Lactococcus Cremoris Research Articles

High-throughput ecological interaction mapping of dairy microorganisms

To engineer efficient microbial management strategies in the food industry, a comprehensive understanding of microbial interactions is crucial. Microorganisms live in communities where they influence each other in several ways. Although much attention has been paid to the production of antagonistic metabolites in lactic acid bacteria (LAB), research that accounts for the complexity of their ecological interactions and their dynamics remains limited. This study explores binary interactions within a mock community of 94 strains, including 23 LAB from culture collections and 71 isolated from dairy products. Using a colony-picking robot and image analysis, bidirectional interactions were measured at high throughput on solid media, where one test strain was challenged against other mock community members as the target strains. Assays of 15 test strains (14 LAB and one yeast) yielded 1,142 bidirectionally mapped interactions, classified by ecological type over seven days. The results showed variation in the strength, directionality, and type of interactions depending on the origin of the target strains. Cooperation rates increased for strains isolated from raw milk to pasteurized milk and cheese, while exploitation was more common in raw milk strains. Cooperating strains also exhibited more similar ecological behaviors than competing strains, suggesting the presence of microbial cliques. Interestingly, Lactococcus cremoris ATCC 19257 developed pink-red pigmentation when co-cultured with Pseudomonas aeruginosa. Overall, these findings present an unprecedented exploratory data set that significantly improves our understanding of microbial interactions at the system level, with potential applications in strain selection for food processes such as fermentation, bioprotection, and probiotics.

Impact of a teat disinfectant based on Lactococcus cremoris on the cow milk proteome

BackgroundDairy cow milking practices require cleaning and disinfection of the teat skin before and after milking to ensure the safety and quality of milk and prevent intramammary infections. Antimicrobial proteins of natural origin can be valuable alternatives to traditional disinfectants. In a recent field trial, we demonstrated that a teat dip based on a nisin A-producing Lactococcus cremoris (L) had comparable efficacy to conventional iodophor dip (C) in preventing dairy cow mastitis. Here, we present the differential shotgun proteomics investigation of the milk collected during the trial.MethodsFour groups of quarter milk samples with low (LSCC) and high somatic cell count (HSCC) collected at the beginning (T0) and end (TF) of the trial were analyzed for a total of 28 LSCC (14 LSCC T0 and 14 LSCC TF) and 12 HSCC (6 HSCC T0 and 6 HSCC TF) samples. Milk proteins were digested into peptides, separated by nanoHPLC, and analyzed by tandem mass spectrometry (LC-MS/MS) on an Orbitrap Fusion Tribrid mass spectrometer. The proteins were identified with MaxQuant and interaction networks of the differential proteins were investigated with STRING. The proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD045030.ResultsIn healthy milk (LSCC), we detected 90 and 80 differential proteins at T0 and TF, respectively. At TF, the Lactococcus group showed higher levels of antimicrobial proteins. In mastitis milk (HSCC), we detected 88 and 106 differential proteins at T0 and TF, respectively. In the Lactococcus group, 14 proteins with antimicrobial and immune defense functions were enriched at TF vs. 4 proteins at T0. Cathelicidins were among the most relevant enriched proteins. Western immunoblotting validation confirmed the differential abundance.ConclusionsAt T0, the proteomic differences observed in healthy milk of the two groups were most likely dependent on physiological variation. On the other hand, antimicrobial and immune defense functions were higher in the milk of cows with mammary gland inflammation of the Lactococcus-treated group. Among other factors, the immunostimulatory action of nisin A might be considered as a contributor.

