一株溶藻细菌的分离及其对铜绿微囊藻的溶解效果研究

发布时间：2018-08-17 14:11

【摘要】：随着全球经济的快速发展,各国普遍存在的水体富营养化现象日趋严重,藻类作为水生生态系统中重要的初级生产力,藻类水华的频繁爆发已成为严重危害生态平衡,影响人类生产生活的重大环境污染问题[1]。也是现阶段影响较大,危害范围遍及全世界每一个水域的世界性问题[2]。因此,对于藻类水华及其引起的灾害问题展开控制技术研究是极其迫切的。2015年,我国的115个湖泊中,处于富营养状态的湖泊有90个,占比高达78.3%[3],不但严重破坏了水生态平衡,同时水体富营养状态下,水华藻类的爆发使藻类代谢产生的藻毒素急剧增加,给人类健康和正常的生产生活带来巨大威胁[4]。溶藻细菌等微生物是水生态系统中的重要组成部分之一,还具有天然易得、经济、环境友好、作用明显等特点,在控制藻类生长,维持其数量平衡上起重要作用[5],也受到越来越多的关注[6-7]。研究溶藻微生物对水华藻类的去除机理,筛选复配出高效溶藻细菌,研发溶藻微生物药剂,对富营养化水体的预防和治理具有重要意义[8]。从本地富营养化水体中分离纯化出7株细菌,编号S1～S7,其中3株(S4、S_5、S7)能使铜绿微囊藻液黄化,最后选择溶藻效果最好,藻液黄化程度最显著的S_5细菌作为本文的研究菌株。对S_5菌株进行初步鉴定和毒性检测,然后将其作用于铜绿微囊藻,验证其溶藻效果,继而对其溶藻机理进行简单探究,最后将其应用于人工采集的天然富养化水体中。对实验数据进行总体归纳,得出以下几项结论。1、结合细菌形态观察、染色试验和生理生化试验等方法,对菌株S_5进行初步鉴定,依据《常见细菌系统鉴定手册》和《伯杰氏细菌鉴定手册》,初步鉴定S_5菌株为芽孢杆菌属,具体种属有待在分子水平上继续研究。2、使溶藻细菌S_5作用于蓝斑马鱼,对其进行生态毒理安全性检测,检测证实,溶藻细菌S_5对作用对象的生长无影响,且没有遗传毒性。3、将溶藻细菌S_5按3%、6%、9%和12%的体积比加入到铜绿微囊藻液中,结果表明,相比对照组,在菌液投加第7天时,9%组藻细胞数和叶绿素a含量分别降低82.65%和74.71%;第9天时,12%组两指标分别降低82.56%和77.14%。且S_5细菌对藻细胞数量和叶绿素a含量的影响大致相同,一定浓度范围内,菌液浓度越高,对铜绿微囊藻的溶解效果越明显。但综合考虑操作步骤及成本,建议实际处理中,使S_5菌液投加量为铜绿微囊藻液体积的9%,且第7天即可达明显的除藻效果。4、分别向铜绿微囊藻中加入不同方式处理得到的菌悬液、高温处理菌悬液、上清液、高温处理上清液、细菌液体培养基及S_5原菌液,探究S_5细菌的溶藻方式,实验证明S_5是通过释放热稳定的胞外物质来间接溶藻的。然后通过上清液的呈色反应对溶藻活性物质进行研究,结果表明,S_5释放的胞外物质为热稳定的糖类物质。5、向富营养状态的天然采集水体中投加9%(v/v)的S_5菌液,15天后分析藻类群落结构发现,加入S_5后水体藻类的总密度由1.75×106个/L下降到1.09×106个/L,藻类去除率达62.29%。进一步分析藻类的组成:水体优势种由最初单一且绝对优势的斜生栅藻变为由四尾栅藻、斜生栅藻和不定微囊藻共同构成优势种,三者共占藻类总量的56.61%,同时多样性指数和均匀度指数均呈上升趋势,而不加菌的反应池中两个指数均有所下降,且优势种种类不变。
[Abstract]:With the rapid development of the global economy, the eutrophication of water bodies is becoming more and more serious. As an important primary productivity in aquatic ecosystem, algae blooms have become a serious ecological balance and a major environmental pollution problem affecting human production and life. In 2015, 90 of the 115 lakes in China were in eutrophic state, accounting for 78.3%[3], which not only seriously destroyed the water ecological balance, but also the water body. Under the eutrophic condition, algae blooms increase algae toxins produced by algae metabolism dramatically and pose a great threat to human health and normal production and life [4].Algolytic bacteria and other microorganisms are one of the important components of the aquatic ecosystem, and also have the characteristics of natural, economic, environment-friendly and obvious role in controlling algae. Growth and quantity balance play an important role [5], but also attracted more and more attention [6-7].To study the removal mechanism of algae by algae-lysing microorganisms, to screen and compound high-efficient algae-lysing bacteria, and to develop algae-lysing microbial agents are of great significance for the prevention and control of eutrophic water bodies [8]. Seven strains of bacteria were purified and numbered S1-S7. Among them, three strains (S4, S 5, S7) could cause the chlorosis of Microcystis aeruginosa. Finally, the S_5 bacterium with the best algal lysis effect and the most significant algal chlorosis degree was selected as the research strain. The algae-dissolving mechanism was explored and applied to the naturally enriched water. The experimental data were summarized and the following conclusions were drawn. 1. Strain S_5 was preliminarily identified by combining the methods of bacterial morphology observation, dyeing test and physiological and biochemical test. The results were in accordance with < Handbook for Identification of Common Bacterial Systems > and < Berger's Bacteria Identification Manual >, preliminary identification of Bacillus sp. S_5 strain, specific species to be further studied at the molecular level. 2, the algae-lysing bacteria S_5 acting on blue zebrafish, its ecological toxicological safety testing, detection confirmed that algae-lysing bacteria S_5 has no effect on the growth of the target, and no genetic toxicity. 3, the algae-lysing bacteria will be fine. The results showed that the number of algae cells and the content of chlorophyll a in 9% group decreased by 82.65% and 74.71% respectively on the 7th day, and that in 12% group decreased by 82.56% and 77.14% respectively on the 9th day. But considering the operation steps and cost, it is suggested that the dosage of S_5 should be 9% of the liquid volume of Microcystis aeruginosa, and the algae removal effect could be obvious on the 7th day. The results showed that S_5 could dissolve algae indirectly by releasing heat-stable extracellular substances. Then the algae-dissolving active substances were studied by color reaction of supernatant. The results showed that the extracellular substance released by S_5 was a heat-stable carbohydrate.5, and 9% (v/v) S_5 bacterial solution was added to the natural eutrophic water. After 15 days, the algae community structure was analyzed. The total algae density decreased from 1.75 106/L to 1.09 6550 The dominant species in the water body changed from single and absolute dominant species to four-tailed species. The dominant species consisted of two kinds of algae, namely, four-tailed species, oblique species and adventitious microcystis, accounting for 56.61% of the total algae. Meanwhile, the diversity index and evenness index increased, while the two indexes in the non-bacterial reaction pool decreased, and the dominant species were found. Unchanged.
【学位授予单位】：辽宁大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：X52;X172

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