B-group vitamin production by lactic acid bacteria--current knowledge and potential applications

Although most vitamins are present in a variety of foods, human vitamin deficiencies still occur in many countries, mainly because of malnutrition not only as a result of insufficient food intake but also because of unbalanced diets. Even though most lactic acid bacteria (LAB) are auxotrophic for several vitamins, it is now known that certain strains have the capability to synthesize water-soluble vitamins such as those included in the B-group (folates, riboflavin and vitamin B(12) amongst others). This review article will show the current knowledge of vitamin biosynthesis by LAB and show how the proper selection of starter cultures and probiotic strains could be useful in preventing clinical and subclinical vitamin deficiencies. Here, several examples will be presented where vitamin-producing LAB led to the elaboration of novel fermented foods with increased and bioavailable vitamins. In addition, the use of genetic engineering strategies to increase vitamin production or to create novel vitamin-producing strains will also be discussed. This review will show that the use of vitamin-producing LAB could be a cost-effective alternative to current vitamin fortification programmes and be useful in the elaboration of novel vitamin-enriched products.     

Recent update on lactic acid bacteria producing riboflavin and folates: application for food fortification and treatment of intestinal inflammation

Lactic acid bacteria (LAB), widely used as starter cultures for the fermentation of a large variety of food, can improve the safety, shelf life, nutritional value and overall quality of the fermented products. In this regard, the selection of strains delivering health-promoting compounds is now the main objective of many researchers. Although most LAB are auxotrophic for several vitamins, it is known that certain strains have the capability to synthesize B-group vitamins. This is an important property since humans cannot synthesize most vitamins, and these could be obtained by consuming LAB fermented foods. This review discusses the use of LAB as an alternative to fortification by the chemical synthesis to increase riboflavin and folate concentrations in food. Moreover, it provides an overview of the recent applications of vitamin-producing LAB with anti-inflammatory/antioxidant activities against gastrointestinal tract inflammation. This review shows the potential uses of riboflavin and folates producing LAB for the biofortification of food, as therapeutics against intestinal pathologies and to complement anti-inflammatory/anti-neoplastic treatments.     

Short-chain fatty acid and vitamin production potentials of Lactobacillus isolated from fermented foods of Khasi Tribes,Meghalaya, India

Vitamins and SCFA (short-chain fatty acids) production from Lactobacillus isolates are studied due to its health benefits to the human hosts. Lactobacillus strains are widely used in fermented foods, and few of them are reported with vitamin and SCFA production potential. Therefore, in the present study, vitamins and SCFA production capability of isolates were studied to find the potent Lactobacillus cultures for value-added functional food product development. Five Lactobacillus strains, i.e., KGL2, KGL3A, KGL4, RNS4, and WTS4, were isolated from rice-based traditional fermented foods of Garo Hills, Meghalaya, India. All the well grown isolates were morphologically, physiologically, and genetically characterized. Then, vitamins and SCFA were estimated using HPLC based methods. Vitamins produced in vitamins free assay medium and SCFA in milk medium are produced by Lactobacillus. Lactic acid bacteria produce essential vitamins like riboflavin, folate, cobalamin, and SCFA which have health impacts (anti-obesity, anti-diabetics, anti-microbial, and other chronic diseases prevention) to the host. These vitamins are essential for cellular and metabolic growth of living system. In the study, five potent Lactobacillus isolates viz., KGL2 (Lactobacillus fermentum), KGL3A (Lactobacillus plantarum), KGL4 (Lactobacillus fermentum), RNS4 (Lactobacillus rhamnosus), and WTS4 (Lactobacillus fermentum) were considered for vitamins (B2, B12, and B9) and SCFA productions (lactate, butyrate, and acetate). However, KGL3A had shown highest B2 production (0.7 μg/ml) while KGL2 exhibited maximum B12 production (0.05 μg/ml) after 36 h. Moreover, WTS4 attributed highest folate production (0.09 μg/ml) after 24 h. In addition, RNS4 reported the maximum short-chain fatty acid production (0.77 g/l acetic acid, 0.26 g/l lactic acid, and 0.008 g/l butyric acid respectively). Potent Lactobacillus isolates from traditional fermented foods of Garo Hills, Meghalaya, India (North East Part of India) showed maximum production of B2, B9, and B12 as well as short-chain fatty acids and could be used for their application as health beneficial functional fermented dairy products.     