Assessment of the Biological Activity of a Probiotic Fermented Milk Product with the Addition of Lactobacillus helveticus Cell-Free Supernatant

Products enriched with probiotic microorganisms have proven to possess immunomodulatory, antioxidant, hypo-cholesterolemic, hypotensive, and antimicrobial properties. Biologically active substances, which are by-products of microbial fermentation, have potential applications in various industries. Cell-free supernatants, depending on the microorganisms used and production conditions, can exhibit antimicrobial, antioxidant, bifidogenic, and other biological activities. This paper presents a study on the biological activity of a probiotic fermented milk product, supplemented with 0.01% lyophilized cell-free supernatant from Lactobacillus helveticus. The fermented milk product was developed based on a composition of Lacticaseibacillus rhamnosus F, Lactococcus cremoris CR201, and Propionibacterium shermanii E2. The research evaluated antimicrobial activity, Bifidobacteria growth stimulation, and the content of organic acids, amino acids, and B vitamins. It was found that adding lyophilized cell-free supernatant to the fermented milk product enhanced its biological activity. In particular, the experimental samples showed a threefold increase in vitamin B6 content compared to the control, reaching 22.412 μg/100 g. Additionally, the amino acid content in the experimental samples exhibited a significant increase of more than 100% in the essential amino acid tryptophan compared to the control. Notably, antimicrobial activity increased against several opportunistic strains. The experimental samples also showed a significant increase in lactic and formic acids, which may enhance the product’s inhibitory properties against pathogens. An increase in antioxidant activity was observed, potentially due to the higher content of tryptophan and vitamin B6. The positive effect of adding cell-free supernatant on the growth of Bifidobacteria was also demonstrated. Thus, the findings suggest that this cell-free supernatant can be recommended as an additive in the production of fermented milk products, food additives, dietary supplements, and animal feed.

Assessment of Lactococcus Cremoris preparations for the pre- and post-milking teat disinfection

Good hygienic milking practices including the disinfection of the cow teat skin before and after milking aimed at preventing the occurrence of new intramammary infections (IMI) in dairy cows. This practice is generally performed using disinfectants, and in the current context of attention to the emergence of resistances, it is of greatest interest to evaluate alternative solutions that can expand treatment options. We assessed the efficacy of a pre-dipping and a post-dipping product based on the bacteriocin-containing culture of Lactococcus cremoris strain FT27, as compared to commercial disinfectants. FT27 was selected for the bactericidal activity in vitro against food pathogens. In the present study, it also revealed a high bactericidal activity against the main mastitis pathogens, most likely related to Nisin A production, according to genomic and proteomic analysis. The lactococcus-based preparations were applied in two commercial dairy farms in Northern Italy. Overall, 298 dairy cows were enrolled, 141 in the treated group (TR), and 157 in the control group (CTR). The cows were sampled at quarter level every two weeks for 3 months. During the trial, all cows showed a good health status. The hygiene level for udder, flanks and legs was generally good (on average < 3 score); the udder hygiene was significantly worse (P < 0.01) in the CTR group. The teat apex hyperkeratosis was overall low (on average < 2 score). We recorded no significant differences between the two experimental groups in the Somatic Cell Count (SCC) values and the bacteriological results. The overall frequency of new IMI was low, ranging 0.6 − 0.5% for S. aureus in the TR or CTR group respectively, to 2.6–4.4% for NASM. Regarding Str. spp., the new IMI accounted for 1.7% or 1.9% in the TR or CTR group, respectively. Notably, the incidence of S. aureus new IMI did not increase during the study, even though neither of the two herds segregated the positive cows. The non-inferiority test showed that the Lactococcus-based pre- and post-dipping products had an efficacy comparable to proven commercial disinfectants in maintaining udder health and preventing new IMI, thus representing a possible alternative to current teat dip products.

Novel cell wall polysaccharide genotypes and structures of lactococcal strains isolated from milk and fermented foods

