Search

Some lactic acid bacteria (LAB) have the ability to synthesize riboflavin, a trait linked to the presence of ribG, ribB, ribA and ribH genes located in the rib operon. Previous screening of riboflavin producers identified several LAB strains belonging to different species with this ability, but none of them surpassed 0.25 mg/L production of the vitamin. In this study, we explored two strategies to obtain riboflavin-overproducing strains: by roseoflavin selection of mutants, and by the transformation of selected strains with plasmids pNZ:TuR.rib or pNZ:TuB.rib containing the genes ribG, ribB, ribA and ribH from Lactococcus cremoris MG1363. The resulting riboflavin-overproducing strains were able to produce yields between 0.5 and 6 mg/L in culture media and several of them were selected for the fermentation of soy beverages. Riboflavin in bio-enriched soy beverages was evaluated by direct fluorescence measurement and high-performance liquid chromatography-fluorescence analysis. Soy beverages fermented with the recombinant strains Lactococcus cremoris ESI 277 pNZ:TuB.rib and Lactococcus lactis INIA 12 pNZ:TuR.rib showed the highest riboflavin yields (>5 mg/L) after 24 h fermentation. On the other hand, roseoflavin-resistant mutant Limosilactobacillus fermentum INIA P143R2 was able to enrich fermented soy beverages with 1.5 mg/L riboflavin. Riboflavin-overproducing LAB strains constitute a good option for riboflavin enrichment of soy beverages by fermentation and the commercialization of such beverages could be very useful to prevent riboflavin deficiency.

Soy beverage is a rich source of phytoestrogens isoflavones, with potential benefits on health. The effect of those compounds depends greatly on their bacterial metabolization into their aglycone forms. This study evaluated the health effects of two soy beverages, non-fermented (SB) and fermented with Bifidobacterium pseudocatenulatum INIA P815 (FSB), in acyclic and cyclic C57BL/6J aged female mice as a model of menopause and premenopause, respectively. SB and FSB treatments were administrated for 36 days and, subsequently, body weight, lipid and inflammatory profile and fertility were analyzed and compared. In addition, hepatic gene expression and faecal microbiota composition were also assessed. After fermentation, FSB presented a high content in the aglycones daidzein and genistein and a higher antioxidant activity. FSB treated cyclic mice showed a significant increase in the number of retrieved oocytes and zigotes. Differences in serum lipids were observed in triglycerides, which were lower in FSB than in SB groups. None of the treatments influenced the inflammatory profile or caused a dramatic change in the intestinal microbiota profile or hepatic gene expression in any of the groups. Our data showed that FSB provided greater health benefits than SB in lipid profile and fertility in cyclic mice. These beneficial effects could be attributed to the fermentation process, which produces more bioavailable and bioactive compounds, achieving a greater impact on health.

Some lactic acid bacteria (LAB) strains have the ability to synthesize riboflavin, a trait linked to the presence of ribG, ribB, ribA and ribH genes in the rib operon. Multiple sequence alignments of these genes showed that these sequences are not identical in different LAB species, so primers designed to detect these genes in one species do not always work with others. Therefore, we designed degenerate primers based on sequences from Lactococcus lactis MG1363, Levilactobacillus brevis ATCC 367 and Limosilactobacillus fermentum IFO3956, and established optimal PCR conditions for the detection of rib genes in different LAB species. Simultaneously, we selected riboflavin-producing LAB strains from our bacterial collection belonging to the species L. brevis, L. fermentum, L. lactis, Leuconostoc mesenteroides and Lactiplantibacillus plantarum, and we were able to detect ribG, ribB, ribA and ribH genes in these strains by PCR using the designed primers. Thus, the development of degenerate primers and optimal PCR conditions for the detection of ribG, ribB, ribA and ribH genes in LAB allowed the detection and the selection of potential riboflavin-producing strains of different species, which could be good candidates for the development of riboflavin-enriched functional foods.

Soy beverages can be a source of bioactive isoflavones, with potential human health benefits. In this work, the suitability of three Lacticaseibacillus and three Bifidobacterium probiotic strains as functional starters for soy beverage fermentation were evaluated, alongside with the effect of refrigerated storage on the viability of the strains and the isoflavone composition of the fermented beverages. The three bifidobacteria strains suffered a decrease in their viability during refrigeration and only Bifidobacterium breve INIA P734 produced high concentrations of bioactive isoflavones. Meanwhile, L. rhamnosus GG and L. rhamnosus INIA P344 produced high levels of aglycones and, with L. paracasei INIA P272, maintained their viability during the refrigeration period, constituting promising starters to obtain functional soy beverages that could gather the benefits of the bioactive isoflavone aglycones and the probiotic strains. Moreover, the three lactobacilli caused an increase in the antioxidant capacity of the fermented beverages, which was maintained over the refrigerated storage.

