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Colorectal cancer (CRC) is still the leading cause of cancer death worldwide, but the clinical effect of drug therapy such as irinotecan is not an ideal way at present. In recent years, probiotics have attracted much attention, and the combination of probiotics may play an important role in the prevention and treatment of CRC. This work proposed a cellular chip-MS system, to study the synergistic effects of probiotic Lactobacillus rhamnosus GG (L.GG) and irinotecan on HCT116 cells by cell viability and on-line mass spectrometry (MS) analysis. The double-layer chip sandwiched with a polycarbonate membrane can co-culture HCT116 cells and L.GG. And the solid phase microextraction chip can be used for desalination and concentration. Finally, the extracted chemicals were entered the electrospray ionization quadrupole time-of-flight MS to detect irinotecan metabolites. The results showed that with the increasing concentration of co-cultured L.GG, the percentage of living HCT116 cells decreased, but the relative amount of metabolized SN-38 by HCT116 cells increased. Therefore, the microfluidic system can be used to detect and monitor the synergistic effect of irinotecan-L.GG combination on HCT116 cells. In summary, our study provided experimental evidence for the first time with potential applications of irinotecan-L.GG combination in CRC treatment, and the cellular chip-MS system as a powerful tool can be used in the experiments of probiotics as new drugs.

Verapamil, a calcium channel blocker, has been approved as the first-line drug for treatment of angina pectoris, hypertension and supraventricular tachycardia. Lactobacillus rhamnosus, one of the normal strains in human intestinal tract, is very popular in the probiotic market for conferring a health benefit on the host. This report investigated the potential of gut microbiota-drug interactions between lactobacillus rhamnosus and verapamil via using wild type (WT) and Cyp3a1/2 knockout (KO) rats. In WT rats, administration of Lactobacillus rhamnosus for 14 days decreased systemic exposure of verapamil and increased its metabolite norverapamil in vivo, and resulted in gut microbiota-drug interactions. In Cyp3a1/2 KO rats, however, this interaction disappeared. Further studies found that Lactobacillus rhamnosus induced CYP3A activity and expression, and changed the composition of gut microbiota, thus changing the pharmacokinetics of verapamil. These results demonstrated the interaction between lactobacillus rhamnosus and verapamil, and indicated that the effect of gut microbiota on metabolic enzymes cannot be ignored.

Lactobacillus (L.) rhamnosus CRL1505 accumulates inorganic polyphosphate (polyP) in its cytoplasm in response to environmental stress. The aim of this study was to evaluate the potential effects of polyP from the immunobiotic CRL1505 on an acute respiratory inflammation murine animal model induced by lipopolysaccharide (LPS). First, the presence of polyP granules in the cytoplasm of CRL1505 strain was evidenced by specific staining. Then, it was demonstrated in the intracellular extracts (ICE) of CRL1505 that polyP chain length is greater than 45 phosphate residues. In addition, the functionality of the genes involved in the polyP metabolism (ppk, ppx1 and ppx2) was corroborated by RT-PCR. Finally, the possible effect of the ICE of CRL1505 strain containing polyP and a synthetic polyP was evaluated in vivo using a murine model of acute lung inflammation. It was observed that the level of cytokines pro-inflammatory (IL-17, IL-6, IL-2, IL-4, INF-γ) in serum was normalized in mice treated with ICE, which would indicate that polyP prevents the local inflammatory response in the respiratory tract. The potential application of ICE from L. rhamnosus CRL1505 as a novel bioproduct for the treatment of respiratory diseases is one of the projections of this work. Fil: Correa Deza, Maria Alejandra. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina Fil: Rodríguez de Olmos, Antonieta. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina Fil: Suárez, Nadia Elina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina Fil: Font, Graciela Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina Fil: Salva, Maria Susana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina Fil: Gerez, Carla Luciana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina

BACKGROUND: Gastrointestinal dysfunction is a frequent and disabling manifestation of autoimmune polyendocrine syndrome type 1 (APS-1), a rare monogenic multi-organ autoimmune disease caused by the loss of central AIRE-controlled immune tolerance. OBJECTIVE: We aimed to understand the role of the gut microbiome in APS-1 symptoms and potentially alleviate common gastrointestinal symptoms by probiotic intervention. METHODS: We characterized the fecal microbiomes of 28 APS-1 patients and searched for associations with gastrointestinal symptoms, circulating anti-cytokine autoantibodies and tryptophan-related metabolites. We additionally administered daily doses of the probiotic Lactobacillus rhamnosus GG for three months. RESULTS: Of 581 metagenomic operational taxonomic units (mOTUs) characterized in total, 14 were significantly associated with APS-1 compared to healthy controls, with six mOTUs depleted and eight enriched in APS-1 patients. Four overabundant mOTUs were significantly associated with severity of constipation. We observed phylogenetically conserved microbial associations with autoantibodies against cytokines. After the three-month intervention with the probiotic L. rhamnosus GG, a subset of gastrointestinal symptoms were alleviated. L. rhamnosus GG abundance was increased post-intervention and corresponded with decreased abundances of Alistipes onderdonkii and Collinsella aerofaciens, two species positively associated with severity of diarrhea in APS-1 patients. CONCLUSION: The APS-1 microbiome correlates with several APS-1 symptoms, some of which are alleviated after a three-month L. rhamnosus GG intervention. Autoantibodies against cytokines appear to shape the gut microbiome by positively correlating with a taxonomically consistent group of bacteria. CLINICAL IMPLICATIONS: Administration of L. rhamnosus GG appears to alleviate certain gastrointestinal symptoms of APS-1.

Lacticaseibacillus rhamnosus GR-1 (LGR-1) (previously classified as Lactobacillus rhamnosus GR-1) is the most researched probiotic strain for women's health. Its various urogenital health effects, including a reduction in the recurrence of bacterial vaginosis and urinary-tract infection, are well documented. The strain has also been safely used by HIV-positive subjects, a portion of whom have reported reduced diarrhea and increased CD4 counts. Unlike most probiotic strains used for urogenital health, LGR-1 has been extensively studied for its properties, including its genomic and metabolic traits and its surface properties. This review aims to highlight the totality of research performed with LGR-1, to act as a rigorous scientific benchmark for probiotic microbes, especially for application to women's health.

Imaging flow cytometry (IFC) combines microscopy-based capturing of cell images with conventional flow cytometry-based high-throughput analysis. Thus, IFC approach may limit false-positive results and provide more accurate and unambiguous data. This may be particularly important in the case of the single-cell analysis of relatively small biological objects such as bacteria. The microbial quality control of milk samples is of a special interest during milk processing and several methods have been previously developed to address the presence of bacterial species in milk and other economically important dairy products. However, the vast majority of screening assays is time-consuming and cost-ineffective to be used as a routine test. In the present study, we propose combined acridine orange (AO) staining and IFC-based profiling of microbial species as a novel alternative method for the microbial quality control of milk samples. Eight bacterial profiles (Lactococcus lactis, Lactobacillus rhamnosus GG, Bacillus subtilis, Bacillus cereus, Bacillus megaterium, Staphylococcus aureus, Salmonella typhimurium and Sarcina lutea) were characterized using a number of IFC parameters and IFC approach was also validated using commercially available milk samples and probiotic supplement. The obtained results suggest that IFC-based microbial profiling may be considered as a potentially useful method for rapid screening for selected bacterial species in milk samples.

The demand for "preservative-free" food products is rising, and biopreservation seems to be a potential alternative to replace or reduce the use of chemical preservatives. This study's objective was to assess the antifungal activity of lactic acid bacteria (LAB) (n = 98) and the efficacy and applicability of the chosen bioprotective cultures against fungal spoilers in dairy products. First, 14 strains of antifungal strains were preliminarily screened by in vitro tests against Pichia pastoris D3, Aspergillus niger D1, Geotrichum candidum N1, Kluyveromyces marxianus W1, and Penicillium chrysogenum B1 and validated by challenge tests in yogurts, indicating that the fungal-inhibiting activity of LAB was species specific and yogurts fermented with antifungal LAB cultures were more effective in extending the shelf life. Secondly, the chosen 14 LAB strains were identified by the 16SrDNA sequence analysis and carbohydrate fermentation test. The results were as follows: 9 strains were Lactobacillus plantarum , 3 were Lactobacillus paracasei , 1 was Enterococus faecium , and 1 was Lactobacillus rhamnosus. Among them, active L. plantarum N7 was the chosen and studied factor that affects the antifungal activity using the response surface methodology (RSM). Finally, in situ tests were conducted to validate the activity of L. plantarum N7 in actual dairy products (whey beverage). Physicochemical and microbial indices of whey beverage during storage period exhibited that antifungal L. plantarum N7 could slow the fungal growth and be candidates of interest for industrial applications.

