貝數(shù),拷貝數(shù)(0)計算方法:

Ct

= -

K

log

X

0

+

b

(1)

式中:為標準曲線斜率;為標準曲線截距.

在上海派森諾生物科技股份有限公司運用Illumina Miseq測序平臺進行PCR產(chǎn)物的高通量測序.運用DADA2方法[29]對原始序列進行去引物,質(zhì)量過濾,去噪,拼接和去嵌合體等步驟得到特征序列(ASVs).運用QIIME2(2019.4)將處理好的ASVs與Silva數(shù)據(jù)庫中的參考序列進行比對,對結(jié)果進行物種注釋.用稀疏(Rarefaction)的方法預測各樣本在該測序深度下所能觀測到的ASVs及其相對豐度[30-31],該過程運用QIIME2(2019.4)的qiime feature–table rarefy功能實現(xiàn),抽平深度設(shè)為最低樣本序列量的95%.通過對抽平后的ASV表格進行統(tǒng)計,可以獲得每個樣本中的微生物群落在各分類水平的具體組成.對去除singleton后的特征表進行統(tǒng)計,實現(xiàn)各樣本在屬分類水平上的組成分布的可視化,并以柱狀圖呈現(xiàn)分析結(jié)果表.

1.4 數(shù)據(jù)統(tǒng)計與分析

基于氣相色譜儀測量的平衡瓶中氣相濃度,利用理想氣體狀態(tài)方程以及亨利定律計算水樣溶解CH4和CO2原始濃度[13].CH4總釋放通量(t,μmol/ (m2· h)的計算公式如下[32]:

式中:為培養(yǎng)裝置上方氣相體積,m3;為培養(yǎng)裝置截面積,m2;為培養(yǎng)裝置中CH4濃度隨時間變化的斜率,μmol/(L·h);為培養(yǎng)裝置氣相柱高,m.

運用origin繪制柱形圖和折線圖,并且利用其中的–test 進行對照組和處理組間DO,CH4,CO2濃度的差異統(tǒng)計分析.

2 結(jié)果與分析

2.1 水體及沉積物環(huán)境變化

圖1 上覆水中DO的變化

巢湖西北湖灣沉積物的含水率為(59%±7.6%), TN含量為(2988±481) mg/kg,TP含量為(1062±394) mg/kg,OM為(2.36%±0.31%).在19d的培養(yǎng)過程中,對照組DO在(5.18±0.20)～(7.68±0.27) mg/L之間波動.加藻組DO在第2d為0,然后逐漸升高,在培養(yǎng)結(jié)束時DO升高到(2.43±0.15) mg/L(圖1).加藻組的DO濃度在整個培養(yǎng)過程中都要低于對照組的DO濃度(<0.001),說明藻源性有機質(zhì)的加入大量消耗了水體DO.

用Chl含量表示藻源性有機質(zhì)含量.從5～19d的培養(yǎng)過程中,表層沉積物(0～2cm)Chl含量由31.4μg/g降至12.9μg/g,這標志著藻源性有機質(zhì)的快速分解(圖2).從5～12d 培養(yǎng)過程中加藻組硫酸根離子逐漸降低,12d后略微升高;整個培養(yǎng)過程中對照組間隙水中SO42-均大于加藻組,兩組之間在5,12d差異大,在第19d差異縮小(圖3).

圖2 沉積物中Chl a的垂向分布

圖3 間隙水中硫酸根不同培養(yǎng)時間垂向分布

2.2 CH4含量及釋放通量

沉積物間隙水中CH4的垂直分布呈現(xiàn)先隨深度增加而增大,在8～12cm某一深度達到峰值,然后趨于穩(wěn)定(圖4).加藻組0～8cm處的CH4含量整體大于對照組(圖4).隨著實驗的進行,加藻組,對照組沉積物中CH4濃度逐漸升高.其中加藻組第5,12,19d表層0～2cm沉積物CH4含量分別達到(106±6.2),(162±10.3),(92.3±1.0)μmol/L,對照組表層沉積物CH4含量則分別為(10.5±0.9),(26.1±4.7),(57.7±6.4) μmol/L.

