Biodiversity and technological-functional potential of lactic acid bacteria isolated from spontaneously fermented chia sourdough.

Chia, is a gluten-free, rich in proteins, oilseed that is "on trend" as an alternative ingredient in food production, adding nutritional value. As a reservoir of natural biodiversity, lactic acid bacteria development, during spontaneous chia flour fermentation (sourdough) for 10 days, were investigated by culturing and high throughput sequencing (HTS). Culture-dependent analysis showed a rapid increase in total LAB numbers from the second day of sourdough refreshment. Taxonomical identification of LAB isolates by rep-PCR and further 16S rRNA sequencing was performed. Besides Among identified LAB by culture-dependent approach, species from genus Enterococcus were the most abundant; Lactococcus (Lc. lactis), Lactobacillus (L. rhamnosus) and Weissella (W. cibaria) species were also isolated. By HTS, twelve OTUs belonging to LAB genera were identified during chia sourdough fermentation with an increased Lactobacillus diversity. Enterococcus (E.) faecium, E. mundtii, W. cibaria and L. rhamnosus were detected as dominant species in the final propagation stages while Bacillus and Clostridium were mostly present during first fermentation stages. The investigation of biotechnological and safety traits (acidification ability, protein hydrolysis, exopolysaccharides production, antimicrobial activity and antibiotic resistance) of 15 representative LAB strains was performed. Strains characterization led to the selection of Lc. lactis CH179, L. rhamnosus CH34 and W. cibaria CH28 as candidates to be used as novel functional starter culture for gluten-free chia fermented products. As far as we know, this is the first study providing information on the molecular inventory of LAB population during spontaneous fermentation of chia sourdough.

Characterization of the pattern of alphas1- and beta-casein breakdown and release of a bioactive peptide by a cell envelope proteinase from Lactobacillus delbrueckii subsp. lactis CRL 581.

The cell envelope-associated proteinases (CEPs) of the lactobacilli have key roles in bacterial nutrition and contribute to the development of the organoleptic properties of fermented milk products as well, as they can release bioactive health-beneficial peptides from milk proteins. The influence of the peptide supply, carbohydrate source, and osmolites on the CEP activity of the cheese starter Lactobacillus delbrueckii subsp. lactis CRL 581 was investigated. The CEP activity levels were controlled by the peptide content of the growth medium. The maximum activity was observed in a basal minimal defined medium, whereas in the presence of Casitone, Casamino Acids, or yeast extract, the synthesis of CEP was inhibited 99-, 70-, and 68-fold, respectively. The addition of specific di- or tripeptides containing branched-chain amino acids, such as leucylleucine, prolylleucine, leucylglycylglycine, or leucylproline, to the growth medium negatively affected CEP activity, whereas dipeptides without branched-chain amino acids had no effect on the enzyme's production. The carbon source and osmolites did not affect CEP activity. The CEP of L. delbrueckii subsp. lactis CRL 581 exhibited a mixed-type CEP(I/III) variant caseinolytic specificity. Mass-spectrometric screening of the main peptide peaks isolated by reverse-phase high-pressure liquid chromatography allowed the identification of 33 and 32 peptides in the alpha(s1)- and beta-casein hydrolysates, respectively. By characterizing the peptide sequence in these hydrolysates, a pattern of alpha(s1)- and beta-casein breakdown was defined and is reported herein, this being the first report for a CEP of L. delbrueckii subsp. lactis. In this pattern, a series of potentially bioactive peptides (antihypertensive and phosphopeptides) which are encrypted within the precursor protein could be visualized.

Metabolic engineering of lactic acid bacteria for the production of nutraceuticals.

Lactic acid bacteria display a relatively simple and well-described metabolism where the sugar source is converted mainly to lactic acid. Here we will shortly describe metabolic engineering strategies on the level of sugar metabolism, that lead to either the efficient re-routing of the lactococcal sugar metabolism to nutritional end-products other than lactic acid such as L-alanine, several low-calorie sugars and oligosaccharides or to enhancement of sugar metabolism for complete removal of (undesirable) sugars from food materials. Moreover, we will review current metabolic engineering approaches that aim at increasing the flux through complex biosynthetic pathways, leading to the production of the B-vitamins folate and riboflavin. An overview of these metabolic engineering activities can be found on the website of the Nutra Cells 5th Framework EU-project (www.nutracells.com). Finally, the impact of the developments in the area of genomics and corresponding high-throughput technologies on nutraceutical production will be discussed.