Interaction between lactic acid bacteria and Polygonatum sibiricum saponins and its application to microencapsulated co-delivery.

This study aimed to investigate the interaction between L.casei and L.bulgaricus with Polygonatum sibiricum saponins (PSS) and to explore the co-microencapsulation to reduce their loss rate during storage and consumption. 1% PSS was added to the culture broth, and it was found that the growth and metabolism of the strains were accelerated, especially in the compound probiotic group, indicating that PSS has potential for prebiotics. LC-MS observed significant differences in the composition and content of saponins in PSS. The metabolomics results suggest that the addition of PSS resulted in significant changes in the metabolites of probiotics. In addition, it was found that the combination of probiotics and PSS may have stronger hypoglycemic ability (ɑ-glucosidase, HepG2). Finally, a co-microencapsulated delivery system was constructed using zein and isomaltooligosaccharide. This system can achieve more excellent resistance of probiotics and PSS in gastrointestinal fluids, effectively transporting both to the small intestine.

The effect of Thymus vulgaris and Thymbra spicata essential oils and/or extracts in pectin edible coating on the preservation of sliced bolognas.

The aim of this research was to determine the effect of pectin coating made with essential oils and/or extracts of Thymus vulgaris (thyme) and Thymbra spicata (thymbra) on the preservation of aerobically packaged sliced bolognas during cold storage. The treatment made with essential oils resulted in a reduction of 1.73 log CFU/g of Salmonella typhimurium ATCC 14028. Also, pectin coating made with essential oil-treated sliced bolognas had the lowest total mesophilic bacteria (6.27 log CFU/g), and total lactic acid bacteria (1.72 CFU/g), in comparison to non-treated bolognas, with 7.65 log CFU/g for total mesophilic bacteria and 4.99 log CFU/g for lactic acid bacteria. Application of an emulsion significantly (P < 0.05) affected L*(lightness), a*(redness), and b*(yellowness) values. The essential oil treatment had the highest TBARS values at the end of the storage period. The pH was not affected by the treatment (P > 0.05), but storage had a significant (P < 0.05) effect on the pH values.

Feeding with Sustainably Sourdough Bread Has the Potential to Promote the Healthy Microbiota Metabolism at the Colon Level.

The contribution of sustainably food processing to healthy intestinal microbial functions is of recent acquisition. The sourdough fermentation fits well with the most sustainable bread making. We manufactured baker's yeast (BYB) and sourdough (t-SB30) breads, which first underwent to an in-depth characterization. According to nutritional questionnaires, we selected 40 volunteers adhering to the Mediterranean diet. Data on their fecal microbiota and metabolome allowed the selection of two highly representative fecal donors to separately run the Twin Mucosal-SHIME (Twin M-SHIME) under 2-week feeding with BYB and t-SB30. Bread feeding did not affect the microbial composition at phylum and family levels of both donors, in all Twin M-SHIME colon tracts, and lumen and mucosal compartments. The genus core microbiota showed few significant fluctuations, which regarded the relative abundances of Lactobacillus and Leuconostoc according to feeding with BYB and t-SB30, respectively. Compared with BYB, the content of all short chain fatty acids (SCFA), and isovaleric and 2-methylbutyric acids significantly increased with t-SB30 feeding. This was evident for all Twin M-SHIME colon tracts and both donors. The same was found for the content of Asp, Thr, Glu, GABA, and Orn. The bread characterization made possible to identify the main features responsible for this metabolic response. Compared with BYB, t-SB30 had much higher contents of resistant starch, peptides, and free amino acids, and an inhomogeneous microstructure. We used the most efficient approach to investigate a staple food component, excluding interferences from other dietary factors and attenuating human physiology overlaps. The daily consumption of sourdough bread may promote the healthy microbiota metabolism at colon level. IMPORTANCE Knowledge on environmental factors, which may compose the gut microbiota, and drive the host physiology and health is of paramount importance. Human dietary habits and food compositions are pivotal drivers to assemble the human gut microbiota, but, inevitably, unmapped for many diet components, which are poorly investigated individually. Approximately 30% of the human diet consists of fermented foods and beverages. Bread, a fermented/leavened food, is a basic component of the human diet. Its potential effect on gut microbiota composition and functionality is challenging. In this study, we industrially made baker's yeast and sourdough breads, which were used to feed the Twin Mucosal-SHIME, a worldwide scientifically validated gastrointestinal simulator. Only the consumption of sourdough bread has the potential to enhance the synthesis of short chain fatty acids and free amino acids at the colon level.

