In this study, a novel potential probiotic strain Lactobacillus paracasei K5, previously isolated from traditional Greek Feta cheese and kefir grains, was evaluated as an adjunct culture for ...industrial white brined cheese production. Targeting industrial applications, apart from free cell cultures, a novel ready‐to‐use freeze‐dried immobilized biocatalyst was prepared. The biocatalyst composed of L. paracasei K5 cells immobilized on delignified wheat bran prebiotic carrier and was freeze‐dried without cryoprotectants. The adjunct free or immobilized culture was added separately without prior adaptation during white brined cheese manufacture and the produced cheeses were compared with commercial white brined cheeses. Several parameters that affect the acceptability and quality of the cheeses, including microbial populations, physicochemical parameters, volatile by‐products and organoleptic characteristics, were analyzed through 70 days of storage. Results showed that the viability of the adjunct culture added either free or immobilized remained in high levels (7 to 8 log cfu/g) during maturation and storage. In addition, all white brined cheeses with the adjunct probiotic culture showed a sharp decrease in spoilage and pathogenic microorganisms such as enterobacteria, salmonella, staphylococci and coliforms during cheese maturation, especially when compared with the commercial white brined cheeses. Finally, after maturation time exceeded, all cheeses were characterized as safe for human consumption. Cheeses volatile compounds were significantly enhanced by the incorporated immobilized biocatalysts. These findings indicate the potential industrial use of freeze‐dried ready‐to‐use immobilized lactobacilli as reinforcement adjunct cultures for the production of good‐quality functional cheese products.
Practical Application
The launch on market of novel foods developed by the incorporation of functional ingredients provides potential benefits to consumers’ diet and new business opportunities for producers. Probiotic food products are one significant category of functional foods. Thus, this study focused on the development of a novel ready‐to‐use freeze‐dried potential probiotic biocatalyst for functional white brined cheese production. The potential industrial application of such biocatalysts is highlighted by their incorporation as adjunct cultures that resulted in good‐quality functional cheeses.
Analysing correlations between the observed health effects of ingested probiotics and their survival in digestive tract allows adapting their preparations for food. Tracking ingested probiotic in ...faecal samples requires accurate and specific tools to quantify live vs dead cells at strain level. Traditional culture-based methods are simpler to use but they do not allow quantifying viable but non-cultivable (VBNC) cells and they are poorly discriminant below the species level. We have set up a viable PCR (vPCR) assay combining propidium monoazide (PMA) treatment and either real time quantitative PCR (qPCR) or droplet digital PCR (ddPCR) to quantify a Lactobacillus rhamnosus and two Lactobacillus paracasei subsp. paracasei strains in piglet faeces. Adjustments of the PMA treatment conditions and reduction of the faecal sample size were necessary to obtain accurate discrimination between dead and live cells. The study also revealed differences of PMA efficiency among the two L. paracasei strains. Both PCR methods were able to specifically quantify each strain and provided comparable total bacterial counts. However, quantification of lower numbers of viable cells was best achieved with ddPCR, which was characterized by a reduced lower limit of quantification (improvement of up to 1.76 log10 compared to qPCR). All three strains were able to survive in the piglets' gut with viability losses between 0.78 and 1.59 log10/g faeces. This study shows the applicability of PMA-ddPCR to specific quantification of small numbers of viable bacterial cells in the presence of an important background of unwanted microorganisms, and without the need to set up standard curves. It also illustrates the need to adapt PMA protocols according to the final matrix and target strain, even for closely related strains. The PMA-ddPCR approach provides a new tool to quantify bacterial survival in faecal samples from a preclinical and clinical trial.
•Tracking probiotics in faecal samples requires accurate tools to quantify viable cells.•qPCR and ddPCR combining propidium monoazide were compared to quantify lactobacilli.•Both PCR methods provided comparable total bacterial counts for each strain tested.•ddPCR is well adapted to specifically quantify low amounts of lactobacilli in faeces.•PMA-ddPCR approach provides a new tool to quantify survival of ingested probiotics.
