Cyclopropane fatty acid synthase mutants of probiotic human-derived Lactobacillus reuteri are defective in TNF inhibition

Probiotics are live microorganisms that confer health benefits to the host when administered in adequate amounts. Many probiotic strains are now marketed to consumers and include organisms such as lactobacilli, streptococci, bifidobacteria, Escherichia coli Nissle 1917, and the yeast Saccharomyces cerevisiae strain, “Saccharomyces boulardii”. In general, probiotics are considered safe for human consumption based on previous clinical trials, epidemiological studies, and historical usage in fermented foods. A subset of these probiotics have been derived from commensal bacteria indigenous to Homo sapiens. One such indigenous Lactobacillus species, Lactobacillus reuteri, includes a variety of strains derived from human breast milk and the gastrointestinal tract. The mechanisms by which commensal-derived probiotics promote health and combat diseases are poorly understood. Possible mechanisms include improving nutrient absorption, pathogen exclusion, strengthening intestinal barrier function and regulation of the immune system. A better understanding of how probiotics influence health is critical to fully optimize the integrative physiology of commensal microbial communities and mammalian hosts.     

Survival of Lactobacillus acidophilus and Bifidobacterium sp. in the small intestine following ingestion in fermented milk. A rational basis for the use of probiotics in man]

Oro-ileal intubation was performed in 6 healthy volunteers who ingested, either 100 g of a fermented milk containing 10(8)/g Lactobacillus acidophilus and 10(7)/g Bifidobacterium sp or sterilized fermented milk along with a meal in random order. Lactobacillus acidophilus and Bifidobacterium were counted in the ileal fluid which was aspirated continuously for 8 h, and flow rates were calculated using the constant slow infusion of PEG 4000. After ingestion of fermented milk but not after control, hourly ileal flow rates of Lactobacillus acidophilus and Bifidobacterium increased form 4.8 +/- 0.2 and 4.9 +/- 0.6 to 7.2 +/- 0.3 and 8.0 +/- 0.3, respectively (mean +/- SE log10 CFU). 8.3 +/- 0.2 Lactobacillus acidophilus and 8.8 +/- 0.1 Bifidobacterium were recovered in the ileum which represented 1.5 percent and 37.5 percent of the ingested bacteria, respectively. In conclusion, under usual conditions of fermented milk ingestion, a large number of living Lactobacillus acidophilus and Bifidobacterium pass through the upper gastrointestinal tract and reach the colon.     

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.     

Impact of a probiotic fermented milk in the gut ecosystem and in the systemic immunity using a non-severe protein-energy-malnutrition model in mice

Background

Malnutrition affects the immune response, causing a decrease of defence mechanisms and making the host more susceptible to infections. Probiotics can reconstitute the intestinal mucosa and stimulate local and systemic immunity. The aim of this work was evaluate the effects of a probiotic fermented milk as a complement of a re-nutrition diet, on the recovery of the intestinal barrier, and mucosal and systemic immune functions in a murine model of non-severe protein-energy-malnutrition. Its potential protection against Salmonella enterica serovar Typhimurium (S. Typhimurium) infection was also analyzed.

Methods

Mice were undernourished and divided into 3 groups according to the dietary supplement received during re-nutrition (milk, probiotic fermented milk or its bacterial free supernatant) and compared to well-nourished and malnourished mice. They were sacrificed previous to the re-nutrition and 5 days post re-nutrition. The phagocytic activity of macrophages from spleen and peritoneum and the changes in the intestinal histology and microbiota were evaluated. Different immune cell populations and cytokine productions were analyzed in the small intestine tissues. The effect of the re-nutrition supplements on the systemic immunity using OVA antigen and against an infection with S. Typhimurium was also studied.

Results

Probiotic fermented milk was the most effective re-nutrition diet that improved the intestinal microbiota. Its administration also increased the number of IgA+ cells, macrophages and dendritic cells. The production of different cytokine (IFN-γ, TNF-α, IL-12) by these cells and the phagocytic activity in peritoneum and spleen was also increased. This re-nutrition diet also stimulated the systemic immune response against OVA antigen which was diminished after the malnutrition period and also improved the host response against S. Typhimurium, decreasing the spread of pathogenic bacteria to the liver and the spleen. The importance of the metabolites released during milk fermentation was also demonstrated through the analysis of the bacterial free supernatant obtained from the probiotic fermented milk, but the whole product showed the best effects in the parameters evaluated in this study.