Recent Advances in Lactic Acid Production by Lactic Acid Bacteria.

Lactic acid can synthesize high value-added chemicals such as poly lactic acid. In order to further minimize the cost of lactic acid production, some effective strategies (e.g., effective mutagenesis and metabolic engineering) have been applied to increase productive capacity of lactic acid bacteria. In addition, low-cost cheap raw materials (e.g., cheap carbon source and cheap nitrogen source) are also used to reduce the cost of lactic acid production. In this review, we summarized the recent developments in lactic acid production, including efficient strain modification technology (high-efficiency mutagenesis means, adaptive laboratory evolution, and metabolic engineering), the use of low-cost cheap raw materials, and also discussed the future prospects of this field, which could promote the development of lactic acid industry.

Positive Interactions between Lactic Acid Bacteria Promoted by Nitrogen-Based Nutritional Dependencies.

Nutritional dependencies, especially those regarding nitrogen sources, govern numerous microbial positive interactions. As for lactic acid bacteria (LAB), responsible for the sanitary, organoleptic, and health properties of most fermented products, such positive interactions have previously been studied between yogurt bacteria. However, they have never been exploited to create artificial cocultures of LAB that would not necessarily coexist naturally, i.e., from different origins. The objective of this study was to promote LAB positive interactions, based on nitrogen dependencies in cocultures, and to investigate how these interactions affect some functional outputs, e.g., acidification rates, carbohydrate consumption, and volatile-compound production. The strategy was to exploit both proteolytic activities and amino acid auxotrophies of LAB. A chemically defined medium was thus developed to specifically allow the growth of six strains used, three proteolytic and three nonproteolytic. Each of the proteolytic strains, Enterococcus faecalis CIRM-BIA2412, Lactococcus lactis NCDO2125, and CIRM-BIA244, was cocultured with each one of the nonproteolytic LAB strains, L. lactis NCDO2111 and Lactiplantibacillus plantarum CIRM-BIA465 and CIRM-BIA1524. Bacterial growth was monitored using compartmented chambers to compare growth in mono- and cocultures. Acidification, carbohydrate consumption, and volatile-compound production were evaluated in direct cocultures. Each proteolytic strain induced different types of interactions: strongly positive interactions, weakly positive interactions, and no interactions were seen with E. faecalis CIRM-BIA2412, L. lactis NCDO2125, and L. lactis CIRM-BIA244, respectively. Strong interactions were associated with higher concentrations of tryptophan, valine, phenylalanine, leucine, isoleucine, and peptides. They led to higher acidification rates, lower pH, higher raffinose utilization, and higher concentrations of five volatile compounds. IMPORTANCE Interactions of lactic acid bacteria (LAB) are often studied in association with yeasts or propionibacteria in various fermented food products, and the mechanisms underlying their interactions are being quite well characterized. Concerning interactions between LAB, they have mainly been investigated to test antagonistic interactions. Understanding how they can positively interact could be useful in multiple food-related fields: production of fermented food products with enhanced functional properties or fermentation of new food matrices. This study investigated the exploitation of the proteolytic activity of LAB strains to promote positive interactions between proteolytic and nonproteolytic strains. The results suggest that proteolytic LAB do not equally stimulate nonproteolytic LAB and that the stronger the interactions between LAB are, the more functional outputs we can expect. Thus, this study gives insight into how to create new associations of LAB strains and to guarantee their positive interactions.

Directed Recovery and Molecular Characterization of Antibiotic Resistance Plasmids from Cheese Bacteria.

