Simulator Of The Human Intestinal Microbial Ecosystem Research Articles

Effects of tris (2-chloroethyl) phosphate exposure on gut microbiome using the simulator of the human intestinal microbial ecosystem (SHIME)

Tris (2-chloroethyl) phosphate (TCEP) has been widely used, and its health risk has received increasing attention. However, the rare research has been conducted on the effects of TCEP exposure on changes in the structure of the human gut microbiome and metabolic functions. In this experiment, Simulator of the human intestinal microbial ecosystem (SHIME) was applied to explore the influences of TCEP on the human gut bacteria community and structure. The results obtained from high-throughput sequencing of 16S rRNA gene have clearly revealed differences among control and exposure groups. High-dose TCEP exposure increased the Shannon and Simpson indexes in the results of α-diversity of the gut microbiome. At phylum level, Firmicutes occupied a higher proportion of gut microbiota, while the proportion of Bacteroidetes decreased. In the genus-level analysis, the relative abundance of Bacteroides descended with the TCEP exposure dose increased in the ascending colon, while the abundances of Roseburia, Lachnospira, Coprococcus and Lachnoclostridium were obviously correlated with exposure dose in each colon. The results of short chain fatty acids (SCFAs) showed a remarkable effect on the distribution after TCEP exposure. In the ascending colon, the control group had the highest acetate concentration (1.666 ± 0.085 mg⋅mL-1), while acetate concentrations in lose-dose medium-dose and high-doseTCEP exposure groups were 1.119 ± 0.084 mg⋅mL-1, 0.437 ± 0.053 mg⋅mL-1 and 0.548 ± 0.106 mg⋅mL-1, respectively. TCEP exposure resulted in a decrease in acetate and propionate concentrations, while increasing butyrate concentrations in each colon. Dorea, Fusicatenibacter, Kineothrix, Lachnospira, and Roseburia showed an increasing tendency in abundance under TCEP exposure, while they had a negatively correlation with acetate and propionate concentrations and positively related with butyrate concentrations. Overall, this study confirms that TCEP exposure alters both the composition and metabolic function of intestinal microbial communities, to arouse public concern about its negative health effects.

Novel In Vitro Intestinal Microbiome Model to Study Lipidomic and Metabolomic Adaptations Due to Exposure to Glyphosate, Perfluorooctanoic Acid, and Docusate Sodium.

Cell and animal models have been used to provide insights with regard to physiological changes in intestinal flora due to exposure to drugs and environmental contaminants. Here, a novel in vitro model known as simulator of the human intestinal microbial ecosystem (SHIME) was used to assess the effects of three chemicals of emerging concern, namely glyphosate, perfluorooctanoic acid (PFOA), and docusate sodium (dioctyl sulfosuccinate, DOSS), on the lipidomic and metabolomic profiles of the gut microenvironment in both the proximal and distal colonic compartments. Nontargeted analyses by ultra-high performance liquid chromatography-tandem mass spectrometry and gas chromatography-electron ionization-mass spectrometry revealed minor differences in the lipidomic and metabolomic signatures of the proximal and distal colon following treatment with either glyphosate or PFOA at acceptable human daily intake levels or average daily exposures. However, global dysregulation of lipids and metabolites was observed due to DOSS treatment at conventional prescription doses when indicated as a stool softener. Our findings suggest that the current guidelines for glyphosate and PFOA exposure may be adequate at the level of the lower gut microbiome in healthy adults, but the probable yet uncharacterized off-target effects, safety, and efficacy of long-term DOSS treatment warrants further investigation. Indeed, we highlight the SHIME system as a novel in vitro approach which can be used as a screening tool to assess the impact of drugs and/or chemicals on the gut microbiome, while implementing state-of-the-art and data-driven mass spectrometric workflows to identify toxic lipidomic and metabolomic signatures.

