枯草芽孢杆菌对冰鲜罗非鱼肠道菌群的调节及保鲜作用

采用一株枯草芽孢杆菌(BS08)作为饲料添加剂喂养罗非鱼,研究其对冰鲜罗非鱼肠道菌群的影响,尤其是对鱼体腐败菌的粘附、定植和滋生的影响,并研究枯草芽孢杆菌延缓冰鲜罗非鱼腐败的作用。通过饲料中添加枯草芽孢杆菌,水体中添加腐败菌(希瓦氏菌和假单胞菌)的方式分组喂养罗非鱼后,对罗非鱼进行冰鲜贮藏,通过高通量测序确定枯草芽孢杆菌和腐败菌对冰鲜过程中罗非鱼肠道菌群的影响,并以TVB-N和K值为腐败指标,研究冰鲜鱼腐败的程度。结果表明:枯草芽孢杆菌对罗非鱼的肠道菌群的变化具有阻遏作用,使菌群多样性维持在一个相对稳定的水平。希瓦氏菌在肠道内定植程度大于假单胞菌,枯草芽孢杆菌可定植在鱼肠道内,并抑制希瓦氏菌在鱼肠道内定植,最终显著遏制希瓦氏菌在肠道中的优势富集。以TVB-N和K值为指标判断鱼的鲜度,添加枯草芽孢杆菌可显著延缓冰鲜鱼的腐败程度。     

真空包装盐水鹅贮藏期菌群多样性动态分析

为揭示真空包装盐水鹅在4,25℃和30℃贮藏温度下的微生物菌群变化及优势腐败菌,采用平板计数法和PCR-DGGE(聚合酶链式反应-变性梯度凝胶电泳)技术对各温度下不同贮藏期样品菌落总数和菌相变化进行分析,并对主要条带进行割胶测序及同源性比对。结果表明:菌落总数在贮藏期逐渐上升,贮藏后期菌落总数呈下降趋势;产品贮藏期间的优势腐败菌主要为耐受极端环境的芽孢杆菌和类芽孢杆菌、组织菌属和假单胞菌属。30℃条件下主要优势腐败菌为提歇尔氏菌属、泛酸枝芽孢杆菌和幼虫芽孢杆菌,25℃条件下主要优势腐败菌为短芽孢杆菌属、芽孢杆菌属、提歇尔氏菌属、类芽孢杆菌属和地衣芽孢杆菌,4℃条件下主要优势腐败菌为类芽孢杆菌属和铜绿假单胞菌,它们可导致产品肉质变软、发黏、变色并产生异味。     

传统豆腐干中主要腐败菌的分离及其系统发育

以腐败的豆腐干为材料,应用纯培养手段和基于16S rRNA基因序列的系统发育分析,对豆腐干的腐败菌进行了分离及其系统发育鉴定研究。实验结果表明,获得的腐败菌DFG01为假单胞菌属,与恶臭假单胞菌(Pseudomonas aeruginosa)的同源性为99.7%;另外,腐败菌DFG02为芽孢杆菌属,与枯草芽孢杆菌(Bacillus subtilis)高达99.5%。因此,传统豆腐干中腐败菌主要为恶臭假单胞菌和枯草芽孢杆菌,在豆腐干生产制作过程中要加强这两种菌的污染控制。     

真空包装盐水鹅中腐败细菌的分离鉴定

为了采取有效措施控制盐水鹅中的微生物污染,遂先采用平板划线分离方法分离出盐水鹅中的主要微生物菌群,再根据革兰氏镜检、菌株特性、生理生化实验以及16S r DNA测序的方法对盐水鹅中主要腐败菌进行鉴定。结果表明,引起盐水鹅腐败变质的主要微生物菌群:菌种M为蜡样芽孢杆菌(Bacillus cereus,B.c)、菌种P为荧光假单胞菌(Pseudomonas fluorescens,P.f)、菌种S为成团泛菌(Pantoea agglomerans,P.a)。      