Perspectives of engineering lactic acid bacteria for biotechnological polyol production

Polyols are sugar alcohols largely used as sweeteners and they are claimed to have several health-promoting effects (low-caloric, low-glycemic, low-insulinemic, anticariogenic, and prebiotic). While at present chemical synthesis is the only strategy able to assure the polyol market demand, the biotechnological production of polyols has been implemented in yeasts, fungi, and bacteria. Lactic acid bacteria (LAB) are a group of microorganisms particularly suited for polyol production as they display a fermentative metabolism associated with an important redox modulation and a limited biosynthetic capacity. In addition, LAB participate in food fermentation processes, where in situ production of polyols during fermentation may be useful in the development of novel functional foods. Here, we review the polyol production by LAB, focusing on metabolic engineering strategies aimed to redirect sugar fermentation pathways towards the synthesis of biotechnologically important sugar alcohols such as sorbitol, mannitol, and xylitol. Furthermore, possible approaches are presented for engineering new fermentation routes in LAB for production of arabitol, ribitol, and erythritol.     

Biotechnological and in situ food production of polyols by lactic acid bacteria

Polyols such as mannitol, erythritol, sorbitol, and xylitol are naturally found in fruits and vegetables and are produced by certain bacteria, fungi, yeasts, and algae. These sugar alcohols are widely used in food and pharmaceutical industries and in medicine because of their interesting physicochemical properties. In the food industry, polyols are employed as natural sweeteners applicable in light and diabetic food products. In the last decade, biotechnological production of polyols by lactic acid bacteria (LAB) has been investigated as an alternative to their current industrial production. While heterofermentative LAB may naturally produce mannitol and erythritol under certain culture conditions, sorbitol and xylitol have been only synthesized through metabolic engineering processes. This review deals with the spontaneous formation of mannitol and erythritol in fermented foods and their biotechnological production by heterofermentative LAB and briefly presented the metabolic engineering processes applied for polyol formation.     

Genotypic and phenotypic diversity of Pediococcus pentosaceus strains isolated from food matrices and characterisation of the penocin operon

Lactic acid bacteria (LAB) are widely used in the food industry. Pediococcus spp. belong to the LAB group and include several species that are essential for the quality of fermented food. Pediococcus pentosaceus is the species that is most frequently isolated from fermented food and beverages but its uncontrolled growth during food fermentation processes can contribute to undesired flavours. Hence, the characterisation of these bacteria at the strain level is of great importance for the quality of fermented products. Despite their importance, misidentification at the species level is common for members of the genus Pediococcus. To clarify the taxonomic relationships among strains, a multilocus sequencing approach was developed for the characterisation of a collection of 29 field strains, 1 type strain and 1 reference strain of P. pentosaceus isolated from food. These strains were also tested for several phenotypic properties of technological interest and for the production of bacteriocins. The chromosomal operon involved in the synthesis of the bacteriocin penocin was also investigated. The present study enabled a good genomic characterisation, identifying 17 sequence types, with an overview of phenotypic characteristics related to different technological abilities, and also provides a thorough characterisation of the operon involved in penocin production.     

Bacteriocins from lactic acid bacteria: production, purification, and food applications

In fermented foods, lactic acid bacteria (LAB) display numerous antimicrobial activities. This is mainly due to the production of organic acids, but also of other compounds, such as bacteriocins and antifungal peptides. Several bacteriocins with industrial potential have been purified and characterized. The kinetics of bacteriocin production by LAB in relation to process factors have been studied in detail through mathematical modeling and positive predictive microbiology. Application of bacteriocin-producing starter cultures in sourdough (to increase competitiveness), in fermented sausage (anti-listerial effect), and in cheese (anti-listerial and anti-clostridial effects), have been studied during in vitro laboratory fermentations as well as on pilot-scale level. The highly promising results of these studies underline the important role that functional, bacteriocinogenic LAB strains may play in the food industry as starter cultures, co-cultures, or bioprotective cultures, to improve food quality and safety. In addition, antimicrobial production by probiotic LAB might play a role during in vivo interactions occurring in the human gastrointestinal tract, hence contributing to gut health.     

