Leuconostoc gelidum and Leuconostoc gasicomitatum strains dominated the lactic acid bacterium population associated with strong slime formation in an acetic-acid herring preserve

Spoilage characterised by strong slime and gas formation affected some manufacture lots of an acetic-acid Baltic herring (Culpea haerengus membras) preserve after few weeks of storage at 0-6 degrees C. The product consisted of herring filets in acetic acid marinade containing sugar, salt, allspice and carrot slices. Microbiological analyses of the spoiled product showed high lactic acid bacterium (LAB) levels ranging from 4.5x10(8) to 2.4x10(9) CFU/g. Yeasts were not detected in any of the herring samples. Since LAB contaminants are seldom associated with fresh fish, LAB populations associated with marinade ingredients (carrots, allspice) were also analyzed. The highest LAB levels exceeding 10(7) CFU/g were detected in equilibrium modified atmosphere packaged baby carrots whereas the levels detected in the allspice samples did not exceed 4.3x10(5). A total of 176 randomly selected LAB isolates originating from herring, carrot and allspice samples were further identified to species level using a 16 and 23S rRNA gene RFLP (ribotyping) database. Leuconostoc gelidum and Leuconostoc gasicomitatum strains dominated both in the spoiled herring and carrot samples. These species are heterofermentative-producing CO(2) from glucose and they also produce dextran from sucrose. Inoculation of some commercial-herring products with spoilage-associated L. gelidum and L. gasicomitatum strains verified that these strains have the capability of producing slime and gas in herring preserves although slime formation was not as strong as in the original samples. Since L. gelidum and L. gasicomitatum strains were commonly detected in carrots, carrot slices used for the fish marinade were considered to be the probable source of these specific spoilage organisms.     

Leuconostoc gasicomitatum is the dominating lactic acid bacterium in retail modified-atmosphere-packaged marinated broiler meat strips on sell-by-day

Lactic acid bacteria (LAB) in retail, modified-atmosphere-packaged (MAP), marinated broiler meat strips on sell-by-day were mainly identified as Leuconostoc gasicomitatum. A total of 32 packages, three to five packages of seven differently marinated broiler meat products, were studied at the end of the producer-defined shelf life (at 6 degrees C, 7-9 days depending on the manufacturer). Prior to the microbiological analyses, appearance and smell of the product was checked and pH measured. Bacteria were cultured on MRS and Tomato Juice Agar (TJA), Rogosa SL agar (SLA), Plate Count Agar (PCA) and Streptomycin Thallium Acetate Agar (STAA) for the enumeration of LAB, lactobacilli, total bacterial count and Brochothrix thermosphacta, respectively. The average CFU/g of the 32 packages was 2.3 x 10(8) on PCA. The highest bacterial average, 3.1 x 10(8), was recovered on TJA, the corresponding CFU/g averages on MRS and SLA being 2.3 x 10(8) and 1.3 x 10(8), respectively. Despite the high LAB numbers detected, radical spoilage changes such as unpleasant odor, slime production and formation of gas were not seen. B. thermosphacta did not form a significant part of the bacterial population since none of the levels exceeded the spoilage threshold level of 10(5) CFU/g reported in previous studies for this organism. In order to characterize the dominating LAB population, as many as 85, 85 and 88 colonies from MRS, TJA and SLA, respectively, were randomly picked and cultured pure. LAB were identified to species level using a 16 and 23S rDNA HindIiI RFLP (ribotyping) database. Fifty-six of the 170 isolates picked from the non-selective LAB media (MRS and TJA) were identified as L. gasicomitatum, followed by Carnobacterium divergens (41 isolates), Lactobacillus sakei and Lactobacillus curvatus subsp. melibiosus (31 isolates) and L. curvatus subsp. curvatus (20 isolates) species. SLA proved not to be completely selective for lactobacilli because the growth of Leuconostoc spp. was not inhibited, Carnobacterium spp. were the only species not detected on SLA.     

