臭氧水与副干酪乳杆菌Z21联合处理对绿豆芽中大肠杆菌O157:H7的抑制机制

为研究臭氧水联合副干酪乳杆菌Z21发酵上清液对绿豆芽中大肠杆菌O157:H7的杀菌效果、细胞结构影响和生物膜清除作用,本实验对人工污染大肠杆菌O157:H7的绿豆芽进行联合处理,选出最优的杀菌条件,采用流式细胞仪、扫描电镜、傅里叶红外光谱（Fourier-transform infrared spectroscopy,FT-IR）、拉曼光谱分析臭氧水联合Z21发酵上清液的杀菌机制;通过菌落计数及胞外聚合物分析,研究了臭氧水联合Z21发酵上清液对大肠杆菌O157:H7生物膜的清除效果。结果表明,1.5 mg/L臭氧水联合10%（v/v）Z21发酵上清液处理对大肠杆菌O157:H7杀菌效果最佳,菌落总数减少了2.81 lg CFU/g;与对照组相比,联合处理破坏了大肠杆菌O157:H7细胞壁和细胞膜中的多糖,脂质和蛋白质结构,增加了细胞膜的通透性,改变了菌体形态。联合处理对生物膜有良好的清除效果,显著降低了生物膜的胞外聚合物含量（P<0.05）。本研究为大肠杆菌生物膜的清除及农产品防腐保鲜提供了理论依据。     

等离子体活化水对沙门氏菌的灭活作用及机制研究

等离子体活化水（Plasma-activated water,PAW）是一种新兴的非热杀菌技术,在食品保鲜等领域具有广泛的应用前景。本文研究了PAW处理对沙门氏菌（S. typhimurium）的杀灭效果及其作用机制。将经等离子体放电30、60和90 s得到的PAW分别记为PAW30、PAW60和PAW90。结果表明,PAW对S. typhimurium的杀菌效果随放电时间的延长而逐渐增强。当初始浓度为7.91 lg CFU/mL时,PAW60处理10 min后S. typhimurium活菌数减少了4.22 lg CFU/mL。扫描电镜（Scanning electron microscope,SEM）结果表明,PAW60处理后S. typhimurium细胞形态发生明显变化。经PAW60处理后,S. typhimurium胞外核酸和蛋白含量均显著（P<0.05）升高,表明其细胞膜通透性显著（P<0.05）增强。此外,PAW60处理破坏了S. typhimurium细胞外膜完整性,造成胞内活性氧水平显著（P<0.05）升高。以上实验结果表明,PAW处理能够有效灭活S. typhimurium,其作用机理可能与其破坏细胞结构、增强细胞膜通透性等有关。研究结果为PAW在食品杀菌保鲜中的应用提供了科学理论依据。     

超高压对大肠杆菌O157:H7细胞膜的损伤效应

本研究旨在揭示超高压对食源性致病微生物大肠杆菌O157:H7细胞膜的损伤。研究了200、400、500 MPa不同压力对大肠杆菌O157:H7的灭活作用,通过对菌体细胞核酸类物质、钾离子和镁离子泄漏量、碘化丙啶(propidium iodide,PI)摄入量、细胞膜Na+/K+-ATP酶和Ca~(2+)/Mg~(2+)-ATP酶活性变化的分析研究,评价不同超高压处理压力对大肠杆菌O157:H7膜损伤效应。结果表明,经200、400 MPa压力处理5 min后,大肠杆菌O157:H7菌落总数由初始8.8(lg(CFU/m L))分别下降至8.2(lg(CFU/m L))和6.3(lg(CFU/m L)),500 MPa压力处理后,大肠杆菌O157:H7全部死亡。压力升高,细菌细胞内核酸类物质、K+、Mg~(2+)离子泄漏量、PI摄入量均显著增加,细胞膜上Na+/K+-ATP酶和Ca~(2+)/Mg~(2+)-ATP酶活性显著降低。Ca~(2+)/Mg~(2+)-ATP酶对压力的敏感性更强,500 MPa处理组该酶活性几乎完全丧失。超高压处理引起大肠杆菌O157:H7细胞膜产生显著损伤,细胞膜上Ca~(2+)/Mg~(2+)-ATP酶的失活是导致大肠杆菌O157:H7死亡的主要原因。     

