Modulation of intestinal microbiota and immunometabolic parameters by caloric restriction and lactic acid bacteria

The objective of this work was to evaluate the effect of a caloric restriction diet with and without the administration of Lactobacillus fermentum CRL1446, Lactobacillus casei CRL431 and Lactococcus lactis CRL1434, on immunemetabolic parameters and the composition of intestinal microbiota in mice. The supplementation of the caloric restriction diet with L. fermentum CRL1446 showed a bifidogenic effect and was able to maintain the abundance of the genus Lactobacillus over time. On the other hand, this strain showed hypocholesterolemic and hypoglycemic properties as well as inducing a decrease in plasma leptin levels. L. casei CRL431 administration increased the abundance of the Lactobacillus genera in the intestinal microbiota, which would improve the absorption of nutrients from the diet. This strain restores glucose values decreased by the diet in addition to inducing an increase in leptin and cytokines. Lac. lactis CRL1434 showed greater immunomodulatory capacity, without significantly affecting the composition of the intestinal microbiota. It had hypoglycemic properties and induced a decrease in leptin concentrations. L. fermentum CRL1446 and Lac. lactis CRL1434 could be potentially probiotic strains useful to correct the immuno-metabolic alterations associated with obesity, while L. casei CRL431 is a more suitable strain to be used in cases of malnutrition where it is sought to improve the absorption of nutrients and protection against infections, in addition to the stimulation of the immune system.Fil: Fabersani Marrades, Mario Emanuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán; ArgentinaFil: Russo, Matias Irineo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Márquez, María Antonela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Abeijon Mukdsi, Maria Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad del Norte Santo Tomás de Aquino; ArgentinaFil: Medina, Roxana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán; Argentina. Universidad del Norte Santo Tomás de Aquino; ArgentinaFil: Gauffin Cano, María Paola. Universidad del Norte Santo Tomás de Aquino; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentin

Esterase activities and biochemical properties of lactic acid bacteria isolated from goat´s milk cheese in Argentina

Twenty-two lactic acid bacteria (LAB) strains isolated from Argentinean goat dairy products were evaluated for its biochemical properties and esterase activities relevant to flavor development. Streptococcus thermophiles (UNSE314), Lactobacillus (L.) delbrueckii subsp. bulgaricus (UNSE309), L. rhamnosus (UNSE308), L. plantarum (UNSE287, UNSE316, UNSE317) and Pediococcus pentosaceus (UNSE315) strains presented high acidifying activity. All strains tested metabolized citrate and produced diacetyl-acetoin in goat milk. Based on these results, ten strains with the best performance in diverse technological properties were selected to determine esterolytic activity. In all evaluated strains, esterase specific activity (ESA) was detected on a-naphthyl (a-NA) acetate and B-naphthyl (B-NA) acetate, propionate, caprylate and a-NA butyrate. No activity was detected on B-NA laurate. The highest values were detected when using a-NA instead of B-NA derivatives as substrate. In Pediococcus strains, wide variability in ESA were observed, which were species- and strain-specific. These results allow us to select strains with biochemical properties and esterase activities to design starter and adjunct cultures that contribute to flavor development during cheese ripening, thus preserving the typical organoleptic characteristics of Argentinean goat cheeses. Fil: Taboada, Natalia Verónica. Universidad Nacional de Santiago del Estero. Facultad de Agronomia y Agroindustrias. Instituto de Cs.y Tecnologías Alimentarias; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lopez Alzogaray, Maria Soledad. Universidad Nacional de Santiago del Estero. Facultad de Agronomia y Agroindustrias. Instituto de Cs.y Tecnologías Alimentarias; ArgentinaFil: Abeijon Mukdsi, Maria Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Centro de Referencia Para Lactobacilos; Argentina. Universidad del Norte Santo Tomás de Aquino; ArgentinaFil: Medina, Roxana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucuman. Centro de Referencia Para Lactobacilos; Argentina. Universidad Nacional de Tucumán. Facultad de Agronomía y Zootecnia; Argentin

Draft genome sequence of the feruloyl esterase-producing strain lactobacillus fermentum CRL1446, a probiotic for malnutrition

We report here the draft genome sequence of Lactobacillus fermentum CRL1446 (2,148,781 bp, 51.4% G+C content). This strain exhibits feruloyl esterase activity and important technological and probiotic properties. Because of its proven beneficial effects in vivo, it represents an interesting candidate for the development of functional foods or pharmabiotics for malnutrition.Fil: Abeijon Mukdsi, Maria Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad del Norte Santo Tomás de Aquino; ArgentinaFil: Saavedra, Maria Lucila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Gauffin Cano, María Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad del Norte Santo Tomás de Aquino; ArgentinaFil: Hebert, Elvira Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Medina, Roxana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad del Norte Santo Tomás de Aquino; Argentin