Fil: Tarifa, María Clara. Universidad Nacional de Río Negro, CIT Río Negro. Río Negro, Argentina Fil: Piqueras, Cristian Martín. Planta Piloto de Ingeniería Química (UNS-CONICET). Departamento de Ingeniería Química (UNS). Buenos Aires, Argentina Fil: Genovese, Diego Bautista. Planta Piloto de Ingeniería Química (UNS-CONICET). Departamento de Ingeniería Química (UNS). Buenos Aires, Argentina. Fil: Brugnoni, Lorena Inés. Instituto de Ciencias Biológicas y Biomédicas del Sur (UNS-CONICET). Departamento de Biología, Bioquímica y Farmacia (UNS). Buenos Aires, Argentina The main objective of the research was to evaluate the performance of synbiotic delivery systems using pectin microgels on the protection of two probiotic strains (Lactobacillus casei ATCC 393 and Lactobacillus rhamnosus strain GG [ATCC 53103]) to simulated gastrointestinal digestion (GD) and storage conditions (4 ± 1 °C) in a 42 days trial. Microgel particles were prepared by ionotropic gelation method and three variables were evaluated: incubation time (24 and 48 h), free vs encapsulated cells, and presence or absence of prebiotic (commercial and Jerusalem artichoke inulin). Results demonstrated an encapsulation efficiency of 96±4% into particles with a mean diameter between 56 and 118 μm. The viability of encapsulated cells after 42 days storage stayed above 7 log units, being encapsulated cells in pectin-inulin microgels more resistant to GD compared to non-encapsulated cells or without prebiotics. In all cases incubation time influenced the strains' survival. El objetivo principal de la investigación fue evaluar el rendimiento de sistemas simbióticos utilizando microgeles de pectina para la protección de dos cepas probióticas (Lactobacillus casei ATCC 393 y Lactobacillus rhamnosus cepa GG [ATCC 53103]) frente a dos situaciones: simulación de condiciones de digestión gastrointestinal (DG) y bajo almacenamiento (4 ± 1 ° C) a lo largo de 42 días. Las partículas de microgel se prepararon mediante el método de gelificación ionotrópica y se evaluaron tres variables: tiempo de incubación (24 y 48 h), células libres vs encapsuladas y presencia o ausencia de prebióticos (inulina comercial e inulina de topinambur). Los resultados demostraron una eficiencia de encapsulación de los probióticas del 96 ± 4% en partículas con un diámetro medio entre 56 y 118 μm. La viabilidad de las células encapsuladas al termino de los 42 días de almacenamiento se mantuvo por encima de 7 unidades logarítmicas, siendo las células encapsuladas en los sistemas de microgeles pectina-inulina más resistentes a la DG en comparación con las células no encapsuladas o sin prebióticos. En todos los casos, el tiempo de incubación influyó en la supervivencia de las cepas.

Although human clinical studies have suggested probiotic effects on blood glucose levels, knowledge about molecular mechanisms is still scarce. To test the hypothesis that selected Lactobacillus probiotic bacteria could regulate the activity of enterocyte glucose transporters, we aimed to measure in vitro effects of selected Lactobacillus probiotic bacteria on transcription and translation of intestinal glucose transporters sodium-dependent glucose cotransporter 1 (SGLT1) and glucose transporter 2 (GLUT2) as well as transepithelial glucose transport. Lactobacillus plantarum strains (PCS20 and PCS26), Lactobacillus rhamnosus GG (LGG) (ATCC 53103) and Lactobacillus acidophilus (L acidophilus) (ATCC 4356) were co-cultivated with noncarcinogenic porcine enterocytes (CLAB) and human epithelial colorectal adenocarcinoma cells (Caco-2) (ATCC HTB-37). Changes in transcription and expression of SGLT1 and GLUT2 were strain and cell line-specific. In CLAB, LGG was the most potent SGLT1 up-regulator, and PCS26 the most potent down-regulator of GLUT2 transcription, which was also reflected on the protein level. In Caco-2, all tested strains tended to downregulate GLUT2 gene expression, while L acidophilus most effectively reduced GLUT2 protein levels. Statistically significant effect of PCS26 and L acidophilus on GLUT2 molecular and protein levels in CLAB and Caco-2 cell lines, respectively, was also followed by a decreased rate of transepithelial glucose transport. Careful selection of specific Lactobacillus probiotic strains could be used to downregulate glucose absorption in intestinal epithelial cells and thereby could be beneficial as a support treatment of pathologies related to glucose homeostasis.

The present study was designed to bio-fortify the soymilk (per se a B12-free plant food matrix). The PCR-based screening characterized the human fecal samples (4 out of 15 tested) and correspondingly identified novel lactobacilli isolates (n = 4) for their B12 production potential and rest (n = 62) as negative for this attribute. Further, 3 out of the 4 selected strains showed ability for extracellular vitamin production. The most prolific strain, Lactobacillus reuteri F2, secreted B12 (132.2 ± 1.9 μg/L) in cobalamin-free-medium with the highest ratio ever reported (0.97:1.00; extra-: intra-cellular). In next stage, the soymilk was biofortified in situ with B12 during un-optimized (2.8 ± 0.3 μg/L) and optimized (156.2 ± 3.6 μg/L) fermentations with a ∼54-fold increase at Artificial Neuro Fuzzy Inference System based R value of >0.99. The added-nutrients, temperature and initial-pH were observed to be the most important fermentation variables for maximal B12 biofortification. We report Lactobacillus rhamnosus F5 as the first B12 producing (101.7 ± 3.4 μg/L) strain from this species. The cyanocobalamin was extracted, purified and separated on UFLC as nutritionally-relevant B12. Besides, the vitamin was bioavailable in an auxotrophic-mutant. The lactobacilli fermentation is suggested, therefore, as an effective approach for B12 biofortification of soymilk.

Background & Aims In colorectal cancer, approximately 95% of patients are refractory to immunotherapy because of low antitumor immune responses. Therefore, there is an exigent need to develop treatments that increase antitumor immune responses and decrease tumor burden to enhance immunotherapy. Methods The gut microbiome has been described as a master modulator of immune responses. We administered the human commensal, Lactobacillus rhamnosus GG (LGG), to mice and characterized the changes in the gut immune landscape. Because the presence of lactobacilli in the gut microbiome has been linked with decreased tumor burden and antitumor immune responses, we also supplemented a genetic and a chemical model of murine intestinal cancer with LGG. For clinical relevance, we therapeutically administered LGG after tumors had formed. We also tested for the requirement of CD8 T cells in LGG-mediated modulation of gut tumor burden. Results We detected increased colonic CD8 T-cell responses specifically in LGG-supplemented mice. The CD8 T-cell induction was dependent on dendritic cell activation mediated via Toll-like receptor-2, thereby describing a novel mechanism in which a member of the human microbiome induces an intestinal CD8 T-cell response. We also show that LGG decreased tumor burden in the murine gut cancer models by a CD8 T-cell–dependent manner. Conclusions These data support the potential use of LGG to augment antitumor immune responses in colorectal cancer patients and ultimately for increasing the breadth and efficacy of immunotherapy., Graphical abstract

The impact of secondary polysaccharide, i.e., low methoxyl pectin (LMP) or κ-carrageenan (KC), and its concentration (0.2, 0.4, and 0.6%) on particle size, shape, morphological, textural properties and swelling behavior of sodium alginate (ALG)- based double-network hydrogel particles, as well as the viability of encapsulated probiotics Lactobacillus rhamnosus GG (LGG) in simulated sequential gastrointestinal (GI) digestion was investigated. We found the addition of LMP impaired the sphericity of double-network hydrogel particles, while the incorporation of KC increased the particle size. The FT-IR results indicated the miscibility and cross-linking capacity of the two polysaccharides in forming double-network hydrogel particles. With respect to the swelling behavior in simulated GI digestion, all hydrogel particles shrank in simulated gastric fluid (SGF) but swelled in simulated intestinal fluid (SIF). Among the two types of double-networking, ALG-KC hydrogel particles showed noticeable shrank in SGF in conjunction with the reduced swelling in SIF, which was unfavorable for protection and the controlled release of probiotics. In the case of death rate of encapsulated LGG, the presence of LMP at a lower level (0.2 or 0.4%) exhibited protective effect against LGG death during the sequential GI digestion, while addition of KC demonstrated an opposite role.