Resistance to antimicrobials is a growing problem of worldwide concern. Plasmids are thought to be major drivers of antibiotic resistance spread. The present work reports a simple way to recover replicative plasmids conferring antibiotic resistance from the bacteria in cheese. Purified plasmid DNA from colonies grown in the presence of tetracycline and erythromycin was introduced into plasmid-free strains of Lactococcus lactis, Lactiplantibacillus plantarum and Lacticaseibacillus casei. Following antibiotic selection, the plasmids from resistant transformants were isolated, analyzed by restriction enzyme digestion, and sequenced. Seven patterns were obtained for the tetracycline-resistant colonies, five from L. lactis, and one each from the lactobacilli strains, as well as a single digestion profile for the erythromycin-resistant transformants obtained in L. lactis. Sequence analysis respectively identified tet(S) and ermB in the tetracycline- and erythromycin-resistance plasmids from L. lactis. No dedicated resistance genes were detected in plasmids conferring tetracycline resistance to L. casei and L. plantarum. The present results highlight the usefulness of the proposed methodology for isolating functional plasmids that confer antibiotic resistance to LAB species, widen our knowledge of antibiotic resistance in the bacteria that inhabit cheese, and emphasize the leading role of plasmids in the spread of resistance genes via the food chain.

Effect of Inoculated Lactic Acid Fermentation on the Fermentable Saccharides and Polyols, Polyphenols and Antioxidant Activity Changes in Wheat Sourdough.

Inoculation of sourdough allows the fermentation medium to be dominated by desired microorganisms, which enables determining the kinetics of the conversion of chemical compounds by individual microorganisms. This knowledge may allow the design of functional food products with health features dedicated to consumers with special needs. The aim of the study was to assess the dynamics of transformations of fermentable oligosaccharide, disaccharide, monosaccharide and polyol (FODMAP) compounds from wheat flour as well as their antioxidant activity during inoculated and spontaneous sourdough fermentation. The FODMAP content in grain products was determined by the fructan content with negligible amounts of sugars and polyols. To produce a low-FODMAP cereal product, the fermentation time is essential. The 72 h fermentation time of L. plantarum-inoculated sourdough reduced the FODMAP content by 91%. The sourdough fermentation time of at least 72 h also positively influenced the content of polyphenols and antioxidant activity, regardless of the type of fermentation. The inoculation of both L. plantarum and L. casei contributed to a similar degree to the reduction in FODMAP in sourdough compared to spontaneous fermentation.

Design of an autochthonous starter culture using strains isolated from traditional Matsoni.

Artisanal products support the conservation of the indigenous biodiversity of food microbiomes, although they do not always comply to quality and hygienic requirements for the dairy industry. This study describes the development of an autochthonous starter culture to produce Matsoni, a traditional Georgian fermented milk. To this end, strains of lactic acid bacteria isolated from artisanal Matsoni samples were used to design a starter formulation reproducing the dominant microbial diversity, also preserving quality characteristics and ensuring the safety of the product. As a result, strains that represent the acidifying portion of the starter (Lactobacillus delbrueckii subsp. lactis, L. delbrueckii subsp. bulgaricus and Streptococcus thermophilus) were combined in different ratios and strain combinations, together with cultures of Lactobacillus rhamnosus that were chosen for their potential beneficial traits. The strain association acting better in milk cultures at laboratory scale was selected as starter culture for the production of Matsoni in pilot-scale industrial trials.

Changes in physico-chemical characteristics and viable bacterial communities during fermentation of alfalfa silages inoculated with Lactobacillus plantarum.

This study investigated the effect of inoculating Lactobacillus (L.) plantarum PS-8 in fermentation of alfalfa silages. We monitored the fermentation characteristics and bacterial population dynamics during the ensiling process. PacBio single molecule real time sequencing was combined with propidium monoazide (PMA) treatment to monitor the viable microbiota dynamics. We found that inoculating L. plantarum PS-8 may improve the silage quality by accelerating acidification, reducing the amounts of clostridia, coliform bacteria, molds and yeasts, elevating the protein and organic acid contents (except butyrate), and enhancing lactic acid bacteria (LAB) while suppressing harmful microorganisms. Some significant differential abundant taxa were found between the PMA-treated and non-treated microbiota. For example, the relative abundances of L. brevis, L. plantarum, and Pediococcus pentosaceus were significantly higher in the PMA-treated group than the non-PMA-treated group, suggesting obvious differences between the viable and non-viable microbiota. It would thus be necessary to distinguish between the viable and non-viable microbial communities to further understand their physiological contribution in silage fermentation. By tracking the dynamics of viable microbiota in relation with changes in the physico-chemical parameters, our study provided novel insights into the beneficial effects of inoculating L. plantarum PS-8 in silage fermentation and the physiological function of the viable bacterial communities.

