【摘要】：目前,我國最新頒布的飲用水相關(guān)標(biāo)準(zhǔn)有六項微生物指標(biāo),無法全面地反映水中病原微生物的污染狀況,還缺少易引起腹泄、腸胃炎等腸道疾病且一般氯消毒方法很難全部殺死的病毒類指標(biāo)。因此,有必要開展病毒類病原微生物的研究和監(jiān)測工作。本論文首次建立了PMA-PCR方法用于定性檢測活性的輪狀病毒、星狀病毒和腺病毒,同時也首次建立了PMA-qPCR方法用于定量檢測活性的輪狀病毒,并將這些方法應(yīng)用于篩查重點流域典型水源地相關(guān)病毒的污染情況。實驗結(jié)果顯示利用PMA-PCR法檢測加熱處理和未加熱處理的陽性樣本時表現(xiàn)出顯著差異,表明該方法能夠區(qū)分活性和非活性的輪狀病毒、星狀病毒、腺病毒。實驗結(jié)果也顯示利用PMA-qPCR法檢測加熱處理和未加熱處理的輪狀病毒陽性樣本時表現(xiàn)出顯著差異,表明該方法能夠區(qū)分活性和非活性的輪狀病毒。這些研究結(jié)果證明了PMA-PCR法、PMA-qPCR法的可靠性。在重點流域典型水源地水樣的實際應(yīng)用檢測中,遼河流域的QH采樣點有活性輪狀病毒檢出。經(jīng)PMA-qPCR法定量測定后,該采樣點活性輪狀病毒的濃度為1.24×105Copies/L研究結(jié)果充分證明了PMA-PCR法能夠成功地應(yīng)用于三種活性病毒的檢測,也充分證明了PMA-qPCR法能夠成功地應(yīng)用于輪狀病毒的定量檢測。 另一方面,為了減輕城市用水壓力和美化環(huán)境,大部分北京地表水都接納了污水處理廠的出水。目前,有研究報道中國的14個污水處理廠的活性污泥樣本存在核心菌群和獨有菌群,而接納污水處理廠出水的北京地表水的細(xì)菌群落情況仍未見有報道,因此有必要調(diào)查分析北京地表水細(xì)菌群落的情況。本研究首次利用焦磷酸測序技術(shù)分析5個北京地表水水樣的細(xì)菌多樣性,作為初步監(jiān)測復(fù)雜環(huán)境中細(xì)菌群落的基本參考。經(jīng)過測序得到60810條優(yōu)化序列,平均長度為400bp,并使用Mothur軟件對優(yōu)化序列進(jìn)行歸類。研究結(jié)果充分展示了從門水平到屬水平的地表水菌群組成情況,其中相對豐度較高的門依次為變形菌門(Proteobacteria),其相對豐度分別占總OTU數(shù)目的20.28%～67.89%；放線菌門(Actinobacteria),分別占總OTU數(shù)目的19.78%～53.59%；擬桿菌(Bacteroidetes),分別占總OTU數(shù)目的3.85%～16.14%：另外,還有疣微菌門(Verrucomicrobia)和藍(lán)藻門(Cyanobacteria),分別占總OTU數(shù)目的0.55%-7.25%和0.68%-2.60%等。這5個門分別占總OTU數(shù)目91.70%-98.50%。研究結(jié)果還顯示了5個北京地表水樣本中存在核心菌群,即門水平上有13個核心菌群,綱水平上由23個核心菌群,目水平上有32個核心菌群,科水平上有40個核心菌群,屬水平上有40個核心菌群。還證明了5個處于不同地理位置的地表水樣中存在相對豐度0.5%的獨有菌群,即目水平上有19個獨有菌群,科水平上有49個獨有菌群,屬水平上有100個獨有菌群。通過Weighted Unifrac PCA分析表明處于地表水樣本1和樣本2的細(xì)菌群落結(jié)構(gòu)更相似。此外,研究結(jié)果還說明了污水處理廠活性污泥樣本的細(xì)菌群落多樣性對北京地表水的細(xì)菌群落多樣性有一定的影響。
[Abstract]:At present, the newly-issued drinking water-related standard in China has six microbial indicators, which can not fully reflect the pollution condition of pathogenic microorganisms in water, but also lacks the virus type indexes which are easy to cause intestinal diseases such as abdominal diarrhea and gastroenteritis, and the general chlorine disinfection method is difficult to kill all. Therefore, it is necessary to carry out the research and monitoring of the virus-like pathogenic microorganisms. The first time in this paper, a PMA-PCR method was established for the qualitative detection of the active rotavirus, star-like virus and adenovirus, and the PMA-qPCR method was also first established for the quantitative detection of the active rotavirus, and these methods were used to screen the contamination of the typical water source-related virus in the key watershed. The results show that the method can distinguish the active and inactive rotavirus, star-like virus and adenovirus by using the PMA-PCR method to detect the positive samples of the heat treatment and the unheated treatment. The results of the experiments also showed a significant difference in the detection of the positive samples of rotavirus by the PMA-qPCR method and the untreated rotavirus, indicating that the method was able to distinguish between active and inactive rotavirus. The results of these studies demonstrated the reliability of PMA-PCR and PMA-qPCR. In the actual application of water sample in the typical water source of the key drainage area, the QH sampling point of the Liaohe river basin is detected by the active rotavirus. The results of PMA-qPCR method show that the concentration of the active rotavirus is 1. 24-105Copies/ L. The results of the study show that the PMA-PCR method can be successfully applied to the detection of three kinds of active viruses, and it is also proved that the PMA-qPCR method can be successfully applied to the quantitative detection of rotavirus. On the other hand, in order to reduce the urban water pressure and the beautifying environment, most of the surface water in Beijing has accepted the sewage treatment plant. At present, there is a study on the presence of the core bacteria and the unique flora in the activated sludge samples of the 14 sewage treatment plants in China, while the bacterial community of the Beijing surface water receiving the effluent from the WWTP is still not reported. Therefore, it is necessary to investigate and analyze the bacterial community of surface water in Beijing. In this study, the bacterial diversity of five Beijing surface water samples was analyzed by pyrosequencing technology for the first time, which was used as the basis for preliminary monitoring of the bacterial community in complex environment. Reference. The 60810 optimized sequence was obtained by sequencing, with an average length of 400bp, and the optimized sequence was performed using the Moothur software. The results of the study fully show the composition of the surface water species from the level of the door to the genus. The relative abundance of the relative abundance is 20. 28% to 67. 89% of the total OTU, and 19. 78% to 53. 5, respectively, of the total OTU. Bacteroides (9%) and Bacteroides, respectively, accounted for 3.85% ~ 16.14% of the total OTU. In addition, the number of total OTU was 0.55%-7.25% and 0.68%-2.60, respectively.%, etc. The 5 doors account for the total OTU number of 91.70% -98,5, respectively. The results of the study also show that there are 13 core strains in the five Beijing surface water samples, i.e., there are 13 core bacteria in the level of the phylum, and there are 32 core bacteria in the level. There are 40 core bacteria in the family level, and there are 40 cores in the level. There are also 5 unique bacterial groups with relative abundance of 0. 5% in surface water samples with different geographical locations. The bacterial population of surface water samples 1 and 2 was analyzed by the weighted Unifrac PCA analysis. Similarly, the results of the study also show that the diversity of the bacterial community in the activated sludge sample of the sewage treatment plant has a certain effect on the bacterial community diversity of the surface water in Beijing.
【學(xué)位授予單位】：北京林業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2013
【分類號】：R123.1;X172

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本文編號：2353133