The biosynthetic machinery for cell wall polysaccharide (CWPS) formation in Lactococcus lactis and Lactococcus cremoris is encoded by the cwps locus. The CWPS of lactococci typically consists of a neutral rhamnan component, which is embedded in the peptidoglycan, and to which a surface-exposed side chain oligosaccharide or polysaccharide pellicle (PSP) component is attached. The rhamnan component has been shown for several lactococcal strains to consist of a repeating rhamnose trisaccharide subunit, while the side chain is diverse in glycan content, polymeric status and glycosidic linkage architecture. The observed structural diversity of the CWPS side chain among lactococcal strains is reflected in the genetic diversity within the variable 3′ region of the corresponding cwps loci. To date, four distinct cwps genotypes (A, B, C, D) have been identified, while eight subtypes (C1 through to C8) have been recognized among C-genotype strains. In the present study, we report the identification of three novel subtypes of the lactococcal cwps C genotypes, named C9, C10 and C11. The CWPS of four isolates representing C7, C9, C10 and C11 genotypes were analysed using 2D NMR to reveal their unique CWPS structures. Through this analysis, the structure of one novel rhamnan, three distinct PSPs and three exopolysaccharides were elucidated. Results obtained in this study provide further insights into the complex nature and fascinating diversity of lactococcal CWPSs. This highlights the need for a holistic view of cell wall-associated glycan structures which may contribute to robustness of certain strains against infecting bacteriophages. This has clear implications for the fermented food industry that relies on the consistent application of lactococcal strains in mesophilic production systems.

Effect of freezing temperature on rate of survival of lactococcus cultures during lyophilization

The important component in the activities of the collection of starter microorganisms is the preservation of the pool of the strains valuable for industrial production. Lyophilization (also known as freeze drying) is a widely used method for the long-term storage of the collection cultures. The critical parameter of the process of lyophilization is the pre-freezing temperature. This article brings forward the results of researches of the rate of survival of lactococcus strain cultures during their lyophilization with pre-freezing at the temperatures of minus 20 °C, minus 30 °C and minus 55 °C. Reconstituted 10% skim milk was used as the medium for strain culturing and lyophilization. Different reactions were detected among the microorganisms of Lactococcus lactis subsp. lactis, Lactococcus cremoris and Lactococcus lactis subsp. lactis biovar. diacetylactis for their freezing and lyophilization under the tested conditions. The highest rate of survival of lactococcus strains was observed at the freezing temperature of minus 20 °C. The most resistant cultures appeared to be Lactococcus lactis subsp. lactis biovar. diacetylactis — their rate of survival after lyophilization at minus 20 °C amounted to 42.6-57.9%. The lowest survival rate was recorded for Lactococcus cremoris in all freezing modes (from 3.1 to 15.7%). Lyophilization of lactococcus collection cultures ensures high survival rate of the cells.

Formation of cheese pattern when using monospecies cultures

The article presents the results of a study of the influence of monospecies gas-aroma-forming cultures Lactococcus lactis subsp. lactis biovar. diacetylactis (L. diacetylactis) and Leuconostoc subsp. on the peculiarities of pattern formation in cheeses with a low second heating temperature molded from a layer. The studied cultures were used at a dose of 0.6% of the total milk volume as as single starter microflora in the model cheeses (1-M and 2-M) and additional microflora along with the main lactococcal microflora (Lactococcus lactis subsp. lactis; Lactococcus cremoris) in the control cheeses (1-K and 2-K). The model and control cheeses were subjected to microbiological tests (total number of lactic acid microorganisms, number of L. diacetylactis and Leuconostoc subsp.), physicochemical (mass fraction of moisture, pH), biochemical (mass fraction of lactose) and organoleptic studies after pressing and during ripening at the age of 15, 30, 45, 60 days. It has been established that the use of a culture of citrate-fermenting lactococci L. diacetylactis ensures a stable lactic acid fermentation process during the production and ripening of the model and control cheeses, the formation of an elastic consistency and the desired pattern with eyes of a regular round shape. The use of the hetero-fermentative culture Leuconostoc subsp., as single starter microflora, does not guarantee the required level of lactic acid fermentation during cheese production and leads to an increase in the moisture content of the cheese mass after pressing and overacidification of the cheese in the first stages of ripening, which together contributes to the formation of an overly plastic consistency and an overdeveloped pattern in the form of cracks. The use of Leuconostoc subsp. as a gas-aroma-forming component of a traditional starter culture for semi-hard cheeses, also consisting of mesophilic lactococci Lc. lactis subsp. lactis and Lc. cremoris, causes the development of a nest-like pattern, which does not fully ensure the formation of the desired pattern with regular rounded eyes.