Conclusions

The administration of probiotic fermented milk as a dietary supplement during the re-nutrition process in a murine immunodeficiency model by malnutrition could be a good adjuvant diet to improve the gut and systemic immune response for the protection against Salmonella infection.     

Ileal losses of nitrogen and amino acids in humans and their importance to the assessment of amino acid requirements

BACKGROUND & AIMS: Irreversible amino acid losses at the human ileum are not taken into account when tracer-derived amino acid requirements are calculated because the data available are scarce. We have investigated amino acid losses at the ileal level in humans after ingestion of a protein meal. METHODS: Thirteen volunteers ingested a single meal of 15N milk or soy proteins. The appearance of 15N and 15N amino acids in the ileal effluents collected using an ileal tube was monitored for 8 hours. RESULTS: In the soy group, higher losses of endogenous nitrogen, especially originating from amino acids, were observed, as well as a higher flow rate of dietary non-amino acid nitrogen. With soy protein, the digestibilities of valine, threonine, histidine, tyrosine, alanine, and proline were significantly lower than with milk. Ileal losses of leucine, valine, and isoleucine amounted to 12, 10, and 7 mg x kg(-1) x day(-1), respectively. Threonine ileal loss (9-12 mg x kg(-1) x day(-1)) was particularly high compared with the current amino acid requirement. CONCLUSIONS: Amino acid losses at the human terminal ileum are substantial and depend on the type of dietary protein ingested. Although it remains unclear whether intact amino acids are absorbed in the colon, we suggest that ileal losses should be considered an important component of amino acid requirements.     

Impact of probiotic supplements on microbiome diversity following antibiotic treatment of mice

Shifts in microbial populations of the intestinal tract have been associated with a multitude of nutritional, autoimmune, and infectious diseases. The limited diversity following antibiotic treatments creates a window for opportunistic pathogens, diarrhea, and inflammation as the microbiome repopulates. Depending on the antibiotics used, microbial diversity can take weeks to months to recover. To alleviate this loss of diversity in the intestinal microbiota, supplementation with probiotics has become increasingly popular. However, our understanding of the purported health benefits of these probiotic bacteria and their ability to shape the microbiome is significantly lacking. This study examined the impact of probiotics concurrent with antibiotic treatment or during the recovery phase following antibiotic treatment of mice. We found that probiotics did not appear to colonize the intestine themselves or shift the overall diversity of the intestinal microbiota. However, the probiotic supplementation did significantly change the types of bacteria which were present. In particular, during the recovery phase the probiotic caused a suppression of Enterobacteriaceae outgrowth (Shigella and Escherichia) while promoting a blooming of Firmicutes, particularly from the Anaerotruncus genus. These results indicate that probiotics have a significant capacity to remodel the microbiome of an individual recovering from antibiotic therapy.     

Amelioration of DSS-induced murine colitis by VSL#3 supplementation is primarily associated with changes in ileal microbiota composition.

Inflammatory bowel diseases encompass gastrointestinal illnesses typified by chronic inflammation, loss of epithelial integrity and gastrointestinal microbiota dysbiosis. In an effort to counteract these characteristic perturbations, we used stem cells and/or a probiotic therapy in a murine model of Dextran Sodium Sulfate induced colitis to examine both their efficacy in ameliorating disease and impact on niche-specific microbial communities of the lower GI tract. Colitis was induced in C57BL/6 mice by administering 3% DSS in drinking water for 10 days prior to administering one of three treatment plans: daily probiotic (VSL#3) supplementation for 3 days, a single tail vein injection of 1x10⁶ murine mesenchymal stem cells, or both. Ileal, cecal and colonic sections were collected for microbiota and histological analyses. Microbiota profiling revealed distinct bacterial community compositions in the ileum, cecum and colon of control untreated animals, all of which were predicted in silico to be enriched for a number of discrete KEGG pathways, indicating compositional and functional niche specificity in healthy animals. DSS-treatment perturbed community composition in all three niches with ileal communities exhibiting the greatest change relative to control animals. Each treatment group exhibited treatment-specific alterations in microbiota composition in the lower GI tract, though disease scores were only improved in VSL#3-treated animals. The ileal microbiota were most profoundly altered in composition in this group of animals and characterized by significant Enterobacteriaceae enrichment compared with colitic mice (P < 0.05).     

