Expert consensus document: The International Scientific Association for Probiotics and Prebiotics (ISAPP) consensus statement on the definition and scope of prebiotics.

In December 2016, a panel of experts in microbiology, nutrition and clinical research was convened by the International Scientific Association for Probiotics and Prebiotics to review the definition and scope of prebiotics. Consistent with the original embodiment of prebiotics, but aware of the latest scientific and clinical developments, the panel updated the definition of a prebiotic: a substrate that is selectively utilized by host microorganisms conferring a health benefit. This definition expands the concept of prebiotics to possibly include non-carbohydrate substances, applications to body sites other than the gastrointestinal tract, and diverse categories other than food. The requirement for selective microbiota-mediated mechanisms was retained. Beneficial health effects must be documented for a substance to be considered a prebiotic. The consensus definition applies also to prebiotics for use by animals, in which microbiota-focused strategies to maintain health and prevent disease is as relevant as for humans. Ultimately, the goal of this Consensus Statement is to engender appropriate use of the term 'prebiotic' by relevant stakeholders so that consistency and clarity can be achieved in research reports, product marketing and regulatory oversight of the category. To this end, we have reviewed several aspects of prebiotic science including its development, health benefits and legislation.

Probiotics in dietary guidelines and clinical recommendations outside the European Union.

Fermented foods have been consumed for centuries across many geographical locales and have traditionally been considered healthy foods, partly because of the live microbes contained in them. The concept of "probiotics" further requires that the microbes be defined and their health effects be demonstrated through human intervention studies or other suitable investigations before marketing with corresponding health messages. Here, we review recommendations for fermented foods and probiotics in several countries outside the EU, focusing on food-based dietary guidelines. We emphasize recommendations on yoghurt and probiotics made by expert bodies. We found that dietary guidelines commonly advocate the consumption of yoghurt or similar products, but specific comments on probiotics are rare. Further, we reviewed guidelines from clinical associations. In general, they acknowledge the beneficial effects of probiotics, but often suggest the need for further research. This is true despite good quality evidence supporting the role of probiotics for certain health effects, such as prevention of eczema in infants, management of side effects from antibiotics and alleviation of functional bowel symptoms. Additional research to support future dietary recommendations should focus on determining effect size, identifying responders and non-responders, clarifying strain-specificity of effects and confirming mechanisms.

Removal of cholera toxin from aqueous solution by probiotic bacteria.

Cholera remains a serious health problem, especially in developing countries where basic hygiene standards are not met. The symptoms of cholera are caused by cholera toxin, an enterotoxin, which is produced by the bacterium Vibrio cholerae. We have recently shown that human probiotic bacteria are capable of removing cyanobacterial toxins from aqueous solutions. In the present study we investigate the ability of the human probiotic bacteria, Lactobacillus rhamnosus strain GG (ATCC 53103) and Bifidobacterium longum 46 (DSM 14583), to remove cholera toxin from solution in vitro. Lactobacillus rhamnosus strain GG and Bifidobacterium longum 46 were able to remove 68% and 59% of cholera toxin from aqueous solutions during 18 h of incubation at 37 °C, respectively. The effect was dependent on bacterial concentration and L. rhamnosus GG was more effective at lower bacterial concentrations. No significant effect on cholera toxin concentration was observed when nonviable bacteria or bacterial supernatant was used.

Probiotics modulate the Bifidobacterium microbiota of elderly nursing home residents.

Gut Bifidobacterium microbiota of the elderly has been suggested to differ from that of adults, possibly promoting the risk of infections and gut barrier dysfunction. Specific probiotics may improve the gut barrier. In this randomized, placebo-controlled intervention study, 66 elders consumed a fermented oat drink containing probiotic Bifidobacterium longum 46 and B. longum 2C or a non-fermented placebo oat drink for 6 months. Faecal samples were collected before, during and after the intervention. Levels of faecal bifidobacteria were determined using species-specific quantitative PCR and plate counting. The Bifidobacterium levels in the elderly were high and the species composition diverse. Probiotic intervention increased the levels bifidobacteria significantly. Specifically, the levels of B. catenulatum, B. bifidum and B. breve were enhanced. Consumption of the fermented oat drink itself was also associated with certain changes in microbiota. In conclusion, Bifidobacterium microbiota of elderly subjects may be modulated by probiotic administration. In some healthy elderly populations, Bifidobacterium microbiota may be more abundant and diverse than previously suggested.

