Lactobacillus Rhamnosus R0011 Research Articles

The Bioavailability of Glycyrrhizinic Acid Was Enhanced by Probiotic Lactobacillus rhamnosus R0011 Supplementation in Liver Fibrosis Rats.

Glycyrrhizinic acid (GL) is clinically applied to treat liver injury, and the bioavailability of orally administered GL is closely related to the gut microbiota. Therefore, the dysbiosis of gut flora in liver injury could significantly influence GL bioavailability. Still, less is known about the impact of probiotic supplementation on the bio-absorption process of oral medication, especially under a pathological state. Herein, probiotic L. rhamnosus R0011 (R0011) with a high viability in the harsh gastrointestinal environment was selected, and the effect of R0011 on the GL bioavailability in rats was investigated. Four groups of rats (n = 6 per group) were included: the normal group (N group), the normal group supplemented with R0011 (NLGG group), CCl4-induced chronic liver injury model (M group), and the model group supplemented with R0011 (MLGG group). Our results showed that liver injury was successfully induced in the M and MLGG groups via an intraperitoneal injection of 50% (v/v) CCl4 solution. Healthy rats supplemented with R0011 could increase the bioavailability of GL by 1.4-fold compared with the normal group by plasma pharmacokinetic analysis. Moreover, the GL bioavailability of MLGG group was significantly increased by 4.5-fold compared with the model group. R0011 directly improved gut microbial glucuronidase and downregulated the host intestinal drug transporter gene expression of multidrug resistance protein 2 (MRP2). More critically, R0011 restored the gut microbiota composition and regulated the metabolic function, significantly enhancing the microbial tryptophan metabolic pathway compared with the pathological state, which may indirectly promote the bioavailability of GL. Overall, these data may provide possible strategies by which to address the low bioavailability of traditional medicine through probiotic intervention.

A randomized controlled trial to evaluate the effect of influenza vaccination and probiotic supplementation on immune response and incidence of influenza-like illness in an elderly population in Indonesia.

AimTo investigate the effect of influenza vaccination with or without probiotic supplementation on the immune response and incidence of influenza-like illness (ILI) in the elderly.MethodsA randomized double-blind, placebo-controlled trial with a modified factorial design was conducted in 554 healthy elderly subjects aged 67 ± 5.6 (ranging from 60–90) years old in the Primary Health Care Center (Puskesmas area) of the Pulo Gadung District East Jakarta. Subjects received either a trivalent influenza vaccine or placebo at the start of the study, and a probiotic supplement (Lactobacillus helveticus R0052 and Lactobacillus rhamnosus R0011) or a placebo for 6 months. Subjects were randomly assigned into four intervention groups: influenza vaccine and probiotics (n = 141), influenza vaccine and placebo (n = 136), placebo and probiotics (n = 140), and both placebo (n = 137). The primary outcome was ILI incidence within 6 months. The secondary outcomes were seroprotection and seroconversion rates at 1, 4, and 6 months after administering the interventions.ResultsThis study showed that the trivalent influenza vaccine increased seroprotection (RR 3.6 [95%CI 2.92–4.47]; p<0.010) and seroconversion (RR 29.8 [95%CI 11.1–79.5]; p<0.010) rates 1 month after vaccination in elderly people while the probiotic supplement did not alter influenza antibody titers (p = 1.000 and p = 0.210). The relative ILI incidence risk was similar between vaccinated and non-vaccinated groups, as well as in the probiotic group compared to the non-probiotic group.ConclusionThe tested trivalent influenza vaccine significantly induced seroprotection and seroconversion in the vaccinated subjects, while probiotics administration did not influence these parameters. Vaccinated individuals displayed a similarly low ILI incidence as those in the Control Group. However, the observed trend towards a reduction of ILI incidence with probiotics supplementation warrants further assessments in a larger, at-risk population.Clinical trial registry numberNCT03695432.

Lactobacillus rhamnosus R0011 Treatment Enhanced Efficacy of Capecitabine against Colon Cancer in Male Balb/c Mice