Detection of Leuconostoc strains at a meat processing plant using polymerase chain reaction

To simplify the labor-intensive conventional routine testing of samples to detect Leuconostoc at a meat processing plant, we developed polymerase chain reaction (PCR) primers specific for Leuconostoc from 16S rRNA gene sequences. These primers did not detect other common lactic acid bacteria such as Lactobacillus plantarum, Lact. sake, Lact. fermentum, Lact. acidophilus and Weissella viridescens. PCR with this primer detected all Leuconostoc species tested (Leu. mesenteroides subsp. mesenteroides, Leu. pseudomesenteroides, Leu. carnosum, Leu. lactic, Leu. citreum, Leu. amelibiosum, Leu. gelidum), except for Leu. fallax, and no other lactic acid bacteria on agarose gel electrophoresis. The method could identify areas contaminated with Leuconostoc in a large-scale industrial meat processing plant. Of 69 samples analyzed, 34 were positive for Leuconostoc according to the conventional culture method (isolation of LAB producing dextran) and PCR, whereas 29 were negative according to both. Six samples were culture-negative but positive by PCR. No false negative results were generated by PCR. The method is rapid and simple, is useful for routinely monitoring areas contaminated with Leuconostoc in meat processing plants, and could help to prevent the spoilage of meat products.     

Bacterial community structure in kimchi, a Korean fermented vegetable food, as revealed by 16S rRNA gene analysis

Kimchi is a traditional Korean food fermented from a variety of vegetables. We elucidated the microbial community structure of five commercially produced kimchis made from Chinese cabbage by examining culture-independent 16S rRNA gene clone libraries. Most of the clones (347 out of 348) belonged to lactic acid bacteria and included several species of the genera Lactobacillus, Leuconostoc and Weissella. Weissella koreensis was found in all the samples and predominated in three of them (42.6-82%). Leuconostoc gelidum, Leuconostoc gasicomitatum and Lactobacillus sakei were common in the remaining kimchi clone libraries (>34%). The composition of bacterial phylotypes in kimchi varied between samples. Our approach revealed different community structures from those reported in previous culture-dependent studies based on phenotypic identification methods. The culture-independent method used here proved to be efficient and accurate and showed that the bacterial communities in kimchi differ from those in other fermented vegetable foods.     

Identification of Leuconostoc species associated with the spoilage of vacuum-packaged Vienna sausages by DNA-DNA hybridization

Approximately 36% of predominant microbiological spoilage populations from vacuum-packaged Vienna sausages consist of leuconostocs which are difficult to identify phenotypically. Seven leuconostocs selected on a pro-rata basis as representatives of three phenotypically different groups from this environment were identified by DNA-DNA hybridization as Leuconostoc mesenteroides subsp. mesenteroides (3), Leuconostoc gelidum (1) and Leuconostoc paramesenteroides (3). The presence of high proportions of Leuconostoc paramesenteroides and absence of Leuconostoc carnosum in this spoilage association was noteworthy. DNA-DNA hybridization represented a reliable identification method for meat spoilage leuconostocs.     

Selection and Use of Phytate-Degrading LAB to Improve Cereal-Based Products by Mineral Solubilization During Dough Fermentation

ABSTRACT:  New producers of phytate-degrading enzymes, especially lactic acid bacteria (LAB), were used to improve mineral solubilization during dough fermentation. In all, among strains from different sources by microorganisms (150 lactic acid bacteria, 36 yeasts), 38 (24%) exhibited a clear zone around the colonies by hydrolyzing hexacalcium phytate contained in solid medium. When phytase-positive strains from plate assay were tested for phytase activity in liquid medium, 6 of the strains (37%) exhibited phytate-degrading activity in at least one of the 3 different media used. Of the LAB, the highest phytase values were found for  Enterococcus faecium  A86 (0.74 U/mL) and  Lactobacillus plantarum  H5 (0.71 U/mL). Two different starter cultures obtained by combinations of phytase-positive (phy+:  L. plantarum  H5 and L3,  Leuconostoc gelidum  A16, and  E. faecium  A86) or phytase-negative (phy−:  L. gelidum  LM249,  L. plantarum  H19, and  L. plantarum  L8) selected LAB strains, were used to measure mineral concentrations of iron, zinc, and manganese during dough fermentation. Although the 2 kinds of starter showed similar acidic values, the presence of phytate-degrading LAB strains increased mineral solubilization in comparison to the starter phy−.     