Lactose-reduced infant formula with added corn syrup solids is associated with a distinct gut microbiota in Hispanic infants

ABSTRACT

Infant formula feeding, compared with human milk, has been associated with development of a distinct infant gut microbiome, but no previous study has examined effects of formula with added sugars. This work examined differences in gut microbiota among 91 Hispanic infants who consumed human milk [at breast (BB) vs. pumped in bottle (BP)] and 2 kinds of infant formula [(traditional lactose-based (TF) vs. lactose-reduced with added sugar (ASF)]. At 1 and 6 months, infant stool was collected to characterize gut microbiota. At 6 months, mothers completed 24-hour dietary recalls and questionnaires to determine infant consumption of human milk (BB vs. BP) or formula (TF vs. ASF). Linear regression models were used to determine associations of milk consumption type and microbial features at 6 months. Infants in the formula groups exhibited a significantly more ‘mature’ microbiome than infants in the human milk groups with the most pronounced differences observed between the ASF vs. BB groups. In the ASF group, we observed reduced log-normalized abundance of Bifidobacteriaceae (TF-BB Mean Difference = ?0.71, ASF-BB Mean Difference = ?1.10), and increased abundance of Lachnospiraceae (TF-BB Mean Difference = +0.89, ASF-BB Mean Difference = +1.20). We also observed a higher Community Phenotype Index of propionate, most likely produced by Lachnospiraceae, in the ASF group (TF-BB Mean Difference = +0.27, ASF-BB Mean Difference = +0.36). This study provides the first evidence that consumption of infant formula with added sugar may have a stronger association than birth delivery mode, infant caloric intake, and maternal BMI on the infant’s microbiome at 6 months of age.     

Effects of probiotic fermented milk on symptoms and intestinal flora in patients with irritable bowel syndrome: A randomized,placebo-controlled trial

Abstract

Objective. The effect of probiotics on IBS symptoms has been mixed, but remains an intriguing treatment option with appeal to the patient. Material and methods. Patients fulfilling the Rome II criteria were randomized double-blind to a daily intake of 500 ml of fermented milk containing at least 5 × 10⁷ CFU/ml of Lactobacillus paracasei ssp paracasei F19, Lactobacillus acidophilus La5 and Bifidobacterium lactis Bb12 or an equal volume of acidified milk for 8 weeks. Symptoms were assessed at baseline and weekly using a disease-specific validated symptom rating scale (IBS-SSI). The predefined primary outcome measure was patient reported adequate symptom relief. Adherence to study protocol were assessed by performing stool samples at the of the treatment period. Results. Eight-one patients were screened. Sixty-four patients were randomized; 18 patients did not complete the study due to protocol violations or withdrew due to lack of effect. Fifty-two patients (13 males) completed the study as per protocol; mean age was 51.3 years (range 29–67). The proportion of patients reporting adequate symptom relief increased in both patient groups, but there was not any statistical difference between the groups. IBS-SSI scores did not differ statistically between the groups at the end of the treatment period, but improved during the study period in both groups. Conclusions. During this 8-week trial gastrointestinal symptoms improved. However, there was no difference between treatment with fermented milk containing probiotics or acidified milk. The effect of probiotics on IBS symptoms remains uncertain and further studies are warranted.     

