熏煮火腿中志贺氏菌的热失活特性

为了研究志贺氏菌不同菌株在熏煮火腿中热失活规律,以55、63℃对接种108 CFU/g志贺氏菌的熏煮火腿进行热处理,测定处理后样品中志贺氏菌的残存菌体浓度。应用Logistic模型和3rd degree polynomial模型拟合在两种温度下志贺氏菌的动力学模型,用D值(decimal reduction time)表示志贺氏菌在熏煮火腿中的耐热情况。结果表明:11株志贺氏菌的热失活曲线运用3rd degree polynomial拟合较好,相关系数均在0.98以上。志贺氏菌不同菌株的D值不同,55℃条件下志贺氏菌在熏煮火腿中的D值为4.46~10.66 min,63℃条件下D值为0.48~3.22 min。不同菌株之间抗热性存在差异,其中菌株S-5的耐热性最强,S-6最弱。     

超高压协同温度处理对烟熏火腿中单增李斯特菌生长预测模型的建立

为了建立超高压协同温度处理烟熏火腿中单增李斯特菌生长预测模型。本研究分别以20、30、40、50℃结合600MPa对接种106cfu/g单增李斯特菌烟熏切片火腿进行5min的超高压处理,4℃贮藏条件下,每10d测定处理后样品中单增李斯特菌的数量。结果表明:当超高压协同处理的温度高于40℃时显著延长了单增李斯特菌在烟熏火腿修复生长的延滞期,所得生长曲线用修正的Gompertz模型进行了拟合,在35、45d进行验证,建立了烟熏火腿中单增李斯特菌在超高压协同温度处理后的生长预测模型,经验证其精确因子(Af)、偏差因子(Bf)、根平均方差(RMSE)和决定系数(R2)均在较好范围内,能较好的模拟单增李斯特菌的生长情况,为低温肉制品中单增李斯特菌的控制提供理论参考。     

热压结合处理对绿色魏斯氏菌生理特性的影响

为了探究热压结合处理在绿色魏斯氏菌致死过程中发挥的作用。本实验采用超高压(350 MPa、25℃、10 min)和热压结合(350 MPa、50℃、10 min)对绿色魏斯氏菌进行压力处理,研究了热压结合处理对绿色魏斯氏菌生理特性、细胞形态特征、细胞膜的通透性等的影响。结果显示,热压结合处理使菌落总数从超高压处理时的7.67(lg(CFU/mL))显著下降到5.71(lg(CFU/mL))(P0.05),协同的热处理使菌体表面出现严重褶皱、菌体内含物变性凝固加剧,使细胞膜损伤,从而改变膜的通透性,导致细胞外紫外吸收物质显著增加(P0.05)。因此,热压结合处理在超高压致死细菌中会加剧细胞膜的形变及损坏,增加细胞膜的通透性,从而导致细胞内紫外吸收物质的流出,实验结果为热压结合灭活绿色魏斯氏菌提供了理论依据。     

应用超高压手段延长低温烟熏火腿的货架期

为揭示超高压处理后低温烟熏火腿货架期、品质及营养指标的变化,以400MPa和600MPa的压力在22℃条件下对切片包装后的烟熏火腿进行10min超高压处理,以未经高压处理样品作对照,4℃贮藏条件下,于0、1、30、90d测定超高压处理样品中的各腐败微生物总数,同时对超高压处理烟熏火腿的脂肪氧化程度、游离脂肪酸含量以及部分感官指标进行评定。结果表明：超高压能够有效杀灭烟熏火腿中的肠杆菌、热杀索丝菌、假单胞菌以及霉菌和酵母,乳酸菌中的部分属较为耐压,是低温烟熏火腿贮藏后期的优势腐败菌;超高压处理组比未处理组样品的饱和脂肪酸含量略有上升,不饱和脂肪酸含量略有下降,且随压力升高饱和脂肪酸增加和不饱和脂肪酸减少的量也相对较大;贮藏初期超高压处理组与未处理组样品TBARS值都没有显著变化,而随贮藏期延长,超高压处理组样品TBARS值比未处理组略有上升。超高压处理能够有效延长低温烟熏火腿的货架期,不会或较小引起游离脂肪酸、贮藏期内色泽和脂肪氧化指标的显著变化,能够较好的保持产品原有游离脂肪酸组成及口感品质。     

