多氯聯(lián)苯高效脫氯培養(yǎng)物的富集及特征研究
發(fā)布時(shí)間：2018-08-04 13:52
【摘要】：多氯聯(lián)苯(Polychlorinated biphenyls,PCBs)是典型的環(huán)境持久性有機(jī)污染物。PCBs污染的微生物修復(fù)是一種有前景的修復(fù)技術(shù),高氯代PCBs厭氧脫氯是其中的關(guān)鍵。PCBs厭氧脫氯菌極難分離,對富集厭氧脫氯培養(yǎng)物的研究有助于我們更好地富集穩(wěn)定的混合培養(yǎng)物、分離高效PCBs脫氯菌。本文以水稻土為富集源,以典型高氯代PCBs混合物Aroclor1260為電子受體,添加乳酸鈉為碳源和電子供體,進(jìn)行富集培養(yǎng),考察培養(yǎng)過程中PCBs脫氯特征變化、PCBs脫氯菌和脫氯基因變化及微生物群落結(jié)構(gòu)的演替,并對高效穩(wěn)定的富集脫氯培養(yǎng)物PSCJ-V進(jìn)行表征。六代富集培養(yǎng)物的培養(yǎng)過程可分為初始、過渡和穩(wěn)定三個階段。主要研究結(jié)果如下:(1)隨著富集過程的進(jìn)行,培養(yǎng)物中的土壤顆粒逐漸減少至無顆粒物,脫氯延滯期和脫氯周期逐漸縮短,氯原子去除速率逐步加快,脫氯程度與脫氯途徑基本無變化;脫氯效果在PSCJ-V中達(dá)到最優(yōu)。與此同時(shí),PCBs脫氯菌Dehalococcoides和脫氯基因A4、A5、Prd11隨著多次的傳代培養(yǎng)逐步增加,且四者有相似的變化,A1、Prd8基因隨著多次的傳代培養(yǎng)逐漸減少,被逐步淘汰。(2)經(jīng)過長期的富集培養(yǎng),本文得到一個高效的Aroclor1260脫氯培養(yǎng)物PSCJ-V。該培養(yǎng)物可以在40天左右的周期內(nèi),以5.14～6.01μmol·L-1·day-1的速率,按途徑N和途徑LP共同對Aroclor1260脫氯,得到的產(chǎn)物單位聯(lián)苯上僅有4.3～5.2個氯原子。該培養(yǎng)物在脫氯程度、脫氯周期、氯原子去除速率方面均優(yōu)于多數(shù)培養(yǎng)物,是目前最高效的Aroclor1260脫氯培養(yǎng)物之一。(2)培養(yǎng)物PSCJ-V中存在與Dehalococcoides mccartyi CG4和Dehalococcoides mccartyi195親緣關(guān)系極近的PCBs脫氯菌PSCJ-Dhc,其細(xì)胞濃度達(dá)到3.98×107 cells/mL左右,是目前脫氯菌細(xì)胞濃度最高的Aroclor1260脫氯培養(yǎng)物之一,僅次于JN培養(yǎng)物。該培養(yǎng)物中某些脫氯菌攜帶了與pcbA5基因序列相似度為98.7%的假定脫氯基因pcbPSCJ,兩者蛋白質(zhì)僅有11個氨基酸的差異。(3)培養(yǎng)物PCSJ-V中古菌豐度為30.45%,以產(chǎn)甲烷菌為主;細(xì)菌豐度為69.55%,以產(chǎn)酸發(fā)酵菌為主;主要存在以下菌屬:脫氯菌Dehalococcoides,產(chǎn)氫產(chǎn)乙酸菌Candidatus Cloacamonas、Smithella、Aminivibrio、Thermovirga、Proteiniphilum、Syntrophomonas、Sphaerochaeta、Youngiibacter、Desulfovibrio、Methanosarcina,產(chǎn)類咕啉菌Methanobacterium、Methanoculleus、Methanothrix、Desulfovibrio、Methanosarcina,以及其他菌屬M(fèi)ethanomassiliicoccus。
[Abstract]:Polychlorinated biphenyls (Polychlorinated) is a typical environmental persistent organic pollutant. Microbial remediation of PCBs is a promising remediation technique. Anaerobic dechlorination of high chlorinated PCBs is the key. The study on the enrichment of anaerobic dechlorination culture is helpful for us to better enrich the stable mixed culture and isolate the high efficiency PCBs dechlorination bacteria. In this paper, paddy soil was used as enrichment source, Aroclor1260 as electron acceptor, sodium lactate as carbon source and electron donor. The characteristics of dechlorination in PCBs were investigated. The changes of dechlorination bacteria and dechlorination genes and the succession of microbial community structure were investigated. The highly efficient and stable enrichment dechlorination culture (PSCJ-V) was characterized. The culture process of six generations of enriched culture can be divided into three stages: initial stage, transition stage and stabilization stage. The main results are as follows: (1) with the enrichment process, soil particles in the culture gradually decreased to no particles, dechlorination delay period and dechlorination cycle gradually shortened, chlorine atom removal rate gradually accelerated. The dechlorination degree and dechlorination pathway had no change, and the dechlorination effect was optimal in PSCJ-V. At the same time, the number of Dehalococcoides and the dechlorination gene A4A5 / Prd11 gradually increased with the passage of several times, and the same changes were observed in the four groups, and the gene of A1 / Prd8 was gradually decreased and phased out with the repeated passage culture. (2) after a long period of enrichment culture, A high efficiency Aroclor1260 dechlorination culture PSCJ-V was obtained. The culture could dechlorinate Aroclor1260 at the rate of 5.14 ~ 6.01 渭 mol L ~ (-1) day-1 at the rate of 5.14 ~ 6.01 渭 mol 路L ~ (-1) day-1 for about 40 days, and only 4.3 ~ 5.2 chlorine atoms could be obtained on the unit biphenyls of the product according to pathway N and path LP. The dechlorination degree, dechlorination period and chlorine atom removal rate of the culture were better than that of most cultures. It is one of the most efficient Aroclor1260 dechlorination cultures at present. (2) there is PSCJ-Dhc, which is closely related to Dehalococcoides mccartyi CG4 and Dehalococcoides mccartyi195 in PSCJ-V, and its cell concentration is about 3.98 脳 10 ~ 7 cells/mL, which is one of the highest Aroclor1260 dechlorination cultures. Second only to JN culture. Some dechlorinated bacteria in the