Studies on an inhibitor produced by lactic acid bacteria of wines on the control of malolactic fermentation

Malolactic fermentation is the microbiological process in wines, where lactic acid bacteria (LAB) govern the process of converting L-malic acid into L-lactic acid. During this process a high microbial load of LAB may lead to an unwanted spoilage phenomena by formation of excessive amounts of undesirable flavor compounds. This study is mainly focused on the isolation of LAB from the native flora of the wine, which has an inhibitory potential against malolactic activity of LAB inherent in wines. An isolate of Leuconostoc mesenteroides subsp. cremoris was found to produce an inhibitory compound against the LAB of wines. This compound was found to be a bacteriocin-like inhibitory substance (BLIS), which has a molecular weight of 32,000 Da, and it was shown that this BLIS was effective in the control of malolactic fermentation.     

Selection of lactic acid bacteria for use in vegetable fermentations

The fermentation of brined vegetables traditionally has depended upon growth of naturally occurring lactic acid bacteria to metabolize the vegetable sugars to organic acids which, together with added salt, results in preservation. Starter cultures have been used only to a limited extent commercially. However, recent efforts to improve fermentation vessels and to develop controlled fermentation methods for fermented vegetables has resulted in an increased interest in developing cultures suitable for application in such methods. Rapid and dominant growth, type and extent of acid production, salt tolerance, temperature range, CO₂ production, cell sedimentation, bacteriophage resistance, nutritional value, and ability to survive as concentrated cultures are factors to consider in developing lactic acid bacterial cultures for use in controlled fermentation of vegetables. Recent examples of efforts to improve cultures include development of nonmalate-decarboxylating strains of lactobacilli for use in cucumber fermentation and isolation of a new species of lactobacillus that produces only the L-isomer of lactic acid from the glucose for use in sauerkraut fermentation. Further improvements in starter cultures for fermented vegetables are likely when suitable genetic transfer systems are developed for selective incorporation or deletion of specific traits.     

Differentiation of mixed lactic acid bacteria communities in beverage fermentations using targeted terminal restriction fragment length polymorphism

Lactic acid bacteria (LAB) are an important group of bacteria in beer and wine fermentations both as beneficial organisms and as spoilage agents. However, sensitive, rapid, culture-independent methods for identification and community analyses of LAB in mixed-culture fermentations are limited. We developed a terminal restriction fragment length polymorphism (TRFLP)-based assay for the detection and identification of lactic acid bacteria and Bacilli during wine, beer, and food fermentations. This technique can sensitively discriminate most species of Lactobacillales, and most genera of Bacillales, in mixed culture, as indicated by both bioinformatic predictions and empirical observations. This method was tested on a range of beer and wine fermentations containing mixed LAB communities, demonstrating the efficacy of this technique for discriminating LAB in mixed culture.     

Occurrence of lactic acid bacteria and biogenic amines in biologically aged wines

Biologically aged sherry-type wines are elaborated by the so-called 'criadera and solera' system, which essentially involves development of the yeast on the wine surface forming a film velum for several years. Lactic acid bacteria can also develop and contribute to sherry-type wine quality, although their presence and role in this enological process have received very little attention. In this study, lactic acid bacteria microbiota and the presence of biogenic amines were investigated throughout the manufacture and biological aging of 36 samples of sherry wines. Malolactic fermentation was found to mainly take place during the first stage of biological ageing. The incidence and populations of lactic acid bacteria in sherry wines were low. The diversity of bacterial species isolated from the wines was greater than previously reported and included species of Lactobacillus, with prevailing Lactobacillus hilgardii, Lactobacillus plantarum and Lactobacillus zeae and Leuconostoc mesenteroides. The biogenic amine-producing capacity of the isolates was also determined. Five strains were putrescine producers, while another strain was shown to produce tyramine and phenylethylamine, simultaneously. L. zeae was one of the predominant species in wines during the biological aging and seemed to be one of the main putrescine producers. The biogenic amine composition of the wines investigated was similar to that reported for other types of wines. Putrescine was the major amine, followed by cadaverine, histamine and tyramine. The amine contents detected were lower than those usually reported in red wines.     

