Jonathan Emiliano Laiño

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 B12 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.

Applicability of a Lactobacillus amylovorus strain as co-culture for natural folate bio-enrichment of fermented milk.

The ability of 55 strains from different Lactobacillus species to produce folate was investigated. In order to evaluate folic acid productivity, lactobacilli were cultivated in the folate-free culture medium (FACM). Most of the tested strains needed folate for growth. The production and the extent of vitamin accumulation were distinctive features of individual strains. Lactobacillus amylovorus CRL887 was selected for further studies because of its ability to produce significantly higher concentrations of vitamin (81.2 ± 5.4 μg/L). The safety of this newly identified folate producing strain was evaluated through healthy experimental mice. No bacterial translocation was detected in liver and spleen after consumption of CRL887 during 7 days and no undesirable side effects were observed in the animals that received this strain. This strain in co-culture with previously selected folate producing starter cultures (Lactobacillus bulgaricus CRL871, and Streptococcus thermophilus CRL803 and CRL415) yielded a yogurt containing high folate concentrations (263.1 ± 2.4 μg/L); a single portion of which would provide 15% of the recommended dietary allowance. This is the first report where a Lactobacillus amylovorus strain was successfully used as co-culture for natural folate bio-enrichment of fermented milk.

Relationships between the genome and some phenotypical properties of Lactobacillus fermentum CECT 5716, a probiotic strain isolated from human milk

Lactobacillus fermentum CECT 5716, isolated from human milk, has immunomodulatory, anti-inflammatory, and anti-infectious properties, as revealed by several in vitro and in vivo assays, which suggests a strong potential as a probiotic strain. In this work, some phenotypic properties of L. fermentum CECT 5716 were evaluated, and the genetic basis for the obtained results was searched for in the strain genome. L. fermentum CECT 5716 does not contain plasmids and showed neither bacteriocin nor biogenic amine biosynthesis ability but was able to produce organic acids, glutathione, riboflavin, and folates and to moderately stimulate the maturation of mouse dendritic cells. No prophages could be induced, and the strain was sensitive to all antibiotics proposed by European Food Safety Authority (EFSA) standards, while no transmissible genes potentially involved in antibiotic resistance were detected in its genome. Globally, there was an agreement between the phenotype properties of L. fermentum CECT 5716 and the genetic information contained in its genome.

Ancestral Andean grain quinoa as source of lactic acid bacteria capable to degrade phytate and produce B-group vitamins

The lactic acid bacteria (LAB) microbiota of quinoa grains (QG) and spontaneous sourdough (QSS) was evaluated. Different strains of Lactobacillus (L.) plantarum (7), L. rhamnosus (5), L. sakei (1), Pediococcus (Ped.) pentosaceus (9), Leuconostoc (Leuc.) mesenteroides (1), Enterococcus (E.) casseliflavus (2), E. mundtii (3), E. hirae (1), E. gallinarum (12), Enterococcus sp. (1), and E. hermanniensis (2) were isolated, identified and characterized. Only four strains isolated from QSS and eight strains isolated from QG showed amylolytic activity. L. plantarum CRL 1973 and CRL 1970, L. rhamnosus CRL 1972 and L. sakei CRL 1978 produced elevated concentrations of folate with strain CRL 1973 producing the highest concentration (143±6ng/ml). L. rhamnosus, isolated from QSS, was the LAB species that produced the most elevated concentrations of total riboflavin (>270ng/ml) with strain CRL 1963 producing the highest amounts (360±10ng/ml). Phytase activity, evaluated in forty-four LAB strains from quinoa, was predominantly detected in L. rhamnosus and Enterococci strains with the highest activities observed in E. mundtii CRL 2007 (957±25U/ml) followed by E. casseliflavus CRL 1988 (684±38U/ml), Leuc. mesenteroides CRL 2012 (617±38U/ml) and L. rhamnosus CRL 1983 (606±79U/ml). In conclusion, this study shows that a diverse LAB microbiota is present in quinoa with important properties; these microorganisms could be used as potential starter cultures to increase the nutritional and functional properties of Andean grains based foods.

Modulation of the immuno-coagulative response in a pneumococcal infection in malnourished mice nasally treated with Lactobacillus casei

We studied the systemic effects of the intranasal administration of Lactobacillus casei on the immuno-coagulative response in pneumoccocal infection in immunocompromised mice. Weaned mice consumed a protein-free diet (PFD) for 21 days and were therefore malnourished. Malnourished mice were fed a balanced conventional diet (BCD) for 7 days (BCD group) or a BCD for 7 days with nasal administration of viable L. casei on days 6 and 7 (BCD+LcN group). The malnourished control mice (MNC) received a PFD, whereas the well-nourished control mice (WNC) continually consumed a BCD. At the end of the treatment period, the mice were infected with Streptococcus pneumoniae. At different times after infection, we analysed the following parameters: global coagulation system, activation of coagulation, coagulation inhibitors, platelet count, leukocyte count and myeloperoxidase (MPO) activity, total proteins, albumin and acute phase proteins (APPs). The MNC group showed greater impairment in the coagulation tests and an increase in the positive APPs. These parameters were normalized by the L. casei treatment. However, the number of leukocytes, decreased by malnutrition, was improved only by the administration of L. casei. After infection, the BCD+LcN group showed similar results to those of the WNC group for most of the haemostatic parameters. The BCD+LcN group did not show significant variations in the prothrombin time or in the level of anticoagulant protein C, but showed higher levels of fibrinogen, platelets, albumin, leukocytes and MPO activity compared with the different experimental groups. The intranasal administration of L. casei was effective in modulating the pro-inflammatory aspects of coagulation without affecting coagulation itself.