Bacteriocins are a subclass of antibacterial peptides considered to be the most promising alternative to antibiotics. A large number of unknown bacteriocins are hidden in lactic acid bacteria. In this study, by combining the genome with LC-MS/MS, 14 novel bacteriocins produced by Lactobacillus rhamnosus LS-8 were detected. Moreover, these bacteriocins were successfully cloned via plasmid pET-28a(+) and pET-30a(+) and heterologously expressed in Escherichia coli BL21. Escherichia coli ATCC25922 and Staphylococcus aureus ATCC25923 were used to confirm their antibacterial activity. Subsequently, the four bacteriocins (pH 25, S68, S81, and S137) with the strongest antibacterial ability were selected, and their expression conditions were optimized. Purification was performed by cation exchange chromatography and high performance liquid chromatography, and the active parts were collected and analyzed by mass spectrometry. The mass spectrometry analysis revealed that peptide coverage was >71.39%. The MICs of the four bacteriocins against four pathogenic bacteria ranged from 5.38 to 19.84 μg/mL. In addition, these bacteriocins significantly inhibited the growth of four standard pathogenic bacteria. They also exhibited broad-spectrum bacteriostasis on Gram-positive and Gram-negative bacteria. Therefore, these new bacteriocins have great potential in the study of alternative antibiotics.

Aflatoxins are common fungal toxins in foods that cause health problems for humans. The aim of this study was to use Saccharomyces cerevisiae and Lactobacillus rhamnosus cell walls immobilized on nano-silica entrapped in alginate as aflatoxin M1 (AFM1) binders. In this study, microbial walls were disrupted using a three-step mechanical technique including autoclave, thermal shock, and ultrasound. Dynamic light scattering (DLS) results proved size reduction in microbial walls ranging 75.8–91.4 nm. Disrupted walls were immobilized on nano-silica to enhance the efficiency of AFM1 adsorption. Then, to prevent the release of the nano-silica or cell walls into the reaction medium, they were entrapped into alginate gel beads. Fourier transform infrared spectrometer (FT-IR) and scanning electron microscopy (SEM) micrographs confirmed the immobilization and entrapment process. Individual and mixtures of free cell walls, immobilized-entrapped walls, alginate bead and nano-silica were contacted with AFM1 for 15 min and 24 h. AFM1 reduction ability was evaluated using high performance liquid chromatography (HPLC). The results showed an AFM1 reduction ranging 53–87% for free cell walls mixture at 15 min and alginate bead respectively. Also, it was possible to reuse immobilized-entrapped walls as binders with an efficiency of about 85%.

Major depressive disorder is a stress-related disease associated with brain metabolic dysregulation in the glutamine-glutamate/γ-aminobutyric acid (Gln-Glu/GABA) cycle. Recent studies have demonstrated that microbiome-gut-brain interactions have the potential to influence mental health. The hypothesis of this study was that Lactobacillus rhamnosus JB-1 (LR-JB1™) dietary supplementation has a positive impact on neuro-metabolism which can be quantified in vivo using magnetic resonance spectroscopy (MRS). A rat model of depressive-like disorder, chronic unpredictable mild stress (CUMS), was used. Baseline comparisons of MRS and behavior were obtained in a control group and in a stressed group subjected to CUMS. Of the 22 metabolites measured using MRS, stressed rats had significantly lower concentrations of GABA, glutamate, glutamine + glutathione, glutamate + glutamine, total creatine, and total N-acetylaspartate (tNAA). Stressed rats were then separated into 2 groups and supplemented with either LR-JB1™ or placebo and re-evaluated after 4 weeks of continued CUMS. The LR-JB1™ microbiotic diet restored these metabolites to levels previously observed in controls, while the placebo diet resulted in further significant decrease of glutamate, total choline, and tNAA. LR-JB1™ treated animals also exhibited calmer and more relaxed behavior, as compared with placebo treated animals. In summary, significant cerebral biochemical downregulation of major brain metabolites following prolonged stress were measured in vivo using MRS, and these decreases were reversed using a microbiotic dietary supplement of LR-JB1™, even in the presence of continued stress, which also resulted in a reduction of stress-induced behavior in a rat model of depressive-like disorder.

Over the past decade there has been increasing interest in the involvement of the microbiota-gut-brain axis in mental health. However, there are major gaps in our knowledge regarding the complex signaling systems through which gut microbes modulate the CNS. The immune system is a recognized mediator in the bidirectional communication continuously occurring between gut and brain. We previously demonstrated that Lactobacillus rhamnosus JB-1 (JB-1), a bacterial strain that has anxiolytic- and antidepressant-like effects in mice, modulates the immune system through induction of immunosuppressive T regulatory cells. Here we examined a potential causal relationship between JB-1 induced regulatory T cells and the observed effects on behaviour. We found that depletion of regulatory T cells, via treatment with monoclonal antibody against CD25, inhibited the antidepressant- and anxiolytic-like effects induced by 4-week oral administration of JB-1 in mice. Ly6Chi monocytes were found to be decreased in JB-1 fed mice with intact regulatory T cells, but not in JB-1 fed mice following depletion. Furthermore, adoptive transfer of CD4+CD25+ cells, from JB-1 treated donor mice, but not from controls, induced antidepressant- and anxiolytic-like effects in recipient mice. Ly6Chi monocytes were also significantly decreased in mice receiving CD4+CD25+ cells from JB1 fed donors. This study identifies cells within the CD4+CD25+ population, most likely regulatory T cells, as both necessary and sufficient in JB-1-induced antidepressant- and anxiolytic-like effects in mice, providing novel mechanistic insight into microbiota-gut-brain communication in addition to highlighting the potential for immunotherapy in psychiatric disorders.

It is widely accepted that regulating microbiome could improve human health. We previously observed apple polysaccharide (AP) reversed high-fat-induced microbial dysbiosis, but the mechanism remains to be elucidated. In this study, the function of AP in vitro was evaluated in Bifidobacterium longum (B. longum) and Lactobacillus rhamnosus (L. rhamnosus). The effects of AP on the composition of fecal bacteria of normal SD rats were investigated by qPCR, TA cloning and 16S sequencing. 0.125-2% AP showed no significant effect on the growth of B. longum and L. rhamnosus. DNA concentration of fecal bacteria cultured with 1% AP was significantly higher than that of control group. qPCR revealed that the number of Bifidobacterium and Lactobacillus in fecal flora incubated by 1% AP was significantly higher than that of control group. Three strains of escherichia coli (E. coli) in fecal bacteria were screened out and analyzed. AP can be utilized by one E. coli and the metabolic products of AP could enhance the proliferation of B. longum. These data suggest that AP could promote the growth of B. longum indirectly, and provide another basis to understand the health care function of apple.

Fermented extracts have evolved to be a potential alternative to synthetic chemicals, owing to their anti-inflammatory and anti-bacterial properties. This study intends to assess the potential of fermented Zanthoxylum schinifolium extract for use in biomedical applications. Probiotic bacteria, Lactobacillus rhamnosus A6-5, were deployed as a seed culture for fermentation. The fermented extract showed greater tyrosinase inhibitory activity and reduced melanin production (58.3%) compared with the raw extract. Cytotoxicity assay inferred that 500 μg/mL is the ideal non-toxic concentration with maximum cell viability. In addition, DAPI staining did not show any damage to the chromatin structure of the cells. The anti-aging property of the fermented extract was confirmed by a decrease in IL-6 content. The fermented extract showed lower MIC (40 mg/mL) and MBC (60 mg/mL), indicating greater anti-bacterial activity than the raw extract. The results confirmed that the fermented Z. schinifolium extract has high biomedical properties compared with the raw extract and can be used as an ideal skin whitening agent.