市售臭豆腐中菌种的分离及生化鉴定

本实验通过对市售臭豆腐发酵卤水中细菌多样性分析,分离纯化其中的各种菌株,后对其进行生化鉴定和食用安全性验证试验,从中筛选出适合安全食用的臭豆腐发酵的菌株。结果发现,市售臭豆腐卤水存在多种微生物,它们是乳酸菌(其中包括短乳杆菌、棒状乳杆菌扭曲亚种和植物乳杆菌)、芽孢杆菌(其中有巨大芽孢杆菌、环状芽孢杆菌)、假单胞菌、弧菌未知种。通过安全性试验的菌种有4种,分别为植物乳杆菌、假单胞菌、环状芽孢杆菌以及棒状乳杆菌扭曲亚种。     

造纸法再造薄片浆料中腐败菌的分离鉴定与抑菌研究

采用平板划线分离方法对造纸法再造烟叶上网浆中的主要微生物群进行分离,再根据细菌的菌落形态、菌体形态、生理生化实验和16S r DNA序列分析对菌株进行了鉴定。结果表明,引起造纸法再造烟叶上网浆腐败变质的主要菌群为约氏不动杆菌和恶臭假单胞菌。此外,还考察了山梨酸钾、双乙酸钠、尼泊金酯钠、乙二胺四乙酸二钠和ε-聚赖氨酸等5种防腐剂对这两种腐败菌的抑菌效果。结果表明5种防腐剂的抑菌效果强弱顺序为ε-聚赖氨酸乙二胺四乙酸二钠尼泊金酯钠双乙酸钠山梨酸钾,其中ε-聚赖氨酸对约氏不动杆菌和恶臭假单胞菌的最小抑菌质量浓度分别为0.075 mg/m L和0.05 mg/m L。     

冰鲜鸭优势腐败菌的鉴定

结合传统的细菌分离培养法与现代分子生物技术方法16SrDNA菌群分析法对冰鲜鸭中优势腐败菌进行鉴定。传统分离培养检测到了17株优势腐败菌,16SrDNA菌群分析表明,冰鲜鸭中假单胞菌占绝对优势,达到54％;气单胞茵、肠杆菌是仅次于假单胞菌的第二个类群,都分别占15％;乳酸菌、节杆菌、紫色杆菌以及一株未鉴定的属都分别占4％。     

冰鲜鸭优势腐败菌的鉴定

结合传统的细菌分离培养法与现代分子生物技术方法 16S rDNA菌群分析法对冰鲜鸭中优势腐败菌进行鉴定。传统分离培养检测到了17株优势腐败菌,16S rDNA菌群分析表明,冰鲜鸭中假单胞菌占绝对优势,达到54%;气单胞菌、肠杆菌是仅次于假单胞菌的第二个类群,都分别占15%;乳酸菌、节杆菌、紫色杆菌以及一株未鉴定的属都分别占4%。     

北豆腐中主要腐败微生物的分离与鉴定

从东北和华北腐败变质的豆腐中分离得到2株主要腐败细菌xy-1、xy-2。对这两种菌株进行了菌落形态、生理生化特征和16S rDNA序列分析。结果表明：导致豆腐腐败的主要微生物为短小芽孢杆菌（Bacillus pumilus）和恶臭假单胞菌（Pseudomonas putida）。     

