Co-culture experiments demonstrate the usefulness of Lactobacillus sakei 10A to prolong the shelf-life of a model cooked ham

This study investigated the usefulness of two selected lactic acid bacteria, Lactobacillus sakei subsp. carnosus (10A) and the lactocin S producing L. sakei 148 (LS5), to extend the shelf-life of cooked meat products. The interaction between these potential protective cultures and the spoilage organisms, Leuconostoc mesenteroides (LM4) and Brochothrix thermosphacta (BT1), were examined in co-culture studies on a model cooked ham product at 7 degrees C under vacuum packaged conditions. Furthermore, the influence of the glucose content of the model cooked ham on the interaction phenomena was investigated. When artificially contaminating the model cooked ham with BT1 at 10(2) cfu/g in combination with 10A at 10(5) cfu/g, the growth of BT1 was significantly slower compared to a simultaneous mono-culture experiment. In a similar experiment with LM4, LM4 reached a level of 10(7) cfu/g +/-14 days later when LM4 grew together with 10A compared to its growth in mono-culture. The lactocin S producing LS5 did not demonstrate an inhibitory action towards LM4 or BT1 and is therefore not useful as protective culture on cooked meat products. The glucose level of the model cooked ham had no influence on the observed antagonistic interactions of 10A towards LM4 or BT1, indicating that the action of the biopreservative 10A in cooked meat products is independent of the substrate glucose.     

剑门火腿菌相构成及主要腐败菌特性分析

采用选择性培养基法对剑门火腿中的菌相构成进行了分析。结果表明,剑门火腿中的微生物主要为葡萄球菌、微球菌和酵母菌,其表层数量明显高于内部,乳酸菌主要分布于火腿内部,霉菌主要存在于火腿表层。剑门火腿中主要细菌的耐盐、耐酸、耐亚硝酸盐及致腐能力分析结果显示,葡萄球菌和乳酸菌的某些菌株是导致其腐败的主要微生物。     

Microbial inactivation after high-pressure processing at 600 MPa in commercial meat products over its shelf life

The behaviour of spoilage and pathogenic microorganisms was evaluated after high-pressure treatment (600 MPa 6 min, 31 °C) and during chilled storage at 4 °C for up to 120 days of commercial meat products. The objective was to determine if this pressure treatment is a valid process to reduce the safety risks associated with Salmonella and Listeria monocytogenes, and if it effectively avoids or delays the growth of spoilage microorganisms during the chilled storage time evaluated. The meat products covered by this study were cooked meat products (sliced cooked ham, pH 6.25, a_w 0.978), dry cured meat products (sliced dry cured ham, pH 5.81, a_w 0.890), and raw marinated meats (sliced marinated beef loin, pH 5.88, a_w 0.985). HPP at 600 MPa for 6 min was an efficient method for avoiding the growth of yeasts and Enterobacteriaceae with a potential to produce off-flavours and for delaying the growth of lactic acid bacteria as spoilage microorganisms. HPP reduced the safety risks associated with Salmonella and L. monocytogenes in sliced marinated beef loin.     

Selective effect of the product type and the packaging conditions on the species of lactic acid bacteria dominating the spoilage microbial association of cooked meats at 4°C

The product type was shown to strongly affect the growth rate and the composition of the spoilage lactic flora during refrigerated (4°C) storage of cooked, cured meats, sharing their processing plant environment, day of production and film packaging conditions. Growth of lactic acid bacteria (LAB) under vacuum was more prolific on the product in the order: ham>turkey breast fillet>smoked pork loin>pariza>mortadella>bacon, and ham>frankfurters, manufactured in two industrial meat plants A and B, respectively. The Lactobacillus sakei/curvatus group prevailed in all products, except the non-smoked, boiled whole-meats, i.e. cooked ham and turkey breast fillet, where Leuconostoc mesenteroides subsp.mesenteroides predominated. Lactobacillus sakei was by far the most prevalent species in smoked whole-meats, i.e. pork loin and bacon. Emulsion sausages, i.e. pariza, mortadella and frankfurters, contained a more diverse lactic flora.Leuconostoc carnosum and Lc. citreum occurred in boiled, whole-meats and emulsion sausages, respectively.Weissella viridescens was isolated from smoked meat products only. A very good correlation between the LAB growth and types and important intrinsic factors, such as the product pH, moisture, salt (brine) concentration and cooking method could be observed. When ham and frankfurters from plant B were stored in air, yeasts and mainly Brochothrix thermosphacta became important members of the spoilage association. Growth of LAB was faster in air. The presence of oxygen resulted in a replacement ofLc. mesenteroides subsp. mesenteroides by other Leuconostoc spp. in ham, and in a shift of the spoilage flora from homo- to heterofermentative LAB species in frankfurters.     