Effect of Echinostoma caproni on Presumptive Lactic Acid Bacteria Abundance and Salmonella enterica Serovar Typhimurium Colonization in the Mouse Gut.

Co-infections of mammalian hosts with intestinal helminths and bacterial pathogens are common, especially in areas with inadequate sanitation. Interactions between co-infecting species and host microbiota can cause significant changes in host immunity, disease severity, and pathogen transmission, requiring unique treatment for each case. A greater understanding of the influences of parasite-bacteria co-infections will improve diagnosis and therapeutic approaches to control infectious diseases. To study the influence of the trematode parasite Echinostoma caproni on commensal and pathogenic bacteria in the mouse gut, we examined the abundance of intestinal lactic acid bacteria and Salmonella enterica serovar Typhimurium in control mice not exposed to E. caproni (P-) or S. Typhimurium (S-), E. caproni-infected (P+S-), S. Typhimurium-infected (P-S+), and E. caproni-S. Typhimurium co-infected (P+S+) mice, and determined bacterial burdens in the livers and spleens of the P-S+ and P+S+ mice. We also examined a subset of P+S- and P+S+ mice for survival and the relative location of E. caproni in the small intestine. The numbers of presumptive lactic acid bacteria were significantly higher in the P+S+ and P-S+ mice compared to the uninfected mice, and S. Typhimurium colonization in the liver and spleen was significantly reduced in the P+S+ mice compared to the P-S+ mice. Echinostoma caproni were located anteriorly in the intestine of P+S- mice, while in the P+S+ mice, the parasites were distributed more posteriorly. Survival of E. caproni was unaffected in either group. The results of our study suggest that E. caproni facilitates a higher abundance of presumptive lactic acid bacteria in the mouse intestine and reduces colonization of S. Typhimurium in the liver and spleen of the co-infected host.

A refined medium to enhance the antimicrobial activity of postbiotic produced by Lactiplantibacillus plantarum RS5.

Postbiotic RS5, produced by Lactiplantibacillus plantarum RS5, has been identified as a promising alternative feed supplement for various livestock. This study aimed to lower the production cost by enhancing the antimicrobial activity of the postbiotic RS5 by improving the culture density of L. plantarum RS5 and reducing the cost of growth medium. A combination of conventional and statistical-based approaches (Fractional Factorial Design and Central Composite Design of Response Surface Methodology) was employed to develop a refined medium for the enhancement of the antimicrobial activity of postbiotic RS5. A refined medium containing 20 g/L of glucose, 27.84 g/L of yeast extract, 5.75 g/L of sodium acetate, 1.12 g/L of Tween 80 and 0.05 g/L of manganese sulphate enhanced the antimicrobial activity of postbiotic RS5 by 108%. The cost of the production medium was reduced by 85% as compared to the commercially available de Man, Rogosa and Sharpe medium that is typically used for Lactobacillus cultivation. Hence, the refined medium has made the postbiotic RS5 more feasible and cost-effective to be adopted as a feed supplement for various livestock industries.

Selection of Riboflavin Overproducing Strains of Lactic Acid Bacteria and Riboflavin Direct Quantification by Fluorescence.

Riboflavin (vitamin B2) is a vitamin of the B group involved in essential biological pathways, including redox reactions and the electron transport chain. Some lactic acid bacteria (LAB) can synthesize riboflavin and this capability is strain-dependent. In the last years, a growing interest has focused on the selection of riboflavin-overproducing food-grade LAB for the vitamin biofortification of fermented foods, as well as for the formulation of innovative functional products.In this chapter we report fast and inexpensive techniques in order to (1) screen LAB isolates able to produce riboflavin from different matrices, (2) select spontaneous roseoflavin-resistant riboflavin overproducing strains, and (3) quantify vitamin B2 in culture media by fluorescence detection.These protocols could be useful to select new overproducing strains and/or species from different ecological niches, as well as to optimize the conditions for vitamin bioproduction.