#3454 IMPACT OF SYMBIOTICS ON UREMIC TOXINS PRODUCTION, RENAL FUNCTION AND PEW SYNDROME IN SHIME AND CKD MICE MODELS

Abstract Background and Aims Alterations in gut microbiota in CKD patients have been linked to CKD progression, cachexia, and mortality. Gut dysbiosis is associated by modification of gut-microbiota-derived metabolites, including a decrease of short-chain fatty acids (SCFAs), an increase in gut-derived uremic toxins. Therefore, modulation of gut microbiota seems to be an attractive therapeutic approach. In silico studies led to the selection of bacterial species able to increase SCFAs production, improve gut function without having the enzymatic machinery for producing uremic toxins. The propose of this study is to indentify the impact of two different symbiotics containing the selected bacteria: 1) on both uremic toxins and SCFAs production using a Simulator of the Human Intestinal Microbial Ecosystem (SHIME) colonized with fresh feces from CKD or healthy controls (HC) and subsequently 2) on renal function, metabolic parameters, intestinal microbiota and gut-microbiota-derived metabolites in a mouse model of CKD (5/6 nephrectomy). Method 1) The SHIME® system manufactured by ProDigest (Ghent, Belgium) consists of systems comprising five double-wall biovessels simulating the stomach (ST), small intestine (SI), ascending colon (AC), transverse colon (TC), and descending colon (DC). The supernatant of the homogenized fecal slurry was inoculated into the AC (25 mL), TC (40 mL), and DC (30 mL), respectively. After fecal inoculation, SHIME system were fed in ST twice day with two different symbiotics (P1 or P2) for 10 days. After 7 days, we applied a diet amino acid (AA) challenge to all the groups. SCFAs and uremic toxins were quantified every day. 2) CKD mice received either P1 or P2 admix for 6 weeks. We measured food intake, body composition and renal function including renal fibrosis quantification. Plasma metabolomic and feces metagenomic analyses were performed. Results In vitro, the concentration of precursor of uremic toxins (indols and p-cresol) tended to be higher in DC colonized by feces from CKD patients and AA challenge increased uremic toxins production more significantly in CKD compared to HC. CKD microbiome showed a lower butyrate concentration at baseline and higher propionate and acetate production. Both P1 and P2 significantly limited uremic toxins production during AA challenge and increase SCFA production. In vivo, both symbiotics increased body weight (+14%) and food intake (+31%) compared to CKD mice on chow diet. Treatment did not impact lipid or glucose metabolism but fat accretion in epididymal was restored with P1. Both diets, but particularly P1, significantly reduced plasmatic urea, proteinuria, and kidney fibrosis. Symbiotics improved intestinal barrier with a greater expression of Occludin in the ileum. Metagenomic analyses of the gut microbiota indicated that the alfa-diversity was not different across groups while the beta-diversity was similar to the sham group if CKD mice were treated with symbiotics. Both symbiotics reduced significantly plasma levels of deleterious uremic toxins such as p-cresyl sulfate and indoxyl sulfate and symbiotics reduce fecal module of trypotophan degradation and other modules related to AA metabolism. Both diets increased fecal bacteria modules related to SCFAs production. Conclusion We have shown both in vitro and in vivo that a symbiotic association can reduce uremic toxins production and increase SCFA production by inducing a significant change in gut microbiota composition and function. These metabolic changes are associated with improved appetite, decreased uremic cachexia, and preserved renal function. This data needs be confirmed in a human clinical trial.

The regulatory effects of Lonicera japonica flos on fecal microbiota from humans with type 2 diabetes in a SHIME model

IntroductionLonicera japonica flos, a well-known herbal medicine in Asian countries, has been widely used for the treatment of type 2 diabetes (T2D). The gut microbiota and relevant metabolites have been shown to be involved in the development of T2D. This study was designed to investigate the effects of Lonicera japonica flos on fecal microbiota from humans with T2D using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME) system. MethodsAn aqueous extract of Lonicera japonica flos (LJE) was prepared. Colorimetry and high-performance liquid chromatography (HPLC) were used to analyze the chemical composition of LJE. Three colonic compartments of SHIME were inoculated with feces from T2D donors, and LJE was added to SHIME to simulate daily oral administration for 7 days. The microbial community and the concentrations of short-chain fatty acids (SCFAs) of colonic samples were analyzed by 16 S rRNA sequencing technology and gas chromatography, respectively. In addition, polyphenols in LJE before and after fermentation by fecal microbiota were measured using HPLC. ResultsLJE is rich in polyphenols and polysaccharides and alters the microbial community, as indicated by the decreased relative abundance of the detrimental genera Shigella and Lactobacillus and the increased abundance of SCFA-producing bacteria, including the genera Prevotella and Bacteroides. LJE significantly promoted SCFA production, including acetic, propionic, and butyric acids, in each colonic compartment. Furthermore, chlorogenic acid in LJE was biotransformed by fecal microbiota. ConclusionsLJE exerted beneficial regulation on the T2D fecal microbial community. The regulatory effects of gut microbiota may partly contribute to the therapeutic action of Lonicera japonica flos on T2D.