Clostridium difficile is a spore-forming obligate anaerobe that most commonly causes nosocomial disease, such as diarrhea and colitis. Although antibiotic treatment has been used for Clostridium difficile infection (CDI), its use can also lethally cause antibiotic-associated CDI with the risk of antibiotic resistance. Recently, to avoid these problems, probiotics therapies for CDI have been introduced and studied. However, the molecular mechanisms of C. difficile induced by probiotics, such as Lactobacillus rhamnosus GG (LGG), and associated pathways, have rarely been studied. Here, we co-cultured C. difficile and LGG, which is a promising candidate for treating CDI, in a transwell platform to profile metabolic and proteomic changes of C. difficile with mass spectrometric methods. In the co-cultured condition with LGG, energy generation pathways related to C. difficile growth, such as Stickland reactions and butyrate metabolism, were significantly inhibited, resulting in the decrease of growth rate. In particular, the inhibition of Stickland reactions had a negative effect on the toxin production of C. difficile. In addition, metabolic changes in purine biosynthesis that is known to play an important role in the life of bacteria were observed. Finally, we briefly discussed proteins involved in major iron uptake and electron transfer, as well as the chaperone proteins in bacteria that showed changes by LGG. These results demonstrate that metabolic alterations in C. difficile by LGG caused growth inhibition. Our in vitro co-culture model and multi-omics approach will not only provide better understanding of pathogen-probiotics interaction, but will also contribute to functional study, such as probiotics screening or probiotics engineering, for the therapy of disease caused by pathogenic bacteria.

SUMMARY: The primary purpose of this study was 2-fold. First, it was important to investigate the effects of short-term feeding of Lactobacillus species (plantarum, paracasei, and rhamnosus) via water on nitrogen compounds in manure and blood serum. Second, the effects of sunflower seed meal (SFM) on the same compounds were investigated. Components measured were: ammonia (NH3) emissions from manure, and ammonium nitrogen (NH4-N), total Kjeldahl nitrogen (TKN), and total nitrogen (TN) concentrations in the manure. Biological components of serum included NH3, uric acid, blood urea nitrogen, total protein, albumin, and creatine kinase. Probiotics were mixed in chlorinated drinking water at > 23.3 Mil cfu/g product or 3.64 × 106 cfu/L for each Lactobacillus species to produce approximately 70,000,000 cfu/g product or 1.092 × 107 cfu/L in total. Laying hens (65- to 74-wk-old) were randomly assigned to 4 treatments and fed for 4 wk. Diets were a corn-soy based Control, the Control + Probiotics, the Control + 20% SFM + probiotics or the control + 20% SFM. Major effects and interactions were determined for manure and serum measurements at P < 0.05. Probiotics did not significantly affect NH3 emissions, NH4-N, TKN, and TN in manure. SFM significantly reduced NH3 emissions (P < 0.0001), TKN (P < 0.0001), and TN (P < 0.0001) in manure. TKN in manure was significantly decreased across all treatments. All serum measurements remained statistically similar for the major effects of Probiotics and 20% SFM at P < 0.05.

Background & Aims The intestinal epithelium must be resilient to physiochemical stress to uphold the physiological barrier separating the systemic compartment from the microbial and antigenic components of the gut lumen. Identifying proteins that mediate protection and enhancing their expression is therefore a clear approach to promote intestinal health. We previously reported that oral ingestion of the probiotic Lactobacillus rhamnosus GG not only induced the expression of several recognized cytoprotective factors in the murine colon, but also many genes with no previously described function, including the gene encoding proline-rich acidic protein 1 (PRAP1). PRAP1 is a highly expressed protein in the epithelium of the gastrointestinal tract and we sought to define its function in this tissue. Methods Purified preparations of recombinant PRAP1 were analyzed biochemically and PRAP1 antisera were used to visualize localization in tissues. Prap1-/- mice were characterized at baseline and challenged with total body irradiation, then enteroids were generated to recapitulate the irradiation challenge ex vivo. Results PRAP1 is a 17-kilodalton intrinsically disordered protein with no recognizable sequence homology. PRAP1 expression levels were high in the epithelia of the small intestine. Although Prap1-/- mice presented only mild phenotypes at baseline, they were highly susceptible to intestinal injury upon challenge. After irradiation, the Prap1-/- mice showed accelerated death with a significant increase in apoptosis and p21 expression in the small intestinal epithelium. Conclusions PRAP1 is an intrinsically disordered protein highly expressed by the gastrointestinal epithelium and functions at exposed surfaces to protect the barrier from oxidative insult., Graphical abstract

The formulation of bioactive powders is crucial to their property, functionality and bioaccessibility. This study investigated the incorporation of Ca2+ as a protectant aid for producing active dry probiotics via spray drying, and explored the mechanism underlying the positive effect of Ca2+ on cell viability. At inlet temperature of 98 °C and outlet temperature of 65 °C, supplementing 1 mM CaCl2 to either 10 wt% lactose or 10 wt% trehalose solution increased the survival ratio of Lactobacillus rhamnosus GG (LGG) from 5% to 30%. The growth capability of dried LGG was also enhanced in the presence of calcium. Powder properties including particle morphology, amorphous state and infrared spectrum were minimally influenced at such a low Ca2+ loading. By contrast, adding 1 or 100 mM Ca2+ in reconstitution solution or MRS agar plates notably improved the survival of LGG cells during single droplet drying with pure lactose protectant. The findings suggested that the protective mechanism of Ca2+ is based on enhancing the intrinsic tolerance of LGG cells, rather than modifying the powder properties. Given the protective effect of Ca2+ on probiotic cells under stressful conditions, calcium-containing dry probiotic products that confer duel benefits to consumers may be developed as new types of functional foods.

The usefulness, the high production rate and the cost effectiveness make tablets the dosage form of choice for oral probiotics. Nevertheless, probiotic bacteria undergo a lot of mechanical stress during tableting which causes damage to their cell wall and membrane and other bio-active components. This can lead to an inactivation of the probiotic bacteria and therefore in a failure of the probiotic therapy. To obtain a tablet with a sufficient amount of viable cells, research on the influence of formulation and process parameters on bacterial survival is essential. This study aimed to decipher tableting properties of the probiotic powder blends that have a major impact on survival rates. The powder blends consisted of the prototype probiotic strain Lactobacillus rhamnosus GG, a filler-binder and a suitable amount of lubricant. They were manufactured by direct compression at different compression pressures and tableting speeds. The tableting properties were analysed in detail by a 3-D modelling technique, which characterized normalized time, pressure and displacement simultaneously. The results of the 3-D modelling demonstrated the significant effect of the pressure plasticity (e) and the angle of rotation (ω) on the viability of L. rhamnosus GG during direct compression.

Objectives High fructose feeding changes fibroblast growth factor 21 (FGF21) regulation. Lactobacillus rhamnosus GG (LGG) supplementation reduces fructose-induced non-alcoholic fatty liver disease (NAFLD). The aim of this study was to determine the role of FGF21 and underlying mechanisms in the protective effects of LGG. Methods FGF21 knockout (KO) mice and C57BL/6 wild type (WT) mice were fed 30% fructose for 12 weeks. LGG was administered to the mice in the last 4 weeks during fructose feeding. FGF21-adiponectin (ADPN)-mediated hepatic lipogenesis and inflammation were investigated. Results FGF21 expression was robustly increased after 5-weeks of feeding and significantly decreased after 12-weeks of feeding in fructose-induced NAFLD mice. LGG administration reversed the depressed FGF21 expression, increased adipose production of ADPN, and reduced hepatic fat accumulation and inflammation in the WT mice but not in the KO mice. Hepatic nuclear carbohydrate responsive-element binding protein (ChREBP) was increased by fructose and reduced by LGG, resulting in a reduction in the expression of lipogenic genes. The methylated form of protein phosphatase 2A (PP2A) C, which dephosphorylates and activates ChREBP, was upregulated by fructose and normalized by LGG. Leucine carboxyl methyltransferase-1, which methylates PP2AC, was also increased by fructose and decreased by LGG. However, those beneficial effects of LGG were blunted in the KO mice. Hepatic dihydrosphingosine-1-phosphate, which inhibits PP2A, was markedly increased by LGG in the WT mice but attenuated in the KO mice. LGG decreased adipose hypertrophy and increased serum levels of ADPN, which regulates sphingosine metabolism. This beneficial effect was decreased in the KO mice. Conclusion LGG administration increases hepatic FGF21 expression and serum ADPN concentration, resulting in a reduced ChREBP activation through dihydrosphingosine-1-phosphate-mediated PP2A deactivation, and subsequently reversed fructose-induced NAFLD. Thus, our data suggest that FGF21 is required for the beneficial effects of LGG in reversal of fructose-induced NAFLD., Highlights • Lactobacillus rhamnosus GG (LGG) attenuates fructose-induced NAFLD. • LGG increases FGF21 and adiponectin expression. • LGG inhibits fructose-activated ChREBP and reduces hepatic lipogenesis. • FGF21 is required for the therapeutic effects of LGG against fructose-induced NAFLD.

