Automated workflow for characterization of bacteriocin production in natural producers Lactococcus lactis and Latilactobacillus sakei.

BACKGROUND: Lactic acid bacteria are commonly used as protective starter cultures in food products. Among their beneficial effects is the production of ribosomally synthesized peptides termed bacteriocins that kill or inhibit food-spoiling bacteria and pathogens, e.g., members of the Listeria species. As new bacteriocins and producer strains are being discovered rapidly, modern automated methods for strain evaluation and bioprocess development are required to accelerate screening and development processes. RESULTS: In this study, we developed an automated workflow for screening and bioprocess optimization for bacteriocin producing lactic acid bacteria, consisting of microcultivation, sample processing and automated antimicrobial activity assay. We implemented sample processing workflows to minimize bacteriocin adsorption to producer cells via addition of Tween 80 and divalent cations to the cultivation media as well as acidification of culture broth prior to cell separation. Moreover, we demonstrated the applicability of the automated workflow to analyze influence of media components such as MES buffer or yeast extract for bacteriocin producers Lactococcus lactis B1629 and Latilactobacillus sakei A1608. CONCLUSIONS: Our automated workflow provides advanced possibilities to accelerate screening and bioprocess optimization for natural bacteriocin producers. Based on its modular concept, adaptations for other strains, bacteriocin products and applications are easily carried out and a unique tool to support bacteriocin research and bioprocess development is provided.

Screening and evaluating microorganisms with broad-spectrum biogenic amine-degrading ability from naturally fermented dry sausage collected from Northeast China.

This study aimed to screen autochthonous strains with broad-spectrum biogenic amine (BA) degradation ability from traditional dry sausages and to evaluate their BA-degrading ability in dry sausages. A total of 120 strains were isolated from dry sausages collected from various regions in Northeast China, and 35 of 120 isolates were identified as non-BA producing strains by the in vitro agar method. The random amplified polymorphic DNA polymerase chain reaction technique genotyped these 35 isolates into 18 biotypes. Moreover, high performance liquid chromatography (HPLC) quantification showed that six strains (Latilactobacillus sakei MDJ6; Lactiplantibacillus plantarum SH7; Weissella hellenica DQ9; Staphylococcus saprophyticus JX18 and SYS8; and Macrococcus caseolyticus SYS11) of the 18 biotypes exhibited broad-spectrum BA-degrading ability, all of which had various levels of amine oxidase activity with monoamine oxidase and diamine oxidase activities ranged of 6.60-619.04 and 26.32-352.81 U/mg protein, respectively. These six strains were subsequently inoculated into dry sausages and the results showed that they exhibited varying degrees of BA-degrading ability, of which strain Lat. sakei MDJ6 allowed to have less BA production on dry sausage with a final concentration of 61.33 mg/kg.

Companilactobacillus alimentarius: An extensive characterization of strains isolated from spontaneous fermented sausages.

Companilactobacillus alimentarius is a facultatively heterofermentative lactic acid bacterium (LAB) that is a significant constituent within the microbiota of various traditional fermented foods exerting several functions in fermentative or ripening processes. This species has been isolated from Spanish fermented sausages, where its frequency of isolation was comparable to those of Latilactobacillus sakei and Latilactobacillus curvatus. Despite to its presence in several niches, ecological information on this species is still scarce and only few publications report information about its safety features (i.e. antibiotic resistance). Since studies on C. alimentarius concern the analysis of a few individual traits regarding this species, a more extensive work on a larger number of isolates from the same matrix have been performed to allow a clearer interpretation of their phenotypic and technological characteristics. Specifically, 14 strains of C. alimentarius isolated from Mediterranean spontaneously fermented sausages, have been screened for their safety and technological characteristics (such as antibiotic resistance, biogenic amine production, inhibiting potential, growth at different temperatures and NaCl concentrations) and with phenotype microarrays with the aim to elucidate their potential role and contribution to sausage fermentation and ripening. In general, a wide variability was observed in relation to the parameters considered. Several of the tested strains were able to produce histamine, tyramine and putrescine while the antibiotic resistance greatly varied according to the strains, with the exception of vancomycin. In addition, C. alimentarius strains showed a relevant potential to grow in conditions of salt and temperature mimicking those found in fermented foods. In particular, the growth at 10 °C and in the presence of salt can explain the presence of C. alimentarius in sausages and its adaptation to fermented meat environment in which low temperature can be applied during ripening. The differentiation of the phenotypic profile reflected the environmental conditions that influenced the isolation source, including those derived by the raw materials. Given the species frequent association with spontaneous fermentations or the ripening microbiota of various products, despite not being intentionally used as starter cultures, the data presented in this study contribute to a deeper comprehension of their role, both advantageous and detrimental, in numerous significant fermented foods.