Characterization of a Leuconostoc gelidum bacteriophage from pork

A new bacteriophage (phage ggg) and its host, Leuconostoc gelidum LRC-BD, were isolated from vacuum-packaged pork loins. Homogenates of pork loin tissue were enriched with L. gelidum LRC-BD to isolate phages. Cultural, biochemical and genetic methods were used to compare L. gelidum LRC-BD and the type strain, L. gelidum ATCC 49366. The phages were characterized by host range, morphology and phage-bacterial interaction in All Purpose Tween (APT) broth and on pork adipose tissue. With the exception of its inability to produce dextran from sucrose and the fermentation of l-arabinose, L. gelidum LRC-BD was culturally and biochemically similar to L. gelidum ATCC 49366. DNA-relatedness of the strains was confirmed by sequencing of the 16s rRNA gene. Electron microscopic observation revealed that phage ggg was a member of the Siphoviridae. The host range was limited to L. gelidum isolates from meats. Phages were able to replicate and limit the growth of L. gelidum LRC-BD in APT broth incubated aerobically and anaerobically at 4 degrees C, with a multiplicity of infection (MOI) of 0.001. When inoculated pork adipose tissue was stored at 4 degrees C in air or vacuum, phages could multiply but a higher MOI (0.01 to 1000) was necessary to limit the growth of L. gelidum LRC-BD. Naturally occurring phages may affect the numbers of L. gelidum and other lactic acid bacteria residing in meats and thereby alter the storage quality or the preservative potential of competitive strains.     

Comparison of microbial communities in marinated and unmarinated broiler meat by metagenomics

Most raw poultry sold in Finland at the retail level is mixed with marinades containing oil, sugar, spices and acetic acid and packaged under modified atmosphere. Premature spoilage of marinated poultry preparations has been observed and associated with high levels of Leuconostoc spp. in meat. In this study we investigated whether marination of broiler fillet strips increased the proportion of Leuconostoc spp. in the microbial communities. To obtain a comprehensive view of the microbiota, we sequenced total DNA and 16S rRNA gene amplicons from the microbial communities. The lactic acid bacterial communities were characterized also by identification of colonies. The results showed that marinade increased the proportions of the spoilage-associated Leuconostoc gasicomitatum in the communities as well as the proportions of Leuconostoc gelidum and Lactobacillus spp. The proportions of Carnobacterium, Vagococcus, Brochothrix thrermosphacta, Clostridium, Enterobacteriaceae and Vibrio were diminished in marinated meat. Analysis of 16S rRNA gene amplicons resulted in 312 and 284 operational taxonomical units (dissimilarity 0.03) in unmarinated and marinated meat, respectively, indicating that the meat communities were more diverse than hitherto shown. Metagenomic analysis revealed a number of bacterial taxa that have not been associated with late shelf-life meat before, including Vagococcus and Vibrio that belonged to the predominating part of the microbial community in unmarinated meat. According to the functional analysis of the metagenomes, the communities in both marinated and unmarinated poultry were characterized by high proportions (15.6% or 17.9%) of genes involved in carbohydrate metabolism.     