The Effect of Hydrolyzed and Fermented Arabinoxylan-Oligo Saccharides (AXOS) Intake on the Middle-Term Gut Microbiome Modulation and Its Metabolic Answer.

Although fermentation and hydrolyzation are well-known processes to improve the bioavailability of nutrients and enable the fortification with dietary fibers, the effect of such pre-treatments on the prebiotic features of arabinoxylan-oligosaccharides (AXOS) had not been explored. The middle-term in vitro simulation through the Simulator of the Human Intestinal Microbial Ecosystem (SHIME) demonstrated that the feeding with different formulations (namely oat bran, rye bran and wheat bran) containing hydrolyzed AXOS fermented by lactic acid bacteria significantly increased the synthesis of short-chain fatty acids (SCFA) by colon microbiota, with hydrolyzed and fermented rye bran displaying the highest effect. After two weeks from the interruption of intake, SCFA concentrations significantly decreased but remained still significantly higher compared to the original condition. The microbiome was also affected, with a significant abundance increase in Lactobacillaceae taxon after feeding with all fermented and hydrolyzed formulates. Hydrolyzed and fermented rye bran showed the highest changes. The fungal community, even if it had a lower variety compared to bacteria, was also modulated after feeding with AXOS formulations, with an increase in Candida relative abundance and a decrease in Issatchenkia. On the contrary, the intake of non-hydrolyzed and non-fermented wheat bran did not produce relevant changes of relative abundances. After two weeks from intake interruption (wash out period) such changes were mitigated, and the gut microbiome modulated again to a final structure that was more like the original condition. This finding suggests that hydrolyzed AXOS fermented by lactic acid bacteria could have a more powerful prebiotic effect compared to non-hydrolyzed and non-fermented wheat bran, shaping the colon microbiome and its metabolic answer. However, the intake should be continuous to assure persistent effects. Opening a window into the ecological evolutions and plausible underlying mechanisms, the findings reinforce the perspective to explore more in depth the use of hydrolyzed and fermented AXOS as additional ingredient for bread fortification.

Role, relevance, and possibilities of in vitro fermentation models in human dietary, and gut-microbial studies.

Dietary studies play a crucial role in determining the health-benefiting effects of most food substances, including prebiotics, probiotics, functional foods, and bioactive compounds. Such studies involve gastrointestinal digestion and colonic fermentation of dietary substances. In colonic fermentation, any digested food is further metabolized in the gut by the residing colonic microbiota, causing a shift in the gut microenvironment and production of various metabolites, such as short-chain fatty acids. These diet-induced shifts in the microbial community and metabolite production, which can be assessed through in vitro fermentation models using a donor's fecal microbiota, are well known to impact the health of the host. Although in vivo or animal experiments are the gold standard in dietary studies, recent advancements using different in vitro systems, like artificial colon (ARCOL), mini bioreactor array (MBRA), TNO in vitro model of the colon (TIM), Simulator of the Human Intestinal Microbial Ecosystem (SHIME), M-SHIME, Copenhagen MiniGut, and Dynamic Gastrointestinal Simulator,make it easy to study the dietary impact in terms of the gut microbiota and metabolites. Such a continuous in vitro system can have multiple compartments corresponding to different parts of the colon, that is, proximal, transverse, and distal colon, making the findings physiologically more significant. Furthermore, postfermentation samples can be analyzed using metagenomic, metabolomic, quantitative-polymerase chain reaction, and flow-cytometry approaches. Moreover, studies have shown that in vitro results are in accordance with the in vivo findings, supporting their relevance in dietary studies and giving confidence that shifts in metabolites are only due to microbes. This review meticulously describes the recent advancements in various fermentation models and their relevance in dietary studies.