Heat-Treated Paraprobiotic Latilactobacillus sakei KU15041 and Latilactobacillus curvatus KU15003 Show an Antioxidant and Immunostimulatory Effect.

The lactic acid bacteria, including Latilactobacillus sakei and Latilactobacillus curvatus, have been widely studied for their preventive and therapeutic effects. In this study, the underlying mechanism of action for the antioxidant and immunostimulatory effects of two strains of heat-treated paraprobiotics was examined. Heat-treated L. sakei KU15041 and L. curvatus KU15003 showed higher radical scavenging activity in both the 2-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) and 2,2-diphenyl-1-picryl-hydrazyl (DPPH) assays than the commercial probiotic strain LGG. In addition, treatment with these two strains exhibited immunostimulatory effects in RAW 264.7 macrophages, with L. curvatus KU15003 showing a slightly higher effect. Additionally, they promoted phagocytosis and NO production in RAW 264.7 cells without any cytotoxicity. Moreover, the expression of tumor necrosis factor-α, interleukin (IL)-1ß, and IL-6 was upregulated. These strains resulted in an increased expression of inducible nitric oxide synthase and cyclooxygenase-2. Moreover, the nuclear factor-κB and mitogen-activated protein kinase signaling pathways were stimulated by these strains. These findings suggest the potential of using L. sakei KU15041 and L. curvatus KU15003 in food or by themselves as probiotics with antioxidant and immune-enhancing properties.

Multifunctional effects of Lactobacillus sakei HEM 224 on the gastrointestinal tract and airway inflammation.

Mucosal tissues serve as the first defense line and their commensal microbiota play a role in sustaining of host health. This study aimed to isolate and evaluate a putative probiotic strain on various mucosal regions. Lactobacillus sakei HEM 224 was isolated from traditional Korean kimchi and identified. In the safety assessment L. sakei HEM 224 showed negative results for hemolysis, biogenic amine production and transferable antibiotic resistance. The probiotic potential of strain HEM 224 in diverse mucosal areas was shown in two different models, viz. a murine model with colitis induced by dextran sulfate sodium (DSS) and an allergic airway inflammation model induced by ovalbumin (OVA). In the colitis model, oral administration of L. sakei HEM 224 improved colitis physiology with immunomodulation, enhancing barrier components and gut microbiota alteration. In the allergic airway inflammation model, the intranasal administration of the strain decreased type 2 inflammation and enhanced epithelial barrier integrity from the airways. These results demonstrate that L. sakei HEM 224 can ameliorate inflammatory conditions in both the gastrointestinal and respiratory tracts through the reinforcement of the epithelial barrier and immunomodulation.

Prophylactic effects of probiotic bacterium Latilactobacillus sakei on haematological parameters and cytokine profile of mice infected with Plasmodium berghei ANKA during early malaria infection.

Malaria is a deadly parasitic disease caused a by protozoan parasite of the genus plasmodium. The challenges facing by chemotherapy and vector control couple with the lack of vaccine against malaria necessitate an urgent need for the development of alternative treatment regimens to combat this disease. One possible antimalarial treatment regimen is the use of probiotic bacteria as dietary supplements. Traditionally fermented milk is a rich source of probiotic bacteria that up to date, very few studies have been carried out on their immunoprotective effects against early malaria infection in mice. This study sought to assess the prophylactic activities of a probiotic bacterium Latilactobacillus sakei on malaria and inflammation in Plasmodium berghei infected mice. The probiotic bacterium was isolated from the Fulani's traditionally fermented milk and identified using the sequencing of the 16S r RNA gene. The repository activity of L. sakei on malaria was assessed using the method described by Peters with slight modification. Eighty-four BALB/c mice were randomly divided into two sets of seven groups of six mice each. One set received orally different doses of L. sakei Chloroquine and Sulfadoxine/Pyrimethamine for seven days before infection while the other set received for fourteen days before infection with 0.1 mL of 107Plasmodium berghei. Parasitaemia density, haematological parameters and inflammatory cytokines profile were evaluated. Data were presented as Mean ± SEM and analysed using SPSS version 20.0. The results of this study revealed that L. sakei significantly (p < 0.05) reduced in dose dependent manner parasite load, body weight loss and reduction of body temperature in all the treated mice when compare to untreated mice. Leukocytopenia, thrombocytosis and inflammation were also found to be significantly (p < 0.05) prevented in treated mice as compared to untreated mice. This study suggested that L sakei possesses immunomodulation and protective effects on early malaria infection in Plasmodium berghei mice.