在整個培養(yǎng)過程中,對照組上覆水CH4濃度維持在(0.047±0.004)～(0.27±0.30)μmol/L;加藻組CH4濃度呈現(xiàn)先升高后降低的趨勢,在第8d達到峰值(9.95±2.2)μmol/L,之后逐漸降低,第13d后趨于平穩(wěn)(圖5).加藻組的CH4濃度在13d之前濃度都顯著高于對照組(<0.001).加藻組CO2濃度從(109±10.3)μmol/L增加至(135±2.4)μmol/L,整體逐漸升高,顯著高于對照組(<0.001).

對照組CH4釋放通量為(0.02±0.01)～(1.37±0.44)μmol/(m2·h);加藻組的CH4釋放通量為(1.59±0.51)～(98.9±14.3)μmol/(m2·h),在第4d達到最大值,之后逐漸降低.對照組CO2釋放通量維持在(95.3±14.9)～(204±10.8)μmol/(m2·h),呈現(xiàn)上升趨勢;加藻組CO2釋放通量在培養(yǎng)的第2d最大,為(934±86.5)μmol/(m2·h),然后不斷下降最低至(194±70.3)μmol/(m2·h).

如圖6所示,對照組的總CO2釋放當量在(271±34.7)～(581±30.8) μg C-CO2/h波動,變化幅度不大.在培養(yǎng)的第2d,加藻組總CO2釋放當量達到最大值(2830±285)μg C-CO2/h,隨后在培養(yǎng)結(jié)束時降為最低為(558±2328)μg C-CO2/h.在整個培養(yǎng)過程中,加藻組的總CO2釋放當量的均值比對照組高出1.2～10.4倍.其中加藻組CH4的釋放量為(4.54±1.76)～ (281±57.6)μg C-CO2/h,占總CO2釋放當量的0.95%～18.7%,而對照組CH4的釋放量在總CO2釋放當量中僅占0.025%～0.88%.

圖4 各組沉積物中CH4的垂向分布

圖5 培養(yǎng)過程中上覆水CH4和CO2濃度及釋放通量變化

2.3 沉積物中微生物變化及分布

加藻組表層沉積物基因拷貝數(shù)為(3.3±0.65)×106copies/g,比對照組(2.4±0.48)×106copies/g升高37.5%(<0.001,圖7).沉積物中產(chǎn)甲烷菌相對豐度較高的依次為甲烷絲菌屬(),甲烷桿菌屬()和甲烷瘤球菌屬().相較對照組,加藻組產(chǎn)甲烷菌群落中僅甲烷絲菌屬和甲烷桿菌屬相對豐度有增加,甲烷絲菌屬豐度較對照組提高了10.1%,增幅最大.而甲烷瘤球菌屬豐度減少了4.91%,嗜氫型甲烷菌()豐度減少了1.24%,其他產(chǎn)甲烷菌屬(Others)豐度減小了3.64%.