Relation between microbiological quality,metabolite production and sensory quality of equilibrium modified atmosphere packaged fresh-cut produce

The quality of four types of fresh-cut produce, packaged in consumer-sized packages under an equilibrium modified atmosphere and stored at 7 degrees C, was assessed by establishing the relation between the microbial outgrowth and the corresponding production of nonvolatile compounds and related sensory disorders.In vitro experiments, performed on a lettuce-juice-agar, demonstrated the production of nonvolatile compounds by spoilage causing lactic acid bacteria and Enterobacteriaceae. Pseudomonas fluorescens and yeasts, however, were not able to produce detectable amounts of nonvolatile metabolites.The type of spoilage and quality deterioration in vivo depended on the type of vegetable. Mixed lettuce and chicory endives, leafy tissues, containing naturally low concentrations of sugars, showed a spoilage dominated by Gram-negative microorganisms, which are not producing nonvolatile compounds. Sensory problems were associated with visual properties and the metabolic activity of the plant tissue. Mixed bell peppers and grated celeriac, on the other hand, demonstrated a fast and intense growth of spoilage microorganisms, dominated by lactic acid bacteria and yeasts. This proliferation resulted in detectable levels of organic acids and the rejection by the trained sensory panel was based on the negative perception of the organoleptical properties (off-flavour, odour and taste).The applied microbiological criteria corresponded well with detectable changes in sensory properties and measurable concentrations of nonvolatile compounds, surely in the cases where lactic acid bacteria and yeasts were provoking spoilage. Consequently, the freshness of minimally processed vegetables, sensitive for outgrowth of lactic acid bacteria and yeasts (e.g., carrots, celeriac, bell peppers, mixtures with non-leafy vegetables) can be evaluated via analysis of the produced nonvolatile compounds.     

Modeling the growth of lactic acid bacteria in sliced ham processed by high hydrostatic pressure

The main responsible for the spoilage of cooked cured meat products stored under refrigerated and anaerobic conditions are lactic acid bacteria. The application of high hydrostatic pressure (HHP) reduces the lactic acid bacterial growth extending the product shelf-life and preserving natural taste, texture, color and vitamin content. This work studied the influence of pressure level and holding time on the lactic acid bacterial growth in vacuum-packaged sliced ham. Modified Gompertz and Logistic models were used to fit experimental data obtained from post-treatment microbial counts carried out along the product storage. Samples of sliced vacuum-packaged ham treated by HHP and control samples (non-treated) were stored at 8 °C until the microorganism population reached 10⁷ CFU/g. An experimental planning 2² with triplicate at the central point was designed to determine the influence of pressure level (200, 300, and 400 MPa) and holding time (5, 10, and 15 min) on the product shelf-life. The results have shown that the pressure intensity and the holding time significantly influenced microbial population over the product storage. Shelf-life of ham treated at 400 MPa for 15 min was extended from 19 (control samples) to 85 days.     

Shelf-life Reduction as an Emerging Problem in Cooked Hams Underlines the Need for Improved Preservation Strategies

Cooked hams have gained an important position within the delicatessen market. Nowadays, consumers not only demand superior sensory properties but also request low levels of sodium and fat and the absence of conventional chemicals and preservatives used for the increase of the technological yield and shelf-life of the products. As a result, products that apply strict quality certificates or ‘‘clean’’ labels become increasingly important. However, such cooked hams suffer from a limited shelf-life. Besides some physicochemical effects, this is mainly due to microbial impact, despite the application of modified-atmosphere-packaging and chilling. Microbial spoilage is mostly due to the metabolic manifestation of lactic acid bacteria and Brochothrix thermosphacta, although Enterobacteriaceae and yeasts may occur too. Several preservation strategies have been developed to prolong the shelf-life of such vulnerable cooked meat products by targeting the microbial communities, with different rates of success. Whereas high-pressure treatments do not always pose a straightforward solution, a promising strategy relates to the use of bioprotective cultures containing lactic acid bacteria. The latter consist of strains that are deliberately added to the ham to outcompete undesirable microorganisms. Spoilage problems seem, however, to be specific for each product and processing line, underlining the importance of tailor-made solutions.     