Probiotic potential of Lactobacillus sakei L-7 in regulating gut microbiota and metabolism.

A growing body of research suggests that gut microbiota is inextricably linked to host health and disease，so we are committed to finding more probiotic resources that are beneficial to human health. This study evaluated the probiotic properties of Lactobacillus sakei L-7 isolated from home-made sausages. The basic probiotic properties of L. sakei L-7 were evaluated through in vitro tests. The strain showed 89% viability after 7 h of digestion in simulating gastric and intestinal fluid. The hydrophobicity, self-aggregation and co-aggregation of L. sakei L-7 showed it had a strong adhesion ability. C57BL/6 J mice were fed L. sakei L-7 for 4 weeks. 16 S rRNA gene analysis indicated that intake of L. sakei L-7 increased the richness of gut microbiota and abundance of beneficial bacteria Akkermansia, Allobaculum and Parabacteroides. Metabonomics analysis revealed that beneficial metabolite gamma-aminobutyric acid and docosahexaenoic acid increased significantly. While the level of metabolite sphingosine and arachidonic acid significantly decreased. In addition, serum levels of inflammatory cytokines interleukin (IL)- 6 and tumor necrosis factor (TNF)-α were significantly decreased. The results suggested that L. sakei L-7 may promote gut health and reduce the occurrence of inflammatory response, it has the potential to become a probiotic.

Effect of Lactobacillus sakei OK67 in Reducing Body and Visceral Fat in Lifestyle-Modified Overweight Individuals: A 12-Week, Randomized, Double-Blind, Placebo-Controlled Clinical Trial.

Obesity is a global health problem that affects the quality of life. It is a multidimensional chronic risk factor for major medical conditions, such as cardiovascular diseases, diabetes, and cancer. This clinical trial evaluated the efficacy of Lactobacillus sakei OK67 (DW2010), a lactic acid bacterium, in reducing body and visceral fat in overweight individuals (body mass index ≥25 kg/m2 and <30 kg/m2), aged 20-60 years. A total of 100 subjects placed in a lifestyle modification program were randomly assigned to receive either DW2010 (2.0 g/day, 1.0 × 1010 CFU) or a placebo for 12 weeks. The efficacy of DW2010 was evaluated by measuring body fat mass using dual-energy X-ray absorptiometry and visceral fat area using computed tomography. After 12 weeks, the change in body fat in the DW2010 group was not markedly different from that in the placebo group. However, visceral fat area decreased more in the DW2010 group than in the placebo group (p = 0.035). During the clinical trial, no major adverse events were reported. Moreover, no statistical differences were observed in the biochemical parameters of the DW2010 and placebo groups. Overall, we concluded that the intake of DW2010 for 12 weeks is safe and potentially reduces visceral fat in lifestyle-modified overweight subjects.

Effect of high pressure processing and water activity on pressure resistant spoilage lactic acid bacteria (Latilactobacillus sakei) in a ready-to-eat meat emulsion model.

The main use of High Pressure Processing (HPP) in food processing is microorganism inactivation, and studies demonstrated that the characteristics of matrix and microorganisms can interfere on it. As the behavior of lactic acid bacteria exposed to different water activity (aw) levels in a meat product is still unclear, this study aimed to determine the effect of pressure, time, and aw to inactivate Latilactobacillus sakei, a pressure resistant lactic acid bacteria (LAB) in a meat emulsion model through a response surface methodology. The meat emulsion model was designed with adjusted aw (from 0.940 to 0.960) and was inoculated with a pressure resistant LAB and processed varying pressure (400-600 MPa) and time (180-480 s), following the Central Composite Rotational Design (CCRD). The inactivation of the microorganism ranged from 0.99 to 4.12 UFC/g depending on the applied condition. At studied conditions, according to the best fitting and most significant polynomial equation (R2 of 89.73 %), in a meat emulsion model, aw had no influenced on HPP inactivation on LAB (p > 0.05) and only pressure and holding time had significative impact on it. The results of experimental validation of the mathematical model were satisfactory, confirming the suitability of the model. The information obtained in the present study stands out the matrix, microorganism and process effects at HPP efficiency. The answers obtained support food processors in product development, process optimization and food waste reduction.