Isoflavones are phenolic compounds (considered as phytoestrogens) with estrogenic and antioxidant function, which are highly beneficial for human health, especially in the aged population. However, isoflavones in foods are not bioavailable and, therefore, have low biological activity. Additionally, their transformation into bioactive compounds by microorganisms is necessary to obtain bioavailable isoflavones with beneficial effects on human health. Many lactic acid bacteria (LAB) can transform the methylated and glycosylated forms of isoflavones naturally present in foods into more bioavailable aglycones, such as daidzein, genistein and glycitein. In addition, certain LAB strains are capable of transforming isoflavone aglycones into compounds with a greater biological activity, such as dihydrodaidzein (DHD), O-desmethylangolensin (O-DMA), dihydrogenistein (DHG) and 6-hydroxy-O-desmethylangolensin (6-OH-O-DMA). Moreover, Lactococcus garviae 20-92 is able to produce equol. Another strategy in the bioconversion of isoflavones is the heterologous expression of genes from Slackia isoflavoniconvertens DSM22006, which have allowed the production of DHD, DHG, equol and 5-hydroxy-equol in high concentrations by engineered LAB strains. Accordingly, the consequences of isoflavone metabolism by LAB and its application in the development of foods enriched in bioactive isoflavones, as well as health benefits attributed to their consumption, will be addressed in this work.

Among vegetable functional products, soy-based foods stand out for their high content in protein and flavonoids,mainly isoflavones. The fermentation of soy beverages with selected bacterial strains is a way to obtain a product,enriched in isoflavone aglycones, alongside with other isoflavone derivates and other bioactive flavonoids. Two bacterial strains with high activity on the transformation of these compounds, Limosilactobacillus mucosae INIA P508 and Bifidobacterium pseudocatenulatum INIA P815, were tested for the fermentation of soy beverage. Both strains produced high concentration of the aglycones daidzein and genistein, in addition to a significant increase (P < 0.01) of glycitein, O-desmethylangolensin, 6′′ -hydroxy-O-desmethylangolensin, tetrahydrodaidzein and other flavonoid aglycones such as kaempferol, naringenin and luteolin. Two alternatives to conserve the developed functional beverages were explored, cold storage and thermal treatment. Refrigeration did not affect the concentration of isoflavones, even observing an increase in aglycones. Similarly, the thermal treatment and further room temperature storage did not reduce the amounts of daidzein and genistein. Therefore, both strains could be used for the development of soy beverages enriched in bioactive isoflavones and other bioactive flavonoids which would maintain their concentrations in refrigeration or at room temperature after heat treatment.

The genetic engineering of bacteria for food applications has biosafety requirements, including the use of non-antibiotic selectable markers. These can be gene-encoding bacteriocin immunity proteins, such as nisI and pedB, which require the use of promoters to ensure optimal expression. Our aim was to search for promoters for the expression of pediocin (pedB) and nisin (nisI) immunity genes, which could allow the selection of a wide variety of transformed lactic acid bacteria (LAB) and bifidobacteria strains. Eight promoters from LAB or bifidobacteria were initially studied using evoglow-Pp1 as the reporter gene in Lactococcus lactis NZ9000, resulting in the selection of P32, P3N, PTuR and PEF-P, which exhibited a strong constitutive expression. These promoters were further tested for the expression of the food-grade selectable markers pedB and nisI in agar diffusion assays with pediocin and nisin, respectively. The results obtained demonstrated that both the PTuR and PEF-P promoters allowed a good level of expression of nisI and pedB in the LAB and bifidobacteria strains tested. A suitable concentration of nisin or pediocin could be established for the selection of the strains transformed with vectors harbouring the combination of the selected promoters and markers nisI and pedB, and this was successfully applied to different strains of LAB and bifidobacteria. Therefore, PTuR and PEF-P promoters are excellent candidates for the expression of nisI and/or pedB as selectable markers in LAB and bifidobacteria, and they are suitable for use in food grade vectors to allow the selection of genetically engineered strains. KEY POINTS: • Food-grade vectors require non-antibiotic selectable markers such as pedB and nisI. • Eight promoters from LAB or bifidobacteria were initially tested in L. lactis NZ9000. • PTuR and PEF-P efficiently drove the expression of pedB and nisI in LAB and bifidobacteria.

