Proteomic comparison of the cytosolic proteins of three <Emphasis Type="Italic">Bifidobacterium longum</Emphasis> human isolates and <Emphasis Type="Italic">B. longum</Emphasis> NCC2705

Background  

Bifidobacteria are natural inhabitants of the human gastrointestinal tract. In full-term newborns, these bacteria are acquired from the mother during delivery and rapidly become the predominant organisms in the intestinal microbiota. Bifidobacteria contribute to the establishment of healthy intestinal ecology and can confer health benefits to their host. Consequently, there is growing interest in bifidobacteria, and various strains are currently used as probiotic components in functional food products. However, the probiotic effects have been reported to be strain-specific. There is thus a need to better understand the determinants of the observed benefits provided by these probiotics. Our objective was to compare three human B. longum isolates with the sequenced model strain B. longum NCC2705 at the chromosome and proteome levels.     

Antiproliferative effects of homogenates derived from five strains of candidate probiotic bacteria.

Unheated and heat-treated homogenates were separately prepared from candidate probiotic bacteria, including Lactobacillus rhamnosus GG, Bifidobacterium lactis, Lactobacillus acidophilus, Lactobacillus delbrueckii subsp. bulgaricus, and Streptococcus thermophilus. We compared the phytohemagglutinin-induced proliferation of mononuclear cells in the presence of homogenates and in the presence of a control containing no homogenate by assessing thymidine incorporation in cell cultures. All homogenates suppressed proliferation, whether the enzymatic activity was inactivated or not inactivated by heating. When the proliferation assays were repeated with cytoplasmic and cell wall extracts derived from the homogenate of L. rhamnosus GG, the cytoplasmic extract but not the cell wall extract was suppressive. These findings indicate that candidate probiotic bacteria possess a heat-stable antiproliferative component(s). These bacteria may be used to generate microbiologically nonviable yet immunologically active probiotic food products that are easier to store and have a longer shelf life.     

Recovery of Viable Bacteria from Probiotic Products that Target Oral Health

Probiotic therapy has predominantly been directed toward promoting and maintaining intestinal health. In recent years, however, probiotic regimens that target oral health have appeared on the market. These regimens are often delivered in the form of lozenges. Despite the oral health claims made by the manufacturers of these products, there is little independent evidence in the literature to support such claims. In theory, probiotic organisms can be beneficial by several different means including direct inhibition of pathogens and boosting of the host immune response, with the underlying assumption that these mechanisms require a critical number of viable organisms. In this study, five brands of probiotics marketed for oral health were tested for the recovery of viable bacteria. For only one brand could viable bacteria be recovered within one log of the manufacturer’s stated starting amount of bacteria. Nearly, a billion viable bacteria could be recovered from a lozenge of this brand. The other brands claimed similar starting amounts of bacteria at the time of manufacture but at least a three-log drop-off was observed in the amount of viable bacteria recovered from those products. Refrigeration of the probiotics significantly improved the recovery for one brand, but recoveries for all but one brand remained below the recommended daily dosage for probiotic regimens. It is concluded that probiotic brands differ significantly in the quantities of bacteria that remain viable with most failing to meet recommended dosage targets.     

Traditional healthful fermented products of Japan

A variety of fermentation products, such as foods containing probiotic bacteria, black rice vinegar (kurosu), soy sauce (shoyu), soybean-barley paste (miso), natto and tempeh, are sold in food stores in Japan. These fermented food products are produced by traditional methods that exploit mixed cultures of various non-toxic microorganisms. These microorganisms include lactic acid bacteria, acetic acid bacteria, sake yeast, koji molds and natto bacteria. Many traditional fermented foods have been studied and their effects on metabolism and/or immune system have been demonstrated in animal and/or human cells. This review summarizes the scientific basis for the effects of these traditional food products, which are currently produced commercially in Japan.     