Alternative culture medium design for biomass production of autochthonous meat starter Latilactobacillus sakei sp. sakei ACU-2.

The autochthonous strain Latilactobacillus sakei sp. sakei ACU-2 was selected as a meat starter culture for dry sausage production. Transferring this strain from laboratory scale to industry requires an increase in biomass production, while lowering process costs. In this study, a combination of techniques was applied in order to optimize the culture medium composition to enhance biomass production of L. sakei ACU-2. One variable at a time experiments, Plackett-Burman design, and mixture design were performed to fulfill the strain nutritional requirements. Eventually, the optimized formulation contained 19.46 g/L yeast extract; 8.28 g/L whey protein concentrate; 2.26 g/L soy peptone; 30 g/L cerelose; 1 g/L Tween 80; 5 g/L sodium acetate; 0.2 g/L magnesium sulfate and 0.05 g/L manganese sulfate. When L. sakei ACU-2 was cultivated in a bioreactor using the alternative medium, an enhancement of 75.5% of biomass production was achieved, in comparison to its growth in the commercial de Man, Rogosa, and Sharpe medium. Furthermore, a reduction of 62-86% of the cost was also attained. These results support a promising large-scale application of the designed medium for high biomass yields of the starter culture at minor costs.

Latilactobacillus sakei Wikim0066 Protects Skin through MMP Regulation on UVB-Irradiated In Vitro and In Vivo Model.

Ultraviolet (UV) B exposure induces wrinkle formation, collagen fiber breakdown, and transepidermal water loss (TEWL). UVB irradiation induces the expression of mitogen-activated protein kinase (MAPK), activator protein 1 (AP-1), and nuclear factor kappa B (NF-κB), which affect the expression of matrix metalloproteinases (MMP). We confirmed the effects of Latilactobacillus sakei wikim0066 (wikim0066) on UVB-irradiated Hs68 cells and HR-1 hairless mice cells. wikim0066 restored the production of type I procollagen by regulating the expression of MMP-1 and -3, MAPK, AP-1, and NF-κB in UVB-irradiated Hs68 cells and HR-1 mice. Oral administration of wikim0066 alleviates wrinkle formation, epidermal thickness, and TEWL in UVB-irradiated HR-1 hairless mice. These results indicated that wikim0066 has the potential to prevent UVB-induced wrinkle formation.

The impact of different fermenting microbes on residual purine content in fermented lamb jerky following in vitro digestion.

This is the first study to investigate the effect of different lactic acid bacteria fermentation methods on the purine content of mutton jerky and to examine the changes in purine in these fermented mutton jerky samples underan in vitro simulated digestive system to determine an efficient method for regulating exogenous purine intake. According to the analysis, x3-2b Lactobacillus plantarum and composite bacteria can directly reduce the purine content in fermented dried mutton and achieve the purpose of reducing the intake of purine. However, after simulated intestinal digestion, it was observed that the purine content of these different sample groups was significantly decreased, and 37x-3 Pediococcus pentosaceus had the better effect. It has been shown that the 37x-3 Pediococcus pentosaceus, Lactobacillus sake, and composite bacteria significantly enhance the degree to which residual purine is depleted in the large intestine during digestion. In sum, one potentially successful method of regulating exogenous purine consumption is the development of fermented meat products using certain beneficial bacteria as a starter.

A New Bacteriocin from Latilactobacillus sakei: In vitro and In situ Application.

