本文選題：大腸桿菌　切入點(diǎn)：生物被膜　出處：《山東中醫(yī)藥大學(xué)》2010年碩士論文　論文類(lèi)型：學(xué)位論文

【摘要】：目的：通過(guò)分光光度法及麥?zhǔn)媳葷岬确椒▽?duì)大腸桿菌菌液濃度進(jìn)行測(cè)定,了解大腸桿菌不同生長(zhǎng)時(shí)期生長(zhǎng)曲線的特征,分析不同濃度的鐵、鋅離子對(duì)大腸桿菌生長(zhǎng)狀況的影響,為微量元素用于大腸桿菌的體外培養(yǎng)和生物被膜的形成提供參考數(shù)值；通過(guò)藥敏試驗(yàn)了解不同濃度的鐵、鋅離子對(duì)阿奇霉素抗菌活性的影響；通過(guò)生物被膜銀染色觀察、細(xì)菌計(jì)數(shù)等方法并以硅膠為載體建立大腸桿菌生物被膜的體外模型,了解大腸桿菌生物被膜的形成,并研究生物被膜的形成鑒定標(biāo)準(zhǔn)及阿奇霉素與黃連素的聯(lián)合用藥效果,為阿奇霉素和黃連素聯(lián)合應(yīng)用于大腸桿菌生物被膜感染的治療提供依據(jù)。 方法：1分別測(cè)量大腸桿菌不同生長(zhǎng)時(shí)期的菌液的OD值,以時(shí)間為橫坐標(biāo),以O(shè)D值為縱坐標(biāo)建立坐標(biāo)系,繪制出大腸桿菌生長(zhǎng)曲線；給MH培養(yǎng)基中加入不同濃度的鋅、鐵離子,混勻接種之后,利用分光光度法測(cè)定的OD值反應(yīng)大腸桿菌菌液濃度,建立時(shí)間---OD值坐標(biāo)系,通過(guò)曲線的走向來(lái)分析鐵、鋅離子對(duì)大腸桿菌生長(zhǎng)的影響；2通過(guò)紙片擴(kuò)散法的藥敏試驗(yàn),比較不同鐵離子和鋅離子濃度下阿奇霉素對(duì)大腸桿菌抑菌圈直徑的影響；3以硅膠為載體建立大腸桿菌生物被膜模型,用細(xì)菌計(jì)數(shù)法比較阿奇霉素與黃連素的聯(lián)合用藥和單獨(dú)用藥的藥效。 結(jié)果：1大腸桿菌的生長(zhǎng)曲線基本符合微生物的生長(zhǎng)特性,并且呈現(xiàn)二次生長(zhǎng)曲線的特征。從生長(zhǎng)曲線圖來(lái)看,加入鐵離子濃度分別為11,17,27,32μmol/L,各組加入鐵離子的大腸桿菌菌液的OD值都比對(duì)照組高,其中27μmol/L鐵離子濃度下大腸桿菌菌液的OD值顯著高于對(duì)照組,但當(dāng)鐵離子濃度達(dá)到32μmol/L時(shí),其OD值要比鐵離子濃度為27μmol/L時(shí)的OD值要小；即通過(guò)試驗(yàn)比較,在設(shè)計(jì)的不同鐵離子濃度中,鐵離子濃度為27μmol/L時(shí)大腸桿菌菌液的OD值最大。鋅離子濃度分別為6,12,18,52μmol/L時(shí),與對(duì)照組相比,各濃度鋅離子組大腸桿菌菌液的OD值都低于對(duì)照組；但鋅離子濃度在12-18μmol/L濃度范圍內(nèi)菌液OD值幾乎沒(méi)有明顯改變,數(shù)值比較穩(wěn)定。2鐵離子濃度分別為11,17,27,32μmol/L的培養(yǎng)基中大腸桿菌的抑菌圈直徑進(jìn)一步證明了鐵離子對(duì)大腸桿菌生長(zhǎng)的促進(jìn)作用；同樣鋅離子濃度為6,12,18,52μmol/L的培養(yǎng)基中抑菌圈直徑也再次說(shuō)明鋅離子對(duì)大腸桿菌生長(zhǎng)有抑制作用；各濃度鐵、鋅離子對(duì)阿奇霉素的抑菌活性均無(wú)明顯影響。3利用銀染色方法鑒定,成功建立大腸桿菌生物被膜模型,配制0.15,0.3,0.6 mg/L的阿奇霉素溶液與0.2,0.4,0.8mg/L的黃連素溶液,通過(guò)單獨(dú)用藥和兩兩聯(lián)合用藥后的大腸桿菌細(xì)菌計(jì)數(shù)值分析比較,當(dāng)黃連素濃度達(dá)到0.8mg/L才出現(xiàn)明顯的協(xié)同抑菌作用,且阿奇霉素的協(xié)同作用與其濃度無(wú)關(guān)。 結(jié)論：鐵離子對(duì)大腸桿菌的生長(zhǎng)有一定的促進(jìn)作用,但與其濃度有很大關(guān)系。低濃度時(shí)幾乎無(wú)促進(jìn)作用,隨著鐵離子濃度的增加其作用開(kāi)始增強(qiáng),但到達(dá)一定濃度后其作用不但不會(huì)增強(qiáng)反而有所減弱。鋅離子在正常血清濃度11-23μmol/L范圍內(nèi)對(duì)大腸桿菌的生長(zhǎng)有抑制作用,在機(jī)體缺乏鋅離子的情況下,較為容易感染大腸桿菌。鋅離子在12-18μmol/L濃度范圍內(nèi)作用沒(méi)有明顯改變,比較穩(wěn)定；隨著鋅離子濃度的增加,到體內(nèi)最高濃度的兩倍多時(shí),抑制作用才比較顯著。不同濃度的鐵、鋅離子對(duì)阿奇霉素的抑菌活性無(wú)明顯影響。當(dāng)黃連素濃度達(dá)到0.8mg/L時(shí),阿奇霉素與黃連素聯(lián)合用藥對(duì)生物被膜內(nèi)大腸桿菌的抑制具有協(xié)同效應(yīng)。
[Abstract]:Objective: to Escherichia coli bacteria concentration were determined by spectrophotometry and Maxwell turbidimetric method, to understand the characteristics of Escherichia coli in different growth stages of growth curve, analysis of different concentrations of iron, effect of zinc ions on the growth of Escherichia coli, for trace elements for in vitro culture and biological Escherichia coli is provided reference values of film formation; understand the different concentrations of iron through drug sensitivity test, effect of zinc ions on the antibacterial activity of Azithromycin; biofilm by silver staining, the bacterial count method with silica gel as the carrier to establish an in vitro model of Escherichia coli biofilm, understanding of biofilm formation of Escherichia coli, and research biofilm formation and identification standard of azithromycin and berberine combination effect, provide for the treatment of azithromycin and berberine on Escherichia coli biofilm infection Basis.