圖6 總CO2釋放當量及CH4和CO2的占比

圖7 沉積物中產(chǎn)甲烷菌群變化,

3 討論

3.1 藻源性有機質(zhì)對CH4釋放的促進作用

沉積物分析結(jié)果顯示加藻組0～8cm的表層沉積物中CH4含量高于對照組(圖4),說明藻源性有機質(zhì)能促進沉積物產(chǎn)甲烷.本研究將藻源性有機質(zhì)引入到沉積物表面,模擬藍藻沉降過程,因此加藻組Chl a含量增加出現(xiàn)在沉積物表面,下層沉積物沒有受到藻源性有機質(zhì)的直接影響(圖3).表層沉積物有機碳增加,從而導致加藻組表層沉積物更有利于CH4的生成,因此在培養(yǎng)前期碳源充足的情況下,加藻組上層沉積物CH4含量高出對照組10倍左右.Furlanetto等[22]對比研究不同營養(yǎng)水平人工湖,發(fā)現(xiàn)富營養(yǎng)化湖泊表層0～1cm沉積物CH4最高,為寡營養(yǎng)湖泊的8倍左右, Wang等[33]觀察到高濃度加藻處理組表層沉積物CH4比對照組高出約10倍.從Chl a監(jiān)測結(jié)果來看,雖然2～8cm沉積物未直接接觸到沉降的藻源性有機質(zhì)(圖3),但這部分沉積物中CH4含量也大幅升高(圖4).這可能有三方面的原因:一是藻源性有機質(zhì)降解產(chǎn)生的溶解性有機質(zhì)向下遷移,為2～8cm沉積物產(chǎn)甲烷提供了更多的易降解有機質(zhì);二是溶解性有機質(zhì)的向下遷移促進了這部分沉積物中原有有機質(zhì)的降解;三是藻源性有機質(zhì)沉降所引起SO42-含量下降,還原環(huán)境提升(圖3),有利于沉積物產(chǎn)甲烷.因此,本研究表明,藻源性有機質(zhì)沉降不僅可以促進表層沉積物產(chǎn)甲烷,也可以促進深處沉積物產(chǎn)甲烷.

沉積物中CH4含量的升高,造成水體中CH4溶存濃度在藻源性有機質(zhì)的作用下也明顯升高.在培養(yǎng)實驗的第8d,加藻組水體中CH4的溶存濃度達到最大(9.95±2.2) μmol/L,平均為對照組的133倍.巢湖西北湖灣野外調(diào)查中發(fā)現(xiàn),沿岸藍藻水華易發(fā)區(qū)水體CH4濃度也高達4.09μmol/L[34].太湖藍藻聚集區(qū)研究表明CH4濃度高達(1.78±1.43)μmol/L,是開闊區(qū)的190倍[35].對于藻源性有機質(zhì)沉降引起的上覆水中CH4濃度升高,除上述沉積物中CH4含量增大以外,沉積物間隙水中SO42-的減小(圖2),是由于在缺氧條件下沉積物微生物會優(yōu)先利用NO3-,然后會利用SO42-[36],因此SO42-的減小表明藻源性有機質(zhì)沉降減少了間隙水中NO3-,金屬離子等電子受體濃度,從而減緩沉積物中其他微生物對產(chǎn)甲烷菌的底物的競爭以及厭氧CH4氧化作用[5].而水體中DO的降低(圖1),也可能降低上覆水中CH4有氧氧化速率.綜上,藻源性有機質(zhì)沉降對水中CH4含量提升是上述CH4產(chǎn)生與氧化兩方面的共同作用結(jié)果.

何強等通過模擬銅綠微囊藻死亡降解過程,發(fā)現(xiàn)銅綠微囊藻有機質(zhì)的降解可將CH4釋放量提高到1549μmol/(m2·d),為對照組的62倍[37].本實驗?zāi)M藻源性有機質(zhì)大量輸入,使得培養(yǎng)的第4d,加藻組CH4釋放通量達到最大為(98.89±14.30)μmol/(m2·h),與上述兩個巢湖西北湖灣藍藻聚集區(qū)的原位釋放通量結(jié)果相近[32,38].在前10d加藻組CH4釋放通量整體高出對照組三個數(shù)量級.對照組去除了藍藻且覆氧良好,未受到藻源性有機質(zhì)的影響,產(chǎn)甲烷反應(yīng)底物相對較少,制約著產(chǎn)甲烷速率;且良好的溶解氧有利于CH4的氧化,進一步導致釋放通量低,造成加藻組和對照組差異高于前人研究.綜上,藍藻衰亡沉降,為水生生態(tài)系統(tǒng)輸入大量藻源性有機質(zhì),所以衰亡沉降后導致的有機質(zhì)輸入可能是CH4的釋放提高主要原因.