Autoinducer-2-like activity in lactic acid bacteria isolated from minced beef packaged under modified atmospheres

Fifteen fingerprints (assigned to Leuconostoc spp., Leuconostoc mesenteroides, Weissella viridescens, Leuconostoc citreum, and Lactobacillus sakei) of 89 lactic acid bacteria (LAB) isolated from minced beef stored under modified atmospheres at various temperatures were screened for their ability to exhibit autoinducer-2 (AI-2)-like activity under certain growth conditions. Cellfree meat extracts (CFME) were collected at the same time as the LAB isolates and tested for the presence of AI-2-like molecules. All bioassays were conducted using the Vibrio harveyi BAA-1117 (sensor 1(-), sensor 2(+)) biosensor strain. The possible inhibitory effect of meat extracts on the activity of the biosensor strain was also evaluated. AI-2-like activity was observed for Leuconostoc spp. isolates, but none of the L. sakei strains produced detectable AI-2-like activity. The AI-2-like activity was evident mainly associated with the Leuconostoc sp. B 233 strain, which was the dominant isolate recovered from storage at 10 and 15°C and at the initial and middle stages of storage at chill temperatures (0 and 5°C). The tested CFME samples displayed low AI-2-like activity and inhibited AI-2 activity regardless of the indigenous bacterial populations. The LAB isolated during meat spoilage exhibited AI-2-like activity, whereas the LAB strains retrieved depended on storage time and temperature. The production of AI-2-like molecules may affect the dominance of different bacterial strains during storage. The results provide a basis for further research concerning the effect of storage temperature on the expression of genes encoding AI-2 activity and on the diversity of the ephemeral bacterial population.     

高通量测序分析正常及腐败熟成牛肉和牛舌细菌多样性

目的 解析正常与产生腐败异味的熟成牛肉和牛舌的优势细菌。方法 利用高通量测序技术分析熟成牛肉和牛舌的细菌多样性。结果 4组样品共获得有效序列数197144,基于序列16SrRNA基因97%的相似度,可划分为498个操作分类单元(operational taxonomic unit,OTU)。正常熟成牛肉(BG)、腐败熟成牛肉(BB)、正常熟成牛舌(BTG)、腐败熟成牛舌(BTB)样本OTU数分别为82、32、165、219。4组样品的Coverage、Sobs、Ace、Chao1与Shannon指数表明细菌多样性依次为:BTB>BTG>BG>BB。BG的优势菌为厚壁菌门(Firmicutes,占比94.5%)的乳杆菌属(Lactobacillus,占比91.0%);BB的优势菌为变形菌门(Proteobacteria,占比99.0%)的不动杆菌属(Acinetobacter,占比92.3%)。BTG的优势菌为Firmicutes(占比67.6%)的Lactobacillus(占比62.8%); BTB的优势菌为Proteobacteria(占比80%)的假单胞菌属(P...     

Effective use of nisin to control lactic acid bacterial spoilage in vacuum-packed bologna-type sausage.

Lactic acid bacteria (LAB) commonly cause spoilage in minimal heat-treated vacuum-packed cured delicatessen meats. Predominant species are Lactobacillus sake and L. curvatus. LAB strains isolated from spoiled products of this type (liver sausage, ham and bologna sausage) were found to be sensitive to low nisin concentrations (maximum of 1.25 microg g(-1)). Addition of 25 microg g(-1) nisin (as Nisaplin) inhibited the growth of LAB spoilage organisms inoculated into vacuum-packed pasteurized bologna-type sausages stored at 8 degrees C. Control sausages became spoiled (>10(8) LAB CFU g(-1)) by day 7, whereas sausages containing nisin remained unspoiled for >50 days. The effect of three types of phosphates (used as emulsifiers) on nisin activity in the sausages was compared. LAB growth rate was fastest in samples containing orthophosphate, and slowest in sausages containing diphosphate. The shelf life was also greatly extended in the latter. Fat content also affected nisin activity. Nisin activity (as indicated by LAB inhibition) was greatest in samples containing 15% > 25% > 37% (wt/wt) fat. In a sausage formulation containing 37% fat and incorporating diphosphate as emulsifier, levels of nisin as low as 2.5 microg g(-1) showed antibacterial effects. A nisin level of 6.25 microg g(-1) totally inhibited LAB growth for over 4 weeks and 25 microg g(-1) for 5 weeks. Spoilage control was achieved in the same sausage formulation but with 25% (wt/wt) fat; 12.5 microg g(-1) nisin prevented LAB growth for 5 weeks.     

