Lack of Heterogeneity in Bacteriocin Production Across a Selection of Commercial Probiotic Products

Probiotics are live microorganisms that, when administered in adequate amounts, confer a health benefit to the host. Bacteriocin production has often been mooted as a desirable probiotic trait and, in specific cases, has been shown to promote probiotic survival within the gastrointestinal tract, contribute to the control of pathogens and even influence host gene expression in the gut. However, it is not clear what proportion of probiotic strains routinely found in commercial products produces bacteriocins, and additionally, it is not known which bacteriocins are produced most frequently. To address this, we conducted a culture-based assessment of the bacteriocinogenic ability of bacterial strains found in a variety of commercially available probiotic products. We detected eight bacteriocin-producing isolates from 16 tested products. Interestingly, in all cases, the isolates were Lactobacillus acidophilus, and the bacteriocin produced was identified as the narrow spectrum class II bacteriocin, lactacin B. The apparent absence of other bacteriocin-producing strains from across these products suggests a lack of heterogeneity in bacteriocin production within probiotic products and suggests that bacteriocin production is not being optimally harnessed as a probiotic trait.     

Microencapsulation of probiotic bacteria: technology and potential applications

In the recent past, there has been an explosion of probiotic health-based products. Many reports indicated that there is poor survival of probiotic bacteria in these products. Further, the survival of these bacteria in the human gastro-intestinal system is questionable. Providing probiotic living cells with a physical barrier against adverse environmental conditions is therefore an approach currently receiving considerable interest. The technology of micro-encapsulation of probiotic bacterial cells evolved from the immobilised cell culture technology used in the biotechnological industry. Several methods of micro-encapsulation of probiotic bacteria have been reported and include spray drying, extrusion, emulsion and phase separation. None of these reported methods however, has resulted in the large numbers of shelf-stable, viable probiotic bacterial cells necessary for use in industry for development of new probiotic products. The most commonly reported micro-encapsulation procedure is based on the calcium-alginate gel capsule formation. Kappa-carrageenan, gellan gum, gelatin and starch are also used as excipients for the micro-encapsulation of probiotic bacteria. The currently available equipment for micro-encapsulation is not able to generate large quantities of uniform sized micro or nano capsules. There is a need to design and develop equipment that will be able to generate precise and uniform micro or nano capsules in large quantities for industrial applications. The reported food vehicles for delivery of encapsulated probiotic bacteria are yoghurt, cheese, ice cream and mayonnaise. Studies need to be done on the application of micro-encapsulation of probiotic bacteria in other food systems. The number of probiotic supplements will increase in the future. More studies, however, need to be conducted on the efficacy of micro-encapsulation to deliver probiotic bacteria and their controlled or targeted release in the gastrointestinal tract.     

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.     

Culture-Independent Analysis of Probiotic Products by Denaturing Gradient Gel Electrophoresis

In order to obtain functional and safe probiotic products for human consumption, fast and reliable quality control of these products is crucial. Currently, analysis of most probiotics is still based on culture-dependent methods involving the use of specific isolation media and identification of a limited number of isolates, which makes this approach relatively insensitive, laborious, and time-consuming. In this study, a collection of 10 probiotic products, including four dairy products, one fruit drink, and five freeze-dried products, were subjected to microbial analysis by using a culture-independent approach, and the results were compared with the results of a conventional culture-dependent analysis. The culture-independent approach involved extraction of total bacterial DNA directly from the product, PCR amplification of the V3 region of the 16S ribosomal DNA, and separation of the amplicons on a denaturing gradient gel. Digital capturing and processing of denaturing gradient gel electrophoresis (DGGE) band patterns allowed direct identification of the amplicons at the species level. This whole culture-independent approach can be performed in less than 30 h. Compared with culture-dependent analysis, the DGGE approach was found to have a much higher sensitivity for detection of microbial strains in probiotic products in a fast, reliable, and reproducible manner. Unfortunately, as reported in previous studies in which the culture-dependent approach was used, a rather high percentage of probiotic products suffered from incorrect labeling and yielded low bacterial counts, which may decrease their probiotic potential.     

Culture-independent analysis of probiotic products by denaturing gradient gel electrophoresis

In order to obtain functional and safe probiotic products for human consumption, fast and reliable quality control of these products is crucial. Currently, analysis of most probiotics is still based on culture-dependent methods involving the use of specific isolation media and identification of a limited number of isolates, which makes this approach relatively insensitive, laborious, and time-consuming. In this study, a collection of 10 probiotic products, including four dairy products, one fruit drink, and five freeze-dried products, were subjected to microbial analysis by using a culture-independent approach, and the results were compared with the results of a conventional culture-dependent analysis. The culture-independent approach involved extraction of total bacterial DNA directly from the product, PCR amplification of the V3 region of the 16S ribosomal DNA, and separation of the amplicons on a denaturing gradient gel. Digital capturing and processing of denaturing gradient gel electrophoresis (DGGE) band patterns allowed direct identification of the amplicons at the species level. This whole culture-independent approach can be performed in less than 30 h. Compared with culture-dependent analysis, the DGGE approach was found to have a much higher sensitivity for detection of microbial strains in probiotic products in a fast, reliable, and reproducible manner. Unfortunately, as reported in previous studies in which the culture-dependent approach was used, a rather high percentage of probiotic products suffered from incorrect labeling and yielded low bacterial counts, which may decrease their probiotic potential.     