Influence of alcoholic and malolactic starter cultures on bioactive amines in Merlot wines

The influence of alcoholic and malolactic fermentations on the levels of amines in Merlot wines was investigated. Saccharomyces bayanus, S. cerevisiae, Lactobacillus plantarum, Oenococcus oeni (DSM 7008 and 12923) and spontaneous fermentations were used. Four of the 10 amines investigated were detected: spermidine, serotonin, putrescine and cadaverine. When considering the factors independently, the malolactic bacteria significantly affected the levels of serotonin and total amines, whereas the fermentation yeasts significantly affected the levels of spermidine (two way Kruskal–Wallis, p ? 0.05). Spermidine levels were significantly higher in wines produced with S. cerevisiae. Significantly higher serotonin levels were found in wines made with L. plantarum. Putrescine and cadaverine were not detected in wines produced by spontaneous alcoholic fermentation or by L. plantarum. There were significant differences in alcohol content, total and volatile acidity, sulphite levels and taste quality among wines (Tukey test, p ? 0.05).     

Intraspecific genetic diversity of lactic acid bacteria from malolactic fermentation of Cencibel wines as derived from combined analysis of RAPD-PCR and PFGE patterns

Three molecular techniques, randomly amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR), pulsed field gel electrophoresis (PFGE) and differential display-polymerase chain reaction (DD-PCR) have been used to assess the intraspecific diversity of the lactic microbiota responsible for spontaneous malolactic fermentation (MLF) in Cencibel wines made at a cellar in Castilla-La Mancha (Spain). The results indicate that RAPD-PCR and PFGE are of value in typing this microbiota. Better discrimination was achieved by RAPD-PCR. Reproducibility using DD-PCR was not good, which makes this method unsuitable. Combined numerical analysis of the patterns obtained from RAPD-PCR and PFGE allowed a better discrimination; this would therefore be a suitable tool to discriminate the diversity of bacterial communities like those found in MLF of wines. Genetic diversity data from combined numerical analysis suggest that there is considerable microbial diversity within MLF of Cencibel wines, with some genotypes coinciding in the two vinifications analysed.     

干酪加工中的牛奶内源酶和非发酵剂乳酸菌

介绍了干酪加工中起作用的牛奶内源酶及微生物，包括血浆酶、组织蛋白酶D、半胱氨酸蛋白酶、脂蛋白脂肪酶和非发酵剂乳酸菌及它们的作用机制和特性。     

Identification of subdominant sourdough lactic acid bacteria and their evolution during laboratory-scale fermentations

Presumptive lactic acid bacterial cocci were found in six sourdoughs (out of 20) from the Abruzzo region (central Italy) and subjected to phenotypic and genotypic characterization. A total of 21 isolates, recognized as seven strains by randomly amplified polymorphic DNA (RAPD)-polymerase chain reaction (PCR) typing, were identified by a polyphasic approach, consisting of 16S rRNA gene sequencing, multiplex PCR assays and physiological features, as Enterococcus faecium and Pediococcus pentosaceus. Four strains belonging to those species and previously isolated from wheat kernels were inoculated in sterile flour to verify their capacity to grow in sourdough environment. Doughs with several dual bacterial combinations, including Lactobacillus sanfranciscensis, were propagated for 11 days and pH measurements and bacterial counts were carried out.     

Practical implications of lactate and pyruvate metabolism by lactic acid bacteria in food and beverage fermentations

This article reviews the metabolism of pyruvate and lactate by lactic acid bacteria (LAB) involved in food and beverage fermentations with an emphasis on practical implications. First, the formation of pyruvate and lactate from a range of substrates, including carbohydrates, organic acids and amino acids, is briefly described. The catabolism of pyruvate and lactate by LAB is then reviewed. This is followed by a discussion of lactate degradation and racemisation by LAB from specific fermented foods and beverages. Finally, the impact of environmental factors and metabolic engineering on pyruvate and lactate metabolism by LAB is evaluated with regard to practical significance.     