Culture media for non-sporulating gram-positive food spoilage bacteria.

The spoilage association especially of protein-rich foods can be dominated by Gram-positive bacteria, notably lactic acid bacteria (LAB) which affect vacuum packaged refrigerated processed meats and some dairy products. New food ecosystems are being created by novel packaging and processing technologies, resulting in spoilage associations differing from those previously reported. In addition, improvement in identification methods, allow the detection and isolation of 'novel' bacterial groups, e.g., Carnobacterium spp. This review considers the genera Aerococcus, Brevibacterium, Brochothrix, Carnobacterium, Kurthia, Lactobacillus, Leuconostoc, Microbacterium, Micrococcus, Pediococcus and Propionibacterium. Strictly selective procedures, including incubation temperature and atmosphere, are not yet available for the genera Aerococcus, Brevibacterium, Microbacterium and Micrococcus, and only with some limitations for Kurthia and Propionibacterium. On the other hand, a causative role in food spoilage has not been established clearly for all those groups, some of which may be 'opportunistic' in their behaviour. The LAB groups Lactobacillus, Leuconostoc and Pediococcus ('LLP-Group') often share similar habitats and show similar physiological behaviour on a number of elective and selective media. Modifications to increase selectivity have been based mainly on de Man, Rogosa and Sharpe (MRS) or Rogosa agar, and include pH reduction, supplementation with chemical preservatives (e.g., sorbic acid and nitrate) and the use of reduced atmospheres or suboptimal incubation temperatures. Carnobacteria differ from other LAB in their non-aciduric nature, and selective plating procedures use high-pH media (pH 8-9) by which competitors (mainly lactobacilli) are eliminated.     

冷鲜牛肉贮藏中菌群结构及优势菌致腐性的分析

为探究牛肉在0?℃冷藏下微生物菌群变化和优势腐败菌,采用培养依赖的16S rRNA结合高通量测序技术分析牛肉样品的微生物多样性变化。通过定期测定接种牛肉汁的生长、感官、蛋白酶活性、pH值和挥发性盐基氮（total volatile basic nitrogen,TVB-N）值,分析分离株的致腐特征。结果表明,牛肉冷藏中感官品质保持良好,15?d出现异味,18?d有腐臭味。而样品中菌落总数第3天快速上升,15?d菌落总数为8.73（lg（CFU/g））,之后呈现稳定,假单胞菌（Pseudomonas spp.）、热死环丝菌（Brochothrix thermosphacta）、肠杆菌（Enterobacter）和乳酸菌4 种分离培养基中细菌与菌落总数增长趋势相似,其中假单胞菌增长最快,肠杆菌数和乳酸菌数生长最慢。两种菌群鉴定结果显示,冷鲜牛肉初始微生物构成复杂,包括热死环丝菌、不动杆菌属（Acinetobacter spp.）、气单胞菌属（Aeromonas spp.）和假单胞菌等多种菌属构成,而腐败末期菌群构成趋于单一,假单胞菌和热死环丝菌为优势腐败菌,特别是莓实假单胞菌（P. fragi）。将腐败分离株10?株假单胞菌、4?株热死环丝菌和1?株蜂房哈夫尼菌（Hafnia alve）接种于冷藏的牛肉汁,发现假单胞菌和蜂房哈夫尼菌的接种组感官评分、pH值和TVB-N值高于热死环丝菌,且假单胞菌有较强的蛋白酶活性。研究表明,结合感官和微生物评价冷鲜牛肉贮藏期为15?d,且菌群多样性下降,假单胞菌和热死环丝菌是优势腐败菌,其中假单胞菌致腐性较强。     