Microbiome Taxonomic and Functional Differences in C3H/HeJ Mice Fed a Long-Term High-Fat Diet with Beef Protein ± Ammonium Hydroxide Supplementation.

Studies have suggested that alkalinized foods may reduce the effects of the acidogenic Western diet in promoting obesity, metabolic syndrome, type 2 diabetes, cancer, and coronary heart disease. Indeed, a recent study in mice fed a high-fat diet containing dietary beef supplemented with ammonium hydroxide showed improvement in a suite of metabolic outcomes. However, the effects of dietary protein ammonium supplementation on the microbiome remain unknown. In this study, the effects of ammonium supplementation on beef protein towards microbiome taxa and function in a high-fat diet were analyzed. Fecal microbiomes were characterized using a shotgun metagenomic approach for 16-month-old male and female mice after long-term diet treatments. The results for ammoniated diets showed that several bacteria known to be associated with health benefits increased significantly, including Romboutsia, Oscillospiraceae, and Lactococcus cremoris. The beneficial mucin-degrader Akkermansia was especially abundant, with a high prevalence (~86%) in females. Concurrently, the phyla Actinomycetota (Actinobacteria) and Bacteroidota (Bacteroidetes) were significantly reduced. While sex was a confounding factor affecting microbiome responses to ammonium supplementation in dietary protein, it is worth noting that several putatively beneficial microbiome functions increased with ammonium supplementation, such as glycine betaine transport, xenobiotic detoxification, enhanced defense, and others. Conversely, many disease-associated microbiome functions reduced. Importantly, modifying protein pH alone via ammonium supplementation induced beneficial microbiota changes. Taken together, these results suggest that ammonium-supplemented proteins may mediate some negative microbiome-associated effects of high-fat/Western diets.

Metagenomic profiles of the antimicrobial resistance in traditional Chinese fermented meat products: Core resistome and co-occurrence patterns

Antimicrobial resistance (AMR) poses a significant challenge to global health, and the presence of antibiotic resistance genes (ARGs) in food poses a potential threat to public health. Traditional Chinese fermented meat products (FMPs) are highly favored because of their unique flavors and cultural value. However, microbial safety and the potential distribution and composition of AMR in these products remain unclear. In this study, a comprehensive analysis of bacterial composition and antibiotic-resistant populations in 216 samples of traditional fermented meat products from different regions of China was conducted using a metagenomic approach. Staphylococcus was the most abundant genus in the samples, accounting for an average abundance of 29.9 %, followed by Tetragenococcus (17.1 %), and Latilactobacillus (3.6 %). A core resistome of FMP samples was constructed for the first time using co-occurrence network analysis, which revealed the distribution and interrelationships of ARGs and bio/metal-resistant genes (BMRGs). Random forest analysis identified the lincosamide nucleotidyltransferase lnuA and the multidrug and toxic compound extrusion (MATE) transporter abeM as potential indicators for assessing the overall abundance of the core resistome. Additionally, Staphylococcus, Acinetobacter, and Pseudomonas were identified as hosts constituting the core resistome. Despite their low abundance, the latter two still serve as major reservoirs of antibiotic resistance genes. Notably, Lactococcus cremoris was identified as the key host for tetracycline resistance genes in the samples, highlighting the need for enhanced resistance monitoring in lactic acid bacteria. Based on our findings, in the microbial safety assessment of fermented meat products, beyond common foodborne pathogens, attention should be focused on detecting and controlling coagulase-negative Staphylococcus, Acinetobacter, and Pseudomonas, and addressing bacterial resistance. The quantitative detection of lnuA and abeM could provide a convenient and rapid method for assessing the overall abundance of the core resistome. Our findings have important implications for the control of bacterial resistance and prevention of pathogenic bacteria in fermented meat products.

Technological and Enzymatic Characterization of Autochthonous Lactic Acid Bacteria Isolated from Viili Natural Starters.