culture carried a hypothetical dechlorination gene pcbPSCJ whose similarity to pcbA5 gene sequence was 98.7%, and the difference between the two proteins was only 11 amino acids. (3) the abundance of ancient bacteria in PCSJ-V was 30.45%, mainly methanogenic bacteria. The bacterial abundance is 69.55, mainly acid-producing bacteria, mainly in the following genera: Dehalococcus coidesi, Candidatus Cloacamonas Smithellaa, Aminivibrilaa Thermovirgaa Proteiniphila Syntrophia sphaerochaeta Youngiibacterium Desulfoviochaeta, Methanobacterium-like bacteria Methanobacterium Methanothrix Desulfovibrio Methanoscinara, and other bacteria, Methanobacteriumum Methanobacteriumum method ocululus Methanothrix Desulfovibrio Methanoscinara.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：X592;X172
本文編號：2164087
[Abstract]:Polychlorinated biphenyls (Polychlorinated) is a typical environmental persistent organic pollutant. Microbial remediation of PCBs is a promising remediation technique. Anaerobic dechlorination of high chlorinated PCBs is the key. The study on the enrichment of anaerobic dechlorination culture is helpful for us to better enrich the stable mixed culture and isolate the high efficiency PCBs dechlorination bacteria. In this paper, paddy soil was used as enrichment source, Aroclor1260 as electron acceptor, sodium lactate as carbon source and electron donor. The characteristics of dechlorination in PCBs were investigated. The changes of dechlorination bacteria and dechlorination genes and the succession of microbial community structure were investigated. The highly efficient and stable enrichment dechlorination culture (PSCJ-V) was characterized. The culture process of six generations of enriched culture can be divided into three stages: initial stage, transition stage and stabilization stage. The main results are as follows: (1) with the enrichment process, soil particles in the culture gradually decreased to no particles, dechlorination delay period and dechlorination cycle gradually shortened, chlorine atom removal rate gradually accelerated. The dechlorination degree and dechlorination pathway had no change, and the dechlorination effect was optimal in PSCJ-V. At the same time, the number of Dehalococcoides and the dechlorination gene A4A5 / Prd11 gradually increased with the passage of several times, and the same changes were observed in the four groups, and the gene of A1 / Prd8 was gradually decreased and phased out with the repeated passage culture. (2) after a long period of enrichment culture, A high efficiency Aroclor1260 dechlorination culture PSCJ-V was obtained. The culture could dechlorinate Aroclor1260 at the rate of 5.14 ~ 6.01 渭 mol L ~ (-1) day-1 at the rate of 5.14 ~ 6.01 渭 mol 路L ~ (-1) day-1 for about 40 days, and only 4.3 ~ 5.2 chlorine atoms could be obtained on the unit biphenyls of the product according to pathway N and path LP. The dechlorination degree, dechlorination period and chlorine atom removal rate of the culture were better than that of most cultures. It is one of the most efficient Aroclor1260 dechlorination cultures at present. (2) there is PSCJ-Dhc, which is closely related to Dehalococcoides mccartyi CG4 and Dehalococcoides mccartyi195 in PSCJ-V, and its cell concentration is about 3.98 脳 10 ~ 7 cells/mL, which is one of the highest Aroclor1260 dechlorination cultures. Second only to JN culture. Some dechlorinated bacteria in the culture carried a hypothetical dechlorination gene pcbPSCJ whose similarity to pcbA5 gene sequence was 98.7%, and the difference between the two proteins was only 11 amino acids. (3) the abundance of ancient bacteria in PCSJ-V was 30.45%, mainly methanogenic bacteria. The bacterial abundance is 69.55, mainly acid-producing bacteria, mainly in the following genera: Dehalococcus coidesi, Candidatus Cloacamonas Smithellaa, Aminivibrilaa Thermovirgaa Proteiniphila Syntrophia sphaerochaeta Youngiibacterium Desulfoviochaeta, Methanobacterium-like bacteria Methanobacterium Methanothrix Desulfovibrio Methanoscinara, and other bacteria, Methanobacteriumum Methanobacteriumum method ocululus Methanothrix Desulfovibrio Methanoscinara.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：X592;X172
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