Bacterial proteome adaptation during fermentation in dairy environments.

The enzymatic repertoire of starter cultures belonging to the Lactococcus genus determines various important characteristics of fermented dairy products but might change in response to the substantial environmental changes in the manufacturing process. Assessing bacterial proteome adaptation in dairy and other food environments is challenging due to the high matrix-protein concentration and is even further complicated in particularly cheese by the high fat concentrations, the semi-solid state of that matrix, and the non-growing state of the bacteria. Here, we present bacterial harvesting and processing procedures that enable reproducible, high-resolution proteome determination in lactococcal cultures harvested from laboratory media, milk, and miniature Gouda cheese. Comparative proteome analysis of Lactococcus cremoris NCDO712 grown in laboratory medium and milk revealed proteome adaptations that predominantly reflect the differential (micro-)nutrient availability in these two environments. Additionally, the drastic environmental changes during cheese manufacturing only elicited subtle changes in the L. cremoris NCDO712 proteome, including modified expression levels of enzymes involved in flavour formation. The technical advances we describe offer novel opportunities to evaluate bacterial proteomes in relation to their performance in complex, protein- and/or fat-rich food matrices and highlight the potential of steering starter culture performance by preculture condition adjustments.

Phenotypic, Technological, Safety, and Genomic Profiles of Gamma-Aminobutyric Acid-Producing Lactococcus lactis and Streptococcus thermophilus Strains Isolated from Cow's Milk.

Gamma-aminobutyric acid (GABA)-producing lactic acid bacteria (LAB) can be used as starters in the development of GABA-enriched functional fermented foods. In this work, four GABA-producing strains each of Lactococcus lactis and Streptococcus thermophilus species were isolated from cow's milk, and their phenotypic, technological, and safety profiles determined. Genome analysis provided genetic support for the majority of the analyzed traits, namely, GABA production, growth in milk, and the absence of genes of concern. The operon harboring the glutamate decarboxylase gene (gadB) was chromosomally encoded in all strains and showed the same gene content and gene order as those reported, respectively, for L. lactis and S. thermophilus. In the latter species, the operon was flanked (as in most strains of this species) by complete or truncated copies of insertion sequences (IS), suggesting recent acquisition through horizontal gene transfer. The genomes of three L. lactis and two S. thermophilus strains showed a gene encoding a caseinolytic proteinase (PrtP in L. lactis and PrtS in S. thermophilus). Of these, all but one grew in milk, forming a coagulum of good appearance and an appealing acidic flavor and taste. They also produced GABA in milk supplemented with monosodium glutamate. Two L. lactis strains were identified as belonging to the biovar. diacetylactis, utilized citrate from milk, and produced significant amounts of acetoin. None of the strains showed any noticeable antibiotic resistance, nor did their genomes harbor transferable antibiotic resistance genes or genes involved in toxicity, virulence, or pathogenicity. Altogether these results suggest that all eight strains may be considered candidates for use as starters or components of mixed LAB cultures for the manufacture of GABA-enriched fermented dairy products.

Exploring Technological, Safety and Probiotic Properties of Enterococcus Strains: Impact on Rheological Parameters in Fermented Milk.

Enterococci, known for their resilience, are commonly found in food, the environment, and the gastrointestinal tracts of humans and animals. In recent research, six strains of enterococcus were isolated from bat guano. These include Enterococcus mundtii SRBG1, Enterococcus gallinarum SRBG3, Enterococcus faecium SRBG2, Enterococcus casseliflavus EC1, and Enterococcus devriesei CAU 1344. Identification was done using 16S DNA analysis. Each strain underwent evaluation for its technological properties (such as tolerances to various NaCl concentrations and temperatures, as well as amylolytic, ß-galactosidase, lipolytic, and proteolytic activities, and EPS production) and selected probiotic properties (including safety profile, resistance to 0.3 percent bile salts and gastric juice with a pH of 2.5, lysozyme tolerance, and antibacterial and antibiofilm activities against four foodborne pathogens). The results were analyzed using Principal Component Analysis. This analysis revealed that E. mundtii SRBG1 and E. gallinarum SRBG3, followed by E. faecium SRBG2, were most closely associated with a broad range of technological characteristics and were subsequently used for fermenting skimmed milk. The rheological properties of the samples indicated a shear-thinning or non-Newtonian behavior. Furthermore, during storage of the fermented milk at 4 °C over periods of 1, 7, 14, and 21 days, there were no significant changes in bacterial count (at around 7 log10 CFU/mL) and pH when fermented with the three evaluated strains.

Molecular docking studies on α-amylase inhibitory peptides from milk of different farm animals.

Milk-derived peptides have emerged as a popular mean to manage various lifestyle disorders such as diabetes. Fermentation is being explored as one of the faster and efficient way of producing peptides with antidiabetic potential. Therefore, in this study, an attempt was made to comparatively investigate the pancreatic α-amylase (PAA) inhibitory properties of peptides derived from milk of different farm animals through probiotic fermentation. Peptide's identification was carried out using liquid chromatography-quadrupole time-of-flight mass spectrometry and inhibition mechanisms were characterized by molecular docking. Results obtained showed a PAA-IC50 value (the amount of protein equivalent needed to inhibit 50% of enzymes) between 2.39 and 36.1 µg protein equivalent for different fermented samples. Overall, Pediococcus pentosaceus MF000957-derived fermented milk from all animals indicated higher PAA inhibition than other probiotic derived fermented milk (PAA-IC50 values of 6.01, 3.53, 15.6, and 10.8 µg protein equivalent for bovine, camel, goat, and sheep fermented milk). Further, molecular docking analysis indicated that camel milk-derived peptide IMEQQQTEDEQQDK and goat milk-derived peptide DQHQKAMKPWTQPK were the most potent PAA inhibitory peptides. Overall, the study concluded that fermentation derived peptides may prove useful in for managing diabetes via inhibition of carbohydrate digesting enzyme PAA.

Purine nucleosidase (PNase) activity, probiotics potential, and food applicability of a newly-isolated Levilactobacillus brevis LAB42.

Hyperuricemia, a condition characterized by elevated levels of uric acid in the blood, is known as a risk factor for gout disease. In this study, we isolated a total of 72 MRS-grown colonies and evaluated their purine nucleosidase (PNase) activity. Among the isolated bacteria, Levilactobacillus (L.) brevis LAB42 displayed the highest PNase activity. Our findings also indicate that PNase activity can vary among lactic acid bacterial strains and during different growth phases. Based on the kinetics study, LAB42 consistently exhibits the highest PNase activity. Due to its ability to attach to Caco-2 cells and its resistance to acidic environments and bile exposure, L. brevis LAB42 was chosen for further studies and showed that with the right combination of additives, it has the potential to be an appropriate starter for milk fermentation.