Caracterització de Lactobacils d'origen intestinal i avaluació in vivo del seu poder probiòtic en pollastres

Una soca de Lactobacillus salivarius resistent a la rifampicina, CTC2197, es va assajar com a probiòtic en pollastres, estudiant la seva capacitat de prevenir la colonització de Salmonella enteritidis C-114 en pollastres. Quan la soca probiòtica es va administrar via oral juntament amb S.enteritidis C-114 directament al proventricle en pollets Leghorn de 1 dia, el patògen fou eliminat completament després de 21 dies. Els mateixos resultats es van obtenir quan la soca es va administrar a través del menjar i l'aigua a més de la inoculació directa al proventricle. La inclusió de L.salivarius CTC2197 en el menjar del primer dia va mostrar que una concentració de 105 UFC g-1 era suficient per assegurar la colonització dels tracte gastrointestinal dels pollets després de 1 setmana. No obstant, entre els 21 i 28 dies, L.salivarius CTC2197 no va ser detectable en el tracte gastrointestinal d'alguns pollets, mostrant que seria necessària més d'una dosis per assegurar la seva presència fins al final de l'etapa d'engreix. La liofilització i la congelació per glicerol o llet descremada com a agents crioprotector, van semblar mètodes adequats per preservar la soca probiòtica. La inclusió de L.salivarius CTC2197 en un pinso comercial va semblar ser un bon mètode per subministrar-lo en granja, tot i que la soca va mostrar sensibilitat a les temperatures utilitzades durant l'emmagatzematge del pinso i a les incubadores dels pollets. A més, la supervivència va millorar després de diverses reinoculacions en pinso.

Prebiotic effects: metabolic and health benefits

The different compartments of the gastrointestinal tract are inhabited by populations of micro-organisms. By far the most important predominant populations are in the colon where a true symbiosis with the host exists that is a key for well-being and health. For such a microbiota, 'normobiosis' characterises a composition of the gut 'ecosystem' in which micro-organisms with potential health benefits predominate in number over potentially harmful ones, in contrast to 'dysbiosis', in which one or a few potentially harmful micro-organisms are dominant, thus creating a disease-prone situation. The present document has been written by a group of both academic and industry experts (in the ILSI Europe Prebiotic Expert Group and Prebiotic Task Force, respectively). It does not aim to propose a new definition of a prebiotic nor to identify which food products are classified as prebiotic but rather to validate and expand the original idea of the prebiotic concept (that can be translated in 'prebiotic effects'), defined as: 'The selective stimulation of growth and/or activity(ies) of one or a limited number of microbial genus(era)/species in the gut microbiota that confer(s) health benefits to the host.' Thanks to the methodological and fundamental research of microbiologists, immense progress has very recently been made in our understanding of the gut microbiota. A large number of human intervention studies have been performed that have demonstrated that dietary consumption of certain food products can result in statistically significant changes in the composition of the gut microbiota in line with the prebiotic concept. Thus the prebiotic effect is now a well-established scientific fact. The more data are accumulating, the more it will be recognised that such changes in the microbiota's composition, especially increase in bifidobacteria, can be regarded as a marker of intestinal health. The review is divided in chapters that cover the major areas of nutrition research where a prebiotic effect has tentatively been investigated for potential health benefits. The prebiotic effect has been shown to associate with modulation of biomarkers and activity(ies) of the immune system. Confirming the studies in adults, it has been demonstrated that, in infant nutrition, the prebiotic effect includes a significant change of gut microbiota composition, especially an increase of faecal concentrations of bifidobacteria. This concomitantly improves stool quality (pH, SCFA, frequency and consistency), reduces the risk of gastroenteritis and infections, improves general well-being and reduces the incidence of allergic symptoms such as atopic eczema. Changes in the gut microbiota composition are classically considered as one of the many factors involved in the pathogenesis of either inflammatory bowel disease or irritable bowel syndrome. The use of particular food products with a prebiotic effect has thus been tested in clinical trials with the objective to improve the clinical activity and well-being of patients with such disorders. Promising beneficial effects have been demonstrated in some preliminary studies, including changes in gut microbiota composition (especially increase in bifidobacteria concentration). Often associated with toxic load and/or miscellaneous risk factors, colon cancer is another pathology for which a possible role of gut microbiota composition has been hypothesised. Numerous experimental studies have reported reduction in incidence of tumours and cancers after feeding specific food products with a prebiotic effect. Some of these studies (including one human trial) have also reported that, in such conditions, gut microbiota composition was modified (especially due to increased concentration of bifidobacteria). Dietary intake of particular food products with a prebiotic effect has been shown, especially in adolescents, but also tentatively in postmenopausal women, to increase Ca absorption as well as bone Ca accretion and bone mineral density. Recent data, both from experimental models and from human studies, support the beneficial effects of particular food products with prebiotic properties on energy homaeostasis, satiety regulation and body weight gain. Together, with data in obese animals and patients, these studies support the hypothesis that gut microbiota composition (especially the number of bifidobacteria) may contribute to modulate metabolic processes associated with syndrome X, especially obesity and diabetes type 2. It is plausible, even though not exclusive, that these effects are linked to the microbiota-induced changes and it is feasible to conclude that their mechanisms fit into the prebiotic effect. However, the role of such changes in these health benefits remains to be definitively proven. As a result of the research activity that followed the publication of the prebiotic concept 15 years ago, it has become clear that products that cause a selective modification in the gut microbiota's composition and/or activity(ies) and thus strengthens normobiosis could either induce beneficial physiological effects in the colon and also in extra-intestinal compartments or contribute towards reducing the risk of dysbiosis and associated intestinal and systemic pathologies.

