發(fā)布時(shí)間：2017-12-31 16:33

  本文關(guān)鍵詞：微囊藻毒素-LR對(duì)降解菌Pseudomonas putida細(xì)胞影響的研究　出處：《暨南大學(xué)》2015年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 微囊藻毒素-LR 惡臭假單胞菌 細(xì)胞表面特性 細(xì)胞活性 毒性

【摘要】：微囊藻毒素-LR(MC-LR)是富營(yíng)養(yǎng)化污染的次生代謝產(chǎn)物,具有很強(qiáng)烈的生物毒性。本文利用前期分離馴化得到的MC-LR降解菌惡臭假單胞菌(Pseudomonas putida)為研究對(duì)象,考察了MC-LR對(duì)菌體細(xì)胞表面特性的影響,并探討其對(duì)細(xì)胞活性、菌體自身代謝和調(diào)節(jié)能力以及菌體形態(tài)的作用,對(duì)MC-LR的微生物毒性作用進(jìn)行了初步的探索。P.putida細(xì)胞質(zhì)膜通透性因?yàn)镸C-LR的濃度而發(fā)生改變,相對(duì)較高濃度的MC-LR會(huì)使菌體細(xì)胞質(zhì)膜通透性持續(xù)上升。高濃度(2.5 mg·L-1)MC-LR作用下,P.putida細(xì)胞出現(xiàn)破裂死亡情況,導(dǎo)致胞內(nèi)物質(zhì)流出,細(xì)胞完整性遭到破壞,同時(shí)影響離子在細(xì)胞膜兩側(cè)的分布,發(fā)生去極化現(xiàn)象,使細(xì)胞膜電位降低。MC-LR能夠影響P.putida的生物活性,加速菌體細(xì)胞的死亡。Biolog碳源實(shí)驗(yàn)表明,MC-LR作用后的P.putida對(duì)碳源的整體利用能力明顯弱于對(duì)照菌體,其毒害作用使得P.putida代謝能力減弱,活性降低。流式細(xì)胞實(shí)驗(yàn)檢測(cè)發(fā)現(xiàn),在MC-LR毒性作用下,菌體細(xì)胞活性下降并發(fā)生凋亡,2.5 mg·L-1 MC-LR作用5 d,其死亡率比未添加MC-LR的對(duì)照體系增加了近30%,體系中菌體生物量持續(xù)下降。實(shí)驗(yàn)結(jié)果表明,MC-LR能夠影響菌體離子代謝及可溶性蛋白等物質(zhì)的分泌。當(dāng)MC-LR質(zhì)量濃度由0增大到2.5 mg·L-1時(shí),能促進(jìn)菌體分泌和釋放可溶性糖及Na+、Cl-等離子。通過(guò)對(duì)P.putida SOD酶和ATPase酶活性測(cè)定可知,低濃度的MC-LR能夠誘導(dǎo)酶活性的增強(qiáng);高濃度的MC-LR則對(duì)菌體抗氧化和新陳代謝能力都有破壞性的影響,并激發(fā)產(chǎn)生活性氧自由基,使P.putida遭受氧化脅迫,對(duì)SOD酶和ATPase酶有強(qiáng)烈的抑制作用,使得酶活性總體水平不斷降低。掃描電鏡和原子力顯微鏡觀察表明,在MC-LR作用下,菌體細(xì)胞的微觀形貌發(fā)生了改變,細(xì)胞褶皺加深,在2.5 mg·L-1 MC-LR存在下培養(yǎng)5 d后,大部分細(xì)胞破裂,細(xì)胞內(nèi)含物流出,細(xì)胞結(jié)構(gòu)損壞嚴(yán)重。微觀形貌觀察實(shí)驗(yàn)也驗(yàn)證了MC-LR對(duì)菌體細(xì)胞的影響,即MC-LR對(duì)P.putida具有一定的毒害作用,且MC-LR濃度越大,作用時(shí)間越長(zhǎng),細(xì)胞損傷越嚴(yán)重。
[Abstract]:Microcystin -LR (MC-LR) is a secondary metabolite of eutrophication, has very strong biological toxicity. MC-LR degrading bacterium Pseudomonas putida by early isolation and domestication of the (Pseudomonas putida) as the research object, the effects of MC-LR on cell surface properties, and its effect on the cell activity. The cell metabolism and regulation ability and cell morphology, microbial toxicity to MC-LR made a preliminary exploration of.P.putida cell membrane permeability for MC-LR concentration change, relatively high concentration of MC-LR can make the cell membrane permeability increased. The high concentration (2.5 mg L-1) MC-LR under the action of rupture of death P.putida cells, resulting in intracellular substance out of cell integrity is destroyed, while the impact of ion distribution in the cell membrane, occurrence of depolarization phenomenon, cell membrane Potential decreased.MC-LR can affect the biological activity of P.putida,.Biolog showed that accelerated the death of carbon source experimental cell, the overall utilization ability of MC-LR after the action of P.putida on carbon source was weaker than that in the control cells, the toxicity of the metabolic capacity of P.putida decreased, the activity reduced. Flow cytometry assay showed that the toxicity of MC-LR. Cell activity decreased and apoptosis, 2.5 mg - L-1 MC-LR 5 d, the mortality rate increased by nearly 30% compared with the control system without the addition of MC-LR, the biomass decreased continuously in the system. The experimental results show that MC-LR can affect cell metabolism and ion secretion of soluble protein substances. When MC-LR concentration increases from 0 2.5 mg - L-1, can promote the secretion and release of soluble sugar and Na+, Cl-. Through the determination of the plasma cell of P.putida SOD enzyme and ATPase enzyme activity, low concentration of MC-LR can induce The enhanced enzyme activity; high concentrations of MC-LR on antioxidant ability of bacteria and The new supersedes the old. with damaging effects, and stimulate the production of oxygen free radicals, P.putida suffered oxidative stress, has a strong inhibitory effect on SOD enzyme and ATPase enzyme, the overall level of enzyme activity decreased rapidly. Scanning electron microscopy and atomic force microscope observation showed that in the presence of MC-LR changed cell morphology, cell fold deepened, in 2.5 mg L-1 the presence of MC-LR after 5 d culture, most cells rupture, cells containing the logistics, cell structure damaged seriously. The microscopic morphology observation experiments verified the effect of MC-LR on the cell. MC-LR has a toxic effect on P.putida and MC-LR, the greater the concentration, the longer the time, the cell damage was more serious.

【學(xué)位授予單位】：暨南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類號(hào)】：X172

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本文編號(hào)：1360581