Behaviour of Brochothrix thermosphacta in presence of other meat spoilage microbial groups

The microbial flora of fresh meat stored aerobically at 5 degrees C up to spoilage was enumerated and collected in order to have mixed spoilage bacterial groups to be used in competition tests against Brochothrix thermosphacta. The bacterial groups collected as bulk colonies were identified by PCR-DGGE followed by partial 16S rDNA sequencing. The predominant bacteria associated with the spoilage of the refrigerated beef were B. thermosphacta, Pseudomonas spp, Enterobacteriaceae and lactic acid bacteria (LAB). The interactions between B. thermosphacta and the other spoilage microbial groups were studied in vitro at 5 degrees C. The results showed that a decrease of the growth of B. thermosphacta was evidenced in presence of LAB at 5 degrees C while the bacterium is the dominant organism when inoculated with mixtures of Pseudomonas spp., LAB and Enterobacteriaceae. A better understanding of bacterial meat spoilage interactions may lead to improved quality of fresh meat stored in refrigerated conditions.     

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.     

Protective cultures inhibit growth of Listeria monocytogenes and Escherichia coli O157:H7 in cooked, sliced, vacuum- and gas-packaged meat

Contamination of cooked meat products with Listeria monocytogenes poses a constant threat to the meat industry. The aim of this study was therefore to investigate the use of indigenous lactic acid bacteria (LAB) as protective cultures in cooked meat products. Cooked, sliced, vacuum- or gas-packaged ham and servelat sausage from nine meat factories in Norway were inoculated with 10(3) cfu/g of a mixture of three rifampicin resistant (rif-mutant) strains of L. monocytogenes and stored at 8 degrees C for four weeks. Growth of L. monocytogenes and indigenous lactic acid flora was followed throughout the storage period. LAB were isolated from samples where L. monocytogenes failed to grow. Five different strains growing well at 3 degrees C. pH 6.2, with 3% NaCl, and producing moderate amounts of acid were selected for challenge experiments with the rif-resistant strains of L. monocytogenes. a nalidixic acid/streptomycin sulphate-resistant strain of Escherichia coli O157:H7 and a mixture of three rif-resistant strains of Yersinia enterocolitica O:3. All five LAB strains inhibited growth of both L. monocytogenes and E. coli O157:H7. No inhibition of Y. enterocolitica O:3 was observed. A professional taste panel evaluated cooked, sliced, vacuum-packaged ham inoculated with each of the five test strains after storage for 21 days at 8 degrees C. All samples had acceptable sensory properties. The five LAB strains hybridised to a 23S rRNA oligonucleotide probe specific for Lactobacillus sakei. These indigenous LAB may be used as protective cultures to inhibit growth of L. monocytogenes and E. coli O157:H7 in cooked meat products.     

Automated turbidimetry for rapid determination of the bacteriological quality of raw meat and processed meat products

A fully automated centrifugal spectrophotometer for turbidimetric measurements linked to a computer for data processing was used to record and analyze growth of different types of bacteria associated with spoilage of raw meat and processed meat products (sliced meat for sandwiches) in order to predict bacterial counts. Media for this specific analysis were developed to enumerate total viable count, lactic acid bacteria. Brochothrix thermosphacta and Pseudomonas spp. Correlation coefficients of -0.91 to -0.97 were obtained between the conventional plate count technique and the turbidimetric method (linear regression). Lactic acid bacteria in the range 10(3) to 10(9) cfu/g could be detected within 48 h, whereas total viable count, Brochothrix thermosphacta and Pseudomonas spp. generally could be detected within 24 h. Counts exceeding 10(7) cfu/g could be detected within 6 h.     