The microbiome of the intestinal system is well-known as a modulatory factor. Having a balanced status of microbiota could help to prevent diseases, especially cancers related to the gastrointestinal system. We investigated the effects of Lactobacillus rhamnosus (Lr) and capecitabine on tumor size and physiologic features, such as bodyweight, liver enzymes, and blood profile, in a subcutaneously induced cancer model using CT-26 murine colon carcinoma cells. We divided 48 male Balb/c inbred mice into six groups. Lr had been orally pre-inoculated to the mice for 14 day consecutively. CT-26 cells were implanted subcutaneously into the mice’s flank. Following the injection of cancer cells, Lr was inoculated to the mice three times per week for four weeks. Capecitabine was inoculated in the third week after the induction of cancer. The tumor size was significantly decreased in treated groups in comparison to the cancer group (1174.5 ± 63.8, 1119.2 ± 86.3, and 985.6 ± 48 mm3 vs. 1674.2 ± 66 mm3, P < 0.0001). Data showed that Lr and capecitabine enhanced Bax/Bcl-2 ratio and caspase-3 level compared to cancer group (p < 0.0001). White blood cells (WBCs) were significantly decreased in the capecitabine group compared to probiotic group (P < 0.05). Measurement of bodyweight, liver enzymes, and interleukin-6 (IL-6) level showed that Lr, in addition to preventive and therapeutic effects, might have protective effects against chemotherapy side effects. Preventing WBCs’ reduction, protecting mice from losing weight, induction of apoptosis, and enhancing the serum level of IL-6 indicated that Lr might be associated with better management of colorectal cancer and chemotherapy side effects.

Effect of a Symbiotic Mixture on Fecal Microbiota in Pediatric Patients Suffering of Functional Abdominal Pain Disorders

(1) Background: Functional abdominal pain disorders (FAPDs) represent one of the main etiologies of chronic abdominal pain in the pediatric population. A wide spectrum of probiotic or prebiotic mixtures has been evaluated in trials regarding benefits in patients with FAPDs, mainly in the adult population. (2) Methods: This study was interested in evaluating the effect of oral supplementation with a symbiotic mixture on intestinal microbiota in children with functional dyspepsia (FD), irritable bowel syndrome with diarrhea (IBS-D), and irritable bowel syndrome with constipation (IBS-C). A combination of six bacterial strains (Lactobacillus rhamnosus R0011, Lactibacillus casei R0215, Bifidobacterium lactis BI-04, Lactobacillus acidophilus La-14, Bifidobacterium longum BB536, Lactobacillus plantarum R1012) and 210 mg of fructo-oligosaccharides-inulin were administered orally, daily, for 12 weeks and patients were scored for severity of symptoms and fecal microbiota before and after the treatment. (3) Results: The proportion of patients with adequate symptom relief was higher in the IBS-D than in the IBS-C group; however, the difference was not statistically significant (74.4% vs. 61.9%, p = 0.230). There was an increasing proportion of bacterial genera associated with health benefits, for both IBS-C and IBS-D (IBS-C: 31.1 ± 16.7% vs. 47.7 ± 13.5%, p = 0.01; IBS-D: 35.8 ± 16.2% vs. 44.1 ± 15.1%, p = 0.01). (4) Conclusions: Administration of a symbiotic preparation resulted in significant changes to the microbiota and gastrointestinal symptoms in patients with FAPDs.

Подход к профилактике и лечению неалкогольной жировой болезни печени у детей с ожирением с позиции современных международных рекомендаций

Актуальность. Статья посвящена проблеме профилактики развития и терапии неалкогольной жировой болезни печени (НАЖБП) у детей с ожирением по алиментарно-конституциональному типу. Цель: изучение эффективности и безопасности использования препаратов омега-3 полиненасыщенных жирных кислот (ПНЖК) с витамином Е и мультипробиотического комплекса, обогащенного витаминами, для профилактики развития и прогрессирования стеатогепатоза. Материалы и методы. Под наблюдением находились 55 детей с избыточной массой тела. Всем пациентам была назначена медикаментозная терапия с использованием препарата омега-3 ПНЖК Рейтоил и комплексного пробиотика Бревелак (Lactobacillus helveticus R0052, Lactococcus lactis ssp. Lactis R1058, Bifidobacterium longum R0175, Lactobacillus rhamnosus R0011, Bifidobacterium breve R0070, Streptococcus thermophilus R0083, Bifidobacterium bifidum R0071, Lactobacillus casei R0215, Lactobacillus plantarum R1012) на фоне модификации образа жизни. Результаты. Течение НАЖБП у детей в большинстве случаев не имеет характерной клинической симптоматики и выраженного синдрома цитолиза. Структурно-функциональные изменения в печени при НАЖБП могут вызывать метаболические нарушения с формированием выраженной атерогенной дислипидемии. Трехмесячный курс терапии НАЖБП препаратами омега-3 ПНЖК Рейтоил в сочетании с использованием комплексного пробиотического препарата Бревелак показал достоверное снижение активности печеночных цитолитических ферментов и улучшение показателя липидного спектра крови. Выводы. Назначение препаратов омега-3 ПНЖК в сочетании с комплексом штаммов пробиотических культур с доказанной клинической эффективностью при НАЖБП на фоне модификации образа жизни способствует эффективной профилактике и лечению данной патологии у детей.