在培養(yǎng)第4d后,CH4釋放通量快速降低,這可能是缺少底物以及電子受體升高的結(jié)果(圖2,圖3).藍藻分解產(chǎn)生的易降解有機質(zhì)消耗速度很快[39],可能在培養(yǎng)前4d易降解有機質(zhì)被快速消耗,CH4釋放量快速提高.第4d后易降解有機質(zhì)開始限制產(chǎn)甲烷的速率,導致0～2cm沉積物第5～12d的監(jiān)測中CH4含量變化不大(圖4),并且電子受體(DO,NO3-,鐵離子,SO42-)逐漸恢復表層沉積物中的產(chǎn)甲烷菌被抑制,厭氧甲烷氧化菌開始增強氧化作用[5,40],使得表層沉積物的CH4濃度在培養(yǎng)后期降低(圖4).本實驗結(jié)果與在長期監(jiān)測過程中的結(jié)果類似,在藍藻爆發(fā)期CH4高,隨后因缺少底物而降低[41],實驗在有限空間內(nèi)進行,所以CH4釋放量的變化較快.

然而,實際環(huán)境下藍藻的生長,死亡和沉降是一個不斷發(fā)生的過程,其對沉積物CH4釋放的促進作用通常不如本研究及其他培養(yǎng)實驗顯示的這么強烈.生長期的藍藻在光照下有利于CH4氧化的發(fā)生,降低CH4釋放量[42],導致晝夜CH4釋放量有差異[43].藍藻含量高的月份,CH4釋放通量隨之增高[34],在圣奧古斯丁湖藍藻爆發(fā)年份,CH4平均通量是其他年份的2倍左右[16];荀凡等[38]在巢湖西北湖灣近岸進行藍藻連續(xù)打撈,發(fā)現(xiàn)未打撈藍藻區(qū)的CH4平均釋放通量是打撈區(qū)域的2倍.

根據(jù)CO2釋放結(jié)果可知(圖5),加藻組的總CO2釋放當量高于對照組,說明藻源性有機質(zhì)可能還會引起湖泊的總CO2釋放當量增加.雖然,碳釋放整體是以CO2的釋放為主,CH4僅占很小一部分,但是藻源性有機質(zhì)使得碳釋放中CH4的比重顯著提高(圖6),表明藻源性有機質(zhì)會增大碳釋放量更多歸因于CH4產(chǎn)量的增加,Sun等[44]在烏梁素海觀察到的現(xiàn)象與本文一致.因此,藻源性有機質(zhì)會促進CH4釋放量升高,而CO2釋放占比則會下降,從而改變溫室氣體釋放模式,使得湖泊成為CH4的重要自然釋放源.

3.2 藻源性有機質(zhì)沉降促進CH4釋放的微生物機制

藍藻的加入改變了沉積物-水系統(tǒng)的理化性質(zhì),產(chǎn)甲烷菌數(shù)量也隨之增加(圖7).本研究中加藻組沉積物基因為(3.3±0.65)×106copies/g,相比對照組提高了37.5%.Xu等[37]選取御臨河的沉積物利用銅綠微囊藻進行的培養(yǎng)實驗,在第61d培養(yǎng)結(jié)束沉積物基因增加至8.39×106copies/g,提高了2.8倍,與本研究結(jié)果相似;另外在巢湖的原位圍隔實驗中也發(fā)現(xiàn),含高濃度藍藻的下層水基因,在第5d就提高了一個數(shù)量級[45].然而,在美國密歇根州的貧營養(yǎng)化湖引入藍藻進行培養(yǎng)后,雖然CH4產(chǎn)生速率提高了約3倍,但基因拷貝數(shù)與對照組無顯著差異,因為分解菌群落代謝產(chǎn)生的有機質(zhì)能被產(chǎn)甲烷菌利用的有限,使得產(chǎn)甲烷菌活性提高但豐度變化不大[46].因此,藻源性有機質(zhì)的出現(xiàn)通常會增加基因數(shù)量,但同時也受其他因素影響.