Lacticaseibacillus paracasei INIA P272 and Lacticaseibacillus rhamnosus INIA P344, isolated from breast-fed infants, are two promising bacterial strains for their use in functional foods according to their demonstrated probiotic and technological characteristics. To better understand their probiotic characteristics and evaluate their safety, here we report the draft genome sequences of both strains as well as the analysis of their genetical content. The draft genomes of L. paracasei INIA P272 and L. rhamnosus INIA P344 comprise 3.01 and 3.26 Mb, a total of 2994 and 3166 genes and a GC content of 46.27 % and 46.56 %, respectively. Genomic safety was assessed following the EFSA guidelines: the identification of both strains was confirmed through Average Nucleotide Identity, and the absence of virulence, pathogenic and antibiotic resistance genes was demonstrated. The genome stability analysis revealed the presence of plasmids and phage regions in both genomes, however, CRISPR sequences and other mechanisms to fight against phage infections were encoded. The probiotic abilities of both strains were supported by the presence of genes for the synthesis of SCFA, genes involved in resistance to acid and bile salts or a thiamine production cluster. Moreover, the encoded exopolysaccharide biosynthesis genes could provide additional protection against the deleterious gastrointestinal conditions, besides which, playing a key role in adherence and coaggregation of pathogenic bacteria together with the high number of adhesion proteins and domains encoded by both genomes. Additionally, the bacteriocin cluster genes found in both strains, could provide an advantageous ability to compete against pathogenic bacteria. This genomic study supports the probiotic characteristics described previously for these two strains and satisfies the safety requirements to be used in food products.

Fluorescent proteins are widely used for cell and protein tracking. Most of these proteins show a high signal and need the presence of oxygen to emit fluorescence. Among them, the fluorescent protein mCherry stands up because of its bright signal and fast maturation. Furthermore, the anaerobic cyan-green fluorescent protein Evoglow-Pp1 allows fluorescent detection under anaerobic conditions. In this work, we modified the pNZ:TuR.aFP plasmid, which harbors the gene encoding Evoglow-Pp1 and the promoter of elongation factor Tu from Limosilactobacillus reuteri CECT925, to obtain a plasmid containing the mrfp gene encoding the monomeric mCherry (pNZ:TuR.mCherry). Moreover, both genes were cloned together (pNZ:TuR.aFP.mCherry) developing a chimeric protein; and with a stop codon between them (pNZ:TuR.aFP.STOP.mCherry) resulting in the expression of both Evoglow-Pp1 and mCherry proteins separately under the influence of the same promoter. Lactococcus lactis, Lacticaseibacillus casei, Lactiplantibacillus plantarum, Limosilactobacillus fermentum, Lacticaseibacillus rhamnosus, and L. reuteri strains were transformed with the previously mentioned plasmids, showing an excellent red (pNZ:TuR.mCherry), green (pNZ:TuR.aFP), and red combined with green (pNZ:TuR.aFP.mCherry and pNZ:TuR.aFP.STOP.mCherry) fluorescence signal. Both fluorescence emissions were stable in strains transformed with pNZ:TuR.aFP.STOP.mCherry, while differences in the red or green fluorescence emission were observed in some of the strains harboring pNZ:TuR.aFP.mCherry. Moreover, these plasmids allowed strains differentiation in a complex environment, such as fecal microbiota. Hence, we present the plasmid pNZ:TuR.aFP.STOP.mCherry as a useful tool for the labeling of lactobacilli strains, which would be functional under anoxic conditions, thanks to Evoglow-Pp1, while having the high brightness and good photostability of mCherry. KEY POINTS: • LAB transformed with pNZ:TuR.mCherry expressed the red fluorescent p