Досвід використання пробіотичних штамів Lactobacillus acidophilus R0052 та Lactobacillus rhamnosus R0011 у дітей з опіковою травмою

На сьогодні опіки посідають друге місце серед усіх травм дітей. Основною причиною летальних наслідків у випадку виражених та глибоких опіків є інфекція, що виникає у 23–82 % усіх випадків опіків. Раціональна антибактеріальна терапія має велике значення в боротьбі з генералізованою інфекцією. Метою цього дослідження було ретельне вивчення епідеміології та характеристики антибіотик-асоційованої діареї (ААД) у хворих дітей з опіками, а також можливість запобігання цій діареї, що виникала внаслідок антибактеріальної терапії в пацієнтів Обласного опікового відділення м. Запоріжжя. Протягом 2012–2015 років ми спостерігали 438 дітей з опіковими травмами, які приймали антибіотики. У цих госпіталізованих дітей було знайдено специфічні для ААД Clostridium difficile токсини А + В у фекаліях. Це дозволило діагностувати Clostridium difficile-ентероколіт, пов’язаний з прийомом хворими антимікробних препаратів (A4.07, МКБ‑10). Розроблено профілактичні методи AAД, а саме призначення з перших годин перебування в стаціонарі per os пробіотичного засобу Lactobacillus acidophilus R0052 і Lactobacillus rhamnosus R0011 у складі препарату Lacidofil®. Використання пробіотичних штамів з антитоксичною активністю щодо Clostridium difficile скорочує в 3,4 раза число випадків антибіотик-асоційованої діареї в дітей з опіками та зменшує тяжкість її симптомів.

Technological Characterisation of Probiotic Lactic Acid Bacteria as Starter Cultures for Dry Fermented Sausages.

The objective of this study was to investigate probiotic microorganisms for use as starter cultures in dry fermented sausages production. A total of eight strains were studied evaluating technological and safety characteristics including the ability to grow, lactic acid production, gas formation, catalase activity, nitrate reductase activity, proteolytic activity, lipolytic activity, hydrogen peroxide production, salt tolerance, performance at low temperatures, decarboxylation of amino acids and antimicrobial activity against pathogens associated with the product. Lactobacillus rhamnosus R0011, L. rhamnosus Lr-32, Lactobacillus paracasei Lpc-37, Lactobacillus casei Shirota and Enterococcus faecium MXVK29 were good candidates for use as fermented sausages starters cultures because they showed the best technological and safety properties since they did not demonstrate amino acid decarboxylation but showed antimicrobial activity against Listeria monocytogenes, Escherichia coli, Salmonella Dublin and Staphylococcus aureus. L. rhamnosus Lr-32 was the strain best tolerating the levels of salt, nitrate and low pH during the simulated stages of fermentation and ripening of sausage. The strain was thus the most promising of the tested probiotics as sausage starter culture. The findings warrant studies in a meat matrix, such as that of raw-cured sausage, to evaluate the effects of L. rhamnosus Lr-32 under actual conditions.

Temporal transcriptional, metabolic, and functional re-programming of THP-1 macrophages by the Lactobacillus rhamnosus R0011 secretome

Abstract Macrophage responses to activation are fluid and dynamic and their ability to respond appropriately to subsequent challenge is integral to host defence. Recent evidence suggests that bacteria influence macrophage differentiation and subsequent polarization into pro-inflammatory (M1) and immunoregulatory (M2) phenotypes through direct interactions. However, many questions surround indirect communication mechanisms mediated through secretomes derived from gut bacteria, such as lactobacilli. We examined the effects of secretome-mediated conditioning on macrophage phenotype, focusing on the ability of the Lactobacillus rhamnosus R0011 secretome (LrS) to drive macrophage polarization and to prime responses to subsequent challenge with lipopolysaccharide (LPS). Transcriptional profiling revealed increased M2-associated gene transcription (IL10R, CD36, CD163, TLR1, and TLR8) in response to LrS conditioning. Cytokine and chemokine profiling confirmed these results, indicating increased M2-associated chemokine and cytokine production (IL-10, CCL1, 17, 20, CXCL1 and 2). Metabolite utilization assays indicated diminished reliance on glycolysis for energy generation, coupled with increased phagocytic capacity, characteristics of functional M2 activity. LPS challenge of LrS-conditioned THP-1s revealed heightened responsiveness, indicative of innate immune priming. Overall, the LrS conditions THP-1s into M2 macrophages and primes responses to subsequent LPS challenge. Secretome conditioning of macrophages to respond robustly to inflammatory challenge within an M2 phenotype is an uncharacterized and potentially important route through which lactobacilli can train innate immunity.

Secretome-Mediated Interactions with Intestinal Epithelial Cells: A Role for Secretome Components from Lactobacillus rhamnosus R0011 in the Attenuation of Salmonella enterica Serovar Typhimurium Secretome and TNF-α-Induced Proinflammatory Responses.