LED蓝光与柠檬酸对大肠杆菌的协同抗菌作用

为了开发新型食品抗菌方法,研究了LED蓝光辐照强度、柠檬酸质量浓度和温度3个因素对营养肉汤中大肠杆菌O157:H7抗菌性能的影响。结果表明:LED蓝光在低温(12℃)下对大肠杆菌O157:H7的抗菌效果优于室温(25℃),且抗菌效果随着光照剂量的增加而增强;柠檬酸在低温下对大肠杆菌O157:H7的影响较弱,但在室温下可明显减缓其生长速率;与空白对照组相比,在使用4 072.3J/cm~2光照剂量的LED蓝光照射后,大肠杆菌O157:H7的数量在低温下可减少(2.60±0.19)lg CFU/mL,而在室温下仅减少(0.67±0.12)lg CFU/mL;加入柠檬酸后,显著增强了LED蓝光的抗菌效果,在使用2 471.0J/cm~2光照剂量的LED蓝光照射后,大肠杆菌O157:H7的数量在低温下可减少(5.13±0.11)lg CFU/mL,在室温下可减少(3.08±0.11)lg CFU/mL。     

副干酪乳杆菌Z17-壳聚糖复配对大肠杆菌O157:H7的抑菌效果及作用机制

目的：研究副干酪乳杆菌Z17-壳聚糖复配对草莓中大肠杆菌O157:H7抑菌活性及作用机制。方法：采用流式细胞术、傅里叶变换红外光谱、拉曼光谱及扫描电子显微镜技术分析副干酪乳杆菌Z17-壳聚糖对大肠杆菌O157:H7细胞膜的影响。结果：质量分数1.0%壳聚糖溶液与副干酪乳杆菌Z17复配处理能有效去除草莓上的大肠杆菌O157:H7,减菌率达99%;壳聚糖溶液与副干酪乳杆菌Z17共同作用3 h使大肠杆菌O157:H7 DNA胞外释放量达（381.00±3.53）ng/μL,细胞膜破损率为58.3%;细胞壁膜中脂肪酸、蛋白、肽聚糖、糖苷环、多糖结构成分被破坏;细胞膜局部位移变薄,大分子物质黏附于菌体细胞表面,细胞表面出现孔洞,胞内物质泄漏,最终导致菌体死亡。结论：副干酪乳杆菌Z17-壳聚糖能够有效地抑制草莓中大肠杆菌O157:H7,其抑菌作用靶点为大肠杆菌O157:H7的细胞膜,研究可为大肠杆菌O157:H7的生物防治提供参考。     

植物源香辛料对E.coli O157：H7的杀菌效果比较

以常见植物源香辛料包括八角、丁香、桂皮、茴香、芥末、花椒、麻椒、香叶为材料,研究其醇提取物在不同浓度和作用时间下对E.coli O157：H7的杀菌效果,并分析不同产地对香辛料杀菌效果的影响。结果表明：试验用香辛料对E.coli O157：H7均具有一定的杀菌作用,杀菌能力差异显著（p＜0.05）,杀菌效果最好的香辛料为丁香,体积分数为9.1%的丁香提取物作用1 h可杀灭E.coli O157：H7 5 lg（CFU/mL）以上,其次为八角;相比其它香辛料,芥末提取物杀灭E.coli O157：H7的效果不理想,体积分数为50%的3个不同芥末产品提取物处理1 h平均杀灭2.47 lg（CFU/mL）E.coli O157：H7;产地来源对某些香辛料的杀菌能力有影响,如茴香、桂皮、八角（p＜0.05）。     

流式分析技术快速定量检测牛乳中大肠杆菌O157:H7

建立一种基于流式分析技术的快速定量检测牛乳中大肠杆菌O157:H7的方法。用偶联有异硫氰酸荧光素的大肠杆菌O157单克隆抗体对大肠杆菌O157:H7进行特异性标记,通过优化抗体反应条件,建立流式检测方法,然后对磷酸盐缓冲溶液（phosphate buffer saline,PBS）和人工污染牛乳样品中不同浓度的大肠杆菌O157:H7进行定量检测。本研究建立的流式检测方法的在PBS中的检测范围为2.57×103～1.12×108?CFU/mL,灵敏度达到2.57×103?CFU/mL。将所建立的流式检测方法应用于牛乳样品检测,当人工污染牛乳样品中大肠杆菌O157:H7的浓度在2.31×104～1.48×108?CFU/mL之间时,流式检测方法与平板计数方法检测结果基本一致,方法的灵敏度为2.31×104?CFU/mL,检测时间为35?min。该方法能快速、定量地检测出牛乳样品中的大肠杆菌O157:H7,在食源性致病菌的快速筛查和监控中具有重要的应用价值。     