乳酸菌的分类鉴定及在食品工业中的应用

乳酸菌是一类革兰氏阳性杆菌或球菌、可収酵乳糖或葡萄糖产生大量乳酸的细菌的通称,将乳酸菌应用到食品中具有提高其营养价值,改善食品风味,抑制食品中腐败菌的生长,延长食品保质期等作用。因其独特的生理特性,近年来国内外学者对乳酸菌的研究逐渐增多。本文主要论述了乳酸菌的分类和鉴定技术,介绍了传统鉴定斱法和分子生物学鉴定斱法的优缺点以及乳酸菌在食品工业(乳制品、肉制品、果蔬制品、食品保鲜)中的应用,为提高乳酸菌的鉴定效率以及乳酸菌在食品工业、饲料生产和临床医疗上的迚一步应用提供参考。     

贵式肉品中乳酸菌的选育及其对发酵里脊火腿风味品质的影响

以锦屏和榕江酸肉、凯里和遵义香肠、威宁和盘县火腿以及凯里、遵义和毕节风肉9 种贵州发酵肉为对象,对其中乳酸菌分离鉴定并进行蛋白质降解、生理生化特性、生产适应性、安全性研究。根据初筛结果选取1 株蛋白降解能力强且具有肉类发酵剂特性的乳酸菌SJ-4,接种发酵里脊火腿,以未接种火腿为对照,分析该菌株对发酵里脊火腿色泽、质构、电子舌响应值及风味的影响。结果表明：菌株SJ-4能耐受6 g/100 mL NaCl、150 mg/kg NaNO2,不产NH3、H2S、H2O2,无氨基酸脱羧酶活性,具有分解蛋白质的能力但无分解脂肪活性,最适温度37 ℃,最适pH 6.0,培养4 h进入对数生长期,培养18 h pH值可降至3.80,已达到肉类发酵剂条件。经16S rDNA测序鉴定为植物乳杆菌（Lactobacillus plantarum）。菌株SJ-4接种对里脊火腿色泽、硬度、黏聚性均影响显著,添加该菌后有助于产品滋味形成,且关键风味物质如壬醛、3-羟基-2-丁酮等含量增加。植物乳杆菌SJ-4具有作为高活性肉制品发酵剂的潜力。     

Combining reformulation,active packaging and non-thermal post-packaging decontamination technologies to increase the microbiological quality and safety of cooked ready-to-eat meat products

BackgroundCooked ready-to-eat (RTE) meat products are subjected to contamination of spoilage microorganisms such as lactic acid bacteria and pathogens such as Listeria monocytogenes. These microorganisms contaminate cooked RTE meat products after the cooking step and may further grow during shelf-life potentially leading to spoilage or foodborne diseases, respectively. In the current context of salt, fat and chemical preservatives reduction in meat products formulations, a combined strategy that considers the development of more robust formulations, active packaging and the use of non-thermal post-packaging decontamination strategies seems required to ensure shelf-stable and safe RTE cooked food products.Scope and approachThe main objective of this review was to discuss the aspects related to reformulation, active packaging and the application of non-thermal decontamination technologies at the post-packaging step of cooked RTE meat products, their advantages, limitations and main challenges for their implementation.Key findings and conclusionsIn general, post-packaging decontamination technologies aim to reduce or inactivate pathogens and spoilage microorganisms present on the surface of ready-to-eat meat products. Low-temperature plasma, high-pressure processing (HPP), pulsed electric fields, pulsed ultraviolet light and ultrasound are promising alternatives in this segment. However, the choice of the most appropriate approach for post-packaging decontamination of cooked ready-to-eat meat products depends on the type of product and the technological objectives. Meat products formulation and packaging material properties should be considered while defining a post-packaging decontamination approach. Although they are advantageous, non-thermal technologies may present certain limitations such as the increase of oxidative reactions over the shelf-life.     

Impact of slicing hygiene upon shelf life and distribution of spoilage bacteria in vacuum packaged cured meats