超高压处理抑制低温烟熏火腿中的优势腐败菌

研究尝试应用微生物菌体总RNA提取代替DNA提取,进而通过反转录-PCR( RT-PCR),结合变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)技术,以期揭示超高压处理后低温烟熏火腿中腐败微生物的存活情况,探索RNA-DGGE手段判定超高压处理后微生物存活状态的可行性.分别以400 MPa和600 MPa的压力在室温(22℃)条件下,对包装后的烟熏火腿进行10 min超高压处理,未经高压处理样品作对照,于4℃冷藏条件下,贮藏1、15、30、60、90 d,直接提取样品中微生物的总RNA,对其进行RT-PCR和DGGE指纹图谱分析.DGGE指纹图谱显示,超高压处理对烟熏火腿中的优势腐败菌具有较强的抑制作用,且随压力的升高抑菌效应增强;超高压处理后烟熏火腿微生物种群结构变得单一,Weissella viridescens和Leuconostoc mesenteroides是超高压处理后烟熏火腿中的优势腐败菌.基于菌体总RNA提取的DGGE手段能够有效检测超高压处理后微生物的存活状况,揭示超高压对低温烟熏火腿中优势腐败微生物的抑菌效果.     

超高压协同乳酸链球菌素抑制低温火腿中的耐压腐败菌

为探寻低温肉制品中耐压腐败菌的抑制手段,揭示超高压和乳酸链球菌素之间的协同抑菌效应。以肉制品中典型的耐压腐败菌绿色魏斯菌和肠膜明串珠菌为供试材料,评价二者在1～500μg/mL的乳酸链球菌素存在条件下的生长情况,并结合100～600MPa(5min,20℃)的超高压处理研究不同压力条件与乳酸链球菌素之间的协同增效作用。结果表明:随着乳酸链球菌素添加量的增加,两种菌的数量急剧降低,当添加量达到200μg/mL时,对二者的抑制程度达到最大;单独超高压处理时,绿色魏斯菌和肠膜明串珠菌分别在400MPa和300MPa的压力下受到显著抑制。而超高压结合200μg/mL乳酸链球菌素处理时,二者受到显著抑制的压力均降低到100MPa;500MPa,5min,20℃的超高压条件结合200μg/mL乳酸链球菌素处理能够抑制绿色魏斯菌数达9lg(cfu/mL)。超高压和乳酸链球菌素之间存在显著的协同作用,乳酸链球菌素的添加将会有效降低工业化生产中超高压的压力水平。     

食窦魏斯氏菌NCU034005与柠檬明串珠菌NCU027003对泡菜代谢物及风味的影响

在灭菌后的泡菜原材料中分别接种食窦魏斯氏菌NCU034005与柠檬明串珠菌NCU027003,室温纯种发酵7d,期间检测泡菜液中挥发性物质与非挥发性物质(葡萄糖、有机酸、氨基酸等)、pH、风味物质香气活度值。结果表明:食窦魏斯氏菌NCU034005与柠檬明串珠菌NCU027003纯种发酵泡菜中pH不同程度下降,两株乳酸菌对碳水化合物、有机酸与氨基酸的代谢能力不同;主成分分析表明食窦魏斯氏菌NCU034005与柠檬明串珠菌NCU027003纯种发酵泡菜中的代谢物与风味存在显著差异,且有自身的代表性标志代谢物与风味因子。食窦魏斯氏菌NCU034005与柠檬明串珠菌NCU027003对泡菜代谢物与风味有显著影响。     