Survival and therapeutic potential of probiotic organisms with reference to Lactobacillus acidophilus and Bifidobacterium spp

The present paper provides an overview on the use of probiotic organisms as live supplements, with particular emphasis on Lactobacillus acidophilus and Bifidobacterium spp. The therapeutic potential of these bacteria in fermented dairy products is dependent on their survival during manufacture and storage. Probiotic bacteria are increasingly used in food and pharmaceutical applications to balance disturbed intestinal microflora and related dysfunction of the human gastrointestinal tract. Lactobacillus acidophilus and Bifidobacterium spp. have been reported to be beneficial probiotic organisms that provide excellent therapeutic benefits. The biological activity of probiotic bacteria is due in part to their ability to attach to enterocytes. This inhibits the binding of enteric pathogens by a process of competitive exclusion. Attachment of probiotic bacteria to cell surface receptors of enterocytes also initiates signalling events that result in the synthesis of cytokines. Probiotic bacteria also exert an influence on commensal micro-organisms by the production of lactic acid and bacteriocins. These substances inhibit growth of pathogens and also alter the ecological balance of enteric commensals. Production of butyric acid by some probiotic bacteria affects the turnover of enterocytes and neutralizes the activity of dietary carcinogens, such as nitrosamines, that are generated by the metabolic activity of commensal bacteria in subjects consuming a high-protein diet. Therefore, inclusion of probiotic bacteria in fermented dairy products enhances their value as better therapeutic functional foods. However, insufficient viability and survival of these bacteria remain a problem in commercial food products. By selecting better functional probiotic strains and adopting improved methods to enhance survival, including the use of appropriate prebiotics and the optimal combination of probiotics and prebiotics (synbiotics), an increased delivery of viable bacteria in fermented products to the consumers can be achieved.     

Probiotic Formulations: Application and Status as Pharmaceuticals—A Review

The use of probiotics is a new way to control and treat infections in this modern era. Application of beneficial bacteria to protect against detrimental bacteria in the gastrointestinal tract and thus reap a positive health benefit is the basis of probiotic therapy. Probiotics have a long global history of traditional use. They are normally consumed through fermented foods and are currently sold mostly as ingredients in foods or nutritional supplements. They are also supplied as pharma products. Recent research has highlighted the probiotic potential in the treatment or prevention of disease conditions, maintenance of health, improving immunity and in the reduction in the risk of future diseases. But their position in the pharmaceutical industry is still not very clear. Clinical practitioners use probiotic pharma products mostly as supplements. Their status as drugs is still unclear. This review is aimed to analyze probiotics as pharmaceuticals, their current status as dietary supplements and drugs, existing probiotic preparations and future research needs.     

Probiotic bacteria may become dormant during storage

The determination of bacterial viability in probiotic products is of economic, technological, and clinical significance. We compared four methods to enumerate three Bifidobacterium strains in fermented oat products during storage. A subpopulation of nonculturable cells retained a functional cell membrane typical of viable cells, indicating that probiotic bacteria become dormant during storage.     

Utilisation of enterobacterial repetitive intergenic consensus (ERIC) sequence-based PCR to fingerprint the genomes of Bifidobacterium isolates and other probiotic bacteria

Enterobacterial repetitive intergenic consensus based on PCR (ERIC-PCR) was used to generate DNA fingerprints for bifidobacteria and other probiotic bacteria. Two primers (ERIC 1R and ERIC 2) used in ERIC-PCR revealed that all of the probiotic bacteria tested possess enterobacterial repetitive intergenic consensus sequences with the PCR products ranging from 250 bp to 5000 bp. The bacterial strains can be differentiated by comparing fingerprint patterns. The dendrogram of the fingerprints revealed that most of the bifidobacterial wild type strains fell into one cluster at similarity level of approximately 79%.     

Probiotic Bacteria May Become Dormant during Storage

The determination of bacterial viability in probiotic products is of economic, technological, and clinical significance. We compared four methods to enumerate three Bifidobacterium strains in fermented oat products during storage. A subpopulation of nonculturable cells retained a functional cell membrane typical of viable cells, indicating that probiotic bacteria become dormant during storage.     