High ZnPP-forming food-grade lactic acid bacteria as a potential substitute for nitrite/nitrate to improve the color of meat products.

Zinc protoporphyrin IX (ZnPP)-forming food-grade lactic acid bacteria (LAB) were screened from various sources for their ability to improve the color of meat products. The effects of salt and nitrite on the ZnPP-forming ability of these bacteria were also investigated. Finally, these bacteria were applied in salt-added minced meat to assess their ability to improve the color. Twenty-five LAB were screened for their ZnPP-forming ability in pork. Most of the strains exhibited maximum growth anaerobically in 3% salt at 30 °C and grew well at pH 5.5 and 6.5. Moreover, 3% salt slightly retarded ZnPP formation; however, nitrite completely inhibited ZnPP formation in all the ZnPP-forming LAB. Thirteen LAB (avoiding duplication and non-food-grade) could form ZnPP in salt-added minced meat, resulting in improvement of the bright red color, high ZnPP autofluorescence, and increased fluorescence intensity. Finally, considering the safety, Lactobacillus plantarum, Lactococcus lactis subsp. cremoris, and Leuconostoc lactis were suggested as promising candidates to improve the color of meat products.

In vitro characterization of chicken gut bacterial isolates for probiotic potentials.

Probiotics often play an important role in improving gut health in chickens through multiple mechanisms, including enhancement of tight junctions, nutrient acquisition, niche colonization, or coaggregation with enteric pathogens. The objective of this study was to characterize lactic acid bacteria (LAB) isolated from the gut of healthy broiler chickens for a number of phenotypes that might be indicative of good probiotic potentials. A total 40 bacterial isolates were isolated from 3-week-old chickens using Man, Rogosa and Sharpe (MRS) agar plates. The bacterial isolates were evaluated in vitro for motility, autoaggregation, pathogen inhibition, pH of overnight culture, growth on different agar plates, and their impact on gut integrity. Selected isolates were genotyped by sequencing the 16S-23S rRNA gene intergenic region. Based on the phenotype and genotype, we identified 20 potential probiotic (PP) isolates that belong to LAB. Multivariate analysis showed that PP isolates were positively correlated with parameters such as growth on MRS agar plate (pH 5.5), pathogen inhibition, and autoaggregation. However, growth on MacConkey agar plates, supernatant pH, motility, and transepithelial electrical resistance were negatively correlated with the PP isolates. Furthermore, in vivo study needs to be performed for evaluation of the utility of these probiotic candidates in poultry production.

A dynamic resazurin microassay allowing accurate quantification of cells and suitable for acid-forming bacteria.

A resazurin micro-assay was developed to quantify acidifying bacteria. The resorufin fluorescent signal was measured over time and the determined time to reach the max slope (TMS) was plotted against CFU (colony forming unit) counts. This dynamic assay enabled to quantify nine lactic acid bacteria and a Bacillus licheniformis strain despite the increasing acidity of the medium.

Genetic algorithm applied to simultaneous parameter estimation in bacterial growth.

Several mathematical models have been developed to understand the interactions of microorganisms in foods and predict their growth. The resulting model equations for the growth of interacting cells include several parameters that must be determined for the specific conditions to be modeled. In this study, these parameters were determined by using inverse engineering and a multi-objective optimization procedure that allows fitting more than one experimental growth curve simultaneously. A genetic algorithm was applied to obtain the best parameter values of a model that permit the construction of the front of Pareto with 50 individuals or phenotypes. The method was applied to three experimental data sets of simultaneous growth of lactic acid bacteria (LAB) and Listeria monocytogenes (LM). Then, the proposed method was compared with a conventional mono-objective sequential fit. We concluded that the multi-objective fit by the genetic algorithm gives superior results with more parameter identifiability than the conventional sequential approach.

Beetroot and radish powders as natural nitrite source for fermented dry sausages.