Guanosine diphosphatase is required for protein and sphingolipid glycosylation in the Golgi lumen of Saccharomyces cerevisiae

Current models for nucleotide sugar use in the Golgi apparatus predict a critical role for the lumenal nucleoside diphosphatase. After transfer of sugars to endogenous macromolecular acceptors, the enzyme converts nucleoside diphosphates to nucleoside monophosphates which in turn exit the Golgi lumen in a coupled antiporter reaction, allowing entry of additional nucleotide sugar from the cytosol. To test this model, we cloned the gene for the S. cerevisiae guanosine diphosphatase and constructed a null mutation. This mutation should reduce the concentrations of GDP-mannose and GMP and increase the concentration of GDP in the Golgi lumen. The alterations should in turn decrease mannosylation of proteins and lipids in this compartment. In fact, we found a partial block in O- and N-glycosylation of proteins such as chitinase and carboxypeptidase Y and underglycosylation of invertase. In addition, mannosylinositolphosphorylceramide levels were drastically reduced

Specific Strains of Lactic Acid Bacteria Differentially Modulate the Profile of Adipokines In Vitro

Obesity induces local/systemic inflammation accompanied by increases in macrophage infiltration into adipose tissue and production of inflammatory cytokines, chemokines, and hormones. Previous studies have shown that probiotics could improve the intestinal dysbiosis induced by metabolic diseases such as obesity, diabetes, and metabolic syndrome. Microorganisms could (directly or indirectly) affect adipokine levels due to their capacity to induce translocation of several intestinal microbial antigens into systemic circulation, which could lead to metabolic endotoxemia or produce immunomodulation in different organs. The aim of the present study was to select non-inflammatory lactic acid bacteria (LAB) strains with the capacity to modulate adipokine secretion by the adipose tissue. We wish to elucidate the role of potential probiotic strains in the regulation of the cross talking between immune cells such as macrophages and adipose cells. Mouse macrophage cell line RAW 264.7 was used for evaluating the ability of 14 LAB strains to induce cytokine production. The LAB strains were chosen based on their previously studied beneficial properties in health. Then, in murine adipocyte culture and macrophage-adipocyte coculture, we determined the ability of these strains to induce cytokines and leptin secretion. Tumor necrosis factor alpha, interleukin 6 (IL-6), IL-10, monocyte chemoattractant protein-1, and leptin levels were measured in cell supernatants. We also performed the detection and quantification of leptin receptor (Ob-Rb) expression in macrophage cell lines stimulated by these LAB strains. Differential secretion profile of cytokines in macrophage cells induced by LAB strains was observed. Also, the levels of Ob-Rb expression diverged among different LAB strains. In LAB-stimulated coculture cells (adipocytes and macrophages), we observed differential production of leptin and cytokines. Furthermore, we detected lower production levels in single culture than cocultured cells. The principal component analysis showed an association between the four clusters of strains established according to their inflammatory profiles and leptin adipocyte production and leptin receptor expression in macrophages. We conclude that coculture is the most appropriate system for selecting strains with the ability to modulate adipokine secretion. The use of microorganisms with low and medium inflammatory properties and ability to modulate leptin levels could be a strategy for the treatment of some metabolic diseases associated with dysregulation of immune response.Fil: Fabersani Marrades, Mario Emanuel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Abeijon Mukdsi, Maria Claudia. Universidad del Norte Santo Tomás de Aquino; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Ross, Gloria Romina. Universidad del Norte Santo Tomás de Aquino; Argentina. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia; ArgentinaFil: Medina, Roxana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán; ArgentinaFil: González, Silvia. Universidad Nacional de Tucumán; ArgentinaFil: Gauffin-Cano, Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad del Norte Santo Tomás de Aquino; Argentin

New insights about phenotypic heterogeneity within Propionibacterium freudenreichii argue against its division into subspecies