PCR结合生理生化鉴定对冷藏带鱼主要细菌菌相组成分析

将传统的微生物生理生化鉴定技术与16S rDNA分子生物学方法相结合,并通过系统发育树分析,对冷藏带鱼贮藏期间的主要细菌组成比例进行研究。样品在(4±1)℃贮藏,分别于不同贮藏时间取样,对各个阶段的微生物菌落进行分离纯化、形态观察和部分生理生化鉴定,对最终得到的细菌纯培养物直接提取DNA,作16SrDNA PCR扩增,产物经测序后于NCBI与已知序列进行相似性比对,最终得到13株具有典型特征的纯菌株。结果得出,冷藏带鱼在货架期终点时,其主要微生物的种类与所占比例依次为:腐败希瓦氏菌(Shewanella putrefaciens 34.7%)、荧光假单胞菌(Pseudomonas fluorescens 14.5%)、松鼠葡萄球菌(Staphylococcus sciuri 10.2%)、嗜水气单胞菌(Aeromonas hydrophila 8.2%)、弧菌(Vibrio rumoiensis 6.1%)、恶臭假单胞菌(Pseudomonas putida6.1%)、绿色气球菌(Aerococcus viridans 4.1%)、金黄色葡萄球菌(Staphyloccocus aureus 4.1%)、蜡样芽孢杆菌(Bacillus cereus 4.0%)、铜绿假单胞菌(Pseudomonas aeruginosa 2.0%)、嗜冷杆菌(Psychrobacter sp.2.0%)、成团肠杆菌(Enterobacter agglomerans 2.0%)、约氏不动杆菌(Acinetobacter johnsonii 2.0%),其中腐败希瓦氏菌为带鱼冷藏期间的主要优势腐败菌,假单胞菌在带鱼冷藏过程中的所占比例大小依次为荧光假单胞菌>恶臭假单胞菌>铜绿假单胞菌。     

排骨汤的贮藏特性和动力学研究

以猪直排为原料,采用高压四段法制作排骨汤,于4℃贮藏,研究贮藏过程排骨汤体系营养成分、TVB-N、菌落总数和蛋白质消化特性,建立排骨汤的贮藏动力学模型,以预测和控制排骨汤的贮藏品质和货架期。结果表明,贮藏过程中排骨汤体系中肌肉和汤汁的菌落总数有所增加,出现蛋白质降解和营养品质劣化现象。排骨汤的贮藏伴随着多种生物化学变化,用一级化学反应动力学模型拟和贮藏过程中排骨汤体系的特性具有很高的精度。     

凯里地区辣椒酸细菌群落多样性及其乳酸菌分离鉴定

利用Illumina MiSeq高通量测序技术对凯里地区辣椒酸中细菌群落多样性进行研究,并结合传统纯培养技术对样品中乳酸菌进行分离鉴定。结果显示,样品中优势细菌门为厚壁菌门（Firmicutes）和变形菌门（Proteobacteria）,相对含量分别为77.9%和19.59%;优势细菌属为乳酸杆菌（Lactobacillus）、假单胞菌属（Pseudomonas）、肠杆菌属（Enterobacterales）、Lelliottia、芽孢杆菌属（Bacillus）、葡萄球菌属（Staphylococcus）和寡养单胞菌属（Stenotrophomonas）,相对含量分别为72.02%、5.59%、3.97%、5.30%、1.64%、1.07%和1.02%。同时,采用纯培养技术从辣椒酸中分离出了43株细菌,其中34株被鉴定为乳酸杆菌,6株为芽孢杆菌,3株为葡萄球菌。由此可见,乳酸杆菌为凯里地区辣椒酸中的优势细菌,且主要为植物乳杆菌;同时分离出了葡萄球菌,提示传统发酵辣椒酸存在一定的食品安全风险,应引起重视。     

基于模糊数学感官评价和响应面法的红辣椒酸汤发酵工艺研究

以红辣椒为主要原料,通过多菌种乳酸菌协同发酵,以总酸和模糊数学感官评分为考察指标,通过食盐添加量、菌种添加量、发酵时间、发酵温度的单因素实验和响应面优化,确定红辣椒酸汤生产最优发酵工艺条件。结果显示红辣椒酸汤的最佳工艺条件为:食盐添加量0.5%,菌种添加量为5.7%,发酵时间为20 d,发酵温度为30℃。在此工艺条件下,得到红辣椒酸汤的总酸为32.45 g/kg,感官评价总分为8.67分,产品有机酸种类丰富,可为红辣椒酸汤工业化生产提供技术支持。     