Background and aims Probiotic supplementation improves glycemic control in persons with diabetes and the current study examined whether these benefits extend to healthy individuals. Methods The current study was a 90-day placebo-controlled, double-blind, randomized clinical trial of Lactobacillus rhamnosus GG in healthy middle-aged and older adults. Fasting blood glucose and HbA1c were quantified at baseline and follow up. Results ANCOVA controlling for baseline values showed group differences in follow up HbA1c [F (1,90) = 8.44, p = 0.005]; HbA1c values increased in the placebo group, though remained stable in the probiotic group. Conclusions If replicated, Lactobacillus rhamnosus GG may protect against changes in glycemic control.

Lactobacilli rhamnosus has been characterized as a probiotic and plays an important role in human health by stimulating the supplement of nutrients, preventing the colonization of pathogens, and influencing the immune system. This study investigated the anticancer activity of the three Lactobacillus rhamnosus strains SHA111, SHA112, and SHA113 isolated from human breast milk. The cell-free supernatant of a liquid culture of the three strains showed excellent antioxidant activities against DPPH free radicals, superoxide anion radicals, and hydroxyl radicals; furthermore, significant anticancer activity was found on cervix cancer cells (HeLa) via cytotoxicity and induction of apoptosis. RT-qPCR and western blot analysis showed the induction of apoptosis was achieved via the up-regulation of BAD, BAX, Caspase3, Caspase8, Caspase9, and down-regulation of BCL-2 genes in HeLa cells. The results suggest that these strains have potential anticancer capability.

Several factors might impact the proteolysis during cheese manufacture and ripening and, therefore, the release of bioactive peptides. These factors include the heat treatment of the milk, the type of starter and secondary culture used and the ripening time. Thus, the objective of this study was to evaluate the effect of the milk heat treatment and the use of adjunct culture in the development of the peptide profile of Camembert-type cheese during ripening. The cheeses were made from raw and heat-treated milk, with and without the addition of Lactobacillus rhamnosus GG. The results obtained by mass spectrometry (MALDI ToF/MS) and analyzed by chemometrics (PLS-DA) revealed a complex hydrolysis profile of the caseins with 103 peaks found, of which 70 peptides were identified and 15 presented bioactive potential. The potential bioactive peptides important for the separation of cheeses were all derived from β-casein. The heat treatment of the milk, the addition of the adjunct culture and the ripening time affected the peptide profile of the cheeses. At the beginning of ripening the cheeses presented a very similar peptide profile, which differed over time, and this differentiation is clearer for cheeses obtained from raw milk.

The present work was designed to investigate the effect of bound polyphenols on antioxidant and prebiotics properties of carrot dietary fiber (CDF). The polyphenols were removed from CDF by alkaline extraction to obtain dephenolized dietary fiber (CDF-DF). The optimized conditions of extracting polyphenols from CDF were solid-liquid ratio of 1:8.5, the treat temperature of α-amylase 66 °C, the addition of protease of 0.2% and the treat temperature of amyloglucosidase 60 °C, respectively. The FT-IR spectroscopy illustrated that the major functional group of CDF-DF and CDF shared most of the key features, despite changes in the amplitude of some functional groups. Scanning electron microscopy (SEM) was further performed demonstrating no significant structure differences between CDF-DF and CDF. Moreover, the antioxidant properties, including superoxide anion radical, hydroxyl radical and DPPH radical scavenging activity, of CDF-DF were significantly lower than that of CDF. In addition, CDF was more conducive for the growth of Lactobacillus rhamnosus as compared with the CDF-DF. In summary, the above results concluded that there was no significant change of the dietary fiber after removal of bound polyphenols, and the bound polyphenols contributed significantly to the antioxidant and prebiotics properties of dietary fiber from carrot.

This study explored the antagonistic activity of Lactobacillus rhamnosus GG (LGG) (Culturelle®), a commercial probiotic supplement, on foodborne pathogens and its heat sensitivity in soup products. Staphylococcus aureus, Escherichia coli O157:H7, Escherichia coli O104:H4, L. monocytogenes and S. Typhimurium were selected to assess the inhibitory effect of LGG on them and analyze their sensitivities towards LGG. Cream of potato (CP) and vegetable beef (VB) soups were selected as food matrix to determine thermal resistance of LGG between 55 and 62.5 °C. The effectiveness of LGG as a protective culture against the growth of Listeria monocytogenes and Salmonella Typhimurium was evaluated at conditions simulating the commercial application of cook-chill processing in CP soup and stored at 5, 10 and 15 °C for 21 days. Overall, LGG showed inhibitory effect against all pathogens tested on spot agar test. E. coli O157:H7 was the most sensitive, whereas S. Typhimurium was the least sensitive to the inhibitory effect of LGG. D-values obtained in CP soup exceeded those obtained in VB soup at the temperatures tested. D55-62.5 values of LGG ranged between 0.7-24.2 min and 0.5–15 min in CP and VG soup, respectively. The z-value for CP and VB soup was around 5 °C. LGG co-inoculated with either target bacterium led to a reduction in S. Typhimurium or L. monocytogenes to undetectable levels. The probiotic population increased about 1-log unit at 15 °C. At 10 and 5 °C, it was maintained throughout the experiment at around 108 CFU g−1. Results demonstrated the potential of LGG as protective culture in controlling S. Typhimurium and L. monocytogenes growth in cream of potato soup.

Summary: Background & aims: Tungrymbai is a fermented soy product which is a sticky food that exhibits unique flavour and texture which is exclusively made by the local Khasi tribes of Meghalaya (India). A similar kind of product was prepared under laboratory conditions followed by shelf-life study and analysis of its bio-functional attributes with comparison to the traditionally made product. Methods: The traditional practice for the preparation of Tungrymbai was closely monitored and a similar product preparation was made in laboratory conditions using indigenous Lactobacillus cultures. The shelf life study (physicochemical, microbial and organoleptic attributes) was conducted for 4 days for the Tungrymbai products. Further, the bio-functional properties such as ACE-inhibitory, antioxidative activity and biotransformation of Isoflavones were analysed for laboratory made Tungrymbai products with inoculation of Lactobacillus cultures as well as without inoculation (control). Results: Out of all the quality attributes of the various Tungrymbai samples, Tungrymbai with 1% cell biomass of Lactobacillus culture in combinations was mostly preferred by the sensory panellists than other samples with a sensory score >6. ACE inhibitory activity of Tungrymbai prepared under laboratory condition increased from 30.75% at day 0–83.55% at day 5, followed by a reduction after day 10 (36.98%) and the antioxidative activity was maximum at day 0 (99.14%) followed by a decrease at day 2 (81.01%). After fermentation of 4 days, production of soy isoflavones in agyclones were 34.70% genistein and 29.37% daidzein for traditionally made Tungrymbai and 55.44% genistein and 86.58% daidzein for laboratory made Tungrymbai infused with Lactobacillus cultures. Conclusion: Hence, the Lactobacillus culture combinations of Lactobacillus rhamnosus K4E and L. helveticus K14 could serve as potential candidates for the preparation of novel ethnic fermented foods in Meghalaya with more improved bio-functionality. Keywords: Tungrymbai, Lactobacillus, ACE-inhibition, Antioxidation, Isoflavones

Enzymatic NADP+ regeneration is a promising approach to produce valuable chemicals under economic conditions. Among all the enzymatic routes, using water-forming NADH oxidase is an ideal one because there is no by-product. However, most NADH oxidases have a low specific activity to NADPH. In this work, a thermostable NADH oxidase from Lactobacillus rhamnosus (LrNox) was rationally engineered to switch its specificity from NADH to NADPH. The results show that mutants D177A, G178R, D177A/G178R, D177A/G178R/L179S improved the NADPH activity by a factor of 4–6. The highest NADPH catalytic efficiency (Kcat/Km 223.71 S−1 μm−1, 47.6-fold higher than wild-type LrNox) and 51% of NADH activity retention were achieved by replacing the single amino acid Leu179 for serine (L179S) in LrNox. Modeling of L179S-NADPH complex reveals that the phosphate group of NADPH interacts with the hydroxyl of Ser179 with a strong hydrogen bond and several shorter hydrogen bonds with the amino group of Lys185 could stabilize the binding of NADPH in the L179S mutant. This work provides an efficient method for converting NAD(P)H specificity and shows that L179S mutant is a potential and efficient auxiliary enzyme for NADP+ regeneration.