Propionibacterium freudenreichii is widely used in Swiss-type cheese manufacture, where it contributes to flavour and eye development. It is currently divided into two subspecies, according to the phenotype for lactose fermentation and nitrate reduction (lac+/nit- and lac-/nit+ for P. freudenreichii subsp. shermanii and subsp. freudenreichii, respectively). However, the existence of unclassifiable strains (lac+/nit+ and lac-/nit-) has also been reported. The aim of this study was to revisit the relevance of the subdivision of P. freudenreichii into subspecies, by confirming the existence of unclassifiable strains. Relevant conditions to test the ability of P. freudenreichii for lactose fermentation and nitrate reduction were first determined, by using 10 sequenced strains, in which the presence or absence of the lactose and nitrate genomic islands were known. We also determined whether the subdivision based on lac/nit phenotype was related to other phenotypic properties of interest in cheese manufacture, in this case, the production of aroma compounds, analysed by gas chromatography-mass spectrometry, for a total of 28 strains. The results showed that a too short incubation time can lead to false negative for lactose fermentation and nitrate reduction. They confirmed the existence of four lac/nit phenotypes instead of the two expected, thus leading to 13 unclassifiable strains out of the 28 characterized (7 lac+/nit+ and 6 lac-/nit-). The production of the 15 aroma compounds detected in all cultures varied more within a lac/nit phenotype (up to 20 times) than between them. Taken together, these results demonstrate that the division of P. freudenreichii into two subspecies does not appear to be relevant.Fil: de Freitas, Rosangela. Universidade Federal de Viçosa. Departamento de Tecnologia de Alimentos; Brasil. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Madec, Marie Noelle. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Chuat, Victoria. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Maillard, Marie Bernadette. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Abeijon Mukdsi, Maria Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Tucumán. Centro de Referencia para Lactobacilos (i); Argentina. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Falentin, Hélène. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Carvalho, Antonio Fernandes de. Universidade Federal de Viçosa. Departamento de Tecnologia de Alimentos; BrasilFil: Valence, Florence. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; FranciaFil: Thierry, Anne. Institut National de la Recherche Agronomique; Francia. Science et Technologie du Lait et de l; Franci

Oral administration of Lactobacillus fermentum CRL1446 improves biomarkers of metabolic syndrome in mice fed a high-fat diet supplemented with wheat bran

This work aimed to evaluate the effect of oral administration of probiotic Lactobacillus (L.) fermentum CRL1446, with feruloyl esterase (FE) activity, on metabolic biomarkers and intestinal microbiota of mice with high fat diet-induced Metabolic Syndrome (MS) and supplemented with wheat bran as a source ofesterified ferulic acid. Six-week-old male Swiss albino mice developed the components of MS when fed with high fat diet supplemented with wheat bran (HFD + WB) for 14 weeks. Positive impact of L. fermentum CRL1446 administration on these animals was reflected in a decrease in body weight gain and adiposity index compared to the animals that did not receive the probiotic strain. In addition, a decrease in plasma leptin levels, improvement of inflammatory profile, reduction of fatty infiltration in hepatocytes and modification of lipid profile (increased HDL-cholesterol and decreased LDL-cholesteroland triglyceride levels) were observed. On the other hand, L. fermentum CRL1446 reduced fasting glucose and insulin levels, improving the HOMA index in mice with MS. Postprandial glucose levels were also reduced in the oral glucose tolerance test. Consumption of L. fermentum CRL1446 with HFD + WB (HFD + WB-Lf mice group) had a great impact on host metabolism, modulating intestinal microbiota, with an increase in Bacteroidetes and a decrease in Firmicutes abundance being observed. Increased intestinal FE activity, improved oxidative status and increased abundance of 3-hydroxyphenylpropionic acid andbutyric acid concentration in colonic content, were also demonstrated in HFD + WB-Lf mice. Results obtained suggest that supplementation with L. fermentum CRL1446 enhances beneficial effects of a bran diet, attenuating the risk factors associated with MS.Fil: Russo, Matias Irineo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Márquez, María Antonela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Herrera, Héctor Matías. Universidad Nacional de Tucumán. Facultad de Bioquímica, Química y Farmacia; ArgentinaFil: Abeijon Mukdsi, Maria Claudia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Saavedra, Maria Lucila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Hebert, Elvira Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Gauffin Cano, María Paola. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; ArgentinaFil: Medina, Roxana Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Tucumán. Centro de Referencia para Lactobacilos; Argentina. Universidad Nacional de Tucumán. Facultad de Agronomía y Zootecnia; Argentin

CXCL1: A new diagnostic biomarker for human tuberculosis discovered using Diversity Outbred mice.