Recent evidence suggests that lactic acid bacteria communicate with host cells via secretome components to influence immune responses but less is known about gut-pathogen secretomes, impact of lactic acid bacteria secretomes on host-pathogen interactions, and the mechanisms underlying these interactions. Genome-wide microarrays and cytokine profiling were used to interrogate the impact of the Lactobacillus rhamnosus R0011 secretome (LrS) on TNF-α and Salmonella enterica subsp. enterica serovar Typhimurium secretome (STS)-induced outcomes in human intestinal epithelial cells. The LrS attenuated both TNF-α- and STS-induced gene expression involved in NF-κB and MAPK activation, as well as expression of genes involved in other immune-related signaling pathways. Specifically, the LrS induced the expression of dual specificity phosphatase 1 (DUSP1), activating transcription factor 3 (ATF3), and tribbles pseudokinase 3 (TRIB3), negative regulators of innate immune signaling, in HT-29 intestinal epithelial cells challenged with TNF-α or STS. TNF-α- and STS-induced acetylation of H3 and H4 histones was attenuated by the LrS, as was the production of TNF-α- and STS-induced proinflammatory cytokines and chemokines. Interestingly, the LrS induced production of macrophage migration inhibitory factor (MIF), a cytokine involved in host-microbe interactions at the gut interface. We propose that the LrS attenuates proinflammatory mediator expression through increased transcription of negative regulators of innate immune activity and changes in global H3 and H4 histone acetylation. To our knowledge, these findings provide novel insights into the complex multifaceted mechanisms of action behind secretome-mediated interdomain communication at the gut-mucosal interface.

Safety and Effect of a Low- and High-Dose Multi-Strain Probiotic Supplement on Microbiota in a General Adult Population: A Randomized, Double-Blind, Placebo-Controlled Study

Few studies have focused on dose-response analyses of multi-strain probiotics in the general adult population. This study aimed at comparing how a low- and high-dose of a multi-strain probiotic supplement (containing Lactobacillus helveticus R0052, Lactobacillus rhamnosus R0011, Lactobacillus casei R0215, Pediococcus acidilactici R1001, Bifidobacterium breve R0070, Bifidobacterium longum ssp. longum BB536, Lactobacillus plantarum R1012, Lactococcus lactis ssp. lactis R1058) affected microbiota composition, transit persistence and safety in adults. After a 7-d baseline, participants were randomized to receive capsules containing 5 or 25 billion CFU, or placebo daily for 28 days, followed by a 7-d washout. Digestive health and general wellness were assessed. Fecal microbiota composition was analyzed using 16S rRNA gene amplicon sequencing and strain persistence, by qPCR. Participants’ gastrointestinal and general wellbeing were unaffected. No adverse events were associated with either dose. Supplemented strains contributed to the Lactobacillus and Bifidobacterium genera detected in stool, with 0.40 ± 0.11% and 0.51 ± 0.26%, respectively, in the high-dose group. Strain-specific qPCR assays revealed variable levels of post-intervention persistence between strains. Sequencing and composition analyses using the 16S V4 region revealed a decrease in Holdemania and increase in Bacteroidales. The formulation was well tolerated in this sample of the general adult population, even at the higher dose. The strains appear to have influenced microbiota composition minimally, as expected in the absence of dysbiosis, and consistently with the dose administered. Overall, the results provide a rationale to study the effects this formulation on microbiota composition in individuals exhibiting dysbiosis associated with metabolic disorders or obesity.

Milk fermented with Lactobacillus rhamnosus R0011 induces a regulatory cytokine profile in LPS-challenged U937 and THP-1 macrophages

Fermented dairy products have become attractive functional foods for the delivery of probiotics and their biologically active metabolites. The aim of this study was to examine the immunomodulatory activity of milk fermented with the probiotic lactic acid bacterium Lactobacillus rhamnosus R0011 (LrF) on macrophages challenged with lipopolysaccharide (LPS), a potent pro-inflammatory stimulus. To this end, human THP-1 or U937 monocytes were differentiated into resting macrophages then stimulated with LPS and co-incubated with the LrF or with milk controls. Levels of pro-inflammatory and immunoregulatory cytokines were determined by enzyme-linked immunosorbent assays. Culturing of LPS-stimulated U937 macrophages with either the whole or filtered LrF resulted in an increase in Interleukin (IL)-1Ra production relative to the negative control. THP-1 macrophages cultured with the LrF demonstrated an increase in LPS-induced IL-10 and IL-1β production, while production of LPS-induced IL-6, sCD54, IL-8, IL-1β, TNF-α, IL-12p70 and Transforming Growth Factor-β (TGF-β) was unaffected. Further, the LrF induced the expression of DC-SIGN and CD206, markers of immunoregulatory M2 macrophage polarization, in LPS-challenged THP-1 macrophages. Taken together, milk fermented with L. rhamnosus R0011 increased regulatory cytokine production from LPS-challenged U937 and THP-1 macrophages, while simultaneously up-regulating the production of IL-1β and expression of DC-SIGN and CD206, a profile characteristic of polarization into the immunoregulatory M2 macrophage phenotype.