Four lactic acid bacteria strains (Lactobacillus paracasei, Leuconostoc mesenteroides, Lactobacillus rhamnosus GG and Lactobacillus plantarum), possessing β-glucosidase enzyme activity, were investigated for tofu whey fermentation based on the capacity of bioconversion of isoflavones. The results suggested that the mixture of L. rhamnosus GG and L. paracasei had great potential for efficient enrichment of bioactive isoflavone aglycones and the percentage composition (daidzein, genistein) increased from 4.6 to 93.78% after fermentation of tofu whey. At the same time, the viable cell counts of the mixture of L. rhamnosus GG and L. paracasei increased to 9.13 log10 CFU/mL while pH value dropped to 4.48 and titratable acidity was 0.28%. Headspace solid-phase microextraction method followed by gas chromatography-mass spectrometry analysis indicated that the odor compounds, particularly the aldehydes and alcohols, were metabolized to trace or undetected levels after fermentation. This research provided not only a new way to develop a functional probiotic beverage enriched in isoflavone aglycones, but also provided a potentially valuable new use for soy whey protein.

Minimally processed fruits are an alternative to dairy products to deliver probiotics. Bio-protection against several factors that affect their viability has been proposed in the food industry. In this study, probiotic Lactobacillus rhamnosus CECT 8361 was added to alginate-based coatings enriched with inulin and oligofructose and applied on fresh-blueberries. Probiotic viability, microbiological, physicochemical and sensory quality parameters of blueberries were monitored during 21 days of refrigerated storage. Also, L. rhamnosus CECT 8361 was tested for its antagonistic effect against inoculated Listeria innocua and E. coli O157:H7. Advantageously, prebiotic compounds allowed improving probiotic viability with counts above 6.2 log CFU/g for the entire period. Native microbiota counts remained under safe levels. Overall visual quality, odor and flavor were acceptable up to day 14 of storage. Regarding antimicrobial activity, L. rhamnosus CECT 8361 was able to reduce L. innocua counts by 1.7 log in inoculated blueberries. These results encourage further implementation of new fruit-based foods with multifunctional properties. Fil: Bambace, María Florencia. Universidad Nacional de Mar del Plata; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata; Argentina Fil: Alvarez, María Victoria. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Moreira, María del Rosario. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

Early life stress (ELS) can affect brain development and increase lifetime prevalence of psychiatric illnesses. However, the effective therapeutic interventions to ameliorate the deleterious effects of ELS have not yet been well established. Here, we confirmed that maternal separation (MS) for 3 h daily between postnatal days 2–14, a frequently used experimental model of ELS, resulted in early expression of adult-like fear memory retention in male infant rats. Administration of a probiotic formulation, Lacidofil® (95% Lactobacillus rhamnosus R0011 and 5% Lactobacillus helveticus R0052), during the separation period, prevented the precocious transition to adult-like fear memory retention in MS infant rats. Consonant with this effect, probiotic treatment also ameliorated the MS-induced increases in anxiety-like behavior as measured by the elevated plus maze and the light-dark box tests. In addition, probiotic treatment reduced MS-induced increases in neuronal activation and brain-derived neurotrophic factor protein levels in the basolateral nucleus of amygdala (BLA) after auditory fear conditioning. Furthermore, we found that probiotic treatment significantly rescued the heightened hypothalamic-pituitary-adrenal (HPA) axis response to restraint stress in MS infant rats. Taken together, these findings suggest that probiotics can restore normal developmental trajectories of fear memory retention in MS infant rats, at least in part by normalizing HPA axis abnormalities, and that the BLA serves as a critical node to mediate these interventions. Thus, we offer a potential therapeutic intervention to protect children against the harmful effects of ELS.

Cysteine, a critical residue for catalytic process but also vulnerable to oxidative damage, was conventionally expressed as a buried catalytic site in most redox enzymes. In the present work, specific surface-exposed sites of a NADH oxidase from Lactobacillus rhamnosus (LrNox) were selected and mutated to cysteine to investigate its effects on catalytic function because LrNox has a buried catalytic cysteine but no surface-exposed one. The results showed that exception of the sites on dimer interface, the activities of LrNox mutants were improved to vary degrees when the polar uncharged and alanine residues were mutated to cysteine. But the cysteine mutations of polar charged and nonpolar residues except alanine showed obvious decline in catalytic activity. Substituting of Ala85 and Thr96 with other residues suggested that the cysteine mutation showed the highest activity. Structural analysis suggested that even a single cysteine mutation on the specific non-conserved surface area of LrNox could induce changes on the conformation of catalytic cysteine and lower the activation free energy to improve the catalytic activity.

Microbiota in the gut affect brain physiology via various pathways, and dysbiosis seems to play a role in the pathogenesis of Parkinson's disease (PD). Probiotics showed pleiotropic effects on functions of the central nervous system via microbiota-gut-brain axis. However, no studies displayed the neuroprotective effects of probiotics in the Parkinson's disease. This study aimed to test the neuroprotective effects of probiotics in two different models of PD. We evaluated neuroprotective effects of a probiotic cocktail containing Lactobacillus rhamnosus GG, Bifidobacterium animalis lactis, and Lactobacillus acidophilus in PD models induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or rotenone utilizing behavioral tests, immunohistochemistry and neurochemical analysis. To assure the neuroprotection came from increased production of butyrate, we further determined beneficial effects of butyrate in the MPTP-mediated PD model. The probiotic mixture overtly protected the dopaminergic neurons against MPTP neurotoxicity. However, the probiotics downregulated expression of monoamine oxidase (MAO) B in the striatum, which was accompanied by a lower level of 1-methyl-4-phenylpyridinium (MPP+), the main neurotoxic metabolite of MPTP. Thus, we extended the investigation into the rotenone-induced PD model. Rescuing effects of the probiotics were observed in the setup, which came with increased levels of neurotrophic factors and butyrate in the brain. Lactobacillus rhamnosus GG was identified to be a major contributor to the induction of neurotrophic factors and downregulation of MAO B. Finally, we demonstrated that sodium butyrate attenuated MPTP-induced neuronal loss in the nigrostriatal pathway. Probiotics could ameliorate neurodegeneration at least partially by increasing butyrate level. These data highlight the role of probiotics for brain health, and their potential as a preventive measure for neurodegenerative diseases such as PD.

Dynamics of probiotics Lactobacillus rhamnosus LGG, L. reuteri BioGaia, Bifidobacterium animalis subsp. lactis BB12 and B. longum subsp. longum BB536 were investigated during the fermentation of milk supplemented with extracts of Chondrus crispus (CC), Himanthalia elongata (HE), Laminaria ochroleuca (LO), Palmaria palmata (PP), Porphyra umbilicalis (PU), Ulva lactuca (UL) and Undaria pinnatifida (UP). After 24 h at 37 °C, LGG attained counts above 108 cfu/ml in control milk and in milk with all seaweed extracts while BioGaia only attained that level in milk with CC, HE, PP, PU and UL extracts, B12 in milk with UL and UP extracts, and BB536 in milk with CC, HE, PU and UL extracts. In physiological saline, LGG, BioGaia, BB12 and BB536 were respectively stimulated by 4, 0, 5 and 3 seaweed extracts after 24 h at 37 °C and inhibited by 1, 5, 2 and 0 seaweed extracts. Casein and whey proteins added to physiological saline neutralized inhibition caused by UL extract, increasing probiotic counts by 4–5 log cfu/ml, while lactose and citrate had no significant effect. Milk supplementation with seaweed extracts permitted attaining probiotic counts above 108 cfu/ml at the end of fermentation, a result of practical interest for the dairy industry.