肉制品中的绿色魏斯氏菌

尽管魏斯氏菌无处不在,不过魏斯菌属的细菌都没有受到足够的重视。该属的许多成员最初被归为明串珠菌或乳酸菌。随着分子生物学技术的发展,这些系统发育密切相关的细菌被归为一个单独的属,魏斯氏菌属。魏斯氏菌的异型新陈代谢给肉类工业造成相当大的损失。粘液形成和肉制品绿化被认为跟绿色魏斯氏菌密切相关。主要介绍了绿色魏斯氏菌对肉类加工的影响。     

低温条件下即食虾中单增李斯特菌与副溶血性弧菌共存分子预测模型的建立

本研究旨在应用实时荧光定量聚合酶链式反应（quantitative real-time polymerase chain reaction,qPCR）技术描述即食虾中的单增李斯特菌与副溶血性弧菌的生长行为,构建混菌模式下的分子预测模型。将单增李斯特菌与副溶血性弧菌等量（（5.0±0.5）（lg（CFU/g）））混合接种于即食虾中,置于低温环境（4 ℃和10 ℃）下培养,并利用qPCR定量检测单增李斯特菌与副溶血性弧菌数量的动态变化。运用生长模型（修正Gompertz、Logistic、Baranyi）和失活模型（Log-linear、Weibull）分别拟合两株菌的生长和失活趋势。结果表明：低温条件下,修正Gompertz、Logistic和Baranyi模型均可成功拟合单增李斯特菌的生长曲线,其决定系数（R2）均大于0.98。对于副溶血性弧菌,在4 ℃条件下,Log-linear和Weibull模型能够清晰地描述其失活情况,R2分别为0.950和0.945;而10 ℃条件下,2 个失活模型均难以描述其行为变化,R2仅为0.784和0.775。本研究运用分子生物学技术描述即食虾中两种致病菌混合培养的菌量变化,探究混菌模式下微生物的生长失活情况,为分子预测模型的进一步研究提供了新的思路。     

绿色魏斯氏菌实时荧光定量聚合酶链式反应检测方法的建立

以rpoA基因为靶基因,建立绿色魏斯氏菌SYBR Green Ⅰ实时荧光定量聚合酶链式反应（polymerase chain reaction,PCR）快速检测方法。针对rpoA基因设计特异性引物,建立绿色魏斯氏菌实时荧光定量PCR检测体系,通过特异性、灵敏度和重复性实验评价体系的检测效果,同时与常规PCR方法进行比较。结果表明：实时荧光定量PCR方法能够特异性检出绿色魏斯氏菌,对基因组DNA的检测灵敏度达到2.667×10－3 pg/μL,对纯培养物和模拟污染牛肉样品直接检测的灵敏度分别为30 CFU/mL和0.8 CFU/g;与常规PCR相比,实时荧光定量PCR检测的灵敏度是其1 000 倍;不同浓度样品独立重复实验循环阈值的标准差均小于1,变异系数在0.02%～1.28%之间。本研究所建立的绿色魏斯氏菌实时荧光定量PCR检测方法具有特异性好、灵敏度高、重复性好的特点,能够进行准确的定量检测,是快速检测绿色魏斯氏菌的有效手段。     

High hydrostatic pressure inactivation of Lactobacillus viridescens and its effects on ultrastructure of cells

The effect of high hydrostatic pressure treatment on Lactobacillus viridescens, a major spoilage micro-organism of processed meat, was investigated. When pressure was applied at 400, 500 and 600 MPa for 5 min, cell survival in MRS broth was reduced approximately by 2, 7 and 8 log cycles, respectively. The combination of high pressure and temperature increase showed synergistic effects on microbial inactivation. Empty cavities in the cell and DNA denaturation were observed after pressure treatment above 400 MPa. The level of microbial inactivation when Lb. viridescens was pressured in protein-fortified MRS broth and ham were less than half compared with the inactivation achieved in peptone water. Increasing the temperature during high-pressure treatment solved these adverse effects in the ham.     