Characterization of some bacteriocins produced by lactic acid bacteria isolated from fermented foods

Lactic acid bacteria (LAB) isolated from different sources (dairy products, fruits, fresh and fermented vegetables, fermented cereals) were screened for antimicrobial activity against other bacteria, including potential pathogens and food spoiling bacteria. Six strains have been shown to produce bacteriocins: Lactococcus lactis 19.3, Lactobacillus plantarum 26.1, Enterococcus durans 41.2, isolated from dairy products and Lactobacillus amylolyticus P40 and P50, and Lactobacillus oris P49, isolated from bors. Among the six bacteriocins, there were both heat stable, low molecular mass polypeptides, with a broad inhibitory spectrum, probably belonging to class II bacteriocins, and heat labile, high molecular mass proteins, with a very narrow inhibitory spectrum, most probably belonging to class III bacteriocins. A synergistic effect of some bacteriocins mixtures was observed. We can conclude that fermented foods are still important sources of new functional LAB. Among the six characterized bacteriocins, there might be some novel compounds with interesting features. Moreover, the bacteriocin-producing strains isolated in our study may find applications as protective cultures.     

Production of Propionibacterium shermanii biomass and vitamin B12 on spent media

AIMS: The propionibacteria are commercially important due to their use in the cheese industry, and there is a growing interest for their probiotic effects. Stimulatory effects of lactic acid bacteria (LAB) on propionic acid bacteria have been observed. This study was designed to examine the possibility of using spent media previously used to grow LAB for the production of biomass and metabolites of Propionibacterium freudenreichii subsp. shermanii. METHODS AND RESULTS: Seventeen MRS and vegetable juice media were prefermented by various LAB and evaluated for their ability to subsequently support the growth of Propionibacterium, using automated spectrophotometry (AS). Growth of Propionibacterium in spent media was strongly affected by the LAB strain used to produce the spent medium. The native MRS medium (not prefermented) yielded the highest optical density values followed by prefermented media by Lactobacillus acidophilus, Bifidobacterium longum and Lactococcus lactis. Prefermented cabbage juice enabled good growth of Propionibacterium. For the production of organic acids and vitamin B12, cells of Propionibacterium were concentrated and immobilized in alginate beads in the aim of accelerating the bioconversions. More propionic acid was obtained in spent media than in native MRS. The concentration of vitamin B12 was higher in media fermented with free cells than those with immobilized cultures; with the free cells, its concentration varied from 900 to 1800 ng ml(-1) of media. CONCLUSIONS: It was demonstrated that spent media could be recycled for the production of Propionibacterium and metabolites, depending on the LAB strain that was previously grown. Media remediation is needed to improve the production of vitamin B12, especially with immobilized cells. SIGNIFICANCE AND IMPACT OF THE STUDY: This study presents an option for recycling of spent media generated by producers of LAB or producers of fermented vegetables. The propionic fermentation may result in three commercial products: biomass, vitamin B12 or organic acids, which may be used as starters, supplements or food preservatives. It is an attractive process from economical and environmental standpoints.     