The disruption on gut microbiome of Decabromodiphenyl ethane exposure in the simulator of the human intestinal microbial ecosystem (SHIME)

The health risks of Decabromodiphenyl ethane (DBDPE) with its cardiovascular toxicity, liver toxicity and cytotoxicity had been generally acknowledged. However, the influence on gut microbiome and short-chain fatty acids (SCFAs) metabolism caused by DBDPE exposure remained unknown. In this study, three exposure groups (5, 50, 500 mg/L) and control group were used to investigate the effect of DBDPE by using simulator of the human intestinal microbial ecosystem (SHIME). 16S rRNA gene high-throughput sequencing illustrated that high dose DBDPE exposure increased the α-diversity of gut microbiota, while reduced the abundance of Firmicutes and Proteobacteria. In addition, the low dose (5 mg/L) DBDPE inhibited the increasing of SCFAs, but the medium and high dose (50 and 500 mg/L) DBDPE promoted the advancement, especially in ascending colon. Notably, DBDPE exposure lead a similar changing of acetic acid and butyric acid contents in different sections of the colon. This study confirmed the alternation of composition and metabolic function in gut microbial community due to DBDPE exposure, indicating an intestinal damage and appealing for more attention concentrated on the health effects of DBDPE exposure.

Tryptophan Supplementation Increases the Production of Microbial-Derived AhR Agonists in an In Vitro Simulator of Intestinal Microbial Ecosystem.

The aryl hydrocarbon receptor (AhR) plays an important role in intestinal homeostasis, and some microbial metabolites of tryptophan are known AhR agonists. In this study, we assessed the impact of tryptophan supplementation on the formation of tryptophan metabolites, AhR activation, and microbiota composition in the simulator of the human intestinal microbial ecosystem (SHIME). AhR activation, microbial composition, and tryptophan metabolites were compared during high tryptophan supplementation (4 g/L tryptophan), control, and wash-out periods. During tryptophan supplementation, the concentration of several tryptophan metabolites was increased compared to the control and wash-out period, but AhR activation by fermenter supernatant was significantly decreased. This was due to the higher levels of tryptophan, which was found to be an antagonist of AhR signaling. Tryptophan supplementation induced most microbial changes in the transverse colon including increased relative abundance of lactobacillus. We conclude that tryptophan supplementation leads to increased formation of AhR agonists in the colon.

Health Effect of N-Nitroso Diethylamine in Treated Water on Gut Microbiota Using a Simulated Human Intestinal Microbiota System

Chlorination disinfection byproducts (CDBPs) can exert adverse human health effects. Many toxicology-based studies confirmed the health hazards of CDBPs, but little research has been done on gut microbiome. We explored the effect of CDBPs on intestinal microbiota in the Simulator of the Human Intestinal Microbial Ecosystem (SHIME). The results showed that CDBPs slightly inhibited the production of short-chain fatty acids, and the abundance of Actinobacteria decreased in the transverse colon and descending colon. The abundance of Proteobacteria increased in the ascending colon and descending colon, while it decreased in the transverse colon. The abundance of Firmicutes decreased in both the ascending colon and descending colon. In particular, the abundance of Lachnospiraceae members, Bilophila, Oscillospira, Parabacteroides, Desulfovibrio, and Roseburia increased in the ascending colon, while the abundance of Sutterella, Bacteroides, Escherichia, Phascolarctobacterium, Clostridium, Citrobacter, and Klebsiella increased in the descending colon. The Shannon index differed significantly in both the ascending colon and descending colon before and after exposure. Overall, we demonstrate the feasibility of applying the SHIME model to studying the effects of intestinal toxicity on health of chlorinated by-products. The findings of this study improve our understanding of the health impact of CDBPs on the intestinal microbiota and better control of CDBPs in treated water is recommended.

Anti-obesity effects of Chenpi: an artificial gastrointestinal system study.