Leuconostoc carnosum associated with spoilage of refrigerated whole cooked hams in Greece

A polyphasic taxonomic approach was used to identify a major atypical group of gas-forming, arginine-negative lactic acid bacteria associated with spoilage of whole (nonsliced) refrigerated (4 degrees C) cooked hams produced in two Greek industrial meat plants. Biochemical characterization revealed that the ham isolates shared their phenotypic properties with Leuconostoc carnosum, Weissella viridescens, and Weissella hellenica. However, gas chromatographic analysis of cellular fatty acids clearly differentiated the ham isolates from the Weissella spp. None of the isolates contained eicosenoic acid (n-C20:1), which is typically synthesized by W. viridescens, but all strains contained high amounts of C 19cycl acid, which is absent in W. hellenica and has been found in trace amounts in W. viridescens. All strains had similar cellular fatty acid profiles, which were qualitatively similar to those of the cellular fatty acids of L. carnosum. In addition to the phenotypic and chemotaxonomic tests, three representative isolates were studied using a lactic acid bacteria database, which employs 16S and 23S HindIII restriction fragment length polymorphism patterns as operational taxonomic units in a numerical analysis. The isolate patterns were identical to those of the L. carnosum type strain, NCFB 2776T. Based on the polyphasic taxonomic approach, the dominating lactic acid bacteria group was identified as L. carnosum.     

不同储藏时期香肠菌群结构的变化及优势腐败菌研究

以天然肠衣的熏煮香肠为研究对象,将新鲜香肠和腐败香肠分别取样,提取DNA,采用高通量测序技术对两种香肠的菌群结构进行比较研究,并确定优势腐败菌。结果表明:腐败香肠菌群丰度和均匀度较新鲜香肠降低;在属的水平上,新鲜香肠中丰度最高的是Staphylococcus(葡萄球菌属90.64%),而腐败香肠中丰度最高的是Weissella(魏斯氏菌属65.93%),其次是Staphylocococcus(葡萄球菌属33.79%);在种的分类水平上,新鲜香肠中Staphylococcus equorum(马胃葡萄球菌25.59%)和Staphylococcus vitulinus(小牛葡萄球菌18.01%)丰度最高,腐败香肠中丰度最高的是Weissella viridesecens(绿色魏斯氏菌46.04%),其次是Weissella cibaria(食窦魏斯氏菌19.68%)。结果提示:与新鲜香肠的菌群组成相比较,腐败香肠的菌群结构发生了显著的变化;腐败香肠中的优势腐败菌是Weissella viridesecens,该种微生物在香肠储藏过程中大量增殖,主导了产品的腐败;在生产过程中需消除Weissella viridesecens,以延长产品货架期,提高产品品质。     

热压结合处理对低温火腿中耐压腐败菌的抑制作用

本文主要探究热压结合处理(50 ℃,350 MPa保压10 min)对低温火腿中绿色魏斯菌和肠膜明串珠菌的抑制作用。通过薄层平板计数法对热压结合处理的杀菌效果进行判断,使用流式细胞技术判断细胞的生理状态,通过扫描电镜(scanning electron microscope,SEM)和透射电镜(transmission electron microscope,TEM)观察细胞形态结构的变化,利用紫外分光光度法测定热压结合处理对细菌细胞膜通透性的影响。结果表明:经热压结合处理后,绿色魏斯菌和肠膜明串珠菌的存活率分别为-4.74和-6.61lg(CFU/mL),其生长变得迟缓,对数期生长速率降低。扫描电镜结果显示热压结合处理对两种菌的形态造成明显的变形,透射电镜结果显示细胞内部出现团状聚集和透电子区。热压结合处理对细胞膜产生损伤,细胞膜通透性变大,导致细胞外紫外吸收物质也极显著增加(p<0.001),细胞处于亚致死状态。热压结合处理会对细胞膜产生一定的形变及损坏,使得细胞内核酸和蛋白质流出,同时细胞内部也会发生蛋白变性和细胞质皱缩,达到抑制耐压菌生长繁殖的目的。     