More humans have died of tuberculosis (TB) than any other infectious disease and millions still die each year. Experts advocate for blood-based, serum protein biomarkers to help diagnose TB, which afflicts millions of people in high-burden countries. However, the protein biomarker pipeline is small. Here, we used the Diversity Outbred (DO) mouse population to address this gap, identifying five protein biomarker candidates. One protein biomarker, serum CXCL1, met the World Health Organization\u27s Targeted Product Profile for a triage test to diagnose active TB from latent M.tb infection (LTBI), non-TB lung disease, and normal sera in HIV-negative, adults from South Africa and Vietnam. To find the biomarker candidates, we quantified seven immune cytokines and four inflammatory proteins corresponding to highly expressed genes unique to progressor DO mice. Next, we applied statistical and machine learning methods to the data, i.e., 11 proteins in lungs from 453 infected and 29 non-infected mice. After searching all combinations of five algorithms and 239 protein subsets, validating, and testing the findings on independent data, two combinations accurately diagnosed progressor DO mice: Logistic Regression using MMP8; and Gradient Tree Boosting using a panel of 4: CXCL1, CXCL2, TNF, IL-10. Of those five protein biomarker candidates, two (MMP8 and CXCL1) were crucial for classifying DO mice; were above the limit of detection in most human serum samples; and had not been widely assessed for diagnostic performance in humans before. In patient sera, CXCL1 exceeded the triage diagnostic test criteria (\u3e90% sensitivity; \u3e70% specificity), while MMP8 did not. Using Area Under the Curve analyses, CXCL1 averaged 94.5% sensitivity and 88.8% specificity for active pulmonary TB (ATB) vs LTBI; 90.9% sensitivity and 71.4% specificity for ATB vs non-TB; and 100.0% sensitivity and 98.4% specificity for ATB vs normal sera. Our findings overall show that the DO mouse population can discover diagnostic-quality, serum protein biomarkers of human TB

Urine-Based Antigen (Protein) Detection Test for the Diagnosis of Visceral Leishmaniasis

This review describes and appraises a novel protein-based antigen detection test for visceral leishmaniasis (VL). The test detects in patient’s urine six proteins from Leishmania infantum (chagasi) and Leishmania donovani, the etiological agents of VL. The gold standard test for VL is microscopic observation of the parasites in aspirates from spleen, liver, or bone marrow (and lymph node for dogs). Culture of the parasites or detection of their DNA in these aspirates are also commonly used. Serological tests are available but they cannot distinguish patients with active VL from either healthy subjects exposed to the parasites or from subjects who had a successful VL treatment. An antigen detection test based on the agglutination of anti-leishmania carbohydrates antibody coated latex beads has been described. However, the results obtained with this carbohydrate-based test have been conflicting. Using mass spectrometry, we discovered six L. infantum/L. donovani proteins excreted in the urine of VL patients and used them as markers for the development of a robust mAb-based antigen (protein) detection test. The test is assembled in a multiplexed format to simultaneously detect all six markers. Its initial clinical validation showed a sensitivity of 93% and specificity of 100% for VL diagnosis

Topography of initiation of N-glycosylation reactions

Previous studies on the topography of the reactions leading to the formation of dolichol-P-P-Glc-NAc2Man9Glc3 have shown that these occur on both sides of the endoplasmic reticulum membrane (Hirschberg, C. B., and Snider, M. D. (1987) Annu. Rev. Biochem. 56, 63-87). Dolichol-P-P-GlcNAc2Man5 has been detected on the cytoplasmic side of the endoplasmic reticulum membrane while the subsequent dolichol-oligosaccharide intermediates face the lumen. Less clear is the side of the membrane where dolichol-P-P-GlcNAc2 is assembled. We now present evidence strongly suggesting that the active sites of the enzymes catalyzing the synthesis of this latter intermediate are on the cytoplasmic side of the endoplasmic reticulum membrane. In addition, dolichol-P-P-GlcNAc2 has also been detected on this side. Incubations of sealed, right side out rat liver endoplasmic reticulum-derived vesicles with [beta-32P] UDP-GlcNAc in the presence of 5-Br-UMP resulted in the formation of radiolabeled dolichol-P-P-GlcNAc and dolichol-P-P-GlcNAc2 under conditions where there was complete inhibition of transport of the nucleotide sugar. In other experiments with the above radiolabeled nucleotide sugar and sealed vesicles, it was demonstrated that EDTA (a membrane-impermeable reagent) inhibited the N-acetylglucosamine-1-phosphate transferase under conditions where transport of the nucleotide sugar into the lumen was unaffected. Finally, sealed vesicles were first incubated with [32P]UDP-GlcNAc and subsequently with UDP-Gal and soluble galactosyltransferase. This resulted in galactosylation of dolichol-P-P-GlcNAc2. The above results, together with the previous observations, strongly suggest that all reactions leading to this latter dolichol intermediate occur on the cytosolic side of the endoplasmic reticulum membrane