Viili, a Finnish ropy fermented milk, is traditionally manufactured through spontaneous fermentation, by mesophilic lactic acid bacteria and yeast-like fungi, or back-slopping. This study evaluated four natural viili starters as sources of lactic acid bacteria for dairy production. Back-slopping activation of the studied viili samples was monitored through pH and titratable acidity measurements and enumeration of mesophilic lactic acid bacteria. Sixty lactic acid bacteria isolates were collected, molecularly identified, and assayed for acidification performance, enzymatic activities, production of exopolysaccharides (EPSs), presence of the histidine decarboxylase (hdcA) gene of Gram-positive bacteria, and production of bacteriocins. A neat predominance of Lactococcus lactis emerged among the isolates, followed by Enterococcus faecalis, Enterococcus faecium, Enterococcus durans, Enterococcus lactis, and Lactococcus cremoris. Most isolates exhibited proteolytic activity, whereas only a few enterococci showed lipase activity. Five isolates identified as L. cremoris, L. lactis, and E. faecalis showed a good acidification performance. Most of the isolates tested positive for leucine arylamidase, whereas only one E. durans and two L. lactis isolates were positive for valine arylamidase. A few isolates also showed a positive reaction for beta-galactosidase and alpha- and beta-glucosidase. None of the isolates produced EPSs or bacteriocins. The hdcA gene was detected in five isolates identified as L. lactis and E. faecium. A few L. cremoris and L. lactis isolates for potential use as starter or adjunct cultures for dairy processing were finally identified.

Characterization of a Panel of Constitutive Promoters from Lactococcus cremoris for Fine-Tuning Gene Expression.

Lactococcus cremoris (homotypic synonym: Lactococcus lactis) is receiving increasing attention as a prominent vehicle for the delivery of live vaccines. This can hardly be achieved without developing tools for the genetic manipulation of L. cremoris, and the paucity of studies on L. cremoris endogenous promoters has attracted our attention. Here, we report the discovery and characterization of 29 candidate promoters identified from L. cremoris subsp. cremoris NZ9000 by RNA sequencing analysis. Furthermore, 18 possible constitutive promoters were obtained by RT-qPCR screening from these 29 candidate promoters. Then, these 18 promoters were cloned and characterized by a reporter gene, gusA, encoding β-glucuronidase. Eventually, eight endogenous constitutive promoters of L. cremoris were obtained, which can be applied to genetic manipulation of lactic acid bacteria.

Lactococcus cell envelope proteases enable lactococcal growth in minimal growth media supplemented with high molecular weight proteins of plant and animal origin.

Lactic acid bacteria (LAB) have evolved into fastidious microorganisms that require amino acids from environmental sources. Some LAB have cell envelope proteases (CEPs) that drive the proteolysis of high molecular weight proteins like casein in milk. CEP activity is typically studied using casein as the predominant substrate, even though CEPs can hydrolyze other protein sources. Plant protein hydrolysis by LAB has rarely been connected to the activity of specific CEPs. This study aims to show the activity of individual CEPs using LAB growth in a minimal growth medium supplemented with high molecular weight casein or potato proteins. Using Lactococcus cremoris MG1363 as isogenic background to express CEPs, we demonstrate that CEP activity is directly related to growth in the protein-supplemented minimal growth media. Proteolysis is analyzed based on the amino acid release, allowing a comparison of CEP activities and analysis of amino acid utilization by L. cremoris MG1363. This approach provides a basis to analyze CEP activity on plant-based protein substrates as casein alternatives and to compare activity of CEP homologs.

Heterologous expression and purification of the phage lysin-like bacteriocin LysL from Lactococcus lactis LAC460.