AIMS AND BACKGROUND: Natural preservatives are a viable alternative to replace chemical preservatives that have potential toxicity and carcinogenic effects. OBJECTIVE: To prove the effectiveness in increasing the microbiological stability of Minas Frescal cheese with the addition of a bacteriocin obtained from Latilactobacillus sakei as a natural preservative. METHODS: A new broad-spectrum bacteriocin was evaluated for its functional activity in vitro and in situ when applied in the formulation of Minas Frescal cheese. A commercial bacteriocin was used as a positive control. RESULTS: The inhibitory action of the bacteriocin studied was confirmed, with a reduction of 42.86% in the count of coagulase-positive Staphylococcus in relation to the negative control, at the end of the 30 days of study. For the group of thermotolerant coliforms, the bacteriocin studied showed greater efficiency than the commercial preservative. In vitro analyzes showed the inhibitory action of bacteriocin, above 87% inhibition against S. aureus, E. coli and Salmonela enteritidis, and approximately 90% against Listeria monocytogenes. CONCLUSION: It was concluded that the bacteriocin produced by the Latilactobacillus sakei strain has great potential for application in foods such as Minas Frescal cheese.

Autochthonous starter culture selection for Salame Piemonte PGI production.

Salame Piemonte is a dry-fermented meat product typical of the Piedmont region in Italy, manufactured using commercial starter cultures. This study aimed to select autochthonous starter cultures (ASCs) that could be used for sausage fermentation in order to strengthen the link with the geographical area of production and improve the sensory properties of the final product. A culture-dependent approach was adopted during three different spontaneous sausage fermentation processes to isolate and characterise the main bacterial resources involved. Dominant lactic acid bacteria (LAB) in each batch were Pediococcus pentosaceus, Latilactobacillus sakei, and Latilactobacillus curvatus; Staphylococcus xylosus was the most dominant coagulase-negative staphylococci (CNS) in all the studied batches. LAB and presumptive CNS isolates were further evaluated for their physiological properties and biotechnological potential. Thereafter, 11 strains were selected and evaluated for safety. Five selected strains (two P. pentosaceus, two L. sakei, and one S. xylosus strain) were used for pilot-scale Salame Piemonte production with seven different strain combinations. Based on the liking test, three ASC combinations led to the highest liking score compared to industrial products. These three ASCs were then used for the second pilot-scale sausage production confirming the high liking score. In summary, the use of P. pentosaceus and S. xylosus ASC significantly improved product sensory properties compared with that obtained using commercial starter cultures.

Lactobacillus sakei MJM60958 as a Potential Probiotic Alleviated Non-Alcoholic Fatty Liver Disease in Mice Fed a High-Fat Diet by Modulating Lipid Metabolism, Inflammation, and Gut Microbiota.

Non-alcoholic fatty liver disease (NAFLD) is a common liver disease with a rapidly increasing number of cases worldwide. This study aimed to evaluate the effects of Lactobacillus sakei MJM60958 (MJM60958) on NAFLD in vitro and in vivo. In in vitro tests, MJM60958 significantly inhibited lipid accumulation by 46.79% in HepG2 cells stimulated with oleic acid and cholesterol (OA-C). Moreover, MJM60958 showed safe and probiotic characteristics in vitro. In the animal study, MJM60958 administration in a high-fat diet-induced NAFLD mouse model significantly reduced body weight and liver weight, and controlled aspartate aminotransferase (ALT), aspartate transaminase (AST), triglyceride (TG), urea nitrogen (BUN), and uric acid (UA) levels in the blood, which are features of NAFLD. Further, treatment with MJM60958 also reduced steatosis scores in liver tissues, serum leptin and interleukin, and increased serum adiponectin content. Moreover, administration of MJM60958 resulted in a significantly decreased expression of some genes and proteins which are related to lipid accumulation, such as fatty acid synthase (FAS), acetyl-CoA carboxylase (ACC), and sterol regulatory element-binding protein 1 (SREBP-1), and also upregulated genes and protein expression of lipid oxidation such as peroxisome proliferator-activated receptor alpha (PPARα) and carnitine palmitoyltransferase 1a (CPT1A). Administration of MJM60958 increased the relative abundance of specific microbial taxa such as Verrucomicrobia, which are abundant in non-NAFLD mice, and reduced Firmicutes, which are a major group in NAFLD mice. MJM60958 affected the modulation of gut microbiota and altered the strain profile of short-chain fatty acids (SCFAs) production in the cecum by reduced lactic acid and enhanced acetic acid production. Overall, MJM60958 showed potential as a probiotic that can prevent and treat NAFLD.