SummaryThe gut microbiota plays a significant role in human health; however, the complex relationship between gut microbial communities and host health is still to be thoroughly studied and understood. Microbes in the distal gut contribute to host health through the biosynthesis of vitamins and essential amino acids and the generation of important metabolic by‐products from dietary components that are left undigested by the small intestine. Aged citrus peel (Chenpi) is used in traditional Chinese medicine to lower cholesterol, promote weight loss and treat various gastrointestinal symptoms. This study investigated how the microbial community changes during treatment with Chenpi using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME). Two preparations of Chenpi extract were tested: Chenpi suspended in oil only and Chenpi in a viscoelastic emulsion. Short‐chain fatty acids (SCFAs) were measured during treatment to monitor changes in the microbial community of the colon presenting a decrease in production for acetic, propionic and butyric acid (ANOVA (P < 0.001) during the 15 days of treatment. 16S rRNA sequencing of microbial samples showed a clear difference between the two treatments at the different sampling times (ANOSIM P < 0.003; ADOSIM P < 0.002 [R 2 = 69%]). Beta diversity analysis by PcoA showed differences between the two Chenpi formulations for treatment day 6. These differences were no longer detectable as soon as the Chenpi treatment was stopped, showing a reversible effect of Chenpi on the human microbiome. 16S rRNA sequencing of microbial samples from the descending colon showed an increase in Firmicutes for the treatment with the viscoelastic emulsion. At the genus level, Roseburia, Blautia, Subdoligranulum and Eubacterium increased in numbers during the viscoelastic emulsion treatment. This study sheds light on the anti‐obesity effect of a polymethoxyflavone (PMFs)‐enriched Chenpi extract and creates a foundation for the identification of ‘obesity‐prevention’ biomarkers in the gut microbiota.

A novel functional herbal tea containing probiotic Bacillus coagulans GanedenBC30: An in vitro study using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME)

• Herbal tea as carrier of the probiotic Bacillus coagulans GanedenBC 30. • Spores of B. coagulans GanedenBC 30 markedly survive to infusion treatments. • Colonization and persistence in gut ecosystems investigated using the SHIME. • B. coagulans GanedenBC 30 quickly colonizes mucosal compartments and persists. • Viable cells in colon reactors are largely above the probiotic threshold. This study aimed at investigating the potential of a novel functional herbal tea to act as the carrier for the probiotic Bacillus coagulans GanedenBC 30 . Spores markedly survived to infusion treatments and the simulation of the gastrointestinal transit only slightly affected the survival, which was 94.8 ± 2.8%. An in-depth investigation using the Simulator of the Human Intestinal Microbial Ecosystem (SHIME) which incorporated mucin-covered microcosms was performed. Q-PCR with targeted primers demonstrated that the probiotic quickly colonized mucosal compartments and still persisted after 4 days of wash out. As estimated by plate count combined with 16S rRNA sequencing, B. coagulans GanedenBC 30 showed almost the same behavior in luminal compartments both during herbal tea intake and wash out. By summing the luminal and mucosal values and referring to whole volume of colon bioreactors, the content of viable cells of B. coagulans GanedenBC 30 was largely above the probiotic threshold.

The bioavailability of soy isoflavones in vitro and their effects on gut microbiota in the simulator of the human intestinal microbial ecosystem

Since soy isoflavones have many potentially beneficial properties, they have received increasing attention. However, there are few studies on their bioavailability and effects on gut microbiota. In this experiment, the simulator of the human intestinal microbial ecosystem (SHIME) was used to study the bioavailability of soy isoflavones and their effects on gut microbiota. The effects of soy isoflavones on the growth and antibacterial properties of probiotics were also studied in vitro. The results shows that the digestion of soy isoflavones mainly occurs in the colon, and the antioxidant capacity changes with the progress of digestion. Soy isoflavone can regulate gut microbiota, inhibit the growth of harmful bacteria, and affect the growth of obesity-related bacteria. Moreover, soy isoflavones can promote the growth of probiotics and significantly improve their antibacterial ability in vitro. Overall, soy isoflavones have the potential to be a functional food by improving gut microbiota.