Camembert is a white-mould ripened cheese in which intense proteolysis, caused mainly by the fungus Penicillium candidum, among other factors, leads to a substantial production of aromatic and flavor compounds. The aim of this study was to evaluate the sensory profile of the Camembert-type cheese made from raw milk and from pasteurized milk with or without the addition of Lactobacillus rhamnosus GG. The cheeses were analyzed after 50 days of ripening through a combination of Flash-Profile (n = 15) and Check-All-That-Apply (CATA) (n = 80) sensory methods with global acceptance. The 51 sensory attributes obtained allowed the separation of the samples in three clusters. Through the CATA, a significant difference (p

In this study, cream cheese is hypothesized as an alternative dairy product to deliver the probiotic bacteria. The survival of non-encapsulated and encapsulated probiotic L. rhamnosus in the functional cream cheese was studied over 35 days of storage at 4 °C. Further, the microstructure and textural properties of the cream cheese were investigated. L. rhamnosus in both forms remained viable (min 106 CFU/g) in the cream cheese throughout the storage period. Probiotic cream cheese with β-glucan and phytosterol emulsion showed less reduction in their viable counts after 35 days of refrigerated storage. The addition of probiotics either in the non-encapsulated or encapsulated forms did not significantly change the pH, moisture, protein or fat content. The results from SEM analysis indicated that the structure of cream cheese with fat replacers (β-glucan and phytosterol) was open with the probiotic cells distributed in the casein matrix. The addition of encapsulated probiotic led to a firmer and thicker cream cheese compared to non-encapsulated form that also corroborated through the SEM images showing a relatively denser structure for cream cheese with encapsulated probiotic cells.

BACKGROUND: Acute gastroenteritis develops in millions of children in the United States every year, and treatment with probiotics is common. However, data to support the use of probiotics in this population are limited. METHODS: We conducted a prospective, randomized, double-blind trial involving children 3 months to 4 years of age with acute gastroenteritis who presented to one of 10 U.S. pediatric emergency departments. Participants received a 5-day course of Lactobacillus rhamnosus GG at a dose of 1×10(10) colony-forming units twice daily or matching placebo. Follow-up surveys were conducted daily for 5 days and again 14 days after enrollment and 1 month after enrollment. The primary outcome was moderate-to-severe gastroenteritis, which was defined as an illness episode with a total score on the modified Vesikari scale of 9 or higher (scores range from 0 to 20, with higher scores indicating more severe disease), within 14 days after enrollment. Secondary outcomes included the duration and frequency of diarrhea and vomiting, the duration of day-care absenteeism, and the rate of household transmission (defined as the development of symptoms of gastroenteritis in previously asymptomatic household contacts). RESULTS: Among the 971 participants, 943 (97.1%) completed the trial. The median age was 1.4 years (interquartile range, 0.9 to 2.3), and 513 participants (52.9%) were male. The modified Vesikari scale score for the 14-day period after enrollment was 9 or higher in 55 of 468 participants (11.8%) in the L. rhamnosus GG group and in 60 of 475 participants (12.6%) in the placebo group (relative risk, 0.96; 95% confidence interval, 0.68 to 1.35; P = 0.83). There were no significant differences between the L. rhamnosus GG group and the placebo group in the duration of diarrhea (median, 49.7 hours in the L. rhamnosus GG group and 50.9 hours in the placebo group; P = 0.26), duration of vomiting (median, 0 hours in both groups; P = 0.17), or day-care absenteeism (median, 2 days in both groups; P = 0.67) or in the rate of household transmission (10.6% and 14.1% in the two groups, respectively; P = 0.16). CONCLUSIONS: Among preschool children with acute gastroenteritis, those who received a 5-day course of L. rhamnosus GG did not have better outcomes than those who received placebo. (Funded by the Eunice Kennedy Shriver National Institute of Child Health and Human Development and others; ClinicalTrials.gov number, NCT01773967.)

With the growing interest in probiotic microorganisms based on their well established immense health benefits, the present investigation was aimed to assess the adhesion potential and safety of probiotic Lactobacillus rhamnosus MTCC- 5897 (LR) before it can be put into a probiotic formulations. L. rhamnosus showed an adhesion index of 166.7 ± 11, which was further confirmed by scanning electron microscopy and relative expression of mucus binding protein (Mub) and mucus adhesion promoting protein (Map-A) genes. In vitro safety assessment by tetrazolium dye reduction, neutral red and lactate dehydrogenase (LDH) release assays revealed unchanged metabolic activity of Caco-2 cells even when incubated with L. rhamnosus ranged between 106-1010 cfu/mL for 24 h. Similarly, a moderate increase in bile salt hydrolase (bsh) expression (6.84 ± 0.73 and 3.42 ± 0.39 folds in 1% and 3% bile medium respectively) further proved its safety towards normal lipid digestion and absorption. Moreover, L. rhamnosus feeding to mice (107, 109, 1011 and 1013 cfu/animal/d) repetitively for 28 days revealed no adverse effects on parameters of general animal health status including body weight, organ indices, plasma glucose, liver malondialdehyde (MDA), serum aspartate amino transaminase (AST), cholesterol, triglycerides, high-density lipoprotein (HDL). Similarly, significant (p ≤ 0.05) reduced activities of serum alanine amino transaminase (ALT) and LDH on continuous probiotic feeding were also indicative of normal liver/kidney functions as they were in normal range for mice. Further, insignificant changes in macrophage chemoattractant protein (MCP-1) in intestinal fluid irrespective of bacterial dose fed along with significant reduction (p ≤ 0.05) of tumor necrosis factor-α (TNF-α) at much higher dose (1013 cfu/animal/d) also confirmed safe response of probotic L.rhamnosus against inflammation. To conclude, the results obtained under in vitro and in vivo studies has established the Lactobacillus rhamnosus as safe and non-toxic to weaning mice as well as human epithelial cells and thus may be used as a safe food additive.

Dry-fermented sausages are an important and abundant component of the diet of many people. Improving their composition and thus their potential health effects is therefore important. Here we quantified the antioxidant capacity, short chain fatty acids (SCFAs) production, and gut microbiota structure of different salami formulations after in vitro digestion and subsequent fermentation with human gut microbiota. The addition of different types of fiber (citrus fiber, arabinogalactan, and inulin), a probiotic Lactobacillus rhamnosus, and an herbal extract to the salami formulation was tested. Incorporating any dietary fiber into salami formulation increased sausage antioxidant capacity and the amount of SCFAs produced during microbiota fermentation. These effects were highest for salami with citrus fiber and citrus fiber with herbal extract. Presence of fiber in salami also led to an altered gut microbiota structure. Citrus fiber and arabinogalactan but not inulin promoted an increase in the abundance of several known polysaccharide degrading genera and resulted in a reduction in the abundance of Escherichia, a bacterial genus known to contain many human intestinal pathogens. Overall, the addition of dietary fiber to salami formulation prior to curing improved beneficial health markers of this food product.