The benefits to health attributed to the intake of phytoestrogens (PEs) have been demonstrated in previous studies with significant physiological concentrations of bioactive PEs, such as genistein, equol, enterolignans and urolithins in plasma. However, the achievement of high bioactive PE levels in plasma is restricted to a select population group, mainly due to the low intake of plant PEs and/or the absence, or inhibition, of the microbiota capable of producing these bioactive forms. In this study, the intake of plant PEs, the concentration of bioactive PEs in plasma, the ability of the intestinal microbiota to produce bioactive PEs, as well as the different mechanisms used by GRAS bacteria to increase the level of bioactive PEs were evaluated concluding that the use of GRAS bacteria bioactive PE producers and the development of fermented foods enriched in bioactive PEs in addition to a high intake of plant PEs and taking care of the intestinal microbiota, are some of the different strategies to achieve significant physiological concentrations of bioactive PEs in the intestine and, subsequently, in plasma and targets organs which are essential to improve menopausal symptoms or reduce the risk of some pathologies such as breast and colon cancer, or cardiovascular disease.

Three strains with technological and probiotic properties (Lactococcus lactis INIA 650, Lacticaseibacillus paracasei INIA P272 and Bifidobacterium breve INIA P734) were used in different combinations as adjunct cultures for the manufacture of pasteurized ewe's milk. The three strains showed good viability during cheese manufacture and ripening. Moreover, they all were able to survive after the addition of the cheese samples to an in vitro colonic model. The nisin-producing strain L. lactis INIA 650 was able to produce the antimicrobial peptide nisin not only during cheese manufacture and ripening, but also in the colonic model. None of the strains combinations added to the cheese as adjunct cultures had a negative influence in their physicochemical or sensory characteristics when compared to control cheese. This work shows the suitability of cheese as vehicle for a multi-strain combination with potential dual effect in food and gut.

Limosilactobacillus reuteri INIA P572 is a strain able to produce the antimicrobial compound reuterin in dairy products, exhibiting a protective effect against some food-borne pathogens. In this study, we investigated some probiotic properties of this strain such as resistance to gastrointestinal passage or to colonic conditions, reuterin production in a colonic environment, and immunomodulatory activity, using different in vitro and in vivo models. The results showed a high resistance of this strain to gastrointestinal conditions, as well as capacity to grow and produce reuterin in a human colonic model. Although the in vitro assays using the RAW 264.7 macrophage cell line did not demonstrate direct immunomodulatory properties, the in vivo assays using a Dextran Sulphate Sodium (DSS)-induced colitic mice model showed clear immunomodulatory and protective effects of this strain.

Limosilactobacillus reuteri is a beneficial bacterium that inhabits the gastrointestinal tract of different mammals. Diverse beneficial effects have been attributed to specific strains, in part mediated by the production of reuterin. Here, we report the draft genome sequence of L. reuteri INIA P572, a reuterin-producing strain isolated from pig feces.

The draft genome sequence of Bifidobacterium breve INIA P734, a strain shared by mother and child, is reported. It consists of 50 contigs, with 2,391,925 bp, 2,099 genes, and a G+C content of 58.8%. The genome analysis revealed the absence of antibiotic resistance and pathogenicity-related genes.

The isoflavones daidzin and genistin, present in soybeans, can be transformed by the intestinal microbiota into equol and 5-hydroxy-equol, compounds with enhanced availability and bioactivity, although these are only produced by a fraction of the population. Hence, there is an interest in the production of these compounds, although, to date, few bacteria with biotechnological interest and applicability in food have been found able to produce equol. In order to obtain lactic acid bacteria able to produce equol, the daidzein reductase (dzr), dihydrodaidzein reductase (ddr), tetrahydrodaidzein reductase (tdr) and dihydrodaidzein racemase (ifcA) genes, from Slackia isoflavoniconvertens DSM22006, were cloned into the vector pNZ:TuR, under a strong constitutive promoter (TuR). Lactococcus lactis MG1363, Lacticaseibacillus casei BL23, Lactiplantibacillus plantarum WCFS1, Limosilactobacillus fermentum INIA 584L and L. fermentum INIA 832L, harbouring pNZ:TuR.tdr.ddr, were able to produce equol from dihydrodaidzein, while L. fermentum strains showed also production of 5-hydroxy-equol from dihydrogenistein. The metabolization of daidzein and genistein by the combination of strains harbouring pNZ:TuR.dzr and pNZ:TuR.tdr.ddr showed similar results, and the addition of the correspondent strain harbouring pNZ:TuR.ifcA resulted in an increase of equol production, but only in the L. fermentum strains. This pattern of equol and 5-hydroxy-equol production by L. fermentum strains was also confirmed in cow's milk supplemented with daidzein and genistein and incubated with the different combination of strains harbouring the constructed plasmids. Bacteria generally recognized as safe (GRAS), such as the lactic acid bacteria species used in this work, harbouring these plasmids, would be of value for the development of fermented vegetal foods enriched in equol and 5-hydroxy-equol.