Prebiotic interventions for the improvement of gut health

The increasing awareness of the role that the colonic microflora plays in maintaining host health within the gastrointestinal tract and systemically through the absorption of metabolites, has attracted a lot of interest, within the nutritional sciences, in developing dietary tools for controlling the colonic microflora. Among those dietary tools, prebiotics aim to improve health by stimulating numbers and/or activities of the beneficial bacteria in the gut, mainly bifidobacteria and lactobacilli. The ability of incorporating prebiotics in various food processes together with recent developments in understanding how prebiotics are metabolised by health promoting bacteria, allow us to specifically aim such dietary interventions towards selected population groups, such as infants and elderly, and disease states, such as colon cancer and irritable bowel disease.

Modulation of the human gut microflora towards improved health using prebiotics: assessment of efficacy

There is increasing awareness that the human gut microflora plays a critical role in maintaining host health, both within the gastrointestinal tract and, through the absorption of metabolites, systemically. An 'optimal' gut microflora establishes an efficient barrier to the invasion and colonisation of the gut by pathogenic bacteria, produces a range of metabolic substrates which in turn are utilized by the host (e.g. vitamins and short chain fatty acids) and stimulates the immune system in a non-inflammatory manner. Although little is known about the individual species of bacteria responsible for these beneficial activities, it is generally accepted that the bifidobacteria and lactobacilli constitute important components of the beneficial gut microflora. A number of diet-based microflora management tools have been developed and refined over recent decades including probiotic, prebiotic and synbiotic approaches. Each aims to stimulate numbers and/or activities of the bifidobacteria and lactobacilli within the gut microflora. The aim of this article is to examine how prebiotics are being applied to the improvement of human health and to review the scientific evidence supporting their use.

Using probiotics and prebiotics to improve gut health

Recent molecular-based investigations have confirmed the species diversity and metabolic complexity of the human gut microbiota. It is also increasingly clear that the human gut microbiota plays a crucial role in host health, both as a source of infection and environmental insult and, conversely, in protection against disease and maintenance of gut function. Although little is known about the health impact of the dominant groups of gut bacteria it is generally accepted that bifidobacteria and lactobacilli are important components of what might be termed the beneficial gut microbiota. The microbiota management tools of probiotics, prebiotics and synbiotics have been developed and, indeed, commercialized over the past few decades with the expressed purpose of increasing numbers of bifidobacteria and/or lactobacilli within the gastrointestinal tract.

Probiotics and prebiotics in female health

Functional foods such as probiotics, prebiotics and nutraceuticals are of extreme interest to researchers. There is growing evidence that these food ingredients may improve and in some cases treat certain conditions that are implicated in women's health. The use of probiotics (live, beneficial bacteria) in improving gastrointestinal and non-gastrointestinal tract conditions such as irritable bowel syndrome, candidiasis and other female urogenital tract conditions are reviewed. Emphasis is also given to the importance of prebiotics (non-digestible food ingredients) in osteoporosis management and alleviation of menopausal symptoms and reducing the onset of cancer.