Probiotic spectra of lactic acid bacteria (LAB)

Lactic acid bacteria (LAB) and their probio-active cellular substances exert many beneficial effects in the gastrointestinal tract. LAB prevent adherence, establishment, and replication of several enteric mucosal pathogens through several antimicrobial mechanisms. LAB also release various enzymes into the intestinal lumen and exert potential synergistic effects on digestion and alleviate symptoms of intestinal malabsoption. Consumption of LAB fermented dairy products with LAB may elicit antitumor effects. These effects are attributed to the inhibition of mutagenic activity; decrease in several enzymes implicated in the generation of carcinogens, mutagens, or tumor-promoting agents; suppression of tumors; and the epidemiology correlating dietary regimes and cancer. Specific cellular components in LAB strains seem to induce strong adjuvant effects including modulation of cell-mediated immune responses, activation of reticuloendothelial system, augmentation of cytokine pathways and regulation of interleukins, and tumor necrosis factors. Oral administration of LAB is well tolerated and proven to be safe in 143 human clinical trials and no adverse effects were reported in any of the total 7,526 subjects studied during 1961-1998. In an effort to decrease the reliance on synthetic antimicrobials and control the emerging immunocompromised host population, the time has come to carefully explore the prophylactic and therapeutic applications of probiotic LAB.     

Malolactic fermentation of Tempranillo wine: contribution of the lactic acid bacteria inoculation to sensory quality and chemical composition

Malolactic fermentation (MLF) of Tempranillo Rioja wines (Spain) inoculated with two lactic acid bacteria (LAB) strains were studied and compared with spontaneous MLF. Inoculation with selected Oenococcus oeni lyophila shortened MLF duration up to 19 days and lead to wines with more fresh and fruity characters, especially when implantation was 100%. We demonstrated modifications in the concentration of volatile and nitrogenous compounds and a good correlation between analytical and sensory attributes was also noted. In addition, the low initial amino acid concentration and the consumption of these compounds by the inoculated yeast strain during alcoholic fermentation resulted in wines with very low biogenic amines levels (under 3.75 mg L^?1) after MLF and 3 month storage period in all cases. The results showed the significance of choose the most suitable starter to elaborate quality wines and suggest the control of amino acid content in must and wine to prevent the formation of biogenic amines.     

Spontaneous organic cocoa bean box fermentations in Brazil are characterized by a restricted species diversity of lactic acid bacteria and acetic acid bacteria

Spontaneous organic cocoa bean box fermentations were carried out on two different farms in Brazil. Physical parameters, microbial growth, bacterial species diversity [mainly lactic acid bacteria (LAB) and acetic acid bacteria (AAB)], and metabolite kinetics were monitored, and chocolates were produced from the fermented dry cocoa beans. The main end-products of the catabolism of the pulp substrates (glucose, fructose, and citric acid) by yeasts, LAB, and AAB were ethanol, lactic acid, mannitol, and/or acetic acid. Lactobacillus fermentum and Acetobacter pasteurianus were the predominating bacterial species of the fermentations as revealed through (GTG)₅-PCR fingerprinting of isolates and PCR-DGGE of 16S rRNA gene PCR amplicons of DNA directly extracted from fermentation samples. Fructobacillus pseudoficulneus, Lactobacillus plantarum, and Acetobacter senegalensis were among the prevailing species during the initial phase of the fermentations. Also, three novel LAB species were found. This study emphasized the possible participation of Enterobacteriaceae in the cocoa bean fermentation process. Tatumella ptyseos and Tatumella citrea were the prevailing enterobacterial species in the beginning of the fermentations as revealed by 16S rRNA gene-PCR-DGGE. Finally, it turned out that control over a restricted bacterial species diversity during fermentation through an ideal post-harvest handling of the cocoa beans will allow the production of high-quality cocoa and chocolates produced thereof, independent of the fermentation method or farm.     

Dynamics and species diversity of communities of lactic acid bacteria and acetic acid bacteria during spontaneous cocoa bean fermentation in vessels