Probiotic Supplementation is Associated with Increased Antioxidant Capacity and Copper Chelation in C. difficile-Infected Fecal Water.

Probiotic supplementation plays a key role in maintaining intestinal homeostasis due to its ability to modulate gut microbiota. Although their potential as potent antioxidants have previously been explored, their ability to affect the redox status in the gut lumen of healthy subjects or those with gastrointestinal (GI) disorders remains unclear. In our study, we assessed the ability of single strain and multispecies probiotic supplementation to cause a change in the redox status of normal fecal water and in Clostridium (C.) difficile-infected fecal water using a simulated gastrointestinal model. Changes in redox status were assessed by ferric-reducing antioxidant power (FRAP), 2’,2’-diphenyl-1-picrylhydrazyl (DPPH), and iron and copper chelation assays. The findings from our study showed that in normal fecal water, probiotic supplements, apart from Lactobacillus (L.) rhamnosus R0011, showed a significant increase in iron chelation (p < 0.05), which was associated with lower FRAP and copper chelation. In C. difficile-infected fecal water, all probiotic supplements showed a significant increase in FRAP (p < 0.05) and were associated with increased copper chelation. The DPPH assay showed no treatment effect in either fecal water. These findings suggest that C. difficile mediates dysregulation of redox status, which is counteracted by probiotics through ferric-reducing ability and copper chelation.

Effect of Coordinated Probiotic/Prebiotic/Phytobiotic Supplementation on Microbiome Balance and Psychological Mood State in Healthy Stressed Adults

Background: Interest in and knowledge of the gut microbiome has increased exponentially in the past decade. This once overlooked component of the gastrointestinal tract is now implicated in multiple aspects of human health, including mental (e.g. depression, anxiety, stress), metabolic (e.g. diabetes/obesity), neurological (e.g. Alzheimer’s, Parkinson’s, Autism Spectrum Disorder), gastrointestinal (e.g. irritable bowel syndrome, Crohn’s), and immunological (e.g. inflammation, cancer) wellness, among others. Previous research has demonstrated the “strain specificity” of probiotic therapy (e.g. Lactobacillus helveticus R0052 for serotonin/depression; Bifidobacterium longum R0175 for GABA/anxiety; Lactobacillus rhamnosus R0011 for cortisol/stress). Similarly, probiotic bacteria demonstrate different growth trajectories based on availability of preferred fiber substrates (e.g. prebiotics) and phytonutrients such as flavonoids/polyphenols (e.g. phytobiotics). Thus, our objective was two-fold: to determine the change in microbiome ecology/balance and to evaluate the psychological mood state following a coordinated pro-/pre-/phyto-biotic supplementation regimen.Methods: Thirty-two healthy subjects screened for “moderate” levels of psychological stress were randomly assigned to 1-month of Supplement (Amare Fundamentals, N=21) or matching Placebo (N=11). Microbiome balance was assessed in fecal samples using a PCR-based analysis (BiomeTracker) that has previously compared favorably to 16S sequencing for abundance quantification for major phyla/families of bacteria. Psychological mood state parameters were assessed using the validated Profile of Mood States survey (POMS) to generate scores for Global Mood State, and six sub-scales (Depression, Tension, Fatigue, Anger, Confusion, and Vigor).Results: Following supplementation, there was a significant increase in populations of “good” bacteria in the Supplement group (+28% Lactobacillus; +30% Bifidobacterium) and overall composite score (+17%) versus Placebo (p&lt;0.05). Psychological indices were significantly improved in the Supplement group for both positive (+25% Global Mood; +44% Vigor) and negative (-64% Fatigue; -55% Depression; -54% Anger; -45% Tension; -43% Confusion) mood state parameters versus Placebo (p&lt;0.05).Conclusion: The World Health Organization has identified mental wellness issues as the leading contributor to global health burden – highlighting the urgency to develop lifestyle interventions to effectively manage depression, anxiety, and stress. These results demonstrate the close relationship between microbiome balance and psychological parameters – and the utility of targeted supplementation to positively influence the gut-brain-axis for improved mental wellness.Keywords: Lactobacillus helveticus, Bifidobacterium longum, Lactobacillus rhamnosus, depression, anxiety, stress, vigor, mood state, mental wellness

Probiotic treatment restores normal developmental trajectories of fear memory retention in maternally separated infant rats