Evaluation of metabolic activities and probiotic characteristics of two Latilactobacillus sakei strains isolated from pastirma.

The current study was carried out to investigate metabolic activities and main probiotic characteristics of two Latilactobacillus sakei strains (8.P1 and 28.P2) isolated from pastirma, a highly seasoned, air-dried cured beef. Both strains showed antimicrobial activity against important foodborne pathogens like Listeria monocytogenes, Staphylococcus aureus, Pseudomonas aeruginosa and so forth. For the characterization of antimicrobial activity, the effect of various enzymes, temperature and pH were tested. The results of the tests demonstrated that the antimicrobial activity of strains was based on the production of protein-structured compounds such as bacteriocin or bacteriocin like peptides. In metabolic activity studies, amounts of the lactic acid, proteolytic activity and hydrogen peroxide produced by the 8.P1 and 28.P2 were found to range between 16.09 and 17.32 mg/mL, 0.24 and 0.04 mg/mL and 0.98 and 0.04 µg/mL, respectively. It was also observed that neither strain could produce exopolysaccharide. Both strains were found susceptible to vancomycin, chloramphenicol, gentamicin, tetracycline, and netilmicin sulfate. When the strains are evaluated with respect to their probiotic potential, 28.P2 could tolerate acidic conditions, but 8.P1 showed sensitivity. The survival rate of the strains in the simulated gastric juice and their adhesion abilities were found suitable to stay alive in the gastrointestinal tract and to proliferate in the intestine. The evaluation of all the features of both strains demonstrated that both strains had the potential to be used as a protective culture. In addition, it was observed that 8.P1 and 28.P2 were more suitable as a starter culture and a probiotic candidate respectively.

Technological characterization and flavor-producing potential of lactic acid bacteria isolated from traditional dry fermented sausages in northeast China.

Thirty-seven lactic acid bacteria (LAB) strains were isolated from traditional dry sausages collected from Northeast China, including Latilactobacillus sakei (29 strains), Lactiplantibacillus plantarum (4 strains), Latilactobacillus curvatus (2 strains), Weissella hellenica (1 strain), and Lactococcus lactis (1 strain). Some LAB strains had tolerance to high concentrations of sodium chloride (6%), sodium nitrite (150 mg/L NaNO2), and acid (pH 4.0). They showed good growth and acidification properties and antimicrobial activity. Among them, five LAB strains that exhibited the best technological properties were selected and inoculated in the sausage model to explore their roles in flavor development. The contents of total free amino acids (FAAs) decreased ranging from 109.11 mg/g to 58.06 mg/g. A total of 46 volatile compounds were identified and the contents of volatile compounds increased in the sausage model during fermentation. Partial least squares regression analysis showed that Lb. sakei HRB10, Lb. plantarum MDJ2, W. hellenica HRB6, and Lc. lactis HRB0 promoted the generation of FAAs and volatile compounds in the sausage model. These findings demonstrated that the autochthonous LAB species are promising for the production of sausage with better flavor and fermentation performance.

Impact of different sugars and glycosyltransferases on the assertiveness of Latilactobacillus sakei in raw sausage fermentations.

Latilactobacillus sakei comprises a biodiversity of strains, which display different assertiveness upon their application as starter cultures in raw sausage fermentation. While the assertiveness of winning partner strains has been referred to competitive exclusion based on genomic settings enabling occupation of multiple niches of the sausage habitat, single strain assertiveness of L. sakei remained unexplained. In this study we assessed the impact of the expression of a glycosyltransferase enabling the production of a glucan from sucrose to the assertiveness of L. sakei TMW 1.411, which expresses a plasmid-encoded glycosyltransferase (gtf). In a sausage fermentation model wild type L. sakei TMW 1.411 and its plasmid-cured mutant were employed in competition with each other and with other Latilactobacillus sakei strains. To differentiate any effects resulting from general sugar utilization from those of glucan formation, the experiments were carried out with glucose, fructose, and sucrose, respectively. It was shown that the type of sugar affects the individual strains behaviour, and that the wild type was more competitive than the mutant in the presence of any of these sugars. In direct competition between wild type and mutant, a clear competitive advantage could also be demonstrated for the strain possessing the plasmid with the glycosyltransferase. Since this competitive advantage was observed with all sugars, not just sucrose, and Gtf expression has been shown as independent of the employed sugar, it is suggested that possession of the gtf gene-carrying plasmid confers a competitive advantage. It appears that the Gtf contributes to competitive exclusion and the establishment of colonization resistance, to a larger extent by an adhesive functionality of the Gtf on the cellular surface than by the production of glucan. Hence, gtf genes can be used as a possible additional marker for the selection of assertive L. sakei starter strains in sausage fermentation.