Specific strains of probiotic bacteria are efficient in removal of several different cyanobacterial toxins from solution.

The ability of specific strains of probiotic bacteria to remove the pure cyanobacterial peptide toxins microcystin-LR, -RR, -LF, and a combination of microcystins from the cyanobacterial extracts Microcystis PCC 7820 and NIES 107, as well as the cyanobacterial cytotoxin cylindrospermopsin, from aqueous solutions was assessed. The probiotic bacterial strains studied were Lactobacillus rhamnosus strains GG and LC-705, Bifidobacterium lactis strains 420 and Bb12 and Bifidobacterium longum 46, all previously shown to be effective in toxin removal. The maximum removal of microcystin-LR, 60.3%, was observed with L. rhamnosus GG, of microcystin-RR, 62.8%, and microcystin-LF, 77.4%, with L. rhamnosus LC-705, and of cylindrospermopsin, 31.6%, with B. longum 46 (toxin concentration 100mugL(-1), 37 degrees C, 24h). Several microcystins could be removed simultaneously as observed by removal of microcystins present in the cyanobacterial extracts. A combination of three probiotic strains enhanced their removal ability as compared to the removal properties of the individual strains. We conclude that specific strains of probiotic bacteria are effective in elimination of different cyanotoxins from solution.

Effect of glucose in removal of microcystin-LR by viable commercial probiotic strains and strains isolated from dadih fermented milk.

The removal of the cyanobacterial peptide toxin microcystin-LR at 4 and 37 degrees C by six commercial probiotic strains and Lactobacillus plantarum strains IS-10506 and IS-20506 isolated from dadih, Indonesian traditional fermented milk, was assessed in this study. The aim was to evaluate the main factors influencing the viability and metabolic activity of the probiotic strains, as well as their capacity to remove microcystin-LR. Both L. plantarum strains isolated from dadih, as well as Bifidobacterium lactis strains Bb12 and 420, were shown to be more resistant, and >85% remained viable in phosphate-buffered saline (PBS) solution for 48 h of incubation at either temperature, while the viability of the other four commercial bacteria decreased markedly over time. The effect of glucose on viability and removal of toxin was shown to be a strain-specific and strain-dependent property, but in general, the efficiency of microcystin-LR removal increased when glucose was added to the solution. A maximum removal of 95% was observed for L. plantarum strain IS-20506 (37 degrees C, 10 (11) colony-forming units mL(-1)) with 1-2% glucose supplementation and 75% in PBS alone.

Specific Bifidobacterium strains isolated from elderly subjects inhibit growth of Staphylococcus aureus.

Cell-free, pH-controlled supernatants of thirty-eight Bifidobacterium strains isolated from healthy elderly subjects were subjected to antimicrobial activity assay. Bioluminescent indicator strains Staphylococcus aureus RN4220, Escherichia coli K-12, and Salmonella enterica serovar Typhimurium ATCC 14028 were used as targets of antimicrobial activity. The effect of nutrient depletion on the inhibition was eliminated with spent-culture controls. Three out of thirty-eight Bifidobacterium strains were capable of inhibiting the growth of S. aureus. The inhibition was equal to 23.2+/-19.1% to 50.4+/-26.7% of the inhibition caused by 50 IU/ml nisin. Reuterin-producing positive strain Lactobacillus reuteri SD2112 was capable of 86.0+/-24.6% inhibition, but Bifidobacterium lactis Bb-12, a known probiotic strain, showed no inhibition. None of the strains was capable of inhibiting the growth of E. coli or S. enterica. The observed inhibition by bifidobacteria was related to hydrogen peroxide formation and possible production of heat-stable proteinaceous compounds. The results suggest that production of antimicrobial substances other than organic acids is not common among Bifidobacterium strains typical of elderly subjects. However, specific strains were identified which showed considerable inhibitory activity against S. aureus.

Removal of microcystin-LR by strains of metabolically active probiotic bacteria.