Early life stress (ELS) can affect brain development and increase lifetime prevalence of psychiatric illnesses. However, the effective therapeutic interventions to ameliorate the deleterious effects of ELS have not yet been well established. Here, we confirmed that maternal separation (MS) for 3 h daily between postnatal days 2–14, a frequently used experimental model of ELS, resulted in early expression of adult-like fear memory retention in male infant rats. Administration of a probiotic formulation, Lacidofil® (95% Lactobacillus rhamnosus R0011 and 5% Lactobacillus helveticus R0052), during the separation period, prevented the precocious transition to adult-like fear memory retention in MS infant rats. Consonant with this effect, probiotic treatment also ameliorated the MS-induced increases in anxiety-like behavior as measured by the elevated plus maze and the light-dark box tests. In addition, probiotic treatment reduced MS-induced increases in neuronal activation and brain-derived neurotrophic factor protein levels in the basolateral nucleus of amygdala (BLA) after auditory fear conditioning. Furthermore, we found that probiotic treatment significantly rescued the heightened hypothalamic-pituitary-adrenal (HPA) axis response to restraint stress in MS infant rats. Taken together, these findings suggest that probiotics can restore normal developmental trajectories of fear memory retention in MS infant rats, at least in part by normalizing HPA axis abnormalities, and that the BLA serves as a critical node to mediate these interventions. Thus, we offer a potential therapeutic intervention to protect children against the harmful effects of ELS.

Multicenter Trial of a Combination Probiotic for Children with Gastroenteritis

BackgroundGastroenteritis accounts for approximately 1.7 million visits to the emergency department (ED) by children in the United States every year. Data to determine whether the use of probiotics improves outcomes in these children are lacking.MethodsWe conducted a randomized, double-blind trial involving 886 children 3 to 48 months of age with gastroenteritis who presented to six pediatric EDs in Canada. Participants received a 5-day course of a combination probiotic product containing Lactobacillus rhamnosus R0011 and L. helveticus R0052, at a dose of 4.0×109 colony-forming units twice daily or placebo. The primary outcome was moderate-to-severe gastroenteritis, which was defined according to a post-enrollment modified Vesikari scale symptom score of 9 or higher (scores range from 0 to 20, with higher scores indicating more severe disease). Secondary outcomes included the duration of diarrhea and vomiting, the percentage of children who had unscheduled physician visits, and the presence or absence of adverse events.ResultsModerate-to-severe gastroenteritis within 14 days after enrollment occurred in 108 of 414 participants (26.1%) who were assigned to probiotics and 102 of 413 participants (24.7%) who were assigned to placebo (odds ratio, 1.06; 95% confidence interval [CI], 0.77 to 1.46; P=0.72). After adjustment for trial site, age, detection of rotavirus in stool, and frequency of diarrhea and vomiting before enrollment, trial-group assignment did not predict moderate-to-severe gastroenteritis (odds ratio, 1.06; 95% CI, 0.76 to 1.49; P=0.74). There were no significant differences between the probiotic group and the placebo group in the median duration of diarrhea (52.5 hours [interquartile range, 18.3 to 95.8] and 55.5 hours [interquartile range, 20.2 to 102.3], respectively; P=0.31) or vomiting (17.7 hours [interquartile range, 0 to 58.6] and 18.7 hours [interquartile range, 0 to 51.6], P=0.18), the percentages of participants with unscheduled visits to a health care provider (30.2% and 26.6%; odds ratio, 1.19; 95% CI, 0.87 to 1.62; P=0.27), and the percentage of participants who reported an adverse event (34.8% and 38.7%; odds ratio, 0.83; 95% CI, 0.62 to 1.11; P=0.21).ConclusionsIn children who presented to the emergency department with gastroenteritis, twice-daily administration of a combined L. rhamnosus–L. helveticus probiotic did not prevent the development of moderate-to-severe gastroenteritis within 14 days after enrollment. (Funded by the Canadian Institutes of Health Research and others; PROGUT ClinicalTrials.gov number, NCT01853124.)

Suppression of Intestinal Epithelial Cell Chemokine Production by Lactobacillus rhamnosus R0011 and Lactobacillus helveticus R0389 Is Mediated by Secreted Bioactive Molecules.