Effect of high pressure treatment on microbial populations of sliced vacuum-packed cooked ham

In this study, culture-dependent and culture-independent approaches were used to reveal the microbial diversity and dynamic changes occurring in sliced vacuum-packed cooked ham after high pressure processing (HPP, 400MPa or 600MPa for 10min at 22°C) during refrigerated storage over 90days. Direct extraction of genome DNA and total RNA from meat samples, followed by PCR-denaturing gradient gel electrophoresis (DGGE) and RT-PCR-DGGE on 16S rDNA V3 region, was performed to define the structure of the bacterial populations and active species in pressurized cooked ham. Results showed that HPP affected differently the various species detected. The predominant spoilage organisms of cooked ham, such as Lactobacillus sakei and Lactobacillus curvatus, were found to be very sensitive to pressure as they were unable to be detected in HPP samples at any time during refrigerated storage. Weissella viridescens and Leuconostoc mesenteroides survived HPP at 600MPa for 10min at 22°C and were responsible for the final spoilage. An RNA-based DGGE approach clearly has potential for the analysis of active species that have survived in pressurized cooked ham. High pressure processing at 400 or 600MPa for 10min at room temperature (22°C) has a powerful inhibitory effect on the major spoilage bacteria of sliced vacuum-packed cooked ham. High pressure treatment may lead to reduced microbial diversity and improve the products' safety.     

Inactivation of food spoilage bacteria by high pressure processing: Evaluation with conventional media and PCR–DGGE analysis

Microbial diversity and dynamic changes of sliced vacuum-packed cooked ham during refrigerated storage (0–90 days) after high pressure processing (400 MPa at 22 °C for 10 min) was investigated by using culture-dependent and culture-independent approaches. Isolation of genome DNA and total RNA directly from meat samples, followed by PCR–denaturing gradient gel electrophoresis (DGGE) and RT-PCR–DGGE on 16S rDNA V3 region, was performed to describe the structure of the bacterial community and active species in pressurized sliced cooked ham. The DGGE profile showed that most spoilage bacteria including Lactococcus garvieae, Weissella cibaria, Lactobacillus sakei, Lactobacillus curvatus, Weissella paramesenteroides, Leuconostoc carnosum and Lactococcus lactis subsp. lactis were completely inactivated after high pressure processing (HPP), whereas Weissella viridescens and Weissella minor survived HPP and induced the final spoilage. The microbial diversity of HPP samples during the whole refrigerated storage period was extremely simple. Our results clearly indicated that HPP was an efficient method for avoiding the growth of the major spoilage bacteria and could be used to prolong the shelf-life of sliced vacuum-packed cooked ham.     

Influence of Pressurization Rate and Mode on Inactivation of Natural Microorganisms in Purple Sweet Potato Nectar by High Hydrostatic Pressure

High hydrostatic pressure (HHP) processing was applied for inactivation of natural microorganisms, including total aerobic bacteria (TAB), and yeasts and molds (Y&M) in purple sweet potato nectar (PSPN). The pressurization rates were 60 and 120 MPa/min, the pressurization modes were stepwise and linear, the pressure-holding times were 2.5–25 min, and the pressure levels were 400–600 MPa. In all the experimental conditions, the Y&M in PSPN were not detected, the maximum reduction of TAB was greater than 4 log cycles, and the inactivation of TAB was closely related to the HHP parameters. The fast pressurization rate and linear pressurization mode enhanced the inactivation effect of HHP on TAB. With increasing the pressure level and pressure-holding times, the inactivation of TAB was also enhanced. The mathematical models were fitted to the inactivation kinetic data of TAB and fitness of these models was investigated; the Weibull and Biphasic model successfully fitted all the inactivation curves. Pressurization rate and mode had a significant impact on the parameters of the models.