本文選題：金黃色葡萄球菌 + 鈦板��； 參考：《福建醫(yī)科大學(xué)》2015年碩士論文

【摘要】：目的通過(guò)在鈦板表面建立金黃色葡萄球菌細(xì)菌生物膜模型,在激光掃描共聚焦顯微鏡(CLSM)下觀察細(xì)菌生物膜(BBF)的結(jié)構(gòu)、中層及底層的活菌率;研究0.5%、0.25%濃度的聚維酮碘溶液對(duì)金黃色葡萄球菌BBF的體外殺菌效果。方法選取斜面保種的金黃色葡萄球菌(SA),接種于預(yù)先配制的胰蛋白胨大豆肉湯(TSB)中,37℃(5%CO2)的恒溫培養(yǎng)箱中培養(yǎng)16小時(shí)獲得菌液,通過(guò)比濁儀將菌液濃度調(diào)成1.0×106 CFU/ml備用。將15枚清潔、鈍化、消毒過(guò)的鈦板隨機(jī)分裝至3個(gè)25cm2細(xì)胞培養(yǎng)瓶,每個(gè)細(xì)胞培養(yǎng)瓶?jī)?nèi)5枚鈦板、1ml上述調(diào)配好的菌液、20ml TSB,置37℃(5%CO2)的恒溫培養(yǎng)箱中培養(yǎng)24小時(shí)形成成熟BBF。從以上3個(gè)細(xì)胞培養(yǎng)瓶中隨機(jī)取出10枚鈦板在避光環(huán)境下由SYTO9/PI組成的染色劑染色20分鐘,在CLSM下觀察BBF的結(jié)構(gòu)、活菌率,另外5枚在經(jīng)過(guò)漩渦振蕩器、超聲降解、離心機(jī)離心處理后接種至血平板培養(yǎng)基做細(xì)菌培養(yǎng)。根據(jù)上述建模方法,在45枚鈦板表面建立金黃色葡萄球菌BBF模型,取出后用PBS沖洗3次,將45枚鈦板隨機(jī)分為3組,每組15枚,其中兩組分別置于0.5%和0.25%的聚維酮碘溶液中作用5分鐘,通過(guò)由硫代硫酸鈉、吐溫80、卵磷脂組成的中和劑將鈦板表面的碘殘留去除,這兩組中的每組其中10枚鈦板通過(guò)上述染色方法染色20分鐘后在CLSM下觀察BBF的結(jié)構(gòu)、活菌率的變化,另外5枚在經(jīng)過(guò)漩渦振蕩器、超聲清洗、離心機(jī)離心處理后接種至血平板培養(yǎng)基做細(xì)菌培養(yǎng)。另外一組其中10枚鈦板作為對(duì)照組,剩余5枚在經(jīng)過(guò)漩渦振蕩器、超聲清洗、離心機(jī)離心、稀釋處理后接種至血平板培養(yǎng)基做細(xì)菌培養(yǎng)。結(jié)果①BBF在CLSM下呈高度組織化的群體結(jié)構(gòu)。BBF在CLSM下的圖像由橙色或橘黃色熒光、綠色熒光、紅色熒光、暗性區(qū)域組成。其中橙色或橘黃色熒光由活菌和死菌在同一位置疊加而成,綠色熒光代表活菌,紅色熒光代表死菌。②CLSM下的BBF是團(tuán)塊狀的多聚物�；罹蠖啾话谏锬さ闹虚g,死菌大多在BBF同一層面的外圍。③培養(yǎng)24小時(shí)后成熟BBF中層活菌率為(67.35±1.06)%,底層活菌率為(39.42±2.23)%。中層和底層的BBF活菌率具有明顯的統(tǒng)計(jì)學(xué)意義(P0.01)。④0.5%聚維酮碘溶液作用10分鐘后的BBF的中層、底層活菌率分別為(66.65±1.37)%,(38.22±3.15)%。⑤0.25%維酮碘溶液作用10分鐘后的BBF的中層、底層活菌率分別為(66.53±1.00)%,(39.13±2.41)%。⑥0.5%聚維酮碘溶液作用10分鐘后的BBF的中層、底層活菌率與0.25%聚維酮碘溶液作用10分鐘后的BBF的中層、底層活菌率和空白對(duì)照組相比差異都無(wú)統(tǒng)計(jì)學(xué)意義(P0.05)。⑦模型組、0.5%和.25%聚維酮碘組做細(xì)菌培養(yǎng)的血平板培養(yǎng)基內(nèi)均可見(jiàn)菌落形成,經(jīng)鑒定均為SA。結(jié)論①本實(shí)驗(yàn)所采用的建立金黃色葡萄球菌BBF模型方法相對(duì)簡(jiǎn)單、可重復(fù)性強(qiáng),可用于研究同種細(xì)菌不同異物表面細(xì)菌生物膜的形成能力。②在BBF內(nèi),中層活菌率較底層活菌率高,死菌大多出現(xiàn)在活菌同一層面的外圍。③0.5%、0.25%濃度的聚維酮碘溶液對(duì)鈦板表面金黃色葡萄球菌BBF無(wú)明顯殺菌效果。
[Abstract]:Objective to establish the bacterial biofilm model of Staphylococcus aureus on the surface of titanium plate, the structure of bacterial biofilm (BBF), the rate of living bacteria in the middle layer and the bottom were observed under the laser scanning confocal microscope (CLSM), and the bactericidal effect of 0.5%, 0.25% concentration of polyvidone iodine solution on Staphylococcus aureus BBF in vitro was studied. Staphylococcus aureus (SA) was inoculated in the pre prepared tryptone soybean broth (TSB), and cultured for 16 hours in a constant temperature incubator of 37 C (5%CO2), the bacterial liquid concentration was adjusted to 1 x 106 CFU/ml by turbidimetry. 15 clean, passivated and sterilized titanium plates were randomly assigned to 3 25cm2 cell culture bottles and each cell culture was cultured. 5 titanium plates in the bottle were raised in the bottle. 1ml, 20ml TSB, and 37 C (5%CO2) incubator was cultured for 24 hours to form mature BBF., and 10 titanium plates were randomly removed from the 3 cell culture bottles for 20 minutes under the light environment. The structure of BBF, the living bacteria rate, and the other 5 were observed under CLSM. According to the above modeling method, the BBF model of Staphylococcus aureus BBF was established on the surface of 45 titanium plates, and 3 times were washed with PBS, and 45 titanium plates were randomly divided into 3 groups, each group was 15, of which two groups were placed in 0.5% and 0.25% of the polyvidone iodine respectively. In the solution for 5 minutes, the iodine residue on the surface