Host intestinal epithelial cells (IEC) present at the gastrointestinal interface are exposed to pathogenic and non-pathogenic bacteria and their products. Certain probiotic lactic acid bacteria (LAB) have been associated with a range of host-immune modulatory activities including down-regulation of pro-inflammatory gene expression and cytokine production by IEC, with growing evidence suggesting that these bacteria secrete bioactive molecules with immunomodulatory activity. The aim of this study was to determine whether two lactobacilli with immunomodulatory activity [Lactobacillus rhamnosus R0011 (Lr) and Lactobacillus helveticus R0389 (Lh)], produce soluble mediators able to influence IEC responses to Pattern Recognition Receptor (PRR) ligands and pro-inflammatory cytokines [Tumor Necrosis Factor α (TNFα), Interleukin-1β (IL-1β)], signals inducing IEC chemokine production during infection. To this end, the effects of cell-free supernatants (CFS) from Lr and Lh on IEC production of the pro-inflammatory chemokines interleukin (IL)-8 and cytokine-induced neutrophil chemoattractant 1 (CINC-1) induced by a range of host- or pathogen-derived pro-inflammatory stimuli were determined, and the impact on human HT-29 IEC and a primary IEC line (rat IEC-6) was compared. The Lr-CFS and Lh-CFS did not significantly modulate basal IL-8 production from HT-29 IECs or CINC-1 production from IEC-6 cells. However, both Lr-CFS and Lh-CFS significantly down-regulated IL-8 production from HT-29 IECs challenged with varied PRR ligands. Lr-CFS and Lh-CFS had differential effects on PRR-induced CINC-1 production by rat IEC-6 IECs, with no significant down-regulation of CINC-1 observed from IEC-6 IECs cultured with Lh-CFS. Further analysis of the Lr-CFS revealed down-regulation of IL-8 production induced by the pro-inflammatory cytokines IL-1β and TNFα Preliminary characterization of the bioactive constituent(s) of the Lr-CFS indicates that it is resistant to treatment with DNase, RNase, and an acidic protease, but is sensitive to alterations in pH. Taken together, these results indicate that these lactobacilli secrete bioactive molecules of low molecular weight that may modulate host innate immune activity through interactions with IEC.

Gut Bacterial Microbiota and its Resistome Rapidly Recover to Basal State Levels after Short-term Amoxicillin-Clavulanic Acid Treatment in Healthy Adults

Clinical effects of antimicrobials and probiotics in combination have been reported, however, little is known about their impact on gut microbiota and its resistome. In this study 16S rRNA gene amplicon, shotgun metagenomics sequencing and antibiotic resistance (ABR) microarray were used on fecal samples of 70 healthy participants, taken at four time points in probiotic (Lactobacillus rhamnosus R0011 and Lactobacillus helveticus R0052) and placebo groups to profile the gut bacterial microbiota and its resistome following administration of amoxicillin-clavulanic acid for one week. Significant shifts in microbiota family composition caused by the antimicrobial in both groups that included decreases in the proportion of Lachnospiraceae, Coriobacteriaceae and unidentified Clostridiales; and notable increases for the proportion of Enterobacteriaceae, Bacteroidaceae and Porphyromonadaceae compared to baseline levels. Resistome showed a corresponding enrichment of ABR genes compared to baseline from such classes as aminoglycosides and beta-lactams that were linked, by in silico inference, to the enrichment of the family Enterobacteriaceae. Despite perturbations caused by short-term antibiotic treatment, both gut microbiota and resistome showed prompt recovery to baseline levels one week after cessation of the antimicrobial. This rapid recovery may be explained by the hypothesis of community resilience.

Pea-protein alginate encapsulation adversely affects development of clinical signs of Citrobacter rodentium-induced colitis in mice treated with probiotics.

The efficacy of two strains of Lactobacillus probiotics (Lactobacillus rhamnosus R0011 and Lactobacillus helveticus R0052) immobilized in microcapsules composed of pea protein isolate (PPI) and alginate microcapsules was assessed using a mouse model of Citrobacter rodentium-induced colitis. Accordingly, 4-week-old mice were fed diets supplemented with freeze-dried probiotics (group P), probiotic-containing microcapsules (group PE) (lyophilized PPI-alginate microcapsules containing probiotics), or PPI-alginate microcapsules containing no probiotics (group E). Half of the mice (controls, groups P, PE, and E) received C. rodentium by gavage 2 weeks after initiation of feeding. Daily monitoring of disease symptoms (abnormal behavior, diarrhea, etc.) and body weights was undertaken. Histopathological changes in colonic and cecal tissues, cytokine expression levels, and pathogen and probiotic densities in feces were examined, and the microbial communities of the distal colon mucosa were characterized by 16S rRNA sequencing. Infection with C. rodentium led to marked progression of infectious colitis, as revealed by symptomatic and histopathological data, changes in cytokine expression, and alteration of composition of mucosal communities. Probiotics led to changes in most of the disease markers but did not have a significant impact on cytokine profiles in infected animals. On the basis of cytokine expression analyses and histopathological data, it was evident that encapsulation materials (pea protein and calcium alginate) contributed to inflammation and worsened a set of symptoms in the cecum. These results suggest that even though food ingredients may be generally recognized as safe, they may in fact contribute to the development of an inflammatory response in certain animal disease models.