Encapsulation with edible films is a promising approach that may solve the disadvantages associated with the use of bioactive compounds as food additives. This is particularly relevant in the case of probiotics, since their stability in food matrices and in the gastrointestinal tract may be rather poor. Therefore, new cellulose-based edible films have been successfully developed and characterized. Sodium carboxymethyl cellulose (CMC) and hydroxyethyl cellulose (HEC) were used for the film preparation and cross-linked with citric acid (CA) under reasonably mild conditions. Model probiotic bacteria (Lactobacillus rhamnosus GG) were incorporated in the films either during the film formation and casting or after the film synthesis, via bacteria diffusion and adsorption. The later approach could efficiently entrap and preserve viable bacteria. The mechanical properties and swelling ability could be tuned by varying the HEC/CMC ratio and the amount of CA. Moreover, the surface area and total pore volume of the films considerably decreased after cross-linking. Overall, these novel films are regarded as promising inexpensive and friendly matrices for food protection and packaging applications. People Program (Marie Curie Actions) of the European Union's Seventh Framework Program FP7/2007-2013/under REAEuropean Union (EU) [606713 BIBAFOODS] Portuguese Foundation for Science and Technology (FCT)Portuguese Foundation for Science and Technology [PTDC/AGR-TEC/4814/2014, IF/01005/2014] FCT Portuguese Foundation for Science and Technology [PEstOE/QUI/UI0313/2013]

International audience; In order to encourage Western populations to increase their consumption of vegetables, we suggest turning legumes into novel, healthy foods by applying an old, previously widespread method of food preservation: fermentation. In the present study, a total of 55 strains from different microbial species (isolated from cheese or plants) were investigated for their ability to: (i) grow on a emulsion containing 100% pea proteins and no carbohydrates or on a 50:50 pea:milk protein emulsion containing lactose, (ii) increase aroma quality and reduce sensory off-flavors; and (iii) compete against endogenous micro organisms. The presence of carbohydrates in the mixed pea:milk emulsion markedly influenced the fermentation by strongly reducing the pH through lactic fermentation, whereas the absence of carbohydrates in the pea emulsion promoted alkaline or neutral fermentation. Lactic acid bacteria assigned to Lactobacillus plantarum, Lactobacillus rhamnosus, Lactococcus lactis and Lactobacillus casei species grew well in both the pea and pea:milk emulsions. Most of the fungal strains tested (particularly those belonging to the Mucor and Geotrichum genera) were also able to grow on both emulsions. Although most Actinobacteria and Proteobacteria did not compete with endogenous microbiota (Bacillus), some species such as Hafnia alvei, Acinetobacter johnsonii and Glutamicibacter arilaitensis grew strongly and appeared to restrict the development of the endogenous microbiota when the pea emulsion was inoculated with a combination of three to nine strains. In the mixed emulsions, lactic fermentation inhibited Actinobacteria and Proteobacteria (e.g. Brevibacterium casei, Corynebacterium casei, Staphylococcus lentos) to the greatest extent but also inhibited Bacillus (e.g. Bacillus subtilis and Bacillus licheniformis). Overall, this procedure enabled us to select two microbial consortia able to colonize pea-based products and positively influence the release of volatile compounds by generating a roasted/grilled aroma for the 100% pea emulsion, and a fruity, lactic aroma for the 50:50 pea:milk emulsion. Moreover, the fermentation in the pea-based emulsions reduced the level of hexanal, which otherwise leads to an undesired green pea aroma. Our present results show how the assembly of multiple microbial cultures can help to develop an innovative food product.

Synbiotics, a synergistic combination of probiotics and prebiotics, are currently regarded as one of the most practical nutritional supplements in tilapia farms. In this study, the effect of supplementing the diet of red tilapia (Oreochromis spp.) with Jerusalem artichoke (Helianthus tuberosus) and Lactobacillus rhamnosus GG (LGG) was evaluated. Growth performance, serum biochemical parameters, intestinal morphology, goblet cell counts, immune parameters and protection against Aeromonas veronii challenge were determined. The results showed that fish fed with synbiotic-supplemented diets had a significantly higher (P

In the present study, mulberry oligosaccharides were produced from mulberry polysaccharide by physical ultrasonic hydrolysis, chemical acid hydrolysis and enzymatic hydrolysis. The impact of the oligosaccharides on the growth four probiotic bacteria—Lactobacillus plantarum, Leuconostoc mesenteroides, Lactobacillus casei and Lactobacillus rhamnosuswas examined. The oligosaccharides produced from β-mannanase hydrolysis had the greatest effect on the growth of L. rhamnosus (252% of the number of cells compared with medium without the hydrolyzate). The enzymatically-prepared mulberry oligosaccharides (EMPS) were superior to the untreated mulberry polysaccharide and commercially available prebiotics (isomaltooligosaccharides and galactooligosaccharides). L. rhamnosus was cultured with EMPS and then subjected to tolerance tests with artificial gastrointestinal fluids. Bacteria remained viable. Subsequently, simulated-digested L. rhamnosus cultured with EMPS was used in in vitro fermentation with human gut microbiota from fecal inoculum. After 48 h of the incubation, in the experimental group (with pretreated L. rhamnosus), the ratio of Bacteroidetes to Firmicutes was 0.848, compared with 0.659 in the control group (saline instead of L. rhamnosus), suggesting that L. rhamnosus and constituents in EMPS were capable of regulating the growth of intestinal microorganisms. Our results inform the potential for the development of mulberry oligosaccharides as a prebiotics or synbiotics.

Probiotic dietary supplementation plays a crucial role in shaping gut microbiota and boost immune-responses. We herein investigated the potential effect of long-term dietary-supplementation of Lactobacillus rhamnosus (LBR) on gut microbiome structure, metabolome functions, and serum-profile. Our results revealed, LBR long-term supplementation reduces the body weight gain, ameliorates serum cytokines and attenuates serum lipoproteins profile. Moreover, LBR enhances biofilms-aerobics. Gut microbiome 16S rRNA analysis revealed LBR intervention significantly increased the relative abundance of Limnobacter, Turicibacterales, Enterococcus, and Vagococcus. On the contrary, the abundance of Helicobacteraceae, Rikenellaceae, Roseburia, Dorea, Anaerostipes, Coprococcus, Lachnospira, Oscillospira, Ruminococcaceae, Clotridiales, S24_7, Bacteroidia and Bacteroidetes found decreased in LBR treated group. Functional comparison revealed LBR exhibited higher level of energy production and conversion, amino acid transport and metabolism functions. Our findings demonstrate that the LBR dietary supplementation found to be beneficially modulating the immune-response, enhances lipoprotein metabolism, and ameliorates the microbial composition structure and functions.

This research verified the behavior of Lactobacillus rhamnosus and Lactobacillus casei added to ice cream in free form or encapsulated with alginate-chitosan by comparing their: (1) resistance to low temperature; (2) encapsulation efficiency and (3) cell survivability in a simulated gastrointestinal environment. Calcium alginate-chitosan capsules were produced by extrusion method. The ice cream samples with probiotics were stored at −18 °C for 150 days, and the pH, titratable acidity, the cell viability of the probiotics and the gastrointestinal simulation were analyzed. Free L. casei (−1.64 log) were more resistant to freezing than free L. rhamnosus (−1.92 log). L. rhamnosus and L. casei presented encapsulation efficiencies of 73.8% and 79.5%, respectively. In the gastrointestinal simulation, 86.2% L. rhamnosus (−0,83 log) and 84% L. casei (−1,3 log) were protected by the alginate-chitosan capsules. The results demonstrated that microcapsules reduce cell loss in frozen storage and in simulated gastrointestinal conditions. The reduction in cell viability of encapsulated L. rhamnosus was superior to that of its free form, showing that the encapsulation process is not advantageous for all probiotic species. Therefore, considering cellular losses, the best option for preparing functional yellow mombin ice cream is to use free L. rhamnosus or encapsulated L. casei.

Lactobacillus rhamnosus GG is an environmentally and human-friendly bacteria that can be found in probiotics. However, it is incapable of forming a biofilm on metal surfaces, what limits the possibility of using it in bioelectrochemical systems (BESs) as promising and green alternatives. Hence, we propose a simple and cost-effective modification of metal surface with cationic dextran derivative (DEX-G) by using the electrostatically driven self-assembly procedure resulting in the formation of a polymeric monolayer. Modification of metal electrodes was confirmed by using infrared spectroscopy (IR) and atomic force microscopy (AFM). Synthesized metal/polycation electrodes provided a bioactive environment for L. rhamnosus GG enhancing its proliferation and formation of the biofilm as confirmed by SEM imaging. Moreover, using cyclic voltammetry (CV) and chronoamperometry measurements, it was proven that metal/polycation/L. rhamnosus bioelectrodes are electrochemically active in the glucose-rich medium. The electrochemical activity of fabricated bioanodes allows to consider them for their potential use in the bioelectrochemical systems.