Dechlorane Plus exposure on gut microbiome evaluated by using both in vivo and in vitro assays

The gut microbiome is crucial for the health of animals including human and is sensitive to environmental factors. Dechlorane Plus (DP) is a widely used polychlorinated flame retardant in commercial products for which general human population has been exposed to, but its effects on the gut microbiota are largely unknown. In this work, experiments were conducted to examine the effect of DP exposure on the intestinal microbiota by using Sprague-Dawley rats and a simulator of the human intestinal microbial ecosystem (SHIME). 16S rRNA gene high-throughput sequencing revealed that DP exposure in vivo reduced the relative abundance of the genera Prevotella, Quinella, and Ruminococcus, exposure in vitro decreased the relative abundance of Coriobacteriaceae, Parabacteroides, Bilophila, Sporolactobacillus, and Klebsiella. The phyla Synergistetes and Lentisphaerae were only detected in the descending colon. DP exposure also increased the concentrations of short-chain fatty acids (SCFAs) produced by the gut microbe both in vivo and in vitro, but SCFA metabolism was gradually inhibited as the increase of DP concentrations. These results indicated that DP exposure changed both the structure and metabolic function of the gut microbiota, suggesting more attention should be concentrated on the health effects of DP exposure, especially at higher doses.

The bioavailability of soybean polysaccharides and their metabolites on gut microbiota in the simulator of the human intestinal microbial ecosystem (SHIME)

The present study aimed to investigate the bioavailability of soybean polysaccharides and their metabolites on gut microbiota in the simulator of the human intestinal microbial ecosystem (SHIME). The effects of soybean polysaccharides on probiotics and pathogenic bacteria were investigated in vitro. Our results showed that soybean polysaccharides were only partially degraded in the oral, gastric, and small intestinal compartments of the SHIME. Moreover, the polysaccharides could be mainly broken down and utilized by the gut microbiota in the colon of the SHIME. Soybean polysaccharides could significantly reduce the ratio of Firmicutes to Bacteroidetes at the phylum level. Therefore, the number of beneficial bacteria were noticeably enhanced, and the pathogenic bacteria were inhibited. Furthermore, soybean polysaccharides promoted the growth of probiotics and improved the ability of these probiotics to inhibit pathogenic bacteria. Therefore, soybean polysaccharides could potentially be functional food to prevent disease by promoting gut health.

Are silver nanoparticles better than triclosan as a daily antimicrobial? Answers from the perspectives of gut microbiome disruption and pathogenicity

As an alternative to triclosan (TCS), the widespread use of silver nanoparticles (AgNPs) in daily products shows genuine potential. However, information regarding whether AgNPs are substantially better than TCS in their potential disruption of the gut microbiome and health effects is lacking. Using a simulator of the human intestinal microbial ecosystem (SHIME), we systemically compared the effects of TCS and AgNPs (at 1 μg/L and 30 μg/L) on the human gut microbiome in terms of changes in gut homeostasis, microbial community structure, antibiotic resistance profiles and abundances of opportunistic pathogens. Generally, TCS exerted more severe effects than AgNPs on gut disturbances (i.e., decreased production of short-chain fatty acids, increased contents of ammonium and total bile acids, and increased β-glucosidase activities) in a dose-dependent manner, whereas no clear dose effect was observed for the AgNP treatment because of potential nanoparticle transformation. The more serious effect of TCS than AgNPs on the microbiota composition was indicated by the dynamic increase in the Firmicutes/Bacteroidetes ratio determined using 16S rDNA sequencing. Metagenomic analyses revealed a more pronounced effect of TCS than AgNPs on the selection and dissemination of multiple resistance genes to antibiotics, TCS, and even Ag via the enrichment of genes encoding efflux pumps and mobile genetic elements. Consequently, the overgrowth of opportunistic pathogens was observed upon TCS exposure due to an imbalanced microbiome, in contrast to a slight increase in the abundance of some beneficial bacteria (i.e., Bifidobacterium) induced by the AgNP treatment. In conclusion, from the perspective of effects on gut health, AgNPs may prevail over TCS to some extent. However, the stress and potential selection of Ag resistance indicates the need for targeted surveillance of AgNP commercialization for daily use.