贮藏于4℃的家制酸奶中低浓度大肠杆菌O157:H7的存活研究

牛奶预热后分装并分别接种体积分数3%的酸奶和2.15(lg(CFU/mL))的大肠杆菌O157:H7,在45℃下发酵5h后贮藏于4℃的冰箱中。利用Pathatrix大体积循环系统中偶联了抗体的免疫磁珠特异性地捕获这些酸奶中的大肠杆菌O157:H7。免疫磁珠重悬于1%的蛋白胨水后涂布于添加了新生霉素(5mg/L)的山梨醇麦康凯固体培养基中,培养基在37℃恒温培养箱中放置24h。实验结果表明,酸奶中大肠杆菌O157:H7的数量逐渐减少,12d后才检测不到。因此乳制品加工及保存过程中,需要加强对大肠杆菌O157:H7污染的监测,以保证乳制品的安全性。     

出血性大肠杆菌O157∶H7单克隆抗体制备及免疫胶体金试纸条的研制

将骨髓瘤细胞SP2/0与出血性大肠杆菌O157∶H7免疫小鼠得到的脾细胞融合,通过筛选得到3株稳定分泌抗出血性大肠杆菌O157∶H7单克隆抗体的杂交瘤细胞株D3、E7、B9,D3和B9亚类为IgG1,E7亚类为IgG2a,轻链亚型均为κ。交叉反应结果显示这3株单抗仅结合出血性大肠杆菌O157∶H7,对15株其他细菌无反应,测得胶体金标记单抗最佳pH、最佳结合量分别为D3:pH 7.5⁸.0、24μg/mL;B9:pH 7.5、12μg/mL;E7∶pH7.5、18μg/mL,抗体配对选择胶体金结合D3喷涂金标垫、E7以1 mg/mL喷涂NC膜作T线为最优组合,试纸条检测出血性大肠杆菌O157∶H7灵敏度为2.1×106CFU/mL,且仅可检出出血性大肠杆菌O157∶H7,对25株其他细菌均无交叉反应,模拟带菌实验表明,对市售面包、牛奶、果冻各25 g(mL)均添加约200 CFU出血性大肠杆菌O157∶H7,增菌培养8、10、10 h即可检出。研究表明,实验自主制备的抗体性能良好,试纸条特异性、灵敏度达到国外同类产品,可在政府食源性致病菌监管部门、食品企业推广运用。     

大肠杆菌O157:H7量子点免疫层析试纸条的研制

研制一种大肠杆菌O157:H7量子点免疫层析试纸。利用自制水溶性量子点静电偶联大肠杆菌O157:H7单克隆抗体,将大肠杆菌O157:H7单克隆抗体和羊抗兔二抗划线于硝酸纤维素膜分别作为检测线和质控线,制备双抗体夹心法检测大肠杆菌O157:H7的量子点免疫层析试纸。该试纸条能在5min内完成检测,检测限制为1×104 CFU/mL,对常见的8种食源菌无交叉反应。基于量子点的大肠杆菌O157:H7免疫层析试纸操作简便,灵敏度和特异性较好,可用于食品快速检测。     

Biodegradable polylactic acid polymer with nisin for use in antimicrobial food packaging

ABSTRACT:  Biodegradable polylactic acid (PLA) polymer was evaluated for its application as a material for antimicrobial food packaging. PLA films were incorporated with nisin to for control of foodborne pathogens. Antimicrobial activity of PLA/nisin films against Listeria monocytogenes , Escherichia coli O157:H7, and Salmonella Enteritidis were evaluated in culture media and liquid foods (orange juice and liquid egg white). Scanned electron micrograph and confocal laser microscopy revealed that nisin particles were evenly distributed in PLA polymer matrix on the surface and inside of the PLA/nisin films. PLA/nisin significantly inhibited growth of L . monocytogenes in culture medium and liquid egg white. The greatest inhibition occurred at 24 h when the cell counts of L. monocytogenes in the PLA/nisin samples were 4.5 log CFU/mL less than the controls. PLA/nisin reduced the cell population of E. coli O157:H7 in orange juice from 7.5 to 3.5 log at 72 h whereas the control remained at about 6 log CFU/mL. PLA/nisin treatment resulted in a 2 log reduction of S. Enteritidis in liquid egg white at 24 °C. After 21 d at 4 °C the S. Enteritidis population from PLA/nisin treated liquid egg white (3.5 log CFU/mL) was significantly less than the control (6.8 log CFU/mL). E. coli O157:H7 in orange juice was more sensitive to PLA/nisin treatments than in culture medium. The results of this research demonstrated the retention of nisin activity when incorporated into the PLA polymer and its antimicrobial effectiveness against foodborne pathogens. The combination of a biopolymer and natural bacteriocin has potential for use in antimicrobial food packaging.     