Genomic and metabolic features of the Lactobacillus sakei JD10 revealed potential probiotic traits.

Lactic acid bacteria that inhabit in the lung play important roles in maintaining the microbiome balance by interacting with the host immune system. Numerous metabolites (e.g., short chain fatty acids, bacteriocins, and hydrogen peroxide) produced by Lactobacillus sakei possess a special inhibitory spectrum against invading pathogens. In this research, the whole genome of L. sakei JD10 strain isolated from the porcine lung was sequenced and investigated. The whole size of the L. sakei JD10 chromosome was 1,989,921 bp, which encoded a total of 1951 predicted genes. Genome analyses revealed that many genes encoded carbohydrate-active enzymes (CAZymes) were predicted, which were responsible for the carbohydrate degradation and short chain fatty acids production. The metabolic profiles of short chain fatty acids in the L. sakei JD10 culture medium were measured by GC/TOFMS, and their regulatory effects on bacterial phagocytosis of RAW264.7 cells were also determined. The bacteriocin-producing genes of the L. sakei JD10 genome were also predicted, and a bacteriocin gene encoding carnocin was characterized and its molecular structure was analyzed. Two CRISPR-Cas system related genes were identified from the L. sakei JD10 genome, revealed that precise and efficient genome editing technologies could be applied for genetic engineering-manipulation. In all, investigation on the genomic features and metabolic features of L. sakei JD10 showed the potential probiotic traits to fight against pathogenic infection and regulate the host immune function.

A mathematical model to predict the antilisteria bioprotective effect of Latilactobacillus sakei CTC494 in vacuum packaged cooked ham.

Biopreservation is a strategy that has been extensively covered by the scientific literature from a variety of perspectives. However, the development of quantitative modelling approaches has received little attention, despite the usefulness of these tools for the food industry to assess the performance and to set the optimal application conditions. The objective of this study was to evaluate and model the interaction between the antilisteria strain Latilactobacillus sakei CTC494 (sakacin K producer) and Listeria monocytogenes in vacuum-packaged sliced cooked ham. Cooked ham was sliced under aseptic conditions and inoculated with L. monocytogenes CTC1034 and/or L. sakei CTC494 in monoculture and coculture at 10:10, 10:103 and 10:105 cfu/g ratios of pathogen:bioprotective cultures. Samples were vacuum packaged and stored at isothermal temperature (2, 5, 10 and 15 °C). The growth of the two bacteria was monitored by plate counting. The Logistic growth model was applied to estimate the growth kinetic parameters (N0, λ, µmax, Nmax). The effect of storage temperature was modelled using the hyperbola (λ) and Ratkowsky (µmax) models. The simple Jameson-effect model, its modifications including the Ncri and the interaction γ factor, and the predator-prey Lotka Volterra model were used to characterize the interaction between both microorganisms. Two additional experiments at non-isothermal temperature conditions were also carried out to assess the predictive performance of the developed models through the Acceptable Simulation Zone (ASZ) approach. In monoculture conditions, L. monocytogenes and L. sakei CTC494 grew at all temperatures. In coculture conditions, L. sakei CTC494 had an inhibitory effect on L. monocytogenes by lowering the Nmax, especially with increasing levels of L. sakei CTC494 and lowering the storage temperature. At the lowest temperature (2 °C) L. sakei CTC494 was able to completely inhibit the growth of L. monocytogenes when added at a concentration 3 and 5 Log higher than that of the pathogen. The inhibitory effect of the L. sakei CTC494 against L. monocytogenes was properly characterized and modelled using the modified Jameson-effect with interaction γ factor model. The developed interaction model was tested under non-isothermal conditions, resulting in ASZ values ≥83%. This study shows the potential of L. sakei CTC494 in the biopreservation of vacuum-packaged cooked ham against L. monocytogenes. The developed interaction model can be useful for the industry as a risk management tool to assess and set biopreservation strategies for the control of L. monocytogenes in cooked ham.