鲶鱼体表粘液粗提物对铜绿假单胞菌的抑菌机理初探

本文以铜绿假单胞菌为研究对象,探讨鲶鱼体表粘液提取物(Catfish Epidermal Mucus Extracts,CEME)对其的抑制效果及抑菌机理。研究表明:当CEME浓度高于0.15%时,鲶鱼体表粘液提取物对铜绿假单胞菌有显著的抑制作用,通过测定微生物的生长曲线可以看出CEME能够有效减缓铜绿假单胞菌的生长速率。进一步使用扫描电镜发现CEME对铜绿假单胞菌的细胞形态结构能够造成明显的损伤,有效破坏微生物细胞膜的完整性。同时CEME还可以导致铜绿假单胞菌细胞膜的通透性增加,从而导致体内小分子物质以及部分大分子物质向外泄漏。应用SDS-PAGE方法发现CEME处理对铜绿假单胞菌的总蛋白和细菌膜蛋白均有显著的影响,它能够抑制菌体某些蛋白的合成,导致胞内蛋白含量的下降和缺失。上述研究结果表明鲶鱼体表粘液提取物对铜绿假单胞菌有较好的抑制效果。     

丹江镇辣椒红酸汤细菌菌群多样性分析及抗食源性致病菌乳酸菌筛选

以贵州丹江镇传统辣椒红酸汤为研究对象,采用Illumina MiSeq高通量测序技术进行细菌菌群多样性分析,结合微生物纯培养技术和双层平板法从中分离优势乳酸菌,通过分子生物学技术对其进行菌种鉴定,并筛选抗食源性致病菌乳酸菌。结果表明,辣椒红酸汤样品中的优势细菌门为厚壁菌门（Firmicutes）和变形菌门（Proteobacteria）;优势细菌属为乳杆菌属（Lactobacillus）。从辣椒红酸汤样品中共分离得到17株乳酸菌,经鉴定,11株为干酪乳杆菌（Lactobacillus casei）、3株为副干酪乳杆菌（Lactobacillus paracasei）、1株为副干酪乳杆菌亚种（Lactobacillus paracasei subsp.）、1株为植物乳杆菌（Lactobacillus plantarum）、1株为乳酸杆菌（Lactobacillus sp.）,其中4株乳酸菌具有较好的产酸和抑菌性能,且干酪乳杆菌ST7-14的抑菌能力最好,对蜡样芽孢杆菌（Bacillus cereus）ST2-1和阴沟肠杆菌（Enterobacter cloacae）ST2-6的抑菌圈直径分别为37.0 mm和31.5 mm。     

Storage stability of cauliflower soup powder: The effect of lipid oxidation and protein degradation reactions

Soups based on cauliflower soup powders, prepared by dry mixing of ingredients and rapeseed oil, showed a decrease in quality, as evaluated by a sensory panel, during the storage of the soup powder in the dark for up to 12 weeks under mildly accelerated conditions of 40 °C and 75% relative humidity. Antioxidant, shown to be effective in protecting the rapeseed bulk oil, used for the powder preparation, had no effect on storage stability of the soup powder. The freshly prepared soup powder had a relatively high concentration of free radicals, as measured by electron spin resonance spectroscopy, which decreased during storage, and most remarkably during the first two weeks of storage, with only marginal increase in lipid hydroperoxides as primary lipid oxidation products, and without any increase in secondary lipid oxidation products. Analyses of volatiles by SPME-GC–MS revealed a significant increase in concentrations of 2-methyl- and 3-methyl butanals, related to Maillard reactions, together with an increase in 2-acetylpyrrole concentration. The soup powders became more brown during storage, as indicated by a decreasing Hunter L-value, in accord with non-enzymatic browning reactions. A significant increase in the concentrations of dimethyl disulfide in soup powder headspace indicated free radical-initiated protein oxidation. Protein degradation, including Maillard reactions and protein oxidation, is concluded to be more important than lipid oxidation in determining the shelf-life of dry cauliflower soup powder.     