Optically active aromatic alcohols are valuable chiral building blocks of many natural products and chiral drugs. Lactobacillus paracasei BD87E6, which was isolated from a cereal‐based fermented ...beverage, was shown as a biocatalyst for the bioreduction of 1‐(benzofuran‐2‐yl) ethanone to (S)‐1‐(benzofuran‐2‐yl) ethanol with highly stereoselectivity. The bioreduction conditions were optimized using L. paracasei BD87E6 to obtain high enantiomeric excess (ee) and conversion. After optimization of the bioreduction conditions, it was shown that the bioreduction of 1‐(benzofuran‐2‐yl)ethanone was performed in mild reaction conditions. The asymmetric bioreduction of the 1‐(benzofuran‐2‐yl)ethanone had reached 92% yield with ee of higher than 99.9% at 6.73 g of substrate. Our study gave the first example for enantiopure production of (S)‐1‐(benzofuran‐2‐yl)ethanol by a biological green method. This process is also scalable and has potential in application. In this study, a basic and novel whole‐cell mediated biocatalytic method was performed for the enantiopure production of (S)‐1‐(benzofuran‐2‐yl)ethanol in the aqueous medium, which empowered the synthesis of a precious chiral intermediary process to be converted into a sophisticated molecule for drug production.
It is widely assumed that bacterial resistance will be acquired when bacteria are exposed to long-term sublethal concentrations of antibiotics. The objective of this study was to evaluate the ability ...of two bacterial strains
Lactobacillus plantarum
(18A) and
Lactobacillus paracasei
(18C) isolated from the fuel ethanol industry to acquire bacterial resistance during long-term (≥ 14 days) exposure to sublethal concentrations of penicillin G and virginiamycin. Neither strain acquired resistance to virginiamycin after 69 days of exposure, but both strains did acquire resistance to penicillin G after 18 days. Strain 18A appeared to acquire resistance to a penicillin G and virginiamycin mixture after 7 days of exposure, but the incubation period was not long enough to verify. These results indicate that antibiotic resistance in two common
Lactobacillus
strains does not develop from sublethal exposure to virginiamycin after 69 days of exposure, but resistance can be developed with sublethal exposure to penicillin G.
Scope
The present study aims to assess the antidiabetic effect of Lactobacillus paracasei strain NL41 and its potential mechanisms in rats with type 2 diabetes mellitus (T2DM) induced by a high‐fat ...diet and low‐dose streptozotocin administration (HFD/STZ).
Methods and results
Eighteen Sprague–Dawley (SD) rats are randomly assigned to three groups: one control, one HFD/STZ model, and one HFD/STZ‐Lactobacillus protection group with administration of strain NL41 for 12 weeks. Blood is collected for biochemical parameters analysis and tissue samples for histological analysis. Treatment with strain NL41 results in excellent blood glucose regulation and significantly decreases insulin resistance, and HbA1c, glucagon, and leptin levels, accompanied by remarkable improvement of dyslipidemia and oxidative stress status in the animals. Islets of Langerhans, liver, and kidney are significantly protected in the NL41‐treated rats compared to the HFD/STZ‐T2DM model rats. Histochemistry shows that strain NL41 inhibits beta‐cell loss and alpha‐cell expansion, indicating pancreatic islets as the targeted tissues for the primary ameliorative effect of the probiotic strain on HFD/STZ‐T2DM rats. Crosstalk between the gut–liver and liver–pancreas endocrine axes is discussed.
Conclusion
Probiotic strain NL41 prevents HFD/STZ‐T2DM by decreasing insulin resistance and oxidative stress status, and protecting beta‐cell function.
The antidiabetic effect of probiotic Lactobacillus paracasei NL41 and its potential mechanisms in rats with T2DM induced by HFD/STZ are focused on. Treatment with NL41 significantly decreases insulin resistance, protects pancreatic islets function, and remarkably improves dyslipidemia and oxidative stress, which can be a potential probiotic for the primary prevention of T2DM.