A model for studying spoilage in sliced cooked delicatessen meats using conventional plating procedures was characterized by following the transfer of adventitious lactic acid bacteria (LAB), thermotolerant LAB and pediococci from a dry fermented sausage to cooked ham and bologna during a simultaneous slicing procedure. The major group of organisms present in these products was LAB. Each of the three groups of organisms was transferred to the cooked meats, but during storage under vacuum at 7°C, only the adventitious lactobacilli from the sausage appeared to contribute to cooked meat spoilage. Pediococci declined within 28 days of co-slicing, thermotolerant LAB grew slowly and never reached levels of >5 log₁₀cfu cm⁻², but transferred adventitious LAB from the sausage grew and reached peak populations 4 weeks earlier in the co-sliced cooked meats than in controls. During storage at 7°C co-sliced cooked meats spoiled, as assessed by purge (liquid) and ropy slime formation, 2–4 weeks faster than the separately-sliced and vacuum-packaged products. The distribution of organisms on package slices differed. In control cooked products numbers of bacteria present on center package slices were lower than on outside slices. Bacterial numbers were greatest at the meat surface–packaging film interface. In co-sliced cooked products organisms were evenly distributed throughout slices in the packages. Based on findings from use of the co-slice model, in vacuum packages of sliced cured meats where >10⁵cfu bacteria cm₋₂are equally distributed throughout package slices, poor slicing hygiene is the cause. Commercially-sliced bologna was repackaged with allyl isothiocyanate (AIT) to test its ability to delay bacterial growth at the meat surface. The preservative effects of AIT were tested under vacuum or when backflushed with CO₂. CO₂alone was more effective than AIT in delaying LAB growth but neither treatment was particularly effective.     

羊肉微生物相调查及山梨酸钾的防腐作用

7组样品统计分析表明,羊肉微生物相主要有8种类型的革兰氏阳性菌和阴性菌组成;从中选取5株代表性菌株并用山梨酸钾对其进行抑菌试验,结果表明,0.25‰以上山梨酸钾能够有效抑制105cfu/ml革兰氏阳性短芽孢杆菌和革兰氏阴性短杆菌的生长。     

卤制鸭腿中乳酸菌生长预测模型研究

测定了卤制鸭腿中的乳酸菌在5、10、15、20、25℃条件下的生长情况,并利用Gompertz模型拟合了测定温度下的乳酸菌生长一级模型和乳酸菌生长状况。结果表明:模型拟合的R2都大于0.99,利用平方根模型描述温度与最大比生长速率和延滞期的关系,建立了乳酸菌在鸭腿基质上的生长预测二级模型,方差分析表明二级模型拟合显著。模型可用于预测5～25℃范围内乳酸菌在鸭腿上的生长变化情况,为卤制鸭腿中腐败微生物的预测研究提供参考。     

Microbiological ecology of marinated meat products

Marinated meat products are consumed increasingly. In addition to taste, marinating has been considered to increase product safety and shelf life. In Finland, marinades are complex, spiced sauces. They are acidic water-oil emulsions typically containing salt, sugar, sorbate and/or benzoate. Marinated products are usually packaged under modified atmospheres. This results in the growth of psychrotrophic, anaerobic bacteria like lactic acid bacteria (LAB). Marinating did not increase the shelf life of Finnish poultry products and it strongly selected novel spoilage LAB. Surprisingly, it neither had inhibitory effect on Campylobacter. The buffering capability of meat neutralizes the acidic marinade and results in dissociation of the lipophilic acids making their antimicrobial effect nonexistent.     

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.     

Inoculation of lactic acid bacteria on meat surfaces as a means of decontamination in semitropical conditions

Reduction of microbial populations on carcasses has been achieved and reported by some authors by spraying solutions of organic acids, mainly lactic acid, on the meat surface. However, in practice, lactic acid is very expensive. Production of lactic acid in situ by a controlled lactic fermentation seemed to be a feasible answer. The objective of the present study was to explore the viability of this concept at semitropical conditions, i.e. temperatures around 25°C. In a first experiment, seven starters were tested for their ability to produce lactic acid and reducing the growth rate of pseudomonads, taking these microorganisms as indicators of contamination by spoilage microorganisms. The lactic acid bacteria strains were isolated from native Mexican maize-based beverages, and inoculated on the meat surface. Addition of sucrose and wrapping the samples in PVC film in order to induce a microaerophilic environment encouraged the over-growth of L. bulgaricus and P. pentosaceous over pseudomonads. A second experiment was designed to test a mixture of these two strains and a commercial starter with respect to lactic acid production, overgrowth of lactic acid bacteria over pseudomonads, decolouration and oxidation of the meat samples. It was concluded that a commercial starter (L. plantarum + M. kristinae-varians) resulted in a reduction of pseudomonas growth rate, without notably affecting meat colour and degree of oxidation.