Mobilisation of enterocyte fat stores by oral glucose in humans

Background and aims: When a high fat oral load is followed several hours later by further ingestion of nutrients, there is an early postprandial peak in plasma triacylglycerol (TG). The aim of this study was to investigate the location and release of lipid from within the gastrointestinal tract. Methods: Ten healthy patients undergoing oesopho-gastro-duodenoscopy (OGD) were recruited. At t=0, all patients consumed a 50 g fat emulsion and at t=5 hours they consumed either water or a 38 g glucose solution. OGD was performed at t=6 hours and jejunal biopsy samples were evaluated for fat storage. A subgroup of five subjects then underwent a parallel metabolic study in which postprandial lipid and hormone measurements were taken during an identical two meal protocol. Results: Following oral fat at t=0, samples from patients that had subsequently ingested glucose exhibited significantly less staining for lipid within the mucosa and submucosa of the jejunum than was evident in patients that had consumed only water (p=0.028). There was also less lipid storage within the cytoplasm of enterocytes (p=0.005) following oral glucose. During the metabolic study, oral glucose consumed five hours after oral fat resulted in a postprandial peak in plasma TG, chylomicron-TG, and apolipoprotein B48 concentration compared with oral water. Conclusion: After a fat load, fat is retained within the jejunal tissue and released into plasma following glucose ingestion, resulting in a peak in chylomicron-TG which has been implicated in the pathogenesis of atherosclerosis.

Prebiotics

In nutritional sciences there is much interest in dietary modulation of the human gut. The gastrointestinal tract, particularly the colon, is very heavily populated with bacteria. Most bacteria are benign; however, certain gut species are pathogenic and may be involved in the onset of acute and chronic disorders. Bifidobacteria and lactobacilli are thought to be beneficial and are common targets for dietary intervention. Prebiotic is a non-viable food ingredient selectively metabolized by beneficial intestinal bacteria. Dietary modulation of the gut microflora by prebiotics is designed to improve health by stimulating numbers and/or activities of the bifidobacteria and lactobacilli. Having an 'optimal' gut microflora can increase resistance to pathogenic bacteria, lower blood ammonia, increase stimulation of the immune response and reduce the risk of cancer. This chapter examines how prebiotics are being applied to the improvement of human health and reviews the scientific evidence behind their use.

The reaction of flavanols with nitrous acid protects against N-nitrosamine formation and leads to the formation of nitroso derivatives which inhibit cancer cell growth

Studies have suggested that diets rich in polyphenols Such as flavonoids may lead to a reduced risk of gastrointestinal cancers. We demonstrate the ability of monomeric and dimeric flavanols to scavenge reactive nitrogen species derived from nitrous acid. Both epicatechin and dimer B2 (epicatechin dimer) inhibited nitrous acid-induced formation of 3-nitrotyrosine and the formation of the carcinogenic N-nitrosamine, N-nitrosodimethylamine. The reaction of monomeric and dimeric epicatechin with nitrous acid led to the formation of mono- and di-nitroso flavanols, whereas the reaction with hesperetin resulted primarily in the formation of nitrated products. Although, epicatechin was transferred across the jejunum of the small intestine yielding metabolites, its nitroso form was not absorbed. Dimer B2 but not epicatechin monomer inhibited the proliferation of, and triggered apoptosis in, Caco-2 cells. The latter was accompanied by caspase-3 activation and reductions in Akt phosphorylation, suggesting activation of apoptosis via inhibition of prosurvival signaling. Furthermore, the dinitroso derivative of dimer B2, and to a lesser extent the dinitroso-epicatechin, also induced significant toxic effects in Caco-2 cells. The inhibitory effects on cellular proliferation were paralleled by early inhibition of ERK 1/2 phosphorylation and later reductions in cyclin D I levels, indicating modulation of cell cycle regulation in Caco-2 cells. These effects highlight multiple routes in which dietary derived flavanols may exert beneficial effects in the gastrointestinal tract. (c) 2005 Elsevier Inc. All rights reserved.