Lactobacillus paracasei DG is a bacterial strain with recognized probiotic properties and is used in commercial probiotic products. However, the mechanisms underlying its probiotic properties are ...mainly unknown. In this study, we tested the hypothesis that the ability of strain DG to interact with the host is at least partly associated with its ability to synthesize a surface-associated exopolysaccharide (EPS). Comparative genomics revealed the presence of putative EPS gene clusters in the DG genome; accordingly, EPS was isolated from the surface of the bacterium. A sample of the pure EPS from strain DG (DG-EPS), upon nuclear magnetic resonance (NMR) and chemical analyses, was shown to be a novel branched hetero-EPS with a repeat unit composed of l-rhamnose, d-galactose, and N-acetyl-d-galactosamine in a ratio of 4:1:1. Subsequently, we demonstrated that DG-EPS displays immunostimulating properties by enhancing the gene expression of the proinflammatory cytokines tumor necrosis factor alpha (TNF-α) and interleukin 6 (IL-6), and particularly that of the chemokines IL-8 and CCL20, in the human monocytic cell line THP-1. In contrast, the expression of the cyclooxygenase enzyme COX-2 was not affected. In conclusion, DG-EPS is a bacterial macromolecule with the ability to boost the immune system either as a secreted molecule released from the bacterium or as a capsular envelope on the bacterial cell wall. This study provides additional information about the mechanisms supporting the cross talk between L. paracasei DG and the host.
The consumption of food products and supplements called probiotics (i.e., containing live microbial cells) to potentially prevent or treat specific diseases is constantly gaining popularity. The lack of knowledge on the precise mechanisms supporting their potential health-promoting properties, however, greatly limits a more appropriate use of each single probiotic strain. In this context, we studied a well-known probiotic, Lactobacillus paracasei DG, in order to identify the constitutive molecules that can explain the documented health-promoting properties of this bacterium. We found a novel polysaccharide molecule, named DG-EPS, that is secreted by and covers the bacterium. We demonstrated that this molecule, which has a chemical structure never identified before, has immunostimulatory properties and therefore may contribute to the ability of the probiotic L. paracasei DG to interact with the immune system.
Display omitted
•Wild Lb. paracasei enhances flavour of pasteurized cow milk Caciotta cheese.•Experimental production of test and control Caciotta cheeses.•QRT-PCR quantification of starter and ...adjunct culture in curd and cheeses.•Aromatic profile, colour, texture and sensorial perception significantly different.
Caciotta is the name used to define a type of Italian semi-hard cheese Caciotta-type cheese. Due to the short ripening time, pasteurization is necessary to eliminate the potential pathogenic bacteria, which may be present in raw milk, causing also the reduction of ripened cheese flavor. The purpose of this research was to evaluate the effect of a selected wild Lactobacillus paracasei strain experimentally used as adjunct culture to enhance the flavour formation in a short-ripened caciotta-type cheese. An integrated polyphasic approach was used to compare the experimental and control Caciotta produced in a company located in Emilia Romagna region (Italy). It was demonstrated how the L. paracasei 4341 was able to develop in curd and cheese interacting with the acidifying commercial starter. The main acidifying starter species, were differently affected by the presence of the adjunct culture. Streptococcus thermophilus shown comparable behavior in all cheese-making step of control and experimental Caciotta, while Lactobacillus delbrueckii subsp bulgaricus, growth was slowed down by the presence of the adjunct culture during the whole ripening time. The higher amount of volatile compounds and organic acids due to the adjunct L. paracasei 4341 lead to a clear differentiation of the experimental Caciotta respect to the control, in terms of aromatic profile, color, texture and sensorial perception.