Three bifidobacteria strains of human origin (Bifidobacterium pseudolongum INIA P2, Bifidobacterium breve INIA P734, and Bifidobacterium longum INIA P678) were used as potential probiotic adjunct cultures for the manufacture of pasteurized sheep milk cheese. Bifidobacteria were inoculated at 5 to 6 log cfu/mL in milk vats. Microbiological, physicochemical, rheological, color, and sensory characteristics were determined at 7, 28, and 60 d of ripening. Counts of B. pseudolongum INIA P2 remained above 6 log cfu/g during 60 d of ripening as well as after further simulated gastrointestinal digestion of cheeses. Bifidobacterium breve INIA P734 counts remained stable during 28 d and decreased by less than 1 log unit after simulated digestion. Bifidobacterium longum INIA P678 counts dropped sharply during cheese manufacture and ripening and were below detection level after simulated digestion. Addition of bifidobacteria strains did not affect starter viability, cheese pH, dry matter, water activity, or salt content but significantly increased overall proteolysis and the concentration of some free amino acids. Cheeses with bifidobacteria exhibited no significant differences in most sensory characteristics with respect to control cheese. According to our results, B. breve INIA P734 and B. pseudolongum INIA P2 are promising candidates as probiotic adjunct cultures in fresh and semi-hard sheep milk cheese.

Genes involved in the transport and catabolism of carbohydrates are usually controlled through the binding of the catabolite control protein A (CcpA) to the catabolite-responsive elements (cre) of target genes in Gram-positive bacteria. In this work, we show how the elimination of the cre sites in Lactobacillus casei BL23 promoters induced by sorbitol (PgutF), maltose (PmalL), and myo-inositol (PiolT) allowed the induction of gene expression in media supplemented with sorbitol, maltose, and myo-inositol, respectively, even in the presence of glucose. This was studied using plasmids encoding the anaerobic fluorescent protein evoglow-Pp1 as a reporter. In addition, gutF cre site was introduced into a bile inducible promoter (P16090) and into the constitutive promoter of the elongation factor P (PEf-P) of L. casei BL23. The existence of the cre site blocked gene expression in the P16090 inducible promoter in the presence of glucose, while it had no influence on the expression of the PEf-P constitutive one. These results demonstrated that the introduction or elimination of cre sites in inducible promoters allows the control and modification of their heterologous genetic expression, showing how the cre site, the transcriptional regulator, and CcpA interact to control gene expression in inducible genes. KEY POINTS: • Cre sequences regulate gene expression in inducible promoters in L. casei BL23. • Cre sites do not affect gene expression in constitutive promoters in L. casei BL23. • Cre sequences could control heterologous genic expression in lactobacilli.

Detection of bioactive peptides in complex ecosystems like intestinal environment is a difficult task. In this study, we developed two new bioreporters for nisin based on Lactococcus lactis NZ9000 transformed with the vector pNZ:Nis-aFP or pNZ:Nis-mCherry, that encoded for the anaerobic fluorescent protein evoglow-Pp1 (aFP) or the fluorescent protein mCherry, respectively. The biosensors were used to study nisin A production by L. lactis INIA 650 in milk and in a colonic model. The use of L. lactis NZ9000 pNZ:Nis-aFP as a biosensor allowed the detection of nisin produced by L. lactis INIA 650 in milk, but not in the in vitro colonic model. In milk, this reporter was induced by direct addition of 10 ng/ml nisin while, in the colonic model, nisin concentrations of 50 ng/ml were necessary. However, the reporter system based on pNZ:Nis-mCherry showed a higher sensibility, detecting nisin concentrations of 1 ng/ml produced by L. lactis INIA 650 in colonic media using agar diffusion or cross streak bioassays.