Soyamilk fermented with riboflavin-producing Lactobacillus plantarum CRL 2130 reverts and prevents ariboflavinosis in murine models.

It has been previously shown that Lactobacillus plantarum CRL 2130 is able to produce riboflavin in soyamilk. The aim of the present study was to evaluate the efficiency of this riboflavin-bio-enriched soyamilk to revert and/or prevent the nutritional deficiency of riboflavin using different animal models. When used to supplement the diets of previously depleted animals, it was shown that the growth, riboflavin status and morphology of the small intestines reverted to normal parameters and were similar to animals supplemented with commercial riboflavin. In the prevention model, the same tendency was observed, where animals that received soyamilk fermented with L. plantarum CRL 2130 did not show signs of riboflavin deficiency. This new bio-fortified soya-based product could be used as part of normal diets to provide a more natural alternative to mandatory fortification with riboflavin for the prevention of its deficiency.

Draft Genome Sequence of Lactobacillus delbrueckii subsp. bulgaricus CRL871, a Folate-Producing Strain Isolated from a Northwestern Argentinian Yogurt.

ABSTRACT Lactobacillus delbrueckii subsp. bulgaricus CRL871 is the first strain of L. delbrueckii subsp. bulgaricus reported as a folate-producing strain. We report the draft genome sequence of L. delbrueckii subsp. bulgaricus CRL871 (2,063,981 bp, G+C content of 49.1%). This strain is of great biotechnological importance to the dairy industry because it constitutes an alternative to folic acid fortification.

Lactobacillus rhamnosus CRL1505 beneficially modulates the immuno-coagulative response after pneumococcal infection in immunocompromised malnourished mice

This work evaluated the effect of orally or nasally administered Lactobacillus rhamnosus CRL1505 on the resistance of immunocompromised protein-malnourished mice to pneumococcal infection. In particular, we aimed to gain knowledge of the mechanism involved in the immunomodulatory effect of L. rhamnosus CRL1505 in malnourished hosts by evaluating its impact on the immuno-coagulative response. Malnutrition significantly increased lung tissue damage caused by Streptococcus pneumoniae infection. Lung damage was associated with a deregulated activation of coagulation and an altered inflammatory response. Pneumococcal colonization of lung and bacteremia were significantly reduced (p < 0.05) in malnourished mice receiving the CRL1505 strain. Moreover, mice repleted with supplemental L. rhamnosus CRL1505 showed the least alteration of the alveolar-capillary barrier and cell damage in lungs after the infectious challenge, especially when the CRL1505 strain was administered by nasal route. Besides, mice treated with L. rhamnosus CRL1505 showed an improved respiratory innate immune response and a lower activation of coagulation. The results of this work indicate that L. rhamnosus CRL1505 is able to beneficially modulate the inflammation-coagulation interaction after respiratory infections in malnourished hosts.

Characterization of folate production and probiotic potential of Streptococcus gallolyticus subsp. macedonicus CRL415

Mandatory fortification of foods with folic acid is being questioned by many scientists principally because of the potential adverse secondary effects associated with their excessive consumption. It has been shown that selected strains of lactic acid bacteria (LAB) are able to produce natural forms of folate and these could be included in foods to prevent deficiencies without causing adverse effects. The aim of this study was to evaluate folate production and fol gene expression by Streptococcus gallolyticus subsp. macedonicus (S. macedonicus) CRL415 under different growth conditions in vitro and to assess its potential probiotic application. Results showed that glucose as the principal carbon source, and incubation at 42 °C under controlled pH conditions (6.0) increased folate production, fol gene expression, and growth of S. macedonicus CRL415. This strain was able to produce elevated folate concentrations during milk fermentation without the need of prolonged incubation times and was able to resist conditions simulating the gastrointestinal tract. In addition, S. macedonicus was susceptible to all required antibiotics, and had a good adhesion level to intestinal cells in vitro, making it a promising candidate for biotechnological application as functional starter cultures in the dairy industry.

Factors stimulating riboflavin produced by Lactobacillus plantarum CRL 725 grown in a semi‐defined medium

Riboflavin (vitamin B2) is one of the B‐group water‐soluble vitamins and is essential for energy metabolism of the cell. The aim of this study was to determine factors that affect riboflavin production by Lactobacillus (L.) plantarum CRL 725 grown in a semi defined medium and evaluate the expression of its rib genes. The factors found to enhance riboflavin production in this medium were incubation at 30 °C, and the addition of specific medium constituents, such as casamino acids (10 g L−1), guanosine (0.04 g L−1), and sucrose as carbon source (20 g L−1). In these conditions, higher riboflavin concentrations were directly associated with significant increases in the expression of ribA, ribB, and ribC genes. The culture conditions defined in this work and its application to a roseoflavin resistant mutant of L. plantarum allowed for a sixfold increase in riboflavin concentrations in our semi‐defined medium which were also significantly higher than those obtained previously using the same strain to ferment soymilk. These conditions should thus be evaluated to increase vitamin production in fermented foods.