Respiratory tract infections such as flu cause severe morbidity and mortality and are among the leading causes of death in children and adults worldwide. Commensal microbiota is critical for ...orchestrating tissue homeostasis and immunity in the intestine. Probiotics represent an interesting source of immune modulators and several clinical studies have addressed the potential beneficial effects of probiotics against respiratory infections. Therefore, we have investigated the mechanisms of protection conferred by L. paracasei CNCM I-1518 strain in a mouse model of influenza infection. Notably, local myeloid cells accumulation is generated in the lungs after seven days feeding with L. paracasei prior to viral infection. L. paracasei-fed mice showed reduced susceptibility to the influenza infection, associated with less accumulation of inflammatory cells in the lungs, faster viral clearance and general health improvement. Interestingly, Allobaculum was significantly increased in L. paracasei-fed mice 7 days after influenza infection, even if the gut microbiota composition was not altered overall. L. paracasei-purified peptidoglycan partially recapitulated the protective phenotype observed with the entire bacteria. Collectively, our results demonstrate that oral consumption of L. paracasei CNCM I-1518 modulates lung immunity was associated with an improved control of influenza infection. These results further extend the beneficial role for certain lactobacilli to alleviate the burden of respiratory tract infections.
This study aimed to predict the optimal carbon source for higher production of exopolysaccharides (EPS) by Lactobacillus paracasei TD 062, and to evaluate the effect of this carbon source on the ...production and monosaccharide composition of EPS. We evaluated the EPS production capacity of 20 strains of L. paracasei under the same conditions. We further investigated L. paracasei TD 062, which showed the highest EPS-producing activity (0.609 g/L), by examining the associated biosynthesis pathways for EPS. Genomics revealed that fructose, mannose, trehalose, glucose, galactose, and lactose were carbon sources that L. paracasei TD 062 could use to produce EPS. We identified an EPS synthesis gene cluster that could participate in transport, export, and sugar chain synthesis, and generate 6 sugar nucleotides. Experimental results showed that the sugar content of the EPS produced using fermentation with the optimized carbon source (fructose, mannose, trehalose, glucose, galactose, and lactose) increased by 115%. Furthermore, use of the optimized carbon source changed the monosaccharide content of the associated EPS. The results of enzyme activity measurements showed significant increases in the activity of 2 key enzymes involved in the glycoside synthesis pathway. Our study revealed that optimizing the carbon source provided for fermentation not only increased the production of EPS, but also affected the composition of the monosaccharides by increasing enzyme activity in the underlying synthesis pathways, suggesting an important role for carbon source in the production of EPS by L. paracasei TD 062.
Although probiotic lactobacilli and bifidobacteria are generally considered safe by various regulatory agencies, safety properties, such as absence of transferable antibiotic resistance, must still ...be determined for each strain prior to market introduction as a probiotic. Safety requirements for probiotics vary regionally and evaluation methods are not standardized, therefore methodologies are often adopted from food ingredients or chemicals to assess microbial safety. Four individual probiotic strains, Lactobacillus acidophilus NCFM®, Lactobacillus paracasei Lpc-37®, Bifidobacterium animalis subsp. lactis strains Bl-04®, and Bi-07®, and their combination (HOWARU® Restore) were examined for antibiotic resistance by broth microdilution culture, toxin genes by PCR and genome mining, and acute oral toxicity in rats. Only B. lactis Bl-04 exhibited antibiotic resistance above a regulated threshold due to a tetW gene previously demonstrated to be non-transferable. Genomic mining did not reveal any bacterial toxin genes known to harm mammalian hosts in any of the strains. The rodent studies did not indicate any evidence of acute toxicity following a dose of 1.7–4.1 × 1012 CFU/kg body weight. Considering a 100-fold safety margin, this corresponds to 1.2–2.8 × 1012 CFU for a 70 kg human. Our findings demonstrate a comprehensive approach of in vitro, in silico, and in vivo safety testing for probiotics.
•Safety evaluations of individual probiotics are equivalent to combined products.•In vitro and in silico testing sufficiently identifies microbial safety risks.•In vivo toxicology studies of typical probiotic genera may not be necessary.
PDF