Colistin and amoxicillin combinatorial exposure alters the human intestinal microbiota and antibiotic resistome in the simulated human intestinal microbiota

Antibiotics treatment could cause the dysbiosis of human intestinal microbiota and antibiotic resistome. Fecal microbiota transplantation (FMT) has been an efficacious treatment to restore the dysbiosis of intestinal microbiota in a variety of intestinal diseases. However, to data, the effect of the combinatorial antibiotic treatment on microbiota, antibiotic resistome and the FMT for restoration affected by combinatorial antibiotic exposure in the human intestinal microbiota remain unclear. In this study, we systematically investigated the effect of the colistin and amoxicillin combinatorial exposure in the simulator of the human intestinal microbial ecosystem (SHIME) and found that this combinatorial exposure significantly altered (p < 0.05) the human intestinal microbiota and antibiotic resistome. The shift of bacterial community and antibiotic resistome could incompletely recovery to baseline by FMT treatment after combinatorial antibiotic exposure. Additionally, the variance of antibiotic resistome was dominantly driven by the bacterial community (41.18%–68.03%) after the combinatorial antibiotic exposure. Overall, this study first to investigate the influence of the colistin and amoxicillin combinatorial exposure on the intestinal microbiota and antibiotic resistome, and assess the FMT recovery in the simulated human intestinal microbiota, which may potentially provide a correct administration of antibiotics and application of FMT in the clinic.

Evaluation of the fate of Lactobacillus crispatus BC4, carried in Squacquerone cheese, throughout the simulator of the human intestinal microbial ecosystem (SHIME)

Lactobacillus crispatus strain BC4, isolated from the human healthy vaginal environment and characterised by a strong antimicrobial activity against urogenital pathogens and foodborne microorganisms, was employed as a probiotic culture in the cheesemaking of the soft cheese Squacquerone. Such cheese is intended as a “gender food”, that could be used as a hedonistic dietary strategy to reduce the incidence of woman vaginal dysbiosis and infections, given the evidence that a probiotic strain able to survive to the entire digestive process once ingested, can pass from intestine to vagina. This work was aimed to evaluate the resistance of L. crispatus BC4, carried in Squacquerone cheese, to different challenges of the human gastrointestinal tract, including the colon stage. The digestion process was tested using a Simulator of the Human Intestinal Microbial Ecosystem (SHIME®). The viability and metabolic activity of L. crispatus BC4 during the colon simulation were monitored by qPCR and gas chromatography, respectively, also in the presence of a complex microbiota. The results showed that L. crispatus BC4 survival was not affected by the gastric condition, while it was significantly affected by bile salts and pancreatic juice in small intestine conditions, where it decreased of approx. 0.6 log (colony-forming units) CFU/g. Differently, during colon simulation L. crispatus BC4 was able to grow in sterile colon conditions and to maintain viability in the presence of a complex microbiota. Moreover, during colon simulation, L. crispatus BC4 was metabolically active as demonstrated by the higher production of short chain fatty acids (SCFA) and lactate. In the presence of a complex gut microbiota, a decrease of lactate was observed, due to its conversion into propionate (anti-cholesterol activity) and butyrate (anti-inflammatory activity) by cross-feeding. However, no differences in propionate and butyrate production could be observed between control cheese and cheese containing L. crispatus BC4. Despite this may appear as a negative outcome, it must be taken into account that, in this setup, only a single dose of the cheese was tested and the outcome of the colonization and impact of the gut microbiota might be different when daily repeated doses are tested.

Assessment of nutrients effect on the bioaccessibility of Cd and Cu in contaminated soil

Soil is considered as a sink for heavy metals. Human health is severely affected by the continuous intake of toxic heavy metals even in a very low concentration. In the present experiment, we determined the influence of nutritional status including control (fasted condition), glucose (fed state), plant protein (fed state), animal protein (fed state) and calcium (fed state) on soil cadmium (Cd) and copper (Cu) bioaccessibility using physiologically-based extraction test (PBET) method together with simulator of the human intestinal microbial ecosystem (SHIME) model. The bioaccessibility of Cd was 1.06–73.58%, 0.44–54.79% and 0–17.78% and Cu was 3.81–67.32%, 4.98–71.14%, and 0–17.54% in the phase-I, phase-II and Phase-III respectively (in this study gastric phase, small intestinal phase and colon phase were considered as phase-I, phase-II and Phase-III respectively). The outcomes showed that, the average Cd bioaccessibility was higher with animal protein addition compared with other treatments in different phases. So, the effect of animal protein on Cd bioaccessibility was higher than other treatments in the phase-I, phase-II and phase-III. Due to the addition of plant protein, the higher average bioaccessibility of Cu was noticed in phase-I and phase-II in comparison to other treatments. However, in phase-III, the higher average bioaccessibility of Cu was found due to control treatment comparing with other treatments. Therefore, the influence of plant protein was higher than other nutrients on Cu bioaccessibility in the phase-I and phase-II. Moreover, other nutrients showed variable influence on Cd and Cu bioaccessibility. So, nutritional status has a significant effect on bioaccessibility as well as human health risk assessment.