Cheeses manufactured from pasteurized milk supplemented with glycerol and reuterin-producing Lactobacillus reuteri INIA P572 as adjunct to the commercial starter culture were analysed in order to optimize a biopreservation strategy. The highest reuterin concentration determined by a colorimetric assay was detected on day 1 in cheeses with 100–500 mM glycerol. The presence of reuterin was confirmed by a direct detection technique as HPLC. Cheeses made with L. reuteri and 200 or 500 mM glycerol showed a red tendency in color in comparison with control. The results with purified reuterin suggested that the development of slightly rosy colour in cheese was related to some compound produced/overproduced when higher levels of glycerol were present in cheese, but not due to reuterin. Application of L. reuteri INIA P572 as adjunct to the commercial starter with 100 mM glycerol led to such a reuterin concentration in cheese that could control undesirable microorganisms, avoiding the presence of color-changing compounds. © 2017, Association of Food Scientists & Technologists (India).

Age-related degeneration gives rise to a number of pathologies, many of them associated with imbalances of the microbiota and the gut-associated immune system. Thus, the intestine is considered a key target organ to improve the quality of life in senescence. Gut microbiota can have a powerful impact in the deterioration linked to aging by its nutritional and immunomodulatory activity. Reduced numbers of beneficial species and low microbial biodiversity in the elderly have been linked with pathogenesis of many diseases. A healthy lifestyle with an elderly customized diet including probiotics can contribute to reducing the chronic proinflammatory status and other age-related pathologies. Beneficial effects of probiotic lactic acid bacteria and bifidobacteria to alleviate some of these disorders based on their immunomodulatory properties as well as their capacity to produce bioactive metabolites from dietary phytoestrogens are summarized. On one hand, the preservation of gut barrier integrity and an increased ability to fight infections are the main reported immune benefits of probiotics. On the other hand, the intake of a diet rich in phytoestrogens along with the presence of selected probiotic bacteria may lead to the production of equol, enterolignans, and urolithins, which are considered protective against chronic diseases related to aging. © 2017 José Mariá Landete et al.

The food industry seeks alternatives to satisfy consumer demands of safe foods with a long shelf-life able to maintain the nutritional and organoleptic quality. The application of antimicrobial compounds-producing protective cultures may provide an additional parameter of processing in order to improve the safety and ensure food quality, keeping or enhancing its sensorial characteristics. In addition, strong evidences suggest that certain probiotic strains can confer resistance against infection with enteric pathogens. Several mechanisms have been proposed to support this phenomenon, including antimicrobial compounds secreted by the probiotics, competitive exclusion, or stimulation of the immune system. Recent research has increasingly demonstrated the role of antimicrobial compounds as protective mechanism against intestinal pathogens and therefore certain strains could have an effect on both the food and the gut. In this aspect, the effects of the combination of different strains keep unknown. The development of multistrain probiotic dairy products with good technological properties and with improved characteristics to those shown by the individual strains, able to act not only as protective cultures in foods, but also as probiotics able to exert a protective action against infections, has gained increased interest. © 2015 Juan L. Arqués et al.

The genes involved in the glycerol metabolism, glycerol dehydratase (gdh) and two propanediol dehydrogenases (pdh30 and pdh1734), were analyzed in different reuterin- and non-reuterin-producing lactobacilli of biotechnological interest. All the reuterin-producing lactobacilli expressed the gdh, pdh30 and pdh1734, except Lb. coryniformis CECT 5711 which did not contain pdh30. Reuterin production levels in Lb. coryniformis CECT 5711 were much lower than those in reuterin-producing Lb. reuteri. A positive relationship between cobalamin production levels and reuterin production levels was observed in all reuterin-producing lactobacilli tested. Intriguingly, when Lb. coryniformis CECT 5711 was supplemented with cobalamin, a seven times increase in reuterin production was observed. On the other hand, Lb. brevis ESI38 that possess and express gdh, pdh30 and pdh1734, was unable to produce reuterin or cobalamin. To study the role of pdh30 during glycerol metabolism, the gene disruption mutant Lb. brevis INIA ESI38pORI28-pdh30 was constructed. HPLC analysis of the glycerol fermentation products showed an involvement of the pdh30 in the 3-hydroxypropionic acid (3-HP) biosynthesis. However, Lb. coryniformis, that lack pdh30, showed the higher levels of 3-HP, indicating other catalytic mechanisms to produce 3-HP in this strain. The 1,3-propanediol peak was detected in the Lb. reuteri and Lb. coryniformis chromatograms, but not in Lb. brevis, which also confirm divergences in Lactobacillus glycerol metabolism. © 2015, Springer-Verlag Berlin Heidelberg.