The Occurrence of Beer Spoilage Lactic Acid Bacteria in Craft Beer Production

Beer is one of the world's most ancient and widely consumed fermented alcoholic beverages produced with water, malted cereal grains (generally barley and wheat), hops, and yeast. Beer is considered an unfavorable substrate of growth for many microorganisms, however, there are a limited number of bacteria and yeasts, which are capable of growth and may spoil beer especially if it is not pasteurized or sterile‐filtered as craft beer. The aim of this research study was to track beer spoilage lactic acid bacteria (LAB) inside a brewery and during the craft beer production process. To that end, indoor air and work surface samples, collected in the brewery under study, together with commercial active dry yeasts, exhausted yeasts, yeast pellet (obtained after mature beer centrifugation), and spoiled beers were analyzed through culture‐dependent methods and PCR‐DGGE in order to identify the contaminant LAB species and the source of contamination. Lactobacillus brevis was detected in a spoiled beer and in a commercial active dry yeast. Other LAB species and bacteria ascribed to Staphylococcus sp., Enterobaceriaceae, and Acetobacter sp. were found in the brewery. In conclusion, the PCR‐DGGE technique coupled with the culture‐dependent method was found to be a useful tool for identifying the beer spoilage bacteria and the source of contamination. The analyses carried out on raw materials, by‐products, final products, and the brewery were useful for implementing a sanitization plan to be adopted in the production plant.     

The spoilage microflora of cured, cooked turkey breasts prepared commercially with or without smoking

Lactobacillus sakei subsp. carnosus was predominant in the spoilage flora of sliced, vacuum-packed, smoked, oven-cooked turkey breast fillets which developed mild, sour spoilage flavors after 4 weeks storage at 4 degrees C. In contrast, Leuconostoc mesenteroides subsp. mesenteroides predominated in the spoilage flora of sliced, vacuum-packed, unsmoked, boiled turkey breast fillets from the same plant which were also stored at 4 degrees C. The spoilage flora of the unsmoked breasts grew faster than that of the smoked breasts and was more diverse. Lactobacillus sakei, Weissella viridescens and an atypical group of leuconostoc-like bacteria were also members of the unsmoked turkey breasts flora. Consequently, the unsmoked breasts spoiled after 2 weeks at 4 degrees C: the packs swelled and the meat developed strong sour odors and flavors and abundant slime. Except for the unidentified leuconostocs, which apparently survived boiling of the unsmoked turkey, all the spoilage organisms contaminated the meats during the slicing and vacuum packaging operations. From their biochemical reactions and cellular fatty acid profiles, the atypical leuconostocs were more closely related to Leuconostoc carnosum than W. viridescens. Carnobacteria and Brochothrix thermosphacta were present in relatively large numbers on the raw turkey, but were not numerous in the spoilage flora of the cooked, vacuum-packed meat products.     

Contribution of Lactobacillus plantarum and Leuconostoc mesenteroides subsp. mesenteroides to Atypical Odors of Beef Strip Loins

Sterile beef strip loin tissue was inoculated with Lactobacillus plantarum or Leuconostoc mesenteroides subsp. mesenteroides, placed in sterile sample bottles which were purged with CO₂, and stored up to 28 days at 3°C. Volatile compounds detected in the headspace of these samples included acetone, toluene, acetic acid, ethyl acetate, a C₇ hydrocarbon, and trichloromethane. The profile of volatiles in packaged sterile loin tissue stored for up to 28 days was similar. However, the inoculated samples spoiled (soured) at a faster rate.     

Lactic acid bacteria in marinades used for modified atmosphere packaged broiler chicken meat products