Chlorine inactivation of Escherichia coli O157:H7 in water

Six human isolates of Escherichia coli O157:H7 and E. coli (ATCC 11229) were used to determine the concentrations of free chlorine and exposure times required for inactivation. Free chlorine concentrations of 0.25, 0.5, 1.0, and 2.0 ppm at 23 degrees C were evaluated, with sampling times at 0, 0.5, 1.0, and 2.0 min. Results revealed that five of six E. coli O157:H7 isolates and the E. coli control strain were highly susceptible to chlorine, with >7 log10 CFU/ml reduction of each of these strains by 0.25 ppm free chlorine within 1 min. However, comparatively, one of the seven strains was unusually tolerant to chlorine at 23 degrees C for 1 min, with a 4-, 5.5-, 5.8-, and >5.8-log CFU/ml reduction at free chlorine concentrations (ppm) of 0.25, 0.5, 1.0, and 2.0. respectively. Based on these studies most isolates of E. coli O157:H7 have no unusual tolerance to chlorine; however, one strain was exceptional in being recovered after 1-min of exposure of 10(7) CFU/ml to 2.0 ppm of free chlorine. This isolate may be a useful reference strain for future studies on chlorine tolerance of E. coli O157:H7.     

Antimicrobial Activity and Synergistic Effect of Cinnamon with Sodium Benzoate or Potassium Sorbate in Controlling Escherichia coli O157:H7 in Apple Juice

ABSTRACT: Antimicrobial effects of cinnamon, sodium benzoate, potassium sorbate, and combinations were examined against Escherichia coli O157:H7 in apple juice at 8°C and 25°C. E. coli O157:H7 was reduced by 1.6 log colony-forming units (CFU)/mL at 8°C and 2.0 log CFU/mL at 25°C by 0.3% cinnamon. At 8°C, 5.2 log CFU/mL of E. coli O157:H7 was eliminated in 11 d by 0.3% cinnamon with 0.1% sodium benzoate, and in 14 d by 0.3% cinnamon with 0.1% potassium sorbate. At 25°C, 5.3 log CFU/mL E. coli O157:H7 was eliminated in 3 d by the same combinations. A synergistic effect was observed between cinnamon and preservatives against E. coli O157:H7 at 8°C and 25°C.     

Survival of Escherichia coli O157:H7 and Salmonella in apple cider and orange juice as affected by ozone and treatment temperature

Inactivation of Escherichia coli O157:H7 and Salmonella in apple cider and orange juice treated with ozone was evaluated. A five-strain mixture of E. coli O157:H7 or a five-serovar mixture of Salmonella was inoculated (7 log CFU/ml) into apple cider and orange juice. Ozone (0.9 g/h) was pumped into juices maintained at 4 degrees C, ambient temperature (approximately 20 degrees C), and 50 degrees C for up to 240 min, depending on organism, juice, and treatment temperature. Samples were withdrawn, diluted in 0.1% peptone water, and surface plated onto recovery media. Recovery of E. coli O157:H7 was compared on tryptic soy agar (TSA), sorbitol MacConkey agar, hemorrhagic coli agar, and modified eosin methylene blue agar; recovery of Salmonella was compared on TSA, bismuth sulfite agar, and xylose lysine tergitol 4 (XLT4) agar. After treatment at 50 degrees C, E. coli O157:H7 populations were undetectable (limit of 1.0 log CFU/ml; a minimum 6.0-log CFU/ml reduction) after 45 min in apple cider and 75 min in orange juice. At 50 degrees C, Salmonella was reduced by 4.8 log CFU/ml (apple cider) and was undetectable in orange juice after 15 min. E. coli O157:H7 at 4 degrees C was reduced by 4.8 log CFU/ml in apple cider and by 5.4 log CFU/ml in orange juice. Salmonella was reduced by 4.5 log CFU/ml (apple cider) and 4.2 log CFU/ml (orange juice) at 4 degrees C. Treatment at ambient temperature resulted in population reductions of less than 5.0 log CFU/ml. Recovery of E. coli O157:H7 and Salmonella on selective media was substantially lower than recovery on TSA, indicating development of sublethal injury. Ozone treatment of apple cider and orange juice at 4 degrees C or in combination with mild heating (50 degrees C) may provide an alternative to thermal pasteurization for reduction of E. coli O157:H7 and Salmonella in apple cider and orange juice.     