The wild-type Lactococcus lactis strain LAC460 produces two bacteriocin-like phage lysins, LysL and LysP. This study aimed to produce and secrete LysL in various heterologous hosts and an in vitro cell-free expression system for further functional studies. Initially, the lysL gene from L. lactis LAC460 was cloned into Lactococcus cremoris NZ9000 and L. lactis N8 strains, with and without the usp45 signal sequence (SSusp45), under a nisin-inducible promoter. Active LysL was primarily produced intracellularly in recombinant L. lactis N8, with some secretion into the supernatant. Recombinant L. cremoris NZ9000 lysed upon nisin induction, indicating successful lysL expression. However, fusion with Usp45 signal peptide (SPUsp45-LysL) weakened LysL activity, likely due to incomplete signal peptide cleavage during secretion. Active LysL was also produced in vitro, and analysed in SDS-PAGE, giving a 42-kDa band. However, the yield of LysL protein was still low when produced from recombinant lactococci or by in vitro expression system. Therefore, His-tagged LysL was produced in Escherichia coli BL21(DE3). Western blot confirmed the intracellular production of about 44-kDa His-tagged LysL in E. coli. His-tagged active LysL was then purified by Ni-NTA affinity chromatography yielding sufficient 4.34mg of protein to be used in future functional studies.

The microbial and metabolite composition of Gouda cheese made from pasteurized milk is determined by the processing chain

Gouda cheeses of different production batches and ripening times often differ in metabolite composition, which may be due to the starter culture mixture applied or the growth of non-starter lactic acid bacteria (NSLAB) upon maturation. Therefore, a single Gouda cheese production batch was systematically investigated from the thermized milk to the mature cheeses, ripened for up to 100 weeks, to identify the main bacterial species and metabolites and their dynamics during the whole production and ripening. As this seemed to be starter culture strain- and NSLAB-dependent, it requested a detailed, longitudinal, and quantitative investigation. Hereto, microbial colony enumeration, high-throughput full-length 16S rRNA gene sequencing, and a metabolomic approach were combined. Culture-dependently, Lactococcus lactis was the most abundant species from its addition as part of the starter culture up to the first two months of cheese ripening. Afterward, the NSLAB Lacticaseibacillus paracasei became the main species during ripening. The milk was a possible inoculation source for the latter species, despite pasteurization. Culture-independently, the starter LAB Lactococcus cremoris and Lc. lactis were the most abundant species in the cheese core throughout the whole fermentation and ripening phases up to 100 weeks. The cheese rind from 40 until 100 weeks of ripening was characterized by a high relative abundance of the NSLAB Tetragenococcus halophilus and Loigolactobacillus rennini, which both came from the brine. These species were linked with the production of the biogenic amines cadaverine and putrescine. The most abundant volatile organic compound was acetoin, an indicator of citrate and lactose fermentation during the production day, whereas the concentrations of free amino acids were an indicator of the ripening time.

Lactococcus cremoris D2022 alleviates hyperuricemia and suppresses renal inflammation via potential gut-kidney axis.

Hyperuricemia (HUA) is a widespread metabolic disorder. Probiotics have drawn increasing attention as an adjunctive treatment with fewer side effects. However, thus far the effective strains are limited and the mechanisms for their serum uric acid (SUA)-lowering effect are not well understood. Along this line, we conducted the current study using a hyperuricemia mouse model induced by potassium oxonate and adenine. A novel strain of Lactococcus cremoris named D2022 was identified to have significant SUA-lowering capability. Lactococcus cremoris D2022 significantly reduced SUA levels by inhibiting uric acid synthesis and regulating uric acid transportation. It was also found that Lactococcus cremoris D2022 alleviated HUA-induced renal inflammatory injury involving multiple signaling pathways. By focusing on the expression of NLRP3-related inflammatory genes, we found correlations between the expression levels of these genes and free fatty acid receptors (FFARs). In addition, oral administration of Lactococcus cremoris D2022 increased short-chain fatty acids (SCFAs) in cecal samples, which may be one of the mechanisms by which oral probiotics alleviate renal inflammation. Serum untargeted metabolomics showed changes in a variety of serum metabolites associated with purine metabolism and inflammation after oral administration of Lactococcus cremoris D2022, further confirming its systemic bioactivity. Finally, it was proved that Lactococcus cremoris D2022 improved intestinal barrier function. In conclusion, Lactococcus cremoris D2022 can alleviate HUA and HUA-induced nephropathy by increasing the production of SCFAs in the gut and systemic metabolism.

Lactococcus strains with psychobiotic properties improve cognitive and mood alterations in aged mice.