A novel probiotic,Lactobacillus johnsonii 456, resists acid and can persist in the human gut beyond the initial ingestion period

ABSTRACT

Probiotics are considered to have multiple beneficial effects on the human gastrointestinal tract, including immunomodulation, pathogen inhibition, and improved host nutrient metabolism. However, extensive characterization of these properties is needed to define suitable clinical applications for probiotic candidates. Lactobacillus johnsonii 456 (LBJ 456) was previously demonstrated to have anti-inflammatory and anti-genotoxic effects in a mouse model. Here, we characterize its resistance to gastric and bile acids as well as its ability to inhibit gut pathogens and adhere to host mucosa. While bile resistance and in vitro host attachment properties of LBJ 456 were comparable to other tested probiotics, LBJ 456 maintained higher viability at lower pH conditions compared to other tested strains. LBJ 456 also altered pathogen adhesion to LS 174T monolayers and demonstrated contact-dependent and independent inhibition of pathogen growth. Genome analyses further revealed possible genetic elements involved in host attachment and pathogen inhibition. Importantly, we show that ingestion of Lactobacillus johnsonii 456 over a one week yogurt course leads to persistent viable bacteria detectable even beyond the period of initial ingestion, unlike many other previously described probiotic species of lactic acid bacteria.     

Glycated milk protein fermented with Lactobacillus rhamnosus ameliorates the cognitive health of mice under mild-stress condition

ABSTRACT

This study aimed to investigate the effects of glycated milk casein (Gc) fermented with Lactobacillus rhamnosus 4B15 (FGc) on the intestinal microbiota and physiological and behavioral properties in mice under chronic stress. Mice were administered Gc or FGc for 10 weeks and then exposed to unpredictable chronic mild stress (UCMS) for 7 weeks. FGc administration restored alterations of gut microbiota induced by UCMS. Moreover, FGc significantly reduced the stress-induced increase in serum corticosterone and decrease in serotonin levels. Anxiety-like behaviors induced by UCMS were also significantly decreased in the FGc group. UCMS-induced dysregulation of gene and protein expression related to neuroendocrine function, neuronal development, and inflammation, and gut-blood-brain barrier function was controlled by FGc pre-treatment. These results strongly suggest the protective effects of FGc targeting of intestinal microbiota for abnormal brain activity, which is consistent with the view that FGc plays an important role in regulating stress-related gut-brain axis disorders.     

Human milk microbiota development during lactation and its relation to maternal geographic location and gestational hypertensive status

ABSTRACT

Bacteria in human milk could directly seed the infant intestinal microbiota, while information about how milk microbiota develops during lactation and how geographic location, gestational hypertensive status, and maternal age influence this process is limited. Here, we collected human milk samples from mothers of term infants at the first day, 2 weeks, and 6 weeks postpartum from 117 longitudinally followed-up mothers (age: 28.7 ± 3.6 y) recruited from three cities in China. We found that milk microbial diversity and richness were the highest in colostrum but gradually decreased over lactation. Microbial composition changed across lactation and exhibited more discrete compositional patterns in 2-week and 6-week milk samples compared with colostrum samples. At phylum level, the abundance of Proteobacteria increased during lactation, while Firmicutes showed the opposite trend. At genus level, Staphylococcus, Streptococcus, Acinetobacter, Pseudomonas, and Lactobacillus were predominant in colostrum samples and showed distinct variations across lactation. Maternal geographic location was significantly associated with the milk microbiota development and the abundance of predominant genus. In addition, milk from mothers with gestational prehypertension had a different and less diverse microbial community at genus level in early lactation times, and contained less Lactobacillus in the 2-week milk samples than those from normotensive mothers. Findings of our study outlined the human milk microbial diversity and community development over lactation, and underscored the importance of maternal geographic locations and gestational hypertensive status on milk microbiota, which might have important implications in the establishment of the infant intestinal microbiota via breastfeeding.     

Effect of Lactobacillus casei Shirota on colonic transit time in patients with chronic constipation

Background

Slow-transit constipation (STC) is caused by a motility disorder of the colon which leads to delayed transit (>72?h). The probiotic strain Lactobacillus casei Shirota (LcS) has been shown to improve constipation-related symptoms, such as stool frequency and consistency. A randomized double-blind placebo-controlled trial was performed to determine the effect of LcS on the colonic transit time in patients with STC.