The ability of specific strains of probiotic bacteria to remove the cyanobacterial peptide toxin microcystin-LR from aqueous solutions was assessed. Lactobacillus rhamnosus strains GG and LC-705, Bifidobacterium longum 46, Bifidobacterium lactis 420 and Bifidobacterium lactis Bb12 were shown to be the most effective in toxin removal among 11 tested strains. The highest removal percentage of microcystin-LR was 58.1%, observed with B. lactis Bb12 (toxin concentration 100 microg L(-1), 10(10) CFU mL(-1), 37 degrees C, 24 h). Freshly cultured bacteria were shown to be more efficient in microcystin removal than lyophilized or nonviable bacteria. Removal of microcystin-LR was shown to be dependent on both temperature and bacterial concentration. It is concluded that some of the tested strains have good potential in removing microcystins from aqueous solutions.

Comparison of four methods to enumerate probiotic bifidobacteria in a fermented food product.

Four methods of enumeration were compared by monitoring levels of probiotic bifidobacteria in fermented oat drink during storage. Strains of Bifidobacterium longum and B. lactis were quantified by plate counts, fluorescent in situ hybridization (FISH), quantitative real-time PCR and commercial LIVE/DEAD BacLight bacterial viability kit, and the methods were further developed to suit the enumeration of bifidobacteria in fermented foods. Plate counts of both B. lactis and B. longum were lower than the PCR and FISH counts. The LIVE/DEAD counts of B. lactis were comparable to PCR and FISH counts. The plate counts of B. lactis were slightly but significantly lower than LIVE/DEAD counts, suggesting that the cells that were not able to grow on plates may have become dormant. The plate counts of B. longum were several log units lower than LIVE/DEAD counts, suggesting that a remarkable part of the cells were dormant. Real-time PCR and FISH were shown to suit the quantification of the total amount of probiotic bifidobacteria in foods. Plate counts and LIVE/DEAD counts provided conflicting information on viability especially in the case of B. longum. We conclude that the choice of enumeration method for probiotic bacteria may have significant effect on the results of the analysis. The strain-specific properties and the objects of the analysis should be taken into account when enumeration methods for different probiotic strains are chosen.

Intrinsic properties of so-called dormant probiotic bacteria, determined by flow cytometric viability assays.

Plate counting and four culture-independent flow cytometric assays were used to determine the viability and intrinsic properties of three probiotic strains during storage. The strains showed reduction in plate counts but were able to maintain esterase activity, intact cytoplasmic membrane, and pH gradient. The apparently uncultivable probiotic cells were active and stress resistant.

Probiotic supplementation reduces a biomarker for increased risk of liver cancer in young men from Southern China.

BACKGROUND: In vitro and in vivo studies suggest that selected strains of probiotic bacteria can form tight complexes with aflatoxin B(1) and other carcinogens. OBJECTIVE: The aim of the present study was to determine whether administration of probiotic bacteria could block the intestinal absorption of aflatoxin B(1) and thereby lead to reduced urinary excretion of aflatoxin B(1)-N(7)-guanine (AFB-N(7)-guanine), a marker for a biologically effective dose of aflatoxin exposure. Elevated urinary excretion of this aflatoxin-DNA adduct is associated with an increased risk of liver cancer. DESIGN: Ninety healthy young men from Guangzhou, China, were randomly assigned to 2 groups; one group received a mixture of Lactobacillus rhamnosus LC705 and Propionibacterium freudenreichii subsp. shermanii strains 2 times/d for 5 wk, and the other group received a placebo preparation. The subjects provided 4 urine samples: at baseline, at 3 and 5 wk after starting the supplementation, and at the end of the 5-wk postintervention period. RESULTS: The percentage of samples with negative AFB-N(7)-guanine values tended to be higher in the probiotic group than in the placebo group during the 5-wk intervention period (odds ratio: 2.63, P = 0.052), and a statistically significant decrease in the concentration of urinary AFB-N(7)-guanine was observed in the probiotic group. The reduction was 36% at week 3 and 55% at week 5. The geometric means for the probiotic and placebo groups were 0.24 and 0.49 ng AFB-N(7)-guanine/mL, respectively, during the intervention period (P = 0.005). CONCLUSION: A probiotic supplement reduces the biologically effective dose of aflatoxin exposure and may thereby offer an effective dietary approach to decrease the risk of liver cancer.

Probiotics that modify disease risk.