Release studies of Lactobacillus casei strain Shirota from chitosan-coated alginate-starch microcapsules in ex vivo porcine gastrointestinal contents

AIMS: To investigate the release characteristics of Lactobacillus casei strain Shirota (LCS) from chitosan-coated alginate-starch (CCAS) capsule in different regions of ex vivo porcine gastrointestinal (GI) contents. METHODS AND RESULTS: A 0.1 g of CCAS encapsulated bacteria (containing c. 10(8) CFU of LCS) were incubated in different sections of ex vivo porcine GI contents (10 ml) anaerobically at 37 degrees C. Samples were taken from different GI contents at different time intervals (up to 24 h) and estimated for the release of LCS from capsules. There was almost a complete release (1.1 x 10(8) CFU) of LCS from capsules within 8 h of incubation in ileal contents, while it took nearly 12 h to release the completely encapsulated bacteria in colon content under similar conditions. There was only a partial release of encapsulated LCS, incubated in duodenal and jejunal contents, while there was no significant (P > 0.05) release of encapsulated bacteria in gastric contents even after 24 h of incubation. CONCLUSIONS: The capsules were able to release viable probiotic cells completely in ex vivo porcine ileal and colon contents. SIGNIFICANCE AND IMPACT OF THE STUDY: CCAS capsule can be an effective way of protecting probiotic bacteria from adverse gastric conditions and delivering viable bacteria to the host intestinal systems.     

Antiproliferative Effects of Homogenates Derived from Five Strains of Candidate Probiotic Bacteria

Unheated and heat-treated homogenates were separately prepared from candidate probiotic bacteria, including Lactobacillus rhamnosus GG, Bifidobacterium lactis, Lactobacillus acidophilus, Lactobacillus delbrueckii subsp. bulgaricus, and Streptococcus thermophilus. We compared the phytohemagglutinin-induced proliferation of mononuclear cells in the presence of homogenates and in the presence of a control containing no homogenate by assessing thymidine incorporation in cell cultures. All homogenates suppressed proliferation, whether the enzymatic activity was inactivated or not inactivated by heating. When the proliferation assays were repeated with cytoplasmic and cell wall extracts derived from the homogenate of L. rhamnosus GG, the cytoplasmic extract but not the cell wall extract was suppressive. These findings indicate that candidate probiotic bacteria possess a heat-stable antiproliferative component(s). These bacteria may be used to generate microbiologically nonviable yet immunologically active probiotic food products that are easier to store and have a longer shelf life.     

Generic delivery of payload of nanoparticles intracellularly via hybrid polymer capsules for bioimaging applications

Towards the goal of development of a generic nanomaterial delivery system and delivery of the 'as prepared' nanoparticles without 'further surface modification' in a generic way, we have fabricated a hybrid polymer capsule as a delivery vehicle in which nanoparticles are loaded within their cavity. To this end, a generic approach to prepare nanomaterials-loaded polyelectrolyte multilayered (PEM) capsules has been reported, where polystyrene sulfonate (PSS)/polyallylamine hydrochloride (PAH) polymer capsules were employed as nano/microreactors to synthesize variety of nanomaterials (metal nanoparticles; lanthanide doped inorganic nanoparticles; gadolinium based nanoparticles, cadmium based nanoparticles; different shapes of nanoparticles; co-loading of two types of nanoparticles) in their hollow cavity. These nanoparticles-loaded capsules were employed to demonstrate generic delivery of payload of nanoparticles intracellularly (HeLa cells), without the need of individual nanoparticle surface modification. Validation of intracellular internalization of nanoparticles-loaded capsules by HeLa cells was ascertained by confocal laser scanning microscopy. The green emission from Tb(3+) was observed after internalization of LaF(3):Tb(3+)(5%) nanoparticles-loaded capsules by HeLa cells, which suggests that nanoparticles in hybrid capsules retain their functionality within the cells. In vitro cytotoxicity studies of these nanoparticles-loaded capsules showed less/no cytotoxicity in comparison to blank capsules or untreated cells, thus offering a way of evading direct contact of nanoparticles with cells because of the presence of biocompatible polymeric shell of capsules. The proposed hybrid delivery system can be potentially developed to avoid a series of biological barriers and deliver multiple cargoes (both simultaneous and individual delivery) without the need of individual cargo design/modification.     