Patients and methods

Colonic transit time of all consecutive outpatients with chronic constipation was determined by the Hinton test using radiopaque markers. Patients with a transit time longer than 72?h were included in the study. A total of 24 patients received either a dairy drink containing 6.5×10⁹ colony forming units (cfu) of LcS or a placebo daily for 4 weeks. General gastrointestinal symptoms were evaluated weekly by a questionnaire and the measurement of colonic transit time was repeated after the intervention.

Results

The intake of LcS resulted in a significant acceleration of the total colonic transit time from 95.6?h to 76.5?h (p=0.05). This effect was most pronounced in the sigmoid and rectum transit time (p<0.007). In the placebo group no statistically significant change in the total colonic transit time was observed (before: 95.8?h, after: 87.1?h, p=0.282)

Conclusion

The daily intake of a probiotic drink containing LcS significantly reduced the colonic transit time in patients with STC.     

Low dietary fiber intake increases Collinsella abundance in the gut microbiota of overweight and obese pregnant women

The gut microbiota contributes to the regulation of glucose metabolism in pregnancy. Abundance of the genus Collinsella is positively correlated with circulating insulin; however, it is unclear what determines Collinsella abundance. This study aims to validate the correlation between Collinsella and insulin and to elucidate if macronutrient intake alters Collinsella abundance and gut microbiota composition.

Gut microbiota profiles were assessed by 16S rRNA sequencing in 57 overweight and 73 obese pregnant women from the SPRING (Study of PRobiotics IN Gestational diabetes) trial at 16 weeks gestation and correlated with metabolic hormone levels and macronutrient intake. Gut microbiota composition in the top and bottom 10% of dietary fiber intake was evaluated through network analysis.

Collinsella abundance correlated positively with circulating insulin (rho = 0.30, p = 0.0006), independent of maternal BMI, but negatively with dietary fiber intake (rho = ?0.20, p = 0.025) in this cohort. Low dietary fiber intake was associated with a gut microbiota favoring lactate fermentation while high fiber intake promotes short-chain fatty acid-producing bacteria.

Low dietary fiber may enable overgrowth of Collinsella and alter the overall fermentation pattern in gut microbiota. This suggests that dietary choices during pregnancy can modify the nutritional ecology of the gut microbiota, with potential deleterious effects on the metabolic and inflammatory health of the host.

Trial registration: ANZCTR 12611001208998, registered 23/11/2011     

Ability of Lactobacillus kefiri LKF01 (DSM32079) to colonize the intestinal environment and modify the gut microbiota composition of healthy individuals

BackgroundProbiotics have been observed to positively influence the host’s health, but to date few data about the ability of probiotics to modify the gut microbiota composition exist.AimsTo evaluate the ability of Lactobacillus kefiri LKF01 DSM32079 (LKEF) to colonize the intestinal environment of healthy subjects and modify the gut microbiota composition.MethodsTwenty Italian healthy volunteers were randomized in pre-prandial and post-prandial groups. Changes in the gut microbiota composition were detected by using a Next Generation Sequencing technology (Ion Torrent Personal Genome Machine).ResultsL. kefiri was recovered in the feces of all volunteers after one month of probiotic administration, while it was detected only in three subjects belonging to the pre-prandial group and in two subjects belonging to the post-prandial group one month after the end of probiotic consumption. After one month of probiotic oral intake we observed a reduction of Bilophila, Butyricicomonas, Flavonifractor, Oscillibacter and Prevotella. Interestingly, after the end of probiotic administration Bacteroides, Barnesiella, Butyricicomonas, Clostridium, Haemophilus, Oscillibacter, Salmonella, Streptococcus, Subdoligranolum, and Veillonella were significantly reduced if compared to baseline samples.ConclusionL. kefiri LKF01 showed a strong ability to modulate the gut microbiota composition, leading to a significant reduction of several bacterial genera directly involved in the onset of pro-inflammatory response and gastrointestinal diseases.     