Fermentation of heated gluten systems by gut microflora

The Maillard reaction causes changes to protein structure and occurs in foods mainly during thermal treatment. Melanoidins, the final products of the Maillard reaction, may enter the gastrointestinal tract, which is populated by different species of bacteria. In this study, melanoidins were prepared from gluten and glucose. Their effect on the growth of faecal bacteria was determined in culture with genotype and phenotype probes to identify the different species involved. Analysis of peptic and tryptic digests showed that low molecular mass products are formed from the degradation of melanoidins. Results showed a change in the growth of bacteria. This in vitro study demonstrated that melanoidins, prepared from gluten and glucose, affect the growth of the gut microflora.

Porcine models for the metabolic syndrome, digestive and bone disorders: a general overview

The aim of this review article is to provide an overview of the role of pigs as a biomedical model for humans. The usefulness and limitations of porcine models have been discussed in terms of metabolic, cardiovascular, digestive and bone diseases in humans. Domestic pigs and minipigs are the main categories of pigs used as biomedical models. One drawback of minipigs is that they are in short supply and expensive compared with domestic pigs, which in contrast cost more to house, feed and medicate. Different porcine breeds show different responses to the induction of specific diseases. For example, ossabaw minipigs provide a better model than Yucatan for the metabolic syndrome as they exhibit obesity, insulin resistance and hypertension, all of which are absent in the Yucatan. Similar metabolic/physiological differences exist between domestic breeds (e.g. Meishan v. Pietrain). The modern commercial (e.g. Large White) domestic pig has been the preferred model for developmental programming due to the 2- to 3-fold variation in body weight among littermates providing a natural form of foetal growth retardation not observed in ancient (e.g. Meishan) domestic breeds. Pigs have been increasingly used to study chronic ischaemia, therapeutic angiogenesis, hypertrophic cardiomyopathy and abdominal aortic aneurysm as their coronary anatomy and physiology are similar to humans. Type 1 and II diabetes can be induced in swine using dietary regimes and/or administration of streptozotocin. Pigs are a good and extensively used model for specific nutritional studies as their protein and lipid metabolism is comparable with humans, although pigs are not as sensitive to protein restriction as rodents. Neonatal and weanling pigs have been used to examine the pathophysiology and prevention/treatment of microbial-associated diseases and immune system disorders. A porcine model mimicking various degrees of prematurity in infants receiving total parenteral nutrition has been established to investigate gut development, amino acid metabolism and non-alcoholic fatty liver disease. Endoscopic therapeutic methods for upper gastrointestinal tract bleeding are being developed. Bone remodelling cycle in pigs is histologically more similar to humans than that of rats or mice, and is used to examine the relationship between menopause and osteoporosis. Work has also been conducted on dental implants in pigs to consider loading; however with caution as porcine bone remodels slightly faster than human bone. We conclude that pigs are a valuable translational model to bridge the gap between classical rodent models and humans in developing new therapies to aid human health.

Effect of feeding fat or intrajugular infusion of glucagon-like peptide-1 and cholecystokinin on dry matter intake, digestibility, and digesta rate of passage in growing wethers

A cause and effect relationship between glucagon-like peptide 1 (7, 36) amide (GLP-1) and cholecystokinin (CCK) and DMI regulation has not been established in ruminants. Three randomized complete block experiments were conducted to determine the effect of feeding fat or infusing GLP-1 or CCK intravenously on DMI, nutrient digestibility, and Cr rate of passage (using Cr(2)O(3) as a marker) in wethers. A total of 18 Targhee × Hampshire wethers (36.5 ± 2.5 kg of BW) were used, and each experiment consisted of four 21-d periods (14 d for adaptation and 7 d for infusion and sampling). Wethers allotted to the control treatments served as the controls for all 3 experiments; experiments were performed simultaneously. The basal diet was 60% concentrate and 40% forage. In Exp. 1, treatments were the control (0% added fat) and addition of 4 or 6% Ca salts of palm oil fatty acids (DM basis). Treatments in Exp. 2 and 3 were the control and 3 jugular vein infusion dosages of GLP-1 (0.052, 0.103, or 0.155 µg•kg of BW(-1)•d(-1)) or CCK (0.069, 0.138, or 0.207 µg•kg of BW(-1)•d(-1)), respectively. Increases in plasma GLP-1 and CCK concentrations during hormone infusions were comparable with increases observed when increasing amounts of fat were fed. Feeding fat and infusion of GLP-1 tended (linear, P = 0.12; quadratic, P = 0.13) to decrease DMI. Infusion of CCK did not affect (P > 0.21) DMI. Retention time of Cr in the total gastrointestinal tract decreased (linear, P < 0.01) when fat was fed, but was not affected by GLP-1 or CCK infusion. In conclusion, jugular vein infusion produced similar plasma CCK and GLP-1 concentrations as observed when fat was fed. The effects of feeding fat on DMI may be partially regulated by plasma concentration of GLP-1, but are not likely due solely to changes in a single hormone concentration.