The aim of this study was to investigate the use of radish and beetroot powders as potential substitutes of nitrite in fermented dry sausages due to their high nitrate content (around 16,000 and 14,000 mg/kg, respectively). Six treatments were prepared and evaluated during the ripening process and storage time: C1 (control with 150 mg/kg sodium nitrite and 150 mg/kg sodium nitrate), C2 (control without sodium nitrite/nitrate), R05 (0.5% radish powder), R1 (1% radish powder), B05 (0.5% beetroot powder) and B1 (1% beetroot powder). The addition of vegetable powders influenced moisture content, weight loss and water activity of sausages. Nitrite was formed from radish and beetroot powders during the ripening process, especially in R1 and B1 treatments. Beetroot powder affected colour, pigments and lactic acid bacteria counts. The results of pH, colour, lipid oxidation, nitrite and nitrate analysis suggest R1 treatment as a potential nitrite replacer obtained from a simple and feasible drying process.

Effects of lactic acid bacteria fermented yellow whey on the protein coagulation and isoflavones distribution in soymilk.

This study investigated the soymilk coagulation induced by fermented yellow whey (FYW), which is extensively used as a natural tofu coagulant in China. The aggregations involving proteins and isoflavone particles caused by FYW were analyzed using the proteomic technology and high-performance liquid chromatography, respectively. As indicated, the FYW-induced coagulation of soy proteins mainly occurred at pH 5.80-5.90. When the pH of soymilk decreased, the 7S ß, 11S A3 and some of 11S A1a subunits and SBP, Bd, lectin and TA aggregated the earliest, and later did the 11S A4, other 11S A1a, 11S A2 and 11S A1b subunits. The 7S α and α' subunits and TB showed an obvious delay in aggregation. Moreover, isoflavones in the form of aglycones were more likely to coprecipitate with proteins, compared with glycosides. These results could provide an important reference and assistance for future research on the development of traditional FYW-tofu.

Resource allocation explains lactic acid production in mixed-culture anaerobic fermentations.

Lactate production in anaerobic carbohydrate fermentations with mixed cultures of microorganisms is generally observed only in very specific conditions: the reactor should be run discontinuously and peptides and B vitamins must be present in the culture medium as lactic acid bacteria (LAB) are typically auxotrophic for amino acids. State-of-the-art anaerobic fermentation models assume that microorganisms optimise the adenosine triphosphate (ATP) yield on substrate and therefore they do not predict the less ATP efficient lactate production, which limits their application for designing lactate production in mixed-culture fermentations. In this study, a metabolic model taking into account cellular resource allocation and limitation is proposed to predict and analyse under which conditions lactate production from glucose can be beneficial for microorganisms. The model uses a flux balances analysis approach incorporating additional constraints from the resource allocation theory and simulates glucose fermentation in a continuous reactor. This approach predicts lactate production is predicted at high dilution rates, provided that amino acids are in the culture medium. In minimal medium and lower dilution rates, mostly butyrate and no lactate is predicted. Auxotrophy for amino acids of LAB is identified to provide a competitive advantage in rich media because less resources need to be allocated for anabolic machinery and higher specific growth rates can be achieved. The Matlab™ codes required for performing the simulations presented in this study are available at https://doi.org/10.5281/zenodo.4031144.

Dynamic co-culture metabolic models reveal the fermentation dynamics, metabolic capacities and interplays of cheese starter cultures.

In this study, we have investigated the cheese starter culture as a microbial community through a question: can the metabolic behaviour of a co-culture be explained by the characterized individual organism that constituted the co-culture? To address this question, the dairy-origin lactic acid bacteria Lactococcus lactis subsp. cremoris, Lactococcus lactis subsp. lactis, Streptococcus thermophilus and Leuconostoc mesenteroides, commonly used in cheese starter cultures, were grown in pure and four different co-cultures. We used a dynamic metabolic modelling approach based on the integration of the genome-scale metabolic networks of the involved organisms to simulate the co-cultures. The strain-specific kinetic parameters of dynamic models were estimated using the pure culture experiments and they were subsequently applied to co-culture models. Biomass, carbon source, lactic acid and most of the amino acid concentration profiles simulated by the co-culture models fit closely to the experimental results and the co-culture models explained the mechanisms behind the dynamic microbial abundance. We then applied the co-culture models to estimate further information on the co-cultures that could not be obtained by the experimental method used. This includes estimation of the profile of various metabolites in the co-culture medium such as flavour compounds produced and the individual organism level metabolic exchange flux profiles, which revealed the potential metabolic interactions between organisms in the co-cultures.