Methods: 1 od bacteria Escherichia coli were measured at different growth periods, with time as the abscissa, the OD value of the vertical axis coordinate system, draw the growth curve of Escherichia coli; to MH medium added different concentration of zinc, iron, mixing after inoculation, using Escherichia coli od reaction the concentration of spectrophotometry, the establishment of time ---OD coordinates, through the curve to analysis of iron, effect of zinc ions on the growth of Escherichia coli; drug sensitivity test of 2 by disk diffusion method, comparing the different effects of iron ion and zinc ion concentration of azithromycin on Escherichia coli inhibition zone diameter; 3 with silica gel as a vector of Escherichia coli biofilm model, comparison with the bacterial count of azithromycin in combination with berberine and individual drug efficacy.
Results: 1 Escherichia coli growth curve accords with the characteristics of microbial growth, and the growth curve showed two characteristics. From the growth curve, adding iron ion concentration were 11,17,27,32 mol/L, each adding iron ions of Escherichia coli bacteria OD values than the control group, the OD value of 27 mol/L of iron ion the concentration of Escherichia coli was significantly higher than the control group, but when the iron ion concentration reached 32 mol/L, the OD value than the iron ion concentration was 27 mol/L smaller; by comparison, in different concentration of iron ion in the design, the iron ion concentration for Escherichia coli bacteria od the maximum of 27 mol/L. The zinc ion concentration was 6,12,18,52 mol/L, compared with the control group, the OD value of the concentration of zinc ion group of Escherichia coli bacteria were lower than the control group; but the zinc ion concentration at 12-18 mol/L concentration range of bacteria OD The value of almost did not change significantly,.2 values are stable iron ion concentration respectively. The antibacterial circle diameter medium 11,17,27,32 of Escherichia coli mol/L in further proved that the promoting effect of iron ion on the growth of Escherichia coli; the same concentration of zinc ions is bacteriostatic medium 6,12,18,52 mol/L in diameter also showed that zinc ions inhibited again on the growth of Escherichia coli; the concentration of iron and zinc ions on the antibacterial activity of azithromycin were not significantly influenced by.3 silver staining, successfully established Escherichia coli biofilm model, berberine solution of azithromycin solution with 0.2,0.4,0.8mg/L preparation of 0.15,0.3,0.6 mg/ L, by E. coli bacteria alone or the combination of the 22 meter comparative analysis numerical, when berberine concentration of 0.8mg/L apparent only synergistic inhibitory effect, and the synergistic effect of azithromycin and Concentration has nothing to do with it.
Conclusion: there is a certain growth promoting effect of iron ions on Escherichia coli, but a great relationship with its concentration. Low concentration of almost no promotion, with the iron ion concentration started to increase its role, but its role to reach a certain concentration not only will not strengthen but weaken. Some zinc ions on Escherichia coli growth in the normal serum concentration of 11-23 mol/L in the range of inhibition, lack of zinc in the body of the ion, the more susceptible to Escherichia coli. Zinc ions in 12-18 mol/L concentration range function did not change significantly, relatively stable; with the increase of the concentration of zinc ions, to two times for a long time in the highest concentration, inhibition was more significant. Different concentrations of iron, no obvious effect of zinc ions on the antibacterial activity of azithromycin. When berberine concentration of 0.8mg/L, berberine Azithromycin combined with the biofilm The inhibition of internal Escherichia coli has a synergistic effect.

【學(xué)位授予單位】：山東中醫(yī)藥大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R378

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