Probiotics for the treatment of type 2 diabetes: A review of randomized controlled trials

With the increasing prevalence of type 2 diabetes mellitus (T2DM), there is increased interest in probiotic supplementation for improving glycaemic control. This review evaluates nine randomized controlled trials that tested the effects of probiotics on glycaemic outcomes including fasting plasma glucose, fasting plasma insulin, haemoglobin A1c, and homeostatic model assessment of insulin resistance among adults with T2DM. Based on the evidence reviewed, multistrain probiotics that contain seven million to 100 billion colony forming units of Lactobacillus acidophilus, Streptococcus thermophilus, Lactobacillus bulgaricus, and/or Bifidobacterium lactis administered for 6 to 12 weeks may be efficacious for improving glycaemic control in adults with T2DM. Further research is needed to understand the role of the gut microbiota and the probiotic dose, medium, and duration of exposure that is most effective for disease management.     

Prophylactic probiotics reduce cow's milk protein intolerance in neonates after small intestine surgery and antibiotic treatment presenting symptoms that mimics postoperative infection

BackgroundTo examine occurrence of cow's milk protein intolerance (CMPI) in newborns that underwent small intestine surgery and the clinical profiles of those newborns with postoperative CMPI, and to evaluate the preventive effects of probiotics on CMPI.MethodsWe retrospectively reviewed from 2000 to 2009, a total of 30 newborns required surgery on their small intestines. All of these patients had received antibiotics to prevent postoperative infection. Since 2005 we adopted a protocol of targeted probiotic therapy prophylaxis.ResultsEighteen patients received probiotic therapy, while twelve did not. One infant among those eighteen patients and eight patients among those twelve developed CMPI, a significantly lower rate for the group with probiotic therapy than that without it (p < 0.001). Patients with positive cultures for gram positive and gram negative organisms increased in number before and after surgery but then decreased after probiotics treatment. Poor weight gain, gastrointestinal symptoms, and rise in C reactive protein (CRP) levels were observed in all of those nine CMPI patients. Specific IgE antibodies were elevated in four of the nine subjects, and total IgE levels were elevated in seven of them. All CMPI patients had increased level of CRP without proven infections.ConclusionsCMPI was induced in newborns after surgery on their small intestines and antibiotics treatment with presentation of symptoms that mimic postoperative infection. Development of CMPI in this population possibly involves disruption of intestinal flora. Administration of probiotics can reduce the incidence of CMPI after small intestine surgery. The elevated CRP level may be useful in the diagnosis of CMPI.     

Terminal ileal transposition procedure in ileoanal anastomosis following proctocolectomy

We introduced a terminal ileal transposition procedure (TITP) in ileal pouch-anal anastomosis, in which a 50 to 70 cm isolated ileal segment 20 to 40 cm from the ileocecal valve was interposed between the terminal ileum and the anus. Twelve patients underwent this procedure in two or three-staged operations. Mean stool frequency per 24 hours was 4.4±1.7, and stool consistency was formed and soft in all patients at the mean of 13 months after TITP. We observed neither surgical technique-related complications nor metabolic disorders, except for iron deficiency anemia, during and after the operations. The serum level of vitamin B12 significantly increased after the operation in eight patients (P<0.05). TITP has advantages such as preventing the terminal ileum from metabolic dysfunction due to pouchitis, avoiding sacrifice of the terminal ileum in the two-staged operation, and obviating the need for reconstruction of ileostomy in the three-staged operation. It may also promote intestinal absorption and reduce late metabolic complications. Accepted: 18 July 1997     

Intestinal microbiota and blue baby syndrome

Necrotizing enterocolitis (NEC) is the most common intestinal emergency among premature infants. Risk factors in premature infants include immature intestinal immunity and an intestinal microbiota dominated by hospital-acquired bacteria. Some probiotics have been shown to decrease the incidence of NEC in premature infants. Among term infants, NEC is rare. However, among term infants with cyanotic congenital heart disease (CCHD), the incidence of NEC is similar to that of premature infants but with even greater mortality rates. Mechanisms by which NEC occurs in term infants with CCHD are unknown. Of central interest is the potential role of changes in the intestinal microbiota and whether these can be modified with probiotic bacteria; accordingly, we review the literature, propose hypotheses, and present the rationale for future studies involving preliminary probiotic clinical trials.