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.     

Identification method based on PCR combined with automated ribotyping for tracking probiotic Lactobacillus strains colonizing the human gut and vagina

AIMS: A molecular methodology based on PCR-associated automated ribotyping was developed to specifically detect the Lactobacillus strains of two probiotic products (an orally administered lyophilized preparation and vaginal tablets) in human faeces and vaginal swabs. METHODS AND RESULTS: The 16S-23S rDNA sequences and the ribotype profiles of the probiotic lactobacilli were characterized and new species-specific primer sets were designed. The identification of faecal and vaginal lactobacilli isolated from subjects administered with the probiotic products was performed by using PCR with species-specific primers followed by strain-specific automated ribotyping. CONCLUSIONS: The PCR-ribotyping identification allowed to study the colonization patterns of the probiotic lactobacilli in the human gut and vagina evidencing the strains with the best survival capability. SIGNIFICANCE AND IMPACT OF THE STUDY: The proposed molecular method represents a powerful tool of strain-specific identification, useful for differentiating exogenous from indigenous strains in any microbial ecosystem and for rationally choosing probiotic bacteria with the best chance of survival in the host.     

Survival of Mycobacterium avium ssp. paratuberculosis in yoghurt and in commercial fermented milk products containing probiotic cultures

Aims: To assess the survival of Mycobacterium avium ssp. paratuberculosis (MAP) in yoghurt and commercial fermented milk products containing probiotic strains. Methods and Results: Whole and skimmed UHT milk artificially inoculated with MAP were used to manufacture yoghurt, using two different yoghurt starter cultures. Five commercial fermented milk products were inoculated with MAP. Two different MAP strains were studied. The survival of MAP in all products was monitored by culture over a 6‐week storage period at 6°C. In yoghurt, MAP counts did not change appreciably during the storage period. Fat content and type of yoghurt starter culture had no consistent effect on the survival of MAP. In the fermented milk products, survival patterns varied but resulted in a 1·5 to ≥3·8 log reduction for the Niebüll strain and a 1·2–2·2 log reduction for the NIZO strain after 6 weeks, depending on the probiotic starters present in the product. Conclusions: MAP easily survived in yoghurt but MAP numbers decreased in fermented milk products containing probiotic cultures. Significance and Impact of the Study: The results contribute to the lack of knowledge on the behaviour of MAP in yoghurt and fermented milk products containing probiotic cultures. This knowledge is valuable in the context of the risk of MAP transmission to humans via yoghurt and the possible contribution of probiotic fermented milk products to the elimination of MAP.     

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.     

Draft Genome Sequence of Lactobacillus casei W56

We announce the draft genome sequence of Lactobacillus casei W56 in one contig. This strain shows immunomodulatory and probiotic properties. The strain is also an ingredient of commercially available probiotic products.     

The Use of Probiotic-Loaded Single- and Double-Layered Microcapsules in Cake Production

Probiotics and Antimicrobial Proteins - To date, the probiotic product development studies have mostly focused on dairy-based foods. However, endowing bakery products with probiotic properties not...     

Probiotic meat products and human nutrition

Raw cured and ripened meat products have been traditionally manufactured using the fermentation of native or added carbohydrates by lactic acid bacteria found in meat or in its environment. The commercial application of probiotic microorganisms in dry fermented meat products is not yet common. Probiotic bacterial strains that can be used in the manufacturing of dry fermented meat products should be capable of surviving in conditions found in fermented products; furthermore, they should dominate other microorganisms found in the finished product. The initial number of microorganisms in sausage filling or on the surface of ham or loin cannot be reduced as in milk pasteurization, for example. Therefore, the choice of appropriate microorganisms is important. Probiotic meat products are a relatively new and not very well recognized field of meat industry, but the most important issue is to find a compromise between technological aspects, safety, quality and health-beneficial effects of food. Therefore, the object of this review is on the one hand to analyze technological possibilities and quality parameters of probiotic meat products, and on the other hand to discuss risks and benefits of probiotic meat used in human nutrition.     

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.     

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.     

Beneficial microbiota. Probiotics and pharmaceutical products in functional nutrition and medicine

The article is mainly devoted to such representatives of gut microbiota as lactic acid bacteria and bifidobacteria, with minor accent on less frequently used or new probiotic microorganisms. Positive effects in treatment and prevention of diseases by different microbial groups, their metabolites and mechanisms of action, management and market of probiotic products are considered.     