可供產(chǎn)甲烷菌利用的分解產(chǎn)物的增加提高產(chǎn)甲烷菌豐度,而底物的多樣性影響產(chǎn)甲烷菌群落結(jié)構(gòu).本研究中加藻組和對照組群落結(jié)構(gòu)差異不大(圖7),甲烷絲菌屬為二者相對豐度最高的產(chǎn)甲烷菌屬,是一類專性的乙酸型產(chǎn)甲烷菌,只能利用乙酸作為碳源產(chǎn)甲烷[47],該屬在加藻組的相對豐度相較對照組提高了1.27倍;第二優(yōu)勢屬甲烷桿菌屬是氫營養(yǎng)型產(chǎn)甲烷菌,以H2作為電子供體產(chǎn)甲烷[25].加藻組和對照組乙酸營養(yǎng)型產(chǎn)甲烷菌(甲烷絲菌屬與馬澤氏甲烷八疊球菌屬())相對豐度最高為51.6%和40.8%,其中馬澤氏甲烷八疊球菌屬既能利用乙酸也能利用甲醇和甲胺化合物進行產(chǎn)甲烷[48];加藻組和對照組氫營養(yǎng)型產(chǎn)甲烷菌包含甲烷絲菌屬,嗜氫型甲烷菌,甲烷胞菌目(),甲烷微菌(),甲烷袋狀菌屬(),亨氏甲烷螺菌()[49-52]約占34.8%和36.71%.加藻組除上述兩種類型產(chǎn)甲烷菌外其他產(chǎn)甲烷菌相對豐度都略微降低,只有乙酸型產(chǎn)甲烷菌大幅度增加,表明加藻組產(chǎn)甲烷菌的增加主要依賴乙酸型產(chǎn)甲烷菌,其他屬的產(chǎn)甲烷菌可能在絕對豐度上有略微變化,但因乙酸型產(chǎn)甲烷菌的豐度增幅更大使其它屬表現(xiàn)出在相對豐度上減小.

藻源性有機質(zhì)易被微生物分解利用,尤其巢湖的優(yōu)勢種微囊藻(spp.)[53-54].分解時DO快速下降[55],同時伴隨著電子受體(NO3-,SO42-)[56-57]的降低,抑制反硝化菌和硫酸鹽還原菌對有機質(zhì)的利用,促進產(chǎn)甲烷菌的生存.藻源性有機質(zhì)厭氧分解促進產(chǎn)甲烷菌生長繁殖[58],當藍藻完全分解時會產(chǎn)生大量乙酸[59],有利于乙酸營養(yǎng)型途徑,而促進其豐度的增加.本研究中藻源性有機質(zhì)對乙酸型產(chǎn)甲烷菌在沉積物中的促進作用明顯,表明藻源性有機質(zhì)分解產(chǎn)物含大量乙酸,促進乙酸型產(chǎn)甲烷的進行.巢湖采集的原位沉積物中乙酸型及氫營養(yǎng)型產(chǎn)甲烷菌豐度最高(圖7),在額外添加藻源性有機質(zhì)后,優(yōu)勢種群快速利用有機質(zhì)生長繁殖,使得其豐度增加.

在波蘭富營養(yǎng)化水庫研究中發(fā)現(xiàn)表層沉積物乙酸營養(yǎng)途徑占到43%～82%,尤其在夏季乙酸營養(yǎng)型途徑占比更高[60].另外,在德國康斯坦茨湖的培養(yǎng)實驗也發(fā)現(xiàn),藻類提高了底物中乙酸的含量促進乙酸型產(chǎn)甲烷途徑的進行[18].雖然,在太湖藍藻嚴重堆積區(qū)域,藍藻分解會產(chǎn)生甲基硫醚物質(zhì)促進甲基營養(yǎng)型產(chǎn)甲烷,但并沒有取代淡水中優(yōu)勢氫營養(yǎng)型產(chǎn)甲烷[19].Zhu等[61]研究表明藍藻會造成水體中乙酸和甲基物質(zhì)含量不斷升高,甲基型產(chǎn)甲烷菌和乙酸型相對豐度都有所提高.而本實驗在整個培養(yǎng)過程中,僅乙酸型產(chǎn)甲烷菌相對豐度變化明顯.綜上,底物的多樣性影響產(chǎn)甲烷群落結(jié)構(gòu),產(chǎn)甲烷菌可利用的基質(zhì)來源于周圍的分解細菌代謝的產(chǎn)物,所以產(chǎn)甲烷菌群落結(jié)構(gòu)以及產(chǎn)甲烷途徑與周圍的分解細菌群落結(jié)構(gòu)密切相關(guān).