Aging often accompanies cognitive and mood disturbances. Emerging evidence indicates that specific probiotics mitigate cognitive and mood dysfunctions. Strains within Lactococcus, a subgroup of probiotics, including Lactococcus lactis and Lactococcus cremoris are shown beneficial effects on brain functions via the gut microbiota-brain axis (GBA). Our previous study identified two Lactococcus strains (L. lactis WHH2078 and L. cremoris WHH2080) with the ability to promote the secretion of gut 5-hydroxytryptophan (5-HTP), the precursor of the GBA mediator 5-hydroxytryptamine (5-HT). In this study, the modulatory effects of WHH2078 and WHH2080 on cognitive and mood alternations were investigated in aged mice. Oral administration of WHH2078 and WHH2080 (1 × 109 CFU/mL/day) in aged mice (12-month-old) for 12 weeks significantly improved cognitive and depressive-and anxiety-like behaviors. The neuronal loss, the 5-HT metabolism dysfunction, and the neuroinflammation in the hippocampus of aged mice were restored by WHH2078 and WHH2080. the disturbances in the serum tryptophan metabolism in aged mice were unveiled by metabolomics, notably with decreased levels of 5-HT and 5-HTP, and increased levels of kynurenine, 3-hydroxykynurenine, and indolelactic acid, which were reversed by WHH2078 and WHH2080. Regarding the gut microbial community, WHH2078 and WHH2080 restored the increased abundance of Firmicutes, Desulfobacterota, and Deferribacterota and the decreased abundance of Bacteroidota and Actinobacteriota in aged mice. The beneficial effects of the two strains were linked to the modulation of 5-HT metabolism and gut microbiota. Our findings point to the potential role of Lactococcus strains with 5-HTP-promoting abilities as therapeutic approaches for age-related cognitive and mood disorders.

Some enzymatic properties of lactic acid bacteria isolated from dairy products

This article presents data on the study of physiological and biochemical properties, antagonistic and enzymatic activity of lactic acid bacteria isolated from dairy products.9 types of lactic acid bacteria were studied:Lactobacillus bulgaricus GM – 08, Lactobacillus bulgaricus KZh – 01,Lactobacillus bulgaricus GS – 03, Lactococcus cremoris – 6, Lactococcus cremoris – 17, Lactococcus cremoris – 26, Lactococcus lactis – 1,Lactococcus lactis – 15, Lactococcus lactis – 23. These strains were found to have resistance to 2% and 4%-vertical NaCl concentrations, bile and phenol.In addition, the antagonistic activity of Gram-positive and Gram-negative microorganisms in relation to test cultures of Staphylococcus aureus, Salmonella dublin, Escherichia coli, Bacillus subtilis, Sarcina flava was studied.All studied lactic acid bacteria showed activity in Test cultures with different inhibition zones. The Lactococcus lactis – 23 strains showed high activity for all cultures, with an inhibition zone of 17-25 mm.Further, 5 strains were selected from these strains and their aroma-forming properties, the formation of diacetyl and ammonia from arginine, hemolytic and lecithinase activity were studied.Compositions were compiled from these strains to make yeast.The compatibility of strains of lactic acid bacteria was checked with each strain individually and with the duration of milk clotting according to organoleptic indicators compared to the duration of milk clotting.Thus, the most active clot formation was obtained by the Lactococcus lactis – 23 strainof the selected combinations.

Сравнительная оценка динамики развития и кислотообразования лактококков при технически значимых температурах