Dominant lactic acid bacteria and their technological properties isolated from the Himalayan ethnic fermented milk products

Ethnic people of the Himalayan regions of India, Nepal, Bhutan and China consume a variety of indigenous fermented milk products made from cows milk as well as yaks milk. These lesser-known ethnic fermented foods are dahi, mohi, chhurpi, somar, philu and shyow. The population of lactic acid bacteria (LAB) ranged from 10(7) to 10(8) cfu/g in these Himalayan milk products. A total of 128 isolates of LAB were isolated from 58 samples of ethnic fermented milk products collected from different places of India, Nepal and Bhutan. Based on phenotypic characterization including API sugar test, the dominant lactic acid bacteria were identified as Lactobacillus bifermentans, Lactobacillus paracasei subsp. pseudoplantarum, Lactobacillus kefir, Lactobacillus hilgardii, Lactobacillus alimentarius, Lactobacillus paracasei subsp. paracasei, Lactobacillus plantarum, Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. cremoris and Enterococcus faecium. LAB produced a wide spectrum of enzymes and showed high galactosidase, leucine-arylamidase and phosphatase activities. They showed antagonistic properties against selected Gram-negative bacteria. None of the strains produced bacteriocin and biogenic amines under the test conditions used. Most strains of LAB coagulated skim milk with a moderate drop in pH. Some strains of LAB showed a high degree of hydrophobicity, suggesting these strains may have useful adhesive potential. This paper is the first report on functional lactic acid bacterial composition in some lesser-known ethnic fermented milk products of the Himalayas.     

Applications for biotechnology: present and future improvements in lactic acid bacteria

The lactic acid bacteria are involved in the manufacture of fermented foods from raw agricultural materials such as milk, meat, vegetables, and cereals. These fermented foods are a significant part of the food processing industry and are often prepared using selected strains that have the ability to produce desired products or changes efficiently. The application of genetic engineering technology to improve existing strains or develop novel strains for these fermentations is an active research area world-wide. As knowledge about the genetics and physiology of lactic acid bacteria accumulates, it becomes possible to genetically construct strains with characteristics shaped for specific purposes. Examples of present and future applications of biotechnology to lactic acid bacteria to improve product quality are described. Studies of the basic biology of these bacteria are being actively conducted and must be continued, in order for the food fermentation industry to reap the benefits of biotechnology.     

Applications for biotechnology: present and future improvements in lactic acid bacteria

Abstract The lactic acid bacteria are involved in the manufacture of fermented foods from raw agricultural materials such as milk, meat, vegetables, and cereals. These fermented foods are a significant part of the food processing industry and are often prepared using selected strains that have the ability to produce desired products or changes efficiently. The application of genetic engineering technology to improve existing strains or develop novel strains for these fermentations is an active research area world-wide. As knowledge about the genetics and physiology of lactic acid bacteria accumulates, it becomes possible to genetically construct strains with characteristics shaped for specific purposes. Examples of present and future applications of biotechnology to lactic acid bacteria to improve product quality are described. Studies of the basic biology of these bacteria are being actively conducted and must be continued, in order for the food fermentation industry to reap the benefits of biotechnology.     

Microbial analysis of Malaysian tempeh, and characterization of two bacteriocins produced by isolates of Enterococcus faecium

AIMS: Isolation of bacteriocinogenic lactic acid bacteria (LAB) from the Malaysian mould-fermented product tempeh and characterization of the produced bacteriocin(s). METHODS AND RESULTS: LAB were present in high numbers in final products as well as during processing. Isolates, Enterococcus faecium B1 and E. faecium B2 (E. faecium LMG 19827 and E. faecium LMG 19828, respectively) inhibited Gram-positive indicators, including Listeria monocytogenes. Partially purified bacteriocins showed a proteinaceous nature. Activity was stable after heat-treatment except at alkaline pH values. Both strains displayed a bacteriostatic mode of action. Bacteriocin production was associated with late exponential/early stationary growth. Molecular mass, calculated by SDS-PAGE, was 3.4 kDa for B1 bacteriocin, and 3.4 kDa and 5.8 kDa for B2 bacteriocins. PCR screening of enterocin-coding genes revealed three amplified fragments in total genomic DNA that may correspond with PCR signals for enterocin P, enterocin L50A and enterocin L50B. Both B1 and B2 contained a 42-kb plasmid. No differences in bacteriocinogenic capacity were found between wild type strains and plasmid-cured strains. CONCLUSIONS: It was possible to isolate bacteriocinogenic E. faecium active against various Gram-positive bacteria from final products of tempeh. SIGNIFICANCE AND IMPACT OF THE STUDY: A first step in applying biopreservation to fermented South-east Asian foods is to obtain bacteriocinogenic LAB from this source. Such isolates may also be used for biopreservation of mould-fermented foods in general, including various types of mould-ripened cheese.     