4 結(jié)論

4.1 藻源性有機質(zhì)降解過程中水體DO以及間隙水中SO42-含量降低,有利于表層產(chǎn)甲烷活動進行,間接促進了下層沉積物產(chǎn)甲烷.

4.2 藻源性有機物促進沉積物產(chǎn)甲烷菌活動,使得上覆水CH4含量及釋放量快速升高,加藻組甲烷釋放量最高可達(98.9±14.3)μmol/(m2×h),提升總CO2釋放當量及CH4的貢獻作用.

4.3 藻源性有機質(zhì)沉降將表層沉積物中基因數(shù)量提高了37.5%,豐度的增加主要因為乙酸型產(chǎn)甲烷菌(甲烷絲菌屬)的增加.

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Enhancement and mechanism of algal-derived organic matter deposition on lake sediment methane release.

ZHANG Nan1,2, HE Kai2,3, ZHONG Ji-cheng2, AN Yan-fei3, YUAN He-zhong1, ZHANG Lei2*

(1.School of Environmental Science and Engineering, Nanjing University of Information Science & Technology, Nanjing 210044, China；2.State Key Laboratory of Lake Science and Environment, Nanjing Institute of Geography and Limnology, Chinese Academy of Sciences, Nanjing 210008, China；3.School of Resources and Environmental Engineering, Anhui University, Hefei 230601, China)., 2023,43(12)：6641～6650

Lakes are important natural sources of methane (CH4) release. A well-known view is that frequent cyanobacterial blooms promote CH4release in freshwater lakes, but the specific driving process and mechanism are still unclear. In this study, the driving process and effect of algal-derived organic matter deposition on CH4release were simulated and studied by using water and sediment samples from Lake Chaohu. The results showed that the CH4release flux in the group with the treatment of additional added algae was (1.59±0.51)～(98.89±14.30) μmol/(m2·h)) significantly higher than that in the control group (0.02±0.016)～(1.37±0.44) μmol/(m2·h)) (<0.001). During the simulating experiment, the degradation of algal-derived organic matter provided a large number of substrates for methane production while the reduction of electron acceptors such as dissolved oxygen and sulfate created an environment conducive to methanogens, which resulted in a massive proliferation of methanogens, especially acetic acid trophic methanogens. Our study clearly demonstrated that the large input of algal-derived organic matter increased the methane production rate and methane release flux of lake sediments with acetic acid trophic methanogens playing an important role.

emission flux；methanogens；greenhouse gases；carbon dioxide；lake；algal bloom

X511

A

1000-6923(2023)12-6641-10

張 楠,何 凱,鐘繼承,等.藻源性有機質(zhì)沉降對沉積物甲烷釋放促進作用 [J]. 中國環(huán)境科學, 2023,43(12):6641-6650.

Zhang N, He K, Zhong J C, et al. Enhancement and mechanism of algal-derived organic matter deposition on lake sediment methane release [J]. China Environmental Science, 2023,43(12):6641-6650.

2023-04-16

國家自然科學基金資助項目(42177228,41771122);自然資源部國土(耕地)生態(tài)監(jiān)測與修復工程技術(shù)創(chuàng)新中心開放課題(GTST2021- 007)

* 責任作者, 副研究員, leizhang@niglas.ac.cn

本研究在采樣,實驗過程中得到等王洪偉,胡曉康,朱利釗等同學的支持與幫助,在此一并感謝.

張 楠(1999-),女,湖北襄陽人,南京信息工程大學資源與環(huán)境碩士研究生,研究方向為環(huán)境生物地球化學.發(fā)表論文1篇. 1967761818@qq.com.