Mesophilic lactococci strains of Lactococcus lactis subsp. lactis and Lactococcus cremoris are part of many fermented dairy products, including cheeses. As a rule, they serve as the main acid agents in bacterial starter cultures. Throughout all technological stages of cheese production, these microorganisms encounter various factors that affect their growth and metabolism. Despite its widespread industrial use and accumulated scientific knowledge, the question still remains of how lactococci behave during production. This article provides a comparative assessment of temperature-related development processes and acid formation in lactococci Lactococcus lactis subsp. lactis and Lactococcus cremoris. The strains showed no differences in culture development rate or acid-forming activity under optimal thermostating conditions during the deep liquidphase cultivation in milk. These variables were measured by the increase in titratable acidity (°T) and the decrease in active acidity (pH). The lactococci developed and metabolized carbohydrates at (10 ± 1) °C, which allowed for acid formation processes during the cheese ripening stage. Both strains showed significant psychrotrophy. Their development and metabolism were interrupted at (4 ± 1) °C, which made it possible to avoid the risk of reducing the storage capacity of fermented dairy products under the effect of lactococci starter. The greatest risks of using Lactococcus cremoris starter cultures were associated with their low thermal stability, which made the strains sensitive to temperatures above (40 ± 1) °C.

Microbial screening of animal skin bags used in traditional cheesemaking

Bouhezza is a traditional Algerian cheese produced and ripened in goatskin bags called Djeld. The aim of this study was to characterize the microbial ecosystem from Djeld (fresh and dried Djeld for making Bouhezza cheese) and the changes introduced by Lben microflora during its preparation and to identify its role in cheesemaking and its safety. Two replicates of fresh and dried skin bags (FS and DS) were sampled and analyzed before and after contact with Lben. The microbiological results showed no pathogens. Skins observed before the addition of Lben were less populated 2.86 and 3.20 log CFU cm−2 than skins examined after the addition of Lben (approximately 6.0 log CFU cm−2), suggesting a potential role of Lben in releasing some microorganisms into the skin during its time in the Djeld. However, an increase in mesophilic lactic acid bacteria and yeasts was observed in Lben after different periods of interaction with the skin. PCR-TTGE revealed the predominance of lactic acid bacteria (Lactiplantibacillus plantarum, Limosilactobacillus fermentum, Staphylococcus equorum subsp. linens, Lactococcus cremoris, Streptococcus thermophilus) and a few high-GC-content bacteria (Lacticaseibacillus paracasei, Brevibacterium casei). Transfer of several microbial species was observed between the goatskin bag biofilm and Lben during the overnight interaction. Bands corresponding to Lacticaseibacillus paracasei, Brevibacterium casei, and Lactobacillus delbrueckii subsp. lactis were detected in the fresh skin profile and in Lben after contact with the fresh skin. Lacticaseibacillus paracasei was found in dried skin and Lben after contact with dry skin. Lactobacillus helveticus and Enterococcus faecalis appeared in the Lben profile and persisted in Lben and the biofilm-covered dry skin after interaction. These results demonstrate an exchange of specific microbial populations between goatskin bag biofilm and Lben during the traditional preparation method, suggesting that the diversity of goatskin biofilm contributes to the microbial diversity of Lben used in the production of Bouhezza cheese.

Microbial interactions shape cheese flavour formation

Cheese fermentation and flavour formation are the result of complex biochemical reactions driven by the activity of multiple microorganisms. Here, we studied the roles of microbial interactions in flavour formation in a year-long Cheddar cheese making process, using a commercial starter culture containing Streptococcus thermophilus and Lactococcus strains. By using an experimental strategy whereby certain strains were left out from the starter culture, we show that S. thermophilus has a crucial role in boosting Lactococcus growth and shaping flavour compound profile. Controlled milk fermentations with systematic exclusion of single Lactococcus strains, combined with genomics, genome-scale metabolic modelling, and metatranscriptomics, indicated that S. thermophilus proteolytic activity relieves nitrogen limitation for Lactococcus and boosts de novo nucleotide biosynthesis. While S. thermophilus had large contribution to the flavour profile, Lactococcus cremoris also played a role by limiting diacetyl and acetoin formation, which otherwise results in an off-flavour when in excess. This off-flavour control could be attributed to the metabolic re-routing of citrate by L. cremoris from diacetyl and acetoin towards α-ketoglutarate. Further, closely related Lactococcus lactis strains exhibited different interaction patterns with S. thermophilus, highlighting the significance of strain specificity in cheese making. Our results highlight the crucial roles of competitive and cooperative microbial interactions in shaping cheese flavour profile.