代谢工程在维生素生产中的应用及研究进展

维生素是维持人体生命活动必需的一类有机物质,机体本身一般不能合成或合成量不足,因此需经食物或其他强化产品获取。目前,维生素产品已广泛应用于医药、食品添加剂、饲料添加剂、化妆品等领域,而且全球对维生素的需求也是呈逐年增长态势。维生素的生产方法主要包括化学合成法和生物合成法。化学合成法通常安全隐患大、反应条件严苛、废物污染严重,相比之下,代谢工程生产维生素绿色环保安全、能耗低,因此建立微生物细胞工厂具有重大的科学意义和应用需求。文中回顾了近30年来代谢工程在维生素生产领域的研究进展,详细阐述了水溶性维生素(维生素B1、B2、B3、B5、B6、B7、B9、B12和维生素C的前体)和脂溶性维生素(维生素A、维生素D的前体、维生素E和维生素K)的生物合成研究现状,并对其发酵生产的瓶颈进行了探讨,最后对合成生物技术创建维生素生产菌种进行了展望。     

Antilisterial activity of lactic acid bacteria isolated from rigouta, a traditional Tunisian cheese

AIMS: Screening for lactic acid bacteria (LAB) producing bacteriocins and other antimicrobial compounds is of a great significance for the dairy industry to improve food safety. METHODS AND RESULTS: Six-hundred strains of LAB isolated from 'rigouta', a Tunisian fermented cheese, were tested for antilisterial activity. Eight bacteriocinogenic strains were selected and analysed. Seven of these strains were identified as Lactococcus lactis and produced nisin Z as demonstrated by mass spectrometry analysis of the purified antibacterial compound. Polymerase chain reaction experiments using nisin gene-specific primers confirmed the presence of nisin operon. Plasmid profiles analysis suggests the presence of, at least, three different strains in this group. MMT05, the eighth strain of this antilisterial collection was identified, at molecular level, as Enterococcus faecalis. The purified bacteriocin produced by this strain showed a molecular mass of 10 201.33 +/- 0.85 Da. This new member of class III bacteriocins was termed enterocin MMT05. CONCLUSIONS: Seven lactococcal strains producing nisin Z were selected and could be useful as bio-preservative starter cultures. Additional experiments are needed to evaluate the promising strain MMT05 as bio-preservative as Enterococci could exert detrimental or beneficial role in foods. SIGNIFICANCE AND IMPACT OF THE STUDY: Only a few antibacterial strains isolated from traditional African dairy products were described. The new eight strains described herein contribute to the knowledge of this poorly studied environment and constitute promising strains for fermented food safety.     

Immunobiosis and probiosis: antimicrobial activity of lactic acid bacteria with a focus on their antiviral and antifungal properties

Lactic acid bacteria (LAB), a heterogeneous group of bacteria that produce lactic acid as the main product of carbohydrate degradation, play an important role in the production and protection of fermented foods. Moreover, beside the technological use of these microorganisms added to control and steer food fermentations, their beneficial healthy properties are largely overt. Thus, numerous LAB strains have obtained the probiotic status, which entails the ability to maintain and promote a good health of consumers. In particular, increasing consideration is being focused on probiotic microorganisms that can improve the human immune response against dangerous viral and fungal enemies. For such beneficial microbes, the term “immunobiotics” has been coined. Together with an indirect host-mediated adverse effect against undesirable microorganisms, also a direct antagonistic activity of several LAB strains has been largely demonstrated. The purpose of this review is to provide a fullest possible overview of the antiviral and antifungal activities ascribed to probiotic LAB. The interest in this research field is substantiated by a large number of studies exploring the potential application of these beneficial microorganisms both as biopreservatives and immune-enhancers, aiming to reduce and/or eliminate the use of chemical agents to prevent the development of pathogenic, infectious, and/or degrading causes.