Compositions of maple sap microflora and collection system biofilms evaluated by scanning electron microscopy and denaturing gradient gel electrophoresis

The bacterial microflora of maple sap and biofilms in collection system tubing were studied through the use of bacterial counts, scanning electron microscopy (SEM) of surfaces and the analysis of 16S rRNA gene by denaturing gradient gel electrophoresis (DGGE). Samples were taken at five times during the 2002 and 2003 seasons in order to follow the changes in the microflora of this complex ecosystem. Bacterial counts showed the growth of bacterial populations as the season advanced. These populations were mainly composed of psychrotrophic bacteria and Pseudomonas spp. SEM results confirmed the suspected presence of biofilms on the inner surfaces of tubing samples. Bacterial colonization and biofilm formation progressively increased during the season for both lateral and main line surfaces, and biofilms were mainly composed of rod shape bacteria. The bacterial microflora profiles obtained for sap and corresponding biofilm by DGGE showed up to 12 major bands. The Shannon-Weaver index of diversity (H) calculated from DGGE bands were statistically higher for sap samples compared to biofilm. The diversity index was relatively stable or increasing for lateral line sap and biofilm samples during the season while the diversity index for sap and biofilm samples of the main line showed a decreasing profile as the season progressed. Sequence analysis of major DGGE bands revealed the predominance of bacteria from the genera Pseudomonas, Rahnella and another, unidentified genus. The results describe the composition of sap collection system microflora as well as the formation of biofilms and will be useful for further studies on factors affecting maple product quality.     

基于假单胞菌生长模型预测冷却牛肉的货架期

假单胞菌（Pseudomonas spp.）的生长繁殖是影响冷却牛肉货架期的重要因素。为确定假单胞菌为冷却牛肉的特定腐败菌并建立其货架期预测模型,将屠宰后的冷却牛肉4 ℃贮藏,测定了假单胞菌数量与菌落总数、挥发性氨基氮（TVBN）值、颜色明度值（L*）及感官评定分值等品质指标,并确定了冷却牛肉的假单胞菌腐败限控量。将冷却牛肉分别置于0 ℃、5 ℃、10 ℃、15 ℃、20 ℃和25 ℃条件下进行假单胞菌计数,建立Gompertz方程的初级生长模型和二级模型,并进行了模型的验证,建立了货架期预测模型。研究表明,Gompertz预测模型能有效预测4～25 ℃条件下假单胞菌在冷却牛肉中的生长情况,在0 ℃、5 ℃和10 ℃温度条件下,对牛肉货架期进行验证,与实际货架期相比偏差＜1 d,表明货架期预测模型适用。     

The characterisation of Bacillus spores occurring in the manufacturing of (low acid) canned products

Spore-forming bacteria can be a problem in the food industry, especially in the canning industry. Spores present in ingredients or present in the processing environment severely challenge the preservation process since their thermal resistance may be very high. We therefore asked the question which bacterial spore formers are found in a typical soup manufacturing plant, where they originate from and what the thermal resistance of their spores is. To answer these questions molecular techniques for bacterial species and strain identification were used as well as a protocol for the assessment of spore heat stress resistance based on the Kooiman method. The data indicate the existence and physiological cause of the high thermal resistance of spores of many of the occurring species. In particular it shows that ingredients used in soup manufacturing are a rich source of high thermal resistant spores and that sporulation in the presence of ingredients rich in divalent metal ions exerts a strong influence on spore heat resistance. It was also indicated that Bacillus spores may well be able to germinate and resporulate during manufacturing i.e. through growth and sporulation in line. Both these spores and those originating from the ingredients were able to survive certain thermal processing settings. Species identity was confirmed using fatty acid analysis, 16SrRNA gene sequencing and DNA-DNA hybridisation. Finally, molecular typing experiments using Ribotyping and AFLP analysis show that strains within the various Bacillus species can be clustered according to the thermal resistance properties of their spores. AFLP performed slightly better than Ribotyping. The data proofed to be useful for the generation of strain specific probes. Protocols to validate these probes in routine identification and innovation aimed at tailor made heat processing in soup manufacturing have been formulated.