A Review of the Advancements in Probiotic Delivery: Conventional vs. Non-conventional Formulations for Intestinal Flora Supplementation

Probiotic delivery systems are widely used nutraceutical products for the supplementation of natural intestinal flora. These delivery systems vary greatly in effectiveness to exert health benefits for a patient. Probiotic delivery systems can be categorized into conventional, pharmaceutical formulations, and non-conventional, mainly commercial food-based, products. The degree of health benefits provided by these probiotic formulations varies in their ability to deliver viable, functional bacteria in large enough numbers (effectiveness), to provide protection against the harsh effects of the gastric environment and intestinal bile (in vivo protection), and to survive formulation processes (viability). This review discusses the effectiveness of these probiotic delivery systems to deliver viable functional bacteria focusing on the ability to protect the encapsulated probiotics during formulation process as well as against harsh physiological conditions through formulation enhancements using coatings and polymer enhancements. A brief overview on the health benefits of probiotics, current formulation, patient and legal issues facing probiotic delivery, and possible recommendations for the enhanced delivery of probiotic bacteria are also provided. Newer advanced in vitro analyses that can accurately determine the effectiveness of a probiotic formulation are also discussed with an ideal probiotic delivery system hypothesized through a combination of the two probiotic delivery systems described.     

The effects of kefir,koumiss, yogurt and commercial probiotic formulations on PPARα and PPAR-β/δ expressions in mouse kidney

Commercial probiotic capsules that contain probiotic bacteria, kefir, koumiss and yogurt contain beneficial microorganisms that affect cholesterol levels and immune response, and are used for treatment of some diseases. We investigated using immunohistochemistry the effects of kefir, koumiss, yogurt and a commercial probiotic formulation on the expression levels of peroxisome proliferator-activated receptor-α (PPARα) and peroxisome proliferator-activated receptor-β/δ (PPAR-β/δ), which are members of the nuclear steroid hormone receptor superfamily in mouse kidney. Mice were assigned to five groups: group 1, commercial probiotic capsules; group 2, kefir; group 3, koumiss; group 4, yogurt; group 5, control. After oral administration for 15 days, body weights were recorded and kidney tissue samples were obtained. Hematoxylin & eosin staining and the streptavidin-biotin peroxidase complex (ABC) method were applied to tissue sections to examine histology and to determine the localization of PPARα and PPAR-β/δ in the kidneys. We found that the weights of the mice in the kefir, koumiss, yogurt and commercial probiotic capsules groups were increased compared to controls. No differences in kidney histology were observed in any of the experimental groups. Kefir, koumiss, yogurt and the commercial probiotic preparation increased PPARα and PPAR-β/δ expressions.     

Immune response to orally consumed antigens and probiotic bacteria

The gut mucosal system must fulfil conflicting roles in suppressing immune responses against orally fed antigens (tolerance) while still retaining the ability to respond to potential enteric pathogens. It must also, to a large degree, not mount an immune response against commensal enteric bacteria and the administration of large numbers of probiotic bacteria formulated as dietary supplements in food products. Contrary to this dogma, it has been found that feeding ovalbumin as a marker antigen, in association with selected probiotic bacteria, appears to prime for an intestinal immune response that is further augmented by skin vaccination. Skin immunization is known to stimulate a strong innate, humoral and cellular immune response. Such dominant immunogenic signals appear to override tolerogenic signals engendered by oral feeding of antigen. High-dose antigen feeding stimulated a strong Th2-dependent antibody response to skin vaccination but completely suppressed cytotoxic T cell responses. This was true even when ovalbumin was administered in conjunction with various selected probiotic bacteria. However, while yeast appeared to be better at priming for an enhanced humoral response, Lactobacillus fermentum and Staphylococcus carnosus were more effective in enhancing the postvaccinal lymphoproliferative response against ovalbumin.     

Intestinal microflora and homeostasis of the mucosal immune response: implications for probiotic bacteria?

The intestinal microflora can be considered a postnatally acquired organ that is composed of a large diversity of bacteria that perform important functions for the host and can be modulated by environmental factors, such as nutrition. Specific components of the intestinal microflora, including lactobacilli and bifidobacteria, have been associated with beneficial effects on the host, such as promotion of gut maturation and integrity, antagonisms against pathogens and immune modulation. Beyond this, the microflora seems to play a significant role in the maintenance of intestinal immune homeostasis and prevention of inflammation. The contribution of the intestinal epithelial cell in the first line of defense against pathogenic bacteria and microbial antigens has been recognized. However, the interactions of intestinal epithelial cells with indigenous bacteria are less well understood. This review will summarize the increasing scientific attention to mechanisms of the innate immune response of the host towards different components of the microflora, and suggest a potential role for selected probiotic bacteria in the regulation of intestinal inflammation.     