免疫磁分离法高效富集牛奶中大肠杆菌O157:H7

目的建立一种免疫磁分离(immunomagnetic separation,IMS)方法高效富集大肠杆菌O157:H7。方法合成一种核壳型的纳米磁珠(magnetic nanobeads, MNBs),并基于制备的MNBs构建了IMS。通过优化制备免疫MNBs时抗体浓度, IMS过程免疫MNBs的用量和孵育时间,构建了高效的IMS方法。结果构建的IMS方法能够在35 min内完成牛奶中大肠杆菌O157:H7的高效富集,当大肠杆菌O157:H7浓度低于10~5 CFU/m L时,捕获效率高于93.4%,当菌浓度达到10~7CFU/mL,捕获效率仍大于50%。结论该方法简单高效,可被广泛应用于其他食源性致病菌检测的样品前处理。     

基于可视化抗体宏阵列技术的出血性大肠杆菌O157∶H7定量检测

目的：探索建立一种新的出血性大肠杆菌O157∶H7（Escherichia coli O157∶H7）的快速定量检测技术。方法：以硝酸纤维素膜为基片,用生物芯片点样仪制作抗体宏阵列,采用“双抗夹心”原理,对出血性大肠杆菌O157∶H7进行定量检测研究。结果：本方法对出血性大肠杆菌O157∶H7的检出限为3.4×105 CFU/mL,在105～107CFU/mL细菌浓度范围内,宏阵列的灰度值与细菌浓度之间有较好的线性关系。该方法可在2.5 h内同步检测多个样品中出血性大肠杆菌O157∶H7的浓度。结论：通过用标准菌株、模拟带菌等研究显示,用宏阵列技术快速定量检测出血性大肠杆菌O157∶H7,结果肉眼可见,稳定准确,同时操作简便、成本低廉、无需大型设备,制作好的抗体宏阵列可用于快速评估食品中出血性大肠杆菌O157∶H7的污染状况,尤其适合于基层实验室进行快速高通量样品筛查。     

Antimicrobial effect of electrolyzed oxidizing water against Escherichia coli O157:H7 and Listeria monocytogenes on fresh strawberries (Fragaria x ananassa)

ABSTRACT:  Antibacterial activity of electrolyzed oxidizing (EO) water prepared from 0.05% or 0.10% (w/v) sodium chloride (NaCl) solutions against indigenous bacteria associated with fresh strawberries ( Fragaria × ananassa ) was evaluated. The efficacy of EO water and sodium hypochlorite (NaOCl) solution in eliminating and controlling the growth of Listeria monocytogenes and Escherichia coli O157:H7 inoculated onto strawberries stored at 4 ± 1 °C up to 15 d was investigated at exposure time of 1, 5, or 10 min. Posttreatment neutralization of fruit surfaces was also determined. More than 2 log₁₀ CFU/g reductions of aerobic mesophiles were obtained in fruits washed for 10 or 15 min in EO water prepared from 0.10% (w/v) NaCl solution. Bactericidal activity of the disinfectants against L. monocytogenes and E. coli O157:H7 was not affected by posttreatment neutralization, and increasing exposure time did not significantly increase the antibacterial efficacy against both pathogens. While washing fruit surfaces with distilled water resulted in 1.90 and 1.27 log₁₀ CFU/mL of rinse fluid reduction of L. monocytogenes and E. coli O157:H7, respectively, ≥ 2.60 log₁₀ CFU/mL of rinse fluid reduction of L. monocytogenes and up to 2.35 and 3.12 log₁₀ CFU/mL of rinse fluid reduction of E. coli O157:H7 were observed on fruit surfaces washed with EO water and NaOCl solution, respectively. Listeria monocytogenes and E. coli O157:H7 populations decreased over storage regardless of prior treatment. However, EO water and aqueous NaOCl did not show higher antimicrobial potential than water treatment during refrigeration storage.