PL02: Probiotic regimen for outpatient gastroenteritis utility of treatment (PROGUT) study: a multicenter randomized controlled trial

Introduction: Gastroenteritis accounts for 1.7 million emergency department visits by children annually in the United States. We conducted a double-blind trial to determine whether twice daily probiotic administration for 5 days, improves outcomes. Methods: 886 children aged 348 months with gastroenteritis were enrolled in six Canadian pediatric emergency departments. Participants were randomly assigned to twice daily Lactobacillus rhamnosus R0011 and Lactobacillus helveticus R0052, 4.0 x 109 CFU, in a 95:5 ratio or placebo. Primary outcome was development of moderate-severe disease within 14 days of randomization defined by a Modified Vesikari Scale score 9. Secondary outcomes included duration of diarrhea and vomiting, subsequent physician visits and adverse events. Results: Moderate-severe disease occurred in 108 (26.1%) participants administered probiotics and 102 (24.7%) participants allocated to placebo (OR 1.06; 95%CI: 0.77, 1.46; P=0.72). After adjustment for site, age, and frequency of vomiting and diarrhea, treatment assignment did not predict moderate-severe disease (OR, 1.11, 95%CI, 0.80 to 1.56; P=0.53). In the probiotic versus placebo groups, there were no differences in the median duration of diarrhea [52.5 (18.3, 95.8) vs. 55.5 (20.2, 102.3) hours; P=0.31], vomiting [17.7 (0, 58.6) vs. 18.7 (0, 51.6) hours; P=0.18], physician visits (30.2% vs. 26.6%; OR 1.19; 95% CI0.87. 1.62; P=0.27), or adverse events (32.9% vs. 36.8%; OR 0.83; 95%CI 0.62. 1.11; P=0.21). Conclusion: In children presenting to an emergency department with gastroenteritis, twice daily administration of 4.0 x 109 CFU of a Lactobacillus rhamnosus/helveticus probiotic does not prevent development of moderate-severe disease or improvements in other outcomes measured.

Effect of Coordinated Probiotic/Prebiotic/Phytobiotic Supplementation on Microbiome Balance and Psychological Mood State in Healthy Stressed Adults

BackgroundInterest in and knowledge of the gut microbiome has increased exponentially in the past decade. This once overlooked component of the gastrointestinal tract is now implicated in multiple aspects of human health, including mental wellness (e.g. depression, anxiety, stress), metabolic (e.g. diabetes, obesity), neurologic (e.g. Alzheimer's, Parkinson's, Autism Spectrum Disorder), gastrointestinal (e.g. irritable bowel syndrome, Crohn's), and immunologic (e.g. inflammation, cancer), among others.Purpose/ObjectivesPrevious research has demonstrated the “strain specificity” of probiotic therapy – elucidating that targeted health benefits are related to the specific strain of bacteria (e.g. Lactobacillus helveticus R0052 for serotonin/depression; Bifidobacterium longum R0175 for GABA/anxiety; Lactobacillus rhamnosus R0011 for cortisol/stress). Similarly, probiotic bacteria demonstrate different growth trajectories based on availability of preferred fiber substrates (e.g. prebiotics) and phytonutrients such as flavonoids/polyphenols (e.g. phytobiotics). Thus, our objective was two‐fold: to determine the change in microbiome ecology/balance; and to assess the change in psychological mood state (e.g. depression/anxiety/vigor) following a coordinated pro‐/pre‐/phyto‐biotic supplementation regimen.MethodsThirty‐two healthy subjects screened for “moderate” levels of psychological stress were randomly assigned to 30‐days of Supplement (Amare Fundamentals containing specific strains of probiotic bacteria, prebiotic fibers, and phytobiotic plant extracts, N=21) or look‐alike Placebo (N=11). Microbiome balance was assessed in fecal samples using a PCR‐based analysis (BiomeTracker) that has previously compared favorably to 16S sequencing (ACN 2017) for abundance quantification for major phyla/families of bacteria. Psychological mood state parameters were assessed using the validated Profile of Mood States survey (POMS) to generate scores for Global Mood State, and six sub‐scales (Depression, Tension, Fatigue, Anger, Confusion, and Vigor).ResultsFollowing 30‐days of supplementation, there was a significant increase in populations of “good” bacteria in the Supplement group (+28% Lactobacillus; +30% Bifidobacterium) and overall composite score (+17%) versus Placebo (p&lt;0.05). Psychological indices were significantly improved in the Supplement group for both positive (+25% Global Mood; +25% Vigor) and negative (−60% Depression; −54% Tension; −52% Fatigue; −42% Confusion; −39% Anger) parameters versus Placebo (p&lt;0.05).ConclusionsThe World Health Organization has identified mental wellness issues as the leading contributor to global health burden – highlighting the urgency to develop lifestyle interventions to effectively manage depression, anxiety, and stress. These results demonstrate the close relationship between microbiome balance and psychological parameters – and the utility of targeted supplementation to positively influence the gut‐brain‐axis for improved mental wellness.Support or Funding InformationThis trial was funded by Amare Global and conducted jointly by EQQIL and Wasatch ScientificThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.