Lactobacillus reuteri INIA P579 was used for the production and purification of reuterin. The purity of reuterin was assessed by high resolution electrospray ionization mass spectrometry (HRESIMS) and nuclear magnetic resonance (NMR) spectroscopy. After purification, reuterin concentration obtained was 1.3M. The inhibitory activity using Escherichia coli K12 as indicator strain was estimated to be 510 AU/ml. Survival curves in tryptic soy broth revealed that reuterin required to inhibit the growth of three Listeria monocytogenes strains was in the range of 2-4 AU/ml. Purified reuterin (10 AU/g) significantly reduced the growth of L.monocytogenes in cold-smoked salmon kept under moderate or strong temperature abuse conditions. After 15d at 8°C, cold-smoked salmon with added reuterin exhibited L.monocytogenes counts 2.0 log CFU/g lower than control smoked salmon with no reuterin added. At 30°C, reuterin also controlled the growth of the pathogen, with counts 1.4 and 0.9 log CFU/g lower than those observed in control smoked salmon after 24 and 48h, respectively. The addition of purified reuterin might be used as a hurdle technology to improve the safety and extend the shelf-life of lightly preserved seafood products such as cold-smoked salmon. © 2014 Elsevier Ltd.

An efficient method for genetic transformation of lactic acid bacteria (LAB) by electroporation is presented in this work. A comparative survey with other electrotransformation methods already published showed that the method proposed here yields the higher electrotransformation efficiency in the 12 LAB strains tested, which could make the method applicable to other LAB species or genera. © 2014 Elsevier B.V.

Human milk has been traditionally considered sterile; however, recent studies have shown that it represents a continuous supply of commensal, mutualistic and/or potentially probiotic bacteria to the infant gut. Culture-dependent and -independent techniques have revealed the dominance of staphylococci, streptococci, lactic acid bacteria and bifidobacteria in this biological fluid, and their role on the colonization of the infant gut. These bacteria could protect the infant against infections and contribute to the maturation of the immune system, among other functions. Different studies suggest that some bacteria present in the maternal gut could reach the mammary gland during late pregnancy and lactation through a mechanism involving gut monocytes. Thus, modulation of maternal gut microbiota during pregnancy and lactation could have a direct effect on infant health. On the other hand, mammary dysbiosis may lead to mastitis, a condition that represents the first medical cause for undesired weaning. Selected strains isolated from breast milk can be good candidates for use as probiotics. In this review, their potential uses for the treatment of mastitis and to inhibit mother-to-infant transfer of HIV are discussed. © 2012 Elsevier Ltd. All Rights Reserved.

In situ reuterin production during the manufacture and storage of two dairy model systems elaborated from milk supplemented with an optimized concentration of glycerol (50 mM) and inoculated with a commercial starter and with different reuterin-producing Lactobacillus reuteri strains was investigated. L. reuteri was able to survive and to produce reuterin in cheese and yogurt models. The highest reuterin production was achieved by L. reuteri INIA P572 and INIA P579, which displayed reuterin concentrations up to 5.5 mM in cheese and up to 1.5 mM in yogurt. The addition of reuterin-producing L. reuteri and glycerol to milk reduced the viable counts of the cheese starter from day 10 onwards, while did not influence the counts of the yogurt starter, compared to control dairy models without L. reuteri. Strains L. reuteri INIA P572 and INIA P579 could be promising candidates in the development of bioprotective cultures to control pathogenic microorganisms in dairy products due to the potentially inhibitory concentrations of reuterin achieved in situ. © 2013 Elsevier Ltd.

Recent work has shown that human colostrum and milk, which had been traditionally considered sterile, provides a continuous supply of commensal and potential probiotic bacteria to the infant gut. More than 200 different bacterial species, including staphylococci, lactic acid bacteria and bifidobacteria, have been isolated from human milk samples so far, although the cultivable bacterial diversity found in individual samples is much lower (2 to 8 different species per women). Interestingly, the same bacterial strains have been found in both breast milk and infant feces of different mother-infant pairs, confirming the role of human milk on the bacterial colonization of the infant gut. These commensal bacteria could protect the infant gut and direct, at least partly, the maturation of the immune system, among other functions. Different studies suggest that some bacteria present in the maternal gut could reach the mammary gland during late pregnancy and lactation through a mechanism involving gut dendritic cells and macrophages. Thus, modulation of maternal gut microbiota during pregnancy and lactation could have a direct effect on infant health. © 2015.