To speed up research on the usefulness and selection of bacterial starter cultures for cocoa bean fermentation, a benchmark cocoa bean fermentation process under natural fermentation conditions was developed successfully. Therefore, spontaneous fermentations of cocoa pulp-bean mass in vessels on a 20 kg scale were tried out in triplicate. The community dynamics and kinetics of these fermentations were studied through a multiphasic approach. Microbiological analysis revealed a limited bacterial species diversity and targeted community dynamics of both lactic acid bacteria (LAB) and acetic acid bacteria (AAB) during fermentation, as was the case during cocoa bean fermentations processes carried out in the field. LAB isolates belonged to two main (GTG)₅-PCR clusters, namely Lactobacillus plantarum and Lactobacillus fermentum, with Fructobacillus pseudofilculneus occurring occasionally; one main (GTG)₅-PCR cluster, composed of Acetobacter pasteurianus, was found among the AAB isolates, besides minor clusters of Acetobacter ghanensis and Acetobacter senegalensis. 16S rRNA-PCR-DGGE revealed that L. plantarum and L. fermentum dominated the fermentations from day two until the end and Acetobacter was the only AAB species present at the end of the fermentations. Also, species of Tatumella and Pantoea were detected culture-independently at the beginning of the fermentations. Further, it was shown through metabolite target analyses that similar substrate consumption and metabolite production kinetics occurred in the vessels compared to spontaneous cocoa bean fermentation processes. Current drawbacks of the vessel fermentations encompassed an insufficient mixing of the cocoa pulp-bean mass and retarded yeast growth.     

Co‐inoculation of yeast and lactic acid bacteria to improve cherry wines sensory quality

This study examined the effect of co‐inoculation of yeast and lactic acid bacteria (LAB) on the chemical and sensory characteristics of cherry wines, in comparison with a traditional sequential culture. Three LABs were investigated, including two O. oeni (SG26 and Viniflora) and one L. plantarum (PL18). All co‐inoculations significantly shortened the fermentation time (average 8 days earlier) to reach a stable level of residual sugar (<2 g L^?1) and L‐malic acid (<0.5 g L^?1), and no inhibitory effect on the yeast proliferation was observed. For volatiles determined, co‐culture with SG26 produced the greatest amount of volatile components (138.5 mg L^?1), whereas sequential inoculation with PL18 had the lowest level (119.6 mg L^?1). PCA result revealed that different LABs had diverse influences on the volatile profile of cherry wines, and sensory analysis confirmed that these samples presented distinct sensory profiles, and particularly, a stronger note of fruity was perceived when co‐culture was used.     

Characterization of lactic acid bacteria isolated from raw cows’ milk cheeses currently produced in Galicia (NW Spain)

Twenty cheeses belonging to the four Protected Designations of Origin manufactured in Galicia (NW Spain) (6 Arzúa-Ulloa, 4 Tetilla, 6 Cebreiro and 4 San Simón da Costa) were selected from a total of 60 cheeses on basis of their (typical) sensorial profiles. A total of 218 lactic acid bacteria (LAB) isolates were obtained from the predominant microflora of the selected cheeses and were identified as Lactococci (98 isolates), Leuconostocs (56), Mesophilic Lactobacilli (54), Pediococci (8) and Enterococci (2). Eighty-four of the isolates produced mainly malty, spicy or sulfide flavours in pasteurised whole milk, and were not characterized further. Some good producers of diacetyl-acetoin in milk (>100 mg/L) were found among a total of 129 LAB selected, although the isolates were generally less acidifying and less proteolytic than many of those obtained 10-15 years ago. The results suggest that the microflora in cheese-making environments have undergone changes, with the most evident difference being the practical absence of Enterococcal strains among the current isolates.     

乳酸菌及葡萄糖对鱼片中细菌群落及TVBN值的影响

研究了葡萄糖和乳酸菌对罗非鱼片上细菌群落和TVBN值的影响发现经嗜酸乳杆菌CICC6074以及1.5%葡萄糖处理后鱼片的感官性状基本不变。对照鱼片在4℃冷藏6 d,其TVBN值超过国家限量标准;而CICC6074处理的鱼片冷藏12 d后才超过此标准。葡萄糖虽然加速了冷藏初期细菌的生长,但对鱼片上最终的细菌数量没有影响,并可明显降低鱼片的TVBN值。用葡萄糖和CICC6074混合处理鱼片是否可以进一步降低TVBN值则未能确定。PCR-DGGE分析发现,鱼类的特定腐败菌——假单胞菌在CICC6074处理的鱼片其生长被明显延缓,冷藏6 d后数量才明显增加;而对照鱼片仅冷藏3 d其数量已明显增加。鱼片上吸附的CICC6074数量在冷藏初期略有增加,6 d后下降,9 d后消失。结果表明,嗜酸乳杆菌CICC6074可用于鱼片保鲜,而葡萄糖可降低鱼片的TVBN值。     

Characterization of yeast and lactic acid bacterial species in ropy wines

Summary The microflora in ropy sherry-type wines were identified. The genusSaccharomyces were the predominant yeasts (81%), withS. beticus prevailing.Lactobacillus plantarum was the most frequent bacterial species (33.9%), followed byStreptococcus diacetilactis (22%) andS. lactis (16.9%).