Amoxicillin Increased Functional Pathway Genes and Beta-Lactam Resistance Genes by Pathogens Bloomed in Intestinal Microbiota Using a Simulator of the Human Intestinal Microbial Ecosystem.

Antibiotics are frequently used to treat bacterial infections; however, they affect not only the target pathogen but also commensal gut bacteria. They may cause the dysbiosis of human intestinal microbiota and consequent metabolic alterations, as well as the spreading of antibiotic resistant bacteria and antibiotic resistance genes (ARGs). In vitro experiments by simulator of the human intestinal microbial ecosystem (SHIME) can clarify the direct effects of antibiotics on different regions of the human intestinal microbiota, allowing complex human microbiota to be stably maintained in the absence of host cells. However, there are very few articles added the antibiotics into this in vitro model to observe the effects of antibiotics on the human intestinal microbiota. To date, no studies have focused on the correlations between the bloomed pathogens caused by amoxicillin (AMX) exposure and increased functional pathway genes as well as ARGs. This study investigated the influence of 600 mg day–1 AMX on human intestinal microbiota using SHIME. The impact of AMX on the composition and function of the human intestinal microbiota was revealed by 16S rRNA gene sequencing and high-throughput quantitative PCR. The results suggested that: (i) AMX treatment has tremendous influence on the overall taxonomic composition of the gut microbiota by increasing the relative abundance of Klebsiella [linear discriminant analysis (LDA) score = 5.26] and Bacteroides uniformis (LDA score = 4.75), as well as taxonomic diversity (Simpson, P = 0.067, T-test; Shannon, P = 0.061, T-test), and decreasing the members of Parabacteroides (LDA score = 4.18), Bifidobacterium (LDA score = 4.06), and Phascolarctobacterium (LDA score = 3.95); (ii) AMX exposure significantly enhanced the functional pathway genes and beta-lactam resistance genes, and the bloomed pathogens were strongly correlated with the metabolic and immune system diseases gene numbers (R = 0.98, P < 0.001) or bl2_len and bl2be_shv2 abundance (R = 0.94, P < 0.001); (iii) the changes caused by AMX were “SHIME-compartment” different with more significant alteration in ascending colon, and the effects were permanent, which could not be restored after 2-week AMX discontinuance. Overall results demonstrated negative side-effects of AMX, which should be considered for AMX prescription.

Vancomycin exposure caused opportunistic pathogens bloom in intestinal microbiome by simulator of the human intestinal microbial ecosystem (SHIME)

Antibiotics are emerging organic pollutants posing high health risks to humans by causing human intestinal microbial disorders with increasing abundances of opportunistic pathogens, and fecal microbiota transplantation (FMT) has been confirmed to restore the dysbiosis of gut flora in many kinds of intestinal disease. However, to date, few studies have focused on the bloomed opportunistic pathogens associated human disease-related pathways as well as antibiotic resistance genes (ARGs) after vancomycin exposure, and there is limited information on using FMT for restoration of intestinal microbiome affected by antibiotics. Therefore, this study investigated effects of vancomycin on the opportunistic pathogens, human disease-related pathways as well as ARGs in human gut, and the restoration of intestinal microbiome by FMT. Results indicated that vancomycin treatment substantially increased human disease-related pathways and decreased abundances of ARGs. Besides, the bloomed opportunistic pathogens including Achromobacter, Klebsiella, and Pseudomonas, caused by vancomycin exposure, were positively correlated with human disease-related pathways. The microbiota abundance and genes of human disease-related pathways and antibiotic resistance showed a remarkable return towards baseline after FMT, but not for natural recovery. These findings suggest that impacts of vancomycin on human gut are profound and FMT will be a promising strategy in clinical application that can restore the dysbiosis of gut microbiota, which may be valuable for directing future work.