Probiotic bacteria: safety, functional and technological properties

During the past two decades probiotic (health promoting) micro-organisms have been increasingly included in various types of food products, especially in fermented milks. Several aspects, including safety, functional and technological characteristics, have to be taken into consideration in the selection process of probiotic micro-organisms. Safety aspects include specifications such as origin (healthy human GI-tract), non-pathogenicity and antibiotic resistance characteristics. Functional aspects include viability and persistence in the GI-tract, immunomodulation, antagonistic and antimutagenic properties. Before probiotic strains, chosen on the basis of their good safety and functional characteristics, can benefit the consumer, they must first be able to be manufactured under industrial conditions. Furthermore, they have to survive and retain their functionality during storage, and also in the foods into which they are incorporated without producing off-flavours. Factors related to the technological and sensory aspects of probiotic food production are of utmost importance since only by satisfying the demands of the consumer can the food industry succeed in promoting the consumption of functional probiotic products in the future.     

Rapid identification of Lactobacillus and Bifidobacterium in probiotic products using multiplex PCR

Lactic acid bacteria (LAB) are beneficial for the gastrointestinal tract and reinforce immunity in human health. Recently, many functional products using the lactic acid bacteria have been developed. Among these LAB, Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum, and Bifidobacterium bifidum are frequently used for probiotic products. In order to monitor these LAB in commercial probiotic products, a multiplex PCR method was developed. We designed four species-specific primer pairs for multiplex PCR from the 16S rRNA, 16S-23S rRNA intergenic spacer region, and 23S rRNA genes in Lactobacillus acidophilus, Lactobacillus rhamnosus, Bifidobacterium longum, and Bifidobacterium bifidum. Using these primer pairs, 4 different LAB were detected with high specificity in functional foods. We suggest that the multiplex PCR method developed in this study would be an efficient tool for simple, rapid, and reliable identification of LAB used as probiotic strains.     

Influence of a combination of two potential probiotic strains, Lactobacillus rhamnosus IMC 501® and Lactobacillus paracasei IMC 502® on bowel habits of healthy adults

Aims: This study aims to investigate the effect of different kinds of food products enriched with a combination of two potential probiotic strains, Lactobacillus rhamnosus IMC 501^® and Lactobacillus paracasei IMC 502^®, on bowel habits of healthy adults. Methods and Results: Fifty healthy volunteers took part in a double‐blind placebo probiotic feeding study (25 fed probiotics, 25 fed placebo) for 12 weeks. Each volunteer ingested daily one or more food products enriched with a combination of the two potential probiotic strains (probiotic group) or the same food products without the probiotics (control group). Faecal samples were collected before, at the end and 2 weeks later the intervention period, and some of the main groups of faecal bacteria were enumerated by plate count and real‐time PCR. Questionnaires on bowel habits were submitted to volunteers. After the intervention, a significant increase in faecal lactobacilli and bifidobacteria were observed in the probiotic group, and stool frequency and stool volume were higher in the probiotic group than in the placebo group. Conclusions: Daily consumption of food products enriched with the two potential probiotic strains, Lact. rhamnosus IMC 501^® and Lact. paracasei IMC 502^®, contributes to improve intestinal microbiota with beneficial properties and enhances bowel habits of healthy adults. Significance and Impact of the Study: The study revealed that Lact. rhamnosus IMC 501^® and Lact. paracasei IMC 502^® exert a positive effect, in terms of improved bowel habits, on healthy adults.     

Development of Yoghurt with Juçara Pulp (<Emphasis Type="Italic">Euterpe edulis</Emphasis> M.) and the Probiotic <Emphasis Type="Italic">Lactobacillus acidophilus</Emphasis> La5

Yoghurts are dairy products consumed worldwide and can be supplemented with substances that provide extra health benefits as well as probiotic strains. In this context, the present study aimed to prepare a yoghurt added of juçara (Euterpe edulis M.) pulp and the commercial probiotic strain Lactobacillus acidophilus La5. Moreover, the probiotic survival during storage and after in vitro exposure to simulated gastric and enteric conditions was evaluated. Four formulations of yoghurt were prepared: (a) natural yoghurt, (b) yoghurt added of probiotic, (c) yoghurt added of juçara pulp, and (d) yoghurt added of probiotic culture and juçara pulp. The preparations were evaluated for survival of probiotic strain during storage and its tolerance to gastric and enteric conditions in vitro. The probiotic population in yoghurt remained unchanged during 28 days of storage. In addition, juçara pulp increased the probiotic resistance to simulated gastric and enteric conditions in the first day of storage. These data indicate that juçara pulp is a potential ingredient for the production of probiotic yoghurts.     

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.