Probiotics are defined as live bacterial preparations with clinically documented health effects in humans. Probiotics have specific properties and targets in the human intestinal tract and intestinal microbiota. Each probiotic strain, independent of its genus and species is unique and, thus, the properties and the human health effects of each strain have to be assessed in a case-by-case manner. Understanding the mechanisms by which probiotics influence the normal intestinal microbiota and counteract aberrancies in microbiota would facilitate the use of probiotics for both dietary management and reduction in risk of specific diseases. Development of intestinal microbiota is an important factor affecting the health of the newborn. Recent studies suggest that specific bacterial components, especially the bifidobacteria, have a key impact on development of a healthy balanced infant microbiota. The composition of infant and child intestinal microbiota may become aberrant and thus influence the development of diarrheal, inflammatory, and allergic diseases. Based on this understanding, positive health effects of probiotics have been reported in the management of diarrheal, inflammatory, and allergic diseases in infants. Most recently, a reduction in risk of atopic diseases followed early administration of specific probiotics.

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.

Similar bifidogenic effects of prebiotic-supplemented partially hydrolyzed infant formula and breastfeeding on infant gut microbiota.

The aim of the study was to assess the quantitative and qualitative differences of the gut microbiota in infants. We evaluated gut microbiota at the age of 6 months in 32 infants who were either exclusively breast-fed, formula-fed, nursed by a formula supplemented with prebiotics (a mixture of fructo- and galacto-oligosaccharides) or breast-fed by mothers who had been given probiotics. The Bifidobacterium, Bacteroides, Clostridium and Lactobacillus/Enterococcus microbiota were assessed by the fluorescence in situ hybridization, and Bifidobacterium species were further characterized by PCR. Total number of bifidobacteria was lower among the formula-fed group than in other groups (P=0.044). Total amounts of the other bacteria were comparable between the groups. The specific Bifidobacterium microbiota composition of the breast-fed infants was achieved in infants receiving prebiotic supplemented formula. This would suggest that early gut Bifidobacterium microbiota can be modified by special diets up to the age of 6 months.

Probiotic bacteria in the management of atopic disease: underscoring the importance of viability.

OBJECTIVES: The aim of this study was to assess the efficacy of oral supplementation of viable and heat-inactivated probiotic bacteria in the management of atopic disease and to observe their effects on the composition of the gut microbiota. METHODS: The study population included 35 infants with atopic eczema and allergy to cow's milk. At a mean age of 5.5 months, they were assigned in a randomized double-blind manner to receive either extensively hydrolyzed whey formula (placebo group) or the same formula supplemented with viable (viable LGG group) or heat-inactivated Lactobacillus GG (heat-inactivated LGG group), respectively. The changes in symptoms were assessed by the SCORAD method and the presence of some predominant bacterial genera in the feces detected with 16S rRNA-specific probes. RESULTS: The treatment with heat-inactivated LGG was associated with adverse gastrointestinal symptoms and diarrhea. Consequently, the recruitment of patients was stopped after the pilot phase. Within the study population, atopic eczema and subjective symptoms were significantly alleviated in all the groups; the SCORAD scores (interquartile range) decreased from 13 (range, 4-29) to 8 (range, 0-29) units in the placebo group, from 19 (range, 4-47) to 5 (range, 0-18) units in the viable LGG group, and from 15 (range, 0-29) to 7 (range, 0-26) units in the heat-inactivated LGG group. The decrease in the SCORAD scores within the viable LGG group tended to be greater than within the placebo group. The treatments did not appear to affect the bacterial numbers within the genera enumerated. CONCLUSIONS: Supplementation of infant formulas with viable but not heat-inactivated LGG is a potential approach for the management of atopic eczema and cow's milk allergy.

Influence of probiotic supplemented infant formula on composition of plasma lipids in atopic infants.

Probiotic therapy is a new, successful approach to alleviating allergic symptoms. In this study, our aim was to investigate whether the positive results obtained with probiotic therapy would be associated with the differential absorption and utilization of dietary PUFA. 15 infants referred to a pediatric clinic on the basis of atopic eczema were weaned to Bifidobacterium Bb-12 or Lactobacillus GG supplemented infant formula, or to the same formula without probiotics (randomized, placebo-controlled, double blind study design). In plasma neutral lipids, alpha-linolenic acid (18:3 n-3) proportions were reduced by the probiotic supplementation. In phospholipids, Lactobacillus GG supplemented formula did not influence alpha-linolenic acid proportions, while Bifidobacterium Bb-12 supplemented formula increased the proportion of alpha-linolenic acid; from 0.13 +/- 0.03 to 0.24 +/- 0.03 (mean +/- SEM) (P = 0.002). These results show that some physiological effects of probiotics may be associated with physiological interactions between probiotics and dietary PUFA.