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Charakterisierung der Hefe-und der Milchsäurebakterien-Arten in sirupösen Weinen

Zusammenfassung Eine Analyse der Mikroflora in sherryartigen, sirupösen Weinen ergab die Anwesenheit von deckenbildenden Hefen des StammesSaccharomyces (81%), wobeiS. beticus am stärksten vertreten ist. Bei den Bakterienstämmen istLactobacillus plantarum (33,9%) am stärksten vertreten, gefolgt vomStreptococcus diacetilactis (22%) und demS. lactis (16,9%).

     

Growth-inhibition of hiochi bacteria in namazake (raw sake) by bacteriocins from lactic acid bacteria

The bacteriocins produced by Lactococcus lactis subsp. lactis C101910 (C101910) and NBRC 12007 (NBRC 12007) were used to prevent the growth of sake spoiling hiochi bacteria (Lactobacillus hilgardii, Lactobacillus fructivorans, and Lactobacillus paracasei) in namazake, which is raw (unpasteurized) sake. The bacteriocin concentrations required for decreasing the viable cell concentrations of L. hilgardii and L. fructivorans below the detection limit (1.0 × 10² cells/ml) in 24 h from the initial concentration of 4.0–9.5 × 10⁵ cells/ml in the namazake at pH 4.5 and at 4°C, were 18–35 U/ml and 5.6 U/ml for the bacteriocin from C101910 and NBRC 12007, respectively. To decrease the viable cell concentration of L. paracasei from the initial concentration of 7.5 × 10⁵ cells/ml to below the detection limit (1.0 × 10² cells/ml) in 24 h, 350 U/ml bacteriocin from C101910 and 140 U/ml bacteriocin from NBRC 12007 were required. In experiments using McIlvaine buffer (pH 4.5) with 15% ethanol instead of namazake as the medium, the viable cell concentrations of L. hilgardii and L. paracasei decreased to less than 1.0 × 10² cells/ml, whereas those of L. fructivorans decreased to less than 1.0 × 10³ cells/ml, when bacteriocins were added at the concentrations that had proven effective in namazake. The membrane depolarization assay using a fluorescent probe showed that the presence of ethanol stimulated the collapse of the membrane potential induced by bacteriocins. The ethanol induced collapse of the membrane potential suggests that the application of bacteriocins at the storage stage of namazake is more beneficial than when used in other stages of the sake brewing process.     

Fluoride levels in wines of the Canary Islands (Spain)

Fluoride content in bottled wines of the different types and areas of the Canary Islands was studied by direct potentiometry with a fluoride specific electrode using the addition method. We found a mean concentration of 0.15 mg/l and a standard deviation of 0.068. The biggest concentration of fluoride ion (0.50 mg/l) was found in a white wine of Tacoronte-Acentejo. However, all wines analyzed presented concentrations of fluoride lower than the maximum allowed by the International Office of the Vineyard and the Wine. So, the contribution of the Canarian wines to the fluoride daily intake does not represent any risk of fluorosis. Also these concentrations have no influence on the production process of the wine.     

Diversity analysis of lactic acid bacteria in Korean rice wines by culture-independent method using PCR-denaturing gradient gel electrophoresis

A polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) method was used to determine the presence and diversity of lactic acid bacteria (LAB) in takju, a traditional Korean rice wine. Bacterial DNAs were extracted from 15 commercial rice wines and amplicons of partial 16S rRNA genes were separated by DGGE and intense bands were sequenced. Lactobacillus (Lb.) paracasei, Lb. plantarum, and Leuconostoc pseudomesenteroides were detected in all samples and Lb. harbinensis and Lb. parabuchneri were found with above 80% frequency of occurrence. Unknown species of Lactobacillus were also widely detected. This result revealed that, regardless of products and raw materials, the distribution profiles of LAB in takju products have a common pattern comprising of above predominant species and, furthermore, takju can be regarded as a LAB-rich fermented food providing various probiotics.