多重荧光定量聚合酶链反应法检测肉制品中的3种食源性致病菌

摘要：目的 建立多重荧光定量聚合酶链反应法（quantitative polymerase chain reaction,qPCR）快速检测肉制品中沙门氏菌、单增李斯特菌、大肠杆菌O157:H7 3种食源性致病菌的方法。方法 根据沙门氏菌的invA基因、单增李斯特菌的hemolysin基因、大肠杆菌O157:H7的rfbE基因序列分别设计特异性引物及探针,通过优化反应体系,测定其灵敏度、特异性和重复性,建立了可同时检测上述3种食源性致病菌多重qPCR方法。结果 该方法只扩增3种靶细菌,对其他供试菌不扩增;沙门氏菌、单增李斯特菌、大肠杆菌O157:H7的检出限分别为101、102、102拷贝数/μL,并且拥有良好的重复性和特异性。人工污染的猪肉样品经SLE（Salmonella、Listeria monocytogenes and Escherichia coli 3种菌株的共增菌培养基）培养基富集8 h后,分别可以检测到初始菌液浓度为1.56×102 CFU/25 g的沙门氏菌,2.13×102 CFU/25 g的单增李斯特菌,2.32×102 CFU/25 g的大肠杆菌O157:H7。结论 所建立的多重qPCR灵敏度高、特异性强、重复性好,可同时检测肉制品中沙门氏菌、单增李斯特菌和大肠杆菌O157:H7 3种食源性致病菌,在提高食品安全和保护人类健康方面有重要意义。     

基于一步法的金枪鱼生鱼片沙门氏菌生长数值模拟

食物中沙门氏菌的生长是公共健康的重大威胁之一。以生食金枪鱼为研究对象,构建生鱼片中沙门氏菌生长的预测模型。首先,考察恒定温度(8~35℃)条件下沙门氏菌在生鱼片中的生长特性,随机选取两次独立重复试验中一组生长数据,采用一步法同步构建初级模型(Huang模型、Baranyi模型)和二级模型(Huang Square-Root模型),并通过四阶龙格-库塔法联合最小二乘法估计模型参数;其次,选取另一组恒温条件下的独立重复试验数据及波动温度条件下的生长数据,对模型进行验证。结果表明:一步法适用于生鱼片中沙门氏菌的生长曲线分析,同步构建的Huang-HSR模型和Baranyi-HSR模型具有等同的拟合效果,基于Huang模型对迟滞期有着明确的定义,建议选择Huang-HSR模型;通过一步法估计的沙门氏菌的最低生长温度为6.91℃,最大生长浓度为9.15 lg(CFU/g);恒定温度和波动温度验证试验的RMSE分别为0.37 lg(CFU/g)和0.44 lg(CFU/g),其误差分别服从正太分布和拉普拉斯分布。本研究构建的预测模型可用于金枪鱼生鱼片中沙门氏菌的生长预测和风险评估。     

鸡禽肉中的沙门氏菌和现代工业控制其污染的方法

从食品加工的角度论述了鸡在孵化、运输和深度加工过程中受沙门氏菌污染的机制,介绍了工业上去除鸡肉制品上的沙门氏菌污染的几种方法,及使用疫苗培养无沙门氏菌感染的活鸡群,使鸡从根本上摆脱受沙门氏菌感染的方法。     

冻猪肉加工中沙门氏菌的综合控制

强调了沙门氏菌对人畜的危害性和分布的广泛性,着重阐述了生猪屠宰加工分割、包装、冷藏运输各工序对沙门氏菌的综合防控措施.     

MALDI-TOF-MS鉴定2008年辽宁省食源性疾病监测系统检出的沙门菌的初步研究

目的应用MALDI-TOF-MS技术对2008年辽宁省食源性疾病监测检出的24株沙门菌和2株沙门菌标准菌株进行鉴定,并对其中4种血清型的16株沙门菌进行种水平的鉴定。通过MALDI-TOF-MS的相关性分析,研究MALDI-TOF-MS与血清分型结果的相关性及MALDI-TOF-MS对实验的沙门菌的分型能力。方法考察并确定既适合菌株生长又适宜MALDI-TOF-MS分析的沙门菌培养基后,应用MALDI-TOF-MS的相关软件对沙门菌进行数据采集和图谱比对,所得鉴定结果与血清分型结果进行比较。对属于4个种的4株沙门菌的蛋白峰信息进行比较分析。通过生化鉴定及MALDI-TOF-MS的聚类分析及主成分分析讨论4株肠炎沙门菌同源关系。结果 MALDI-TOF-MS技术分别在属和种的水平上准确鉴定了沙门菌株,并且在种的鉴定水平上与血清分型结果具有极大相关性。MALDI-TOF-MS区分了属于4个种的4株沙门菌的蛋白峰之间的微小差异。而且,对相同血清型的4株肠炎沙门菌的聚类分析及主成分分析显示出其同源程度的差异。结论 MALDI-TOF-MS技术在属水平上鉴定沙门菌具有很高的准确性,在种水平上的鉴定也有着很好的应用前景。...     