In vitro bioaccessibility and gut biotransformation of polyphenols present in the water-insoluble cocoa fraction

Scope: Cocoa, especially the water-insoluble cocoa fraction (WICF), is a rich source of polyphenols. In this study, sequential in vitro digestion of the WICF with gastrointestinal enzymes as well as its bacterial fermentation in a human colonic model system were carried out to investigate bioaccessibility and biotransformation of WICF polyphenols, respectively. Methods and results: The yield of each enzymatic digestion step and the total antioxidant capacity (TAC) were measured and solubilized phenols were characterized by MS/MS. Fermentation of WICF and the effect on the gut microbiota, SCFA production and metabolism of polyphenols was analyzed. In vitro digestion solubilized 38.6% of WICF with pronase and Viscozyme L treatments releasing 51% of the total phenols from the insoluble material. This release of phenols does not determine a reduction in the total antioxidant capacity of the digestion-resistant material. In the colonic model WICF significantly increased of bifidobacteria and lactobacilli as well as butyrate production. Flavanols were converted into phenolic acids by the microbiota following a concentration gradient resulting in high concentrations of 3-hydroxyphenylpropionic acid (3-HPP) in the last gut compartment. Conclusion: Data showed that WICF may exert antioxidant action through the gastrointestinal tract despite its polyphenols being still bound to macromolecules and having prebiotic activity.

Production and evaluation of dry alginate-chitosan microcapsules as an enteric delivery vehicle for probiotic bacteria

This study investigates the production of alginate microcapsules, which have been coated with the polysaccharide chitosan, and evaluates some of their properties with the intention of improving the gastrointestinal viability of a probiotic (Bifidobacterium breve) by encapsulation in this system. The microcapsules were dried by a variety of methods, and the most suitable was chosen. The work described in this Article is the first report detailing the effects of drying on the properties of these microcapsules and the viability of the bacteria within relative to wet microcapsules. The pH range over which chitosan and alginate form polyelectrolyte complexes was explored by spectrophotometry, and this extended into swelling studies on the microcapsules over a range of pHs associated with the gastrointestinal tract. It was shown that chitosan stabilizes the alginate microcapsules at pHs above 3, extending the stability of the capsules under these conditions. The effect of chitosan exposure time on the coating thickness was investigated for the first time by confocal laser scanning microscopy, and its penetration into the alginate matrix was shown to be particularly slow. Coating with chitosan was found to increase the survival of B. breve in simulated gastric fluid as well as prolong its release upon exposure to intestinal pH.

Recognition of greater diversity of Bacillus species and related bacteria in human faeces

In a study looking at the culturable, aerobic Actinobacteria associated with the human gastrointestinal tract, the vast majority of isolates obtained from dried human faeces belonged to the genus Bacillus and related bacteria. A total of 124 isolates were recovered from the faeces of 10 healthy adult donors. 16S rRNA gene sequence analyses showed the majority belonged to the families Bacillaceae (n = 81) and Paenibacillaceae (n = 3), with Bacillus species isolated from all donors. Isolates tentatively identified as Bacillus clausii (n = 32) and B. licheniformis (n = 28) were recovered most frequently, with the genera Lysinibacillus, Ureibacillus, Oceanobacillus, Ornithinibacillus and Virgibacillus represented in some donors. Phenotypic data confirmed the identities of isolates belonging to well-characterized species. Representatives of the phylum Actinobacteria were recovered in much lower numbers (n = 11). Many of the bacilli exhibited antimicrobial activity against one or more strains of Clostridium difficile, C. perfringens, Listeria monocytogenes and Staphylococcus aureus, with some (n = 12) found to have no detectable cytopathic effect on HEp-2 cells. This study has revealed greater diversity within gut-associated aerobic spore-formers than previous studies, and suggests that bacilli with potential as probiotics could be isolated from the human gut.