     

Lactobacillaceae and cell adhesion: genomic and functional screening

The analysis of collections of lactic acid bacteria (LAB) from traditional fermented plant foods in tropical countries may enable the detection of LAB with interesting properties. Binding capacity is often the main criterion used to investigate the probiotic characteristics of bacteria. In this study, we focused on a collection of 163 Lactobacillaceace comprising 156 bacteria isolated from traditional amylaceous fermented foods and seven strains taken from a collection and used as controls. The collection had a series of analyses to assess binding potential for the selection of new probiotic candidates. The presence/absence of 14 genes involved in binding to the gastrointestinal tract was assessed. This enabled the detection of all the housekeeping genes (ef-Tu, eno, gap, groEl and srtA) in the entire collection, of some of the other genes (apf, cnb, fpbA, mapA, mub) in 86% to 100% of LAB, and of the other genes (cbsA, gtf, msa, slpA) in 0% to 8% of LAB. Most of the bacteria isolated from traditional fermented foods exhibited a genetic profile favorable for their binding to the gastrointestinal tract. We selected 30 strains with different genetic profiles to test their binding ability to non-mucus (HT29) and mucus secreting (HT29-MTX) cell lines as well as their ability to degrade mucus. Assays on both lines revealed high variability in binding properties among the LAB, depending on the cell model used. Finally, we investigated if their binding ability was linked to tighter cross-talk between bacteria and eukaryotic cells by measuring the expression of bacterial genes and of the eukaryotic MUC2 gene. Results showed that wild LAB from tropical amylaceous fermented food had a much higher binding capacity than the two LAB currently known to be probiotics. However their adhesion was not linked to any particular genetic equipment.     

Production of natural folates by lactic acid bacteria starter cultures isolated from artisanal Argentinean yogurts

Folate is a B-group vitamin that cannot be synthesized by humans and must be obtained exogenously. Although some species of lactic acid bacteria (LAB) can produce folates, little is known about the production of this vitamin by yogurt starter cultures. Lactobacillus delbrueckii subsp. bulgaricus and Streptococcus thermophilus strains were isolated from artisanal Argentinean yogurts and were grown in folate-free culture medium (FACM) and nonfat milk after which intracellular and extracellular folate production were evaluated. From the initial 92 isolated LAB strains, 4 L. delbrueckii subsp. bulgaricus and 32 S. thermophilus were able to grow in the absence of folate. Lactobacillus delbrueckii subsp. bulgaricus CRL 863 and S.?thermophilus CRL 415 and CRL 803 produced the highest extracellular folate levels (from 22.3 to 135?μg/L) in FACM. In nonfat milk, these strains were able to increase the initial folate concentrations by almost 190%. This is the first report where native strains of L. delbrueckii subsp. bulgaricus were shown to produce natural folate. The LAB strains identified in this study could be used in developing novel fermented products bio-enriched in natural folates that could in turn be used as an alternative to fortification with the controversial synthetic chemical folic acid.     

Potential of selected strains of lactic acid bacteria to induce a Th1 immune profile

Lactic acid bacteria (LAB) might switch the Th2 biased immune response in allergic patients towards a balanced Th1/Th2 immune profile, leading to amelioration of allergy. To select strains of LAB that could be of potential application for foods in controlling allergy, 35 bacterial strains were screened in vitro using murine splenocytes and peritoneal exudate cells (PECs). Streptococcus thermophilus AHU1838 (FERM AP-21009), and Lactobacillus paracasei subsp. casei AHU1839 (FERM AP-21010) enhanced the secretion of Th1 cytokines such as interferon-gamma (IFN-gamma) and interleukin-12 (IL-12). The two strains of LAB also up-regulated the expression of CD40, and CD86 in dendritic cells (DCs), and activated cytotoxic T lymphocytes (CTL). These two strains could therefore be used in producing fermented food products that can enhance the Th1 immune profile which is important in ameliorating allergy.