of the titanium plate was removed by the neutralizer composed of sodium thiosulfate, Twain 80 and lecithin, and 10 titanium plates of each of the two groups were stained by the above staining method for 20 minutes to observe the structure of BBF under CLSM, the change of the living bacteria rate, and the other 5 in the vortex oscillator, ultrasonic cleaning, centrifugation. The machine was centrifuged and inoculated to the blood plate culture medium for bacterial culture. The other 10 titanium plates were used as the control group, the remaining 5 were passed through the vortex oscillator, ultrasonic cleaning, centrifuge centrifuge, and then inoculated to the blood plate culture medium for bacterial culture. Results (1) BBF under CLSM, the highly organized group structure.BBF was under CLSM. The images are composed of orange or orange fluorescent, green fluorescence, red fluorescence and dark area. The orange or orange fluorescence is superimposed on the same position by living bacteria and dead bacteria. The green fluorescence represents the living bacteria and the red fluorescence represents the dead bacteria. (2) the BBF under CLSM is a lump polymer. The live bacteria are mostly wrapped in the middle of the biofilm and dead bacteria Mostly at the periphery of the same level of BBF, after 24 hours of culture, the rate of mature BBF medium living bacteria was (67.35 + 1.06)%, and the living bacteria rate at the bottom was (39.42 + 2.23)%. The rate of BBF living bacteria in the middle and bottom layers had significant statistical significance (P0.01). (4) the rate of living bacteria in the middle layer of BBF after 10 minutes' action of 0.5% polyvidone iodine solution was (66.65 + 1.37)%, respectively, 38.2 2 + 3.15)% of the middle layer of BBF in the middle layer after 10 minutes' action of 0.25% dimensional ketone iodine solution (66.53 + 1)% and (39.13 + 2.41)% respectively. 6. 0.5% polyvidone iodine solution in the middle layer of BBF after 10 minutes' action, the bottom living bacteria rate and the middle layer of BBF after the action of 0.25% polyvidone iodine solution for 10 minutes, the difference of the living bacteria rate of the bottom layer and the blank control group There was no statistical significance (P0.05). (7) model group, 0.5% and.25% povidone iodine group had bacterial colony formation in the blood plate culture medium of bacterial culture. The identification was SA. conclusion (1) the method of establishing Staphylococcus aureus BBF model used in this experiment is relatively simple and reproducible, and can be used to study the surface bacteria of different foreign bodies of the same bacteria. The ability to form biofilm. In BBF, the middle lower rate of viable bacteria rate, dead bacteria occurs mostly in the periphery of live bacteria of the same level. In 0.5%, the concentration of 0.25% povidone iodine solution without obvious bactericidal effect on Staphylococcus aureus BBF titanium plate surface.
【學(xué)位授予單位】：福建醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2015
【分類(lèi)號(hào)】：R515

【相似文獻(xiàn)】

相關(guān)期刊論文 前10條

1 白潔;機(jī)組配餐員的金黃色葡萄球菌污染情況[J];國(guó)外醫(yī)學(xué)(衛(wèi)生學(xué)分冊(cè));2002年02期

2 朱自強(qiáng);;金黃色葡萄球菌更具殺傷力的原因在于其金黃色“外殼”[J];基礎(chǔ)醫(yī)學(xué)與臨床;2005年11期

3 杜洪利;張雙雙;歐旭;閆訓(xùn)友;;金黃色葡萄球菌檢測(cè)技術(shù)研究進(jìn)展[J];食品與發(fā)酵科技;2009年05期

4 黃嶺芳;賴(lài)衛(wèi)華;張莉莉;;食品中金黃色葡萄球菌快速檢測(cè)方法的研究進(jìn)展[J];食品與機(jī)械;2009年06期

5 王娉;勞華均;胡s，

本文編號(hào)：2046317