益生菌调控肠道屏障功能预防家禽球虫病的研究进展

球虫病给养禽业带来巨大经济损失,人们对绿色健康食品的迫切需求使球虫病的防控面临新的挑战。伴随世界"禁抗"进程的不断推进,家禽养殖业亟需一种安全有效的新型抗球虫方法。益生菌可竞争性排斥病原菌定殖以防止球虫病继发感染,可刺激宿主抗菌肽、黏蛋白和紧密连接蛋白的分泌以抵御球虫入侵,还可激活免疫反应以增强机体抗球虫感染的能力。本文从调控肠道微生物群、改善肠黏膜屏障和调节免疫系统功能等方面综述了益生菌在预防和控制家禽球虫病中的作用,以期为高效抗球虫益生菌制剂的研发提供参考。     

Role of commercial probiotic strains against human pathogen adhesion to intestinal mucus

AIMS: The aims of this study present were to assess and to evaluate in vitro the abilities of commercial probiotic strains derived from fermented milk products and related sources currently marketed in European countries, to inhibit, compete and displace the adhesion of selected potential pathogens to immobilized human mucus. METHODS AND RESULTS: The adhesion was assessed by measuring the radioactivity of bacteria adhered to the human mucus. We tested 12 probiotic strains against eight selected pathogens. All strains tested were able to adhere to mucus. All probiotic strains tested were able to inhibit and displace (P<0.05) the adhesion of Bacteroides, Clostridium, Staphylococcus and Enterobacter. In addition, the abilities to inhibit and to displace adhered pathogens depended on both the probiotic and the pathogen strains tested suggesting that several complementary mechanisms are implied in the processes. CONCLUSIONS: Our results indicate the need for a case-by-case assessment in order to select strains with the ability to inhibit or displace a specific pathogen. Probiotics could be useful to correct deviations observed in intestinal microbiota associated with specific diseases and also, to prevent pathogen infections. SIGNIFICANCE AND IMPACT OF THE STUDY: The competitive exclusion properties of probiotics as well as their ability to displace and inhibit pathogens are the most importance for therapeutic manipulation of the enteric microbiota. The application of such strategies could contribute to expand the beneficial properties on human health against pathogen infection.     

Identification of Risks in the Life Cycle of Nanotechnology‐Based Products

In order to realize the projected market potential of nanotechnology, the environmental, health, and safety (EHS) uncertainties posed by a nano‐product (i.e., a nanotechnology‐enabled product) need to be characterized through the identification of risks and opportunities in early stages of product development. We present a methodology to identify risks from nano‐products using a scenario analysis approach that allows for expert elicitation on a set of preidentified use and disposal scenarios and what we have labeled “risk triggers” to obtain scores on their likelihood of occurrence and severity. Use and disposal scenarios describe product life‐cycle stages that could result in risk attributed to the nano‐product, whereas risk triggers are particular to nanoparticle properties. These are potential risks, as the risk assessment community is currently debating the specific risks attributed to nanotechnology. Through such a framework, our goal is to identify which products pose greater risks, where these risks occur in the product life cycle, and the impacts of these environmental risks on society. The comparison of risk triggers across nano‐products allows relative risk ranking on axes of exposure‐ and hazard‐related risk triggers. For the specific case of air fresheners, areas of acute risks resulted from bioavailability of nanoparticles in air release and water entrainment exposure scenarios; catalytic activity of nanoparticles in inhalation and air release exposure scenarios; the harmful effects due to the antibacterial property on useful bacteria particularly in susceptible populations; and, finally, risks from the lack of nanoparticle coating stability in air release scenarios.     

Lactococci as probiotic strains: adhesion to human enterocyte-like Caco-2 cells and tolerance to low pH and bile

There have been few studies on the probiotic activity of Lactococcus strains although they are commonly used as starter bacteria in manufacturing many kinds of fermented dairy products. Nine strains of the genus Lactococcus were examined for their probiotic properties, such as adherence to human enterocyte-like Caco-2 cells and tolerance to acid and bile. Six strains were adhesive and the highest adhesion was observed with Lactcoccus lactis ssp. lactis NIAI527. This strain adhered to the microvilli of cells as observed by scanning electron microscopy and also tolerated low pH and bile. These properties should make strain 527 a potential new probiotic strain.