蔗糖癸酸酯对沙门氏菌的抗菌作用及其机理初探

本文研究了蔗糖癸酸酯对沙门氏菌的体外抗菌作用及其机理。采用肉汤稀释法测定了不同p H值下蔗糖癸酸酯对沙门氏菌的最小抑菌浓度(MIC)和最小杀菌浓度(MBC)。通过测定菌体渗漏物、SDS-聚丙烯酰胺凝胶电泳分析、扫描电镜观察、DNA结合试验等,初步研究了蔗糖癸酸酯对沙门氏菌的抗菌机理。结果表明:蔗糖癸酸酯在p H 7.0时对沙门氏菌的MIC为20 m M,在酸性条件下抗菌效果更为显著。蔗糖癸酸酯能够使菌体细胞内核酸、蛋白质和还原糖等外泄,影响细胞膜结构完整性和菌体正常形态;对菌体细胞蛋白(尤其是大分子量蛋白)的合成与积累有干扰作用;同时糖酯的亲水性头基可与DNA磷酸基团以氢键的方式结合,使DNA骨架收缩。因此,可将蔗糖癸酸酯作为兼具乳化和防腐作用的多功能食品添加剂,应用于动物源性食品的加工和储藏过程中。     

Coliphages as an indicator of faecal pollution in water. Its relationship with indicator and pathogenic microorganisms

The study was designed to test the proposal that Escherichia coli specific bacteriophages might serve as universal faecal pollution indicators in water. A highly specific, sensitive and rapid technique for the detection and quantification of these virus particles was developed. The numerical relationship between E. coli and its parasitic phages was investigated in three different aqueous ecosystems such as sea water in the vicinity of sewage outfalls, river water contaminated by domestic and industrial sewage discharges, and estuarine waters, and found to be very close. In addition, the results obtained indicate that the coliphages are good indicators of the presence of the pathogenic microorganisms studied. In nearly all the water samples tested, the results suggest that coliphages are better indicators of faecal pollution than the classical indicator systems currently employed.     

Comparison of Real Time Polymerase Chain Reaction Quantification of Changes in hilA and rpoS Gene Expression of a Salmonella Typhimurium Poultry Isolate Grown at Fast Versus Slow Dilution Rates in an Anaerobic Continuous Culture System

The objective of this study was to determine the genetic responses of a Salmonella enterica Typhimurium poultry isolate during low and high dilution rates (D) in steady state continuous culture (CC) incubations. Samples for genetic analyses were taken from a previous study where S. Typhimurium cells had been grown in two chemostats operated concurrently, which were designated as Trials 1 and 2, with eight dilution rates sampled during steady state. Real-time polymerase chain reaction (PCR) on two target genes (rpoS and hilA) was analyzed as changes in expression of the target gene relative to the reference gene (16S rRNA). At the lowest D (0.0125 h^?1) in Trial 2, rpoS expression was more than twofold higher than the second highest relative expression. In Trial 1 hilA expression was 21.8-, 27.8-, and 21-fold higher in D 0.0125 h^?1, 0.025 h^?1, 0.05 h^?1, respectively, compared to D 0.1 h^?1, 0.54 h^?1, and 1.08 h^?1. In Trial 2, D 0.025 h^?1 and 0.27 h^?1 showed no difference in hilA expression but were significantly higher compared to other D. From these results, low glucose conditions may play an essential role in triggering rpoS induction, as well as contributing to potential virulence.     

Synthesis and expression in Escherichia coli of DNA encoding the murine {lambda}1 chain of a monoclonal antibody specific for Salmonella serotype B O-antigen

A 658 bp DNA sequence corresponding to the murine ₁ chain ofa monoclonal antibody, Se 155-4, specific for the Salmonellaserotype B O-antigen, was designed using Escherichia coli preferredcodons and chemically synthesized by ligation of synthetic fragmentsinto a linearized plasmid followed by transformation into E.coli.A synthetic signal peptide (ompA) was fused to express the Lchain as a free polypeptide into the periplasm of E.coli cells.After isolation and purification, heterologous recombinationof the E.coli L chain with mouse H chain gave an active antigen-bindingprotein. The activity was 15–20% when compared to proteincreated by an equivalent association of isolated natural mouseL and H chains as measured by a direct EIA assay. In inhibitionexperiments with the polysaccharide antigen, the two proteinsshowed identical titration curves and 50% inhibition points,indicating comparable K_A values.     

Induction of Resistance of Salmonella typhimurium to Environmental Stresses by Exposure to Short-Chain Fatty Acids

ABSTRACT: Short-chain fatty acids (SCFA), which are widely used as food preservatives and are also present in the gastrointestinal (GI) tract of animals at high concentrations, may play a role in the persistence of Salmonella typhimurium in the environment. To test the hypothesis, S. typhimurium was adapted to SCFA for 1 h and the % survivors against various stress conditions was determined. For adaptation, the SCFA mixtures at the concentrations found in small (SI) and large intestine (LI) were used. The % survivors against extreme acid (pH 3.0), high osmolarity (2.5 M NaCl), and reactive oxygen (20 mM H₂O₂) was greatly increased by exposure to SCFA LI, but to a much less extent by SCFA SI. The results suggest that encountering SCFA by S. typhimurium in the large intestine of the host food animal or food materials treated with them may increase the persistence of S. typhimurium in food animal pre-and postharvest production by enhancing overall stress resistance.