Lactic acid bacteria (LAB) in some marinades commonly used in Finland for modified atmosphere packaged poultry meat products were enumerated and identified to determine whether the marinades contained LAB species that cause meat spoilage. The concentrations of LAB in 51 marinade samples ranged from less than 100 to 8.0 x 10(5) CFU/ml. Seventeen of the samples produced LAB growth only after enrichment, and in five samples no growth was detected either by direct culturing or enrichment. Eighty-eight randomly selected isolates, 51 from the enumerated plates and 37 from enriched samples, were identified using a database of 16S and 23S rRNA gene HindIII restriction fragment length polymorphism patterns of over 300 type and references LAB strains as operational taxonomic units in numerical analyses. The predominating LAB in the enumerated samples was Lactobacillus plantarum (25 of 51 isolates). Eleven isolates were identified as Lactobacillus paracasei subsp. paracasei, and nine were Lactobacillus parabuchneri. None of these species are considered specific spoilage LAB in marinated modified atmosphere packaged poultry meat products nor have they been reported to dominate in unspoiled late-shelf-life products. These results indicate that even though marinades may contain high numbers of LAB, they are not necessarily sources of specific meat spoilage LAB. Therefore, risks associated with meat quality are not predicted by quantitative enumeration of LAB in marinades.     

Enterococcus species dominating in fresh modified-atmosphere-packaged, marinated broiler legs are overgrown by Carnobacterium and Lactobacillus species during storage at 6 degrees C

In order to show which of the initial lactic acid bacteria (LAB) contaminants are also causing spoilage of a modified-atmosphere-packaged (MAP), marinated broiler leg product at 6 degrees C, LAB were enumerated and identified on the 2nd and 17th days following manufacture. A total of 8 fresh and 13 spoiled packages were studied for LAB levels. In addition, aerobic mesophilic bacteria and Enterobacteriaceae were determined. The average CFU/g values in the 8 fresh packages were 1.3 x 10(3), 9.8 x 10(3) and 2.6 x 10(2) on de Man Rogosa Sharpe agar (MRS), Plate Count Agar (PCA) and Violet Red Bile Glucose agar (VRBG), respectively. The commercial shelf life for the product had been set as 12 days, and all packages analyzed on the 17th day were deemed unfit for human consumption by sensory analysis. The corresponding CFU/g averages in the spoiled product were 1.4 x 10(9), 1.1 x 10(9) and 3.9 x 10(7) on MRS, PCA and VRBG agar, respectively. For characterization of LAB population, 104 colonies originating from the fresh packages and 144 colonies from the spoiled packages were randomly picked, cultured pure and identified to species level using a 16 and 23S rDNA HindIII RFLP (ribotyping) database. The results showed that enterococci (35.7% of the LAB population) were dominating in the fresh product, whereas carnobacteria (59.7%) dominated among the spoilage LAB. Enterococcus faecalis, Carnobacterium piscicola and Carnobacterium divergens were the main species detected. In general, when the initial LAB population is compared to the spoilage LAB, a shift from homofermentative cocci towards carnobacteria, Lactobacillus sakei/curvatus and heterofermentative rods is seen in this marinated product.     

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.     

Monitoring the lactic acid bacterial diversity during shochu fermentation by PCR-denaturing gradient gel electrophoresis

The presence of lactic acid bacteria (LAB) during shochu fermentation was monitored by PCR-denaturing gradient gel electrophoresis (DGGE) and by bacteriological culturing. No LAB were detected from fermented mashes by PCR-DGGE using a universal bacterial PCR primer set. However, PCR-DGGE using a new primer specific for the 16S rDNA of Lactococcus, Streptococcus, Tetragenococcus, Enterococcus, and Vagococcus and two primers specific for the 16S rDNA of Lactobacillus, Pediococcus, Leuconostoc, and Weissella revealed that Enterococcus faecium, Lactobacillus casei, Lactobacillus fermentum, Lactobacillus nagelii, Lactobacillus plantarum, Lactococcus lactis, Leuconostoc citreum, Leuconostoc mesenteroides, and Weissella cibaria inhabited in shochu mashes. It was also found that the LAB community composition during shochu fermentation changed after the main ingredient and water were added during the fermentation process. Therefore, we confirmed that PCR-DGGE using all three primers specific for groups of LAB together was well suited to the study of the LAB diversity in shochu mashes. The results of DGGE profiles were similar to the results of bacteriological culturing. In conclusion, LAB are present during shochu fermentation but not dominant.