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A study was conducted to characterize the bacterial and biochemical composition of the jejunoileal content of veal calves and the effect of pre-slaughter fasting time. At 22 wk of age, 22 preruminant Prim'Holstein calves fed milk replacer and pellets (mainly composed of corn) were slaughtered at 6, 12, or 24 h after their last meal. Chyme samples were collected from the jejunoileal compartment just after slaughter, and pH and redox potential were immediately measured. Culture-based methods were used to determine the concentrations of total anaerobic microflora, lactate-utilizing bacteria, Bacteroides fragilis group, Lactobacilli, Bifidobacteria, Enterococci, and 2 potential pathogenic species, Escherichia coli and Clostridium perfringens. Concentrations of L-lactate, ammonia, and short-chain fatty acids (SCFA) were determined on frozen samples. The biochemical composition (DM, total protein, lactose, galactose, glucose, minerals, AA profile, and fatty acid profile) of the jejunoileal content was determined only on samples from the 6-h fasted group. Microflora concentrations were greater (P < 0.01) in the 6-h fasted group compared with the 12- and 24-h fasted groups, involving a decreased pH (P < 0.05) and greater lactate and SCFA concentrations, both linked directly to the fermentative state of the microorganisms. The 6-h fasted group showed the least interanimal variability in bacterial group levels, except for Cl. perfringens, which presented increased interanimal variability regardless of fasting time. At 6 h postprandial, the jejunoileal content of veal calves seemed to be in a stable state, allowing the creation of a database on its biochemical composition. This study is a key first step in the development of an in vitro system for modeling the jejunoileal ecosystem of veal calves. This model will provide a useful tool for assessing the effects of feed additives on intestinal microflora. Key words: biochemical composition, fasting time, fermentative metabolite, intestinal microflora, jejunoileum, veal calf

The aim of the study was to assess the ability of a dynamic in vitro model to determine the digestibility of OM, CP, and starch compared with a validated, static, in vitro method and in vivo ileal digestibility obtained from growing pigs fitted with a T-cannula. Five experimental diets with different carbohydrate types and level were assessed: a standard cornbased diet (ST) or the same diet with coarse ground corn (CC), 8% sugar beet pulp (BP), 10% wheat bran (WB), or 8% sugar beet pulp and 10% wheat bran (HF). In the in vivo experiment, diets CC and HF reduced (P = 0.015) ileal digestibility of OM compared with the ST diet. The inclusion of sugar beet pulp reduced (P = 0.049) ileal CP digestibility of the BP diet. This reduction was not statistically significant when sugar beet pulp was combined with the wheat bran in the HF diet. No differences were shown for in vivo starch digestibility among diets. With the static in vitro method, the OM disappearance was greater than that observed in the in vivo experiment. In this static method, the BP and HF diets reduced (P = 0.004 and < 0.001, respectively) the disappearance of the OM compared with the ST diet. The coarse grinding of corn did not alter OM digestibility but decreased (P = 0.005) the starch digest ibility. The [R.sup.2] between the in vivo results and the static in vitro methods for OM and starch digestibility was 0.99 when the CC diet was not considered. The dynamic in vitro model yielded OM and CP digestibility coefficients comparable with those obtained in vivo for the ST and CC diets. However, the values were considerably affected by the incorporation of the fibrous ingredients. Diets BP, WB, and HF had decreased (P = 0.009, 0.058, and 0.004, respectively) OM digestibility compared with the ST diet. Protein digestibility was also decreased (P < 0.001, P = 0.019, and P = 0.003, respectively) with the BP, WB, and HF diets compared with the ST diet. However, digestibility was decreased to a greater extent in the BP diet than in the WB and HF diets, both of which contained wheat bran. The [R.sup.2] between the dynamic in vitro model and the in vivo results for CP digestibility was 0.99 when the CC diet was not considered. No differences were detected for starch digestibility among the diets with the dynamic in vitro model. This dynamic in vitro model yielded ileal digestibility results comparable with those obtained in vivo for CP and OM with a corn-soybean diet, or with a diet including coarse corn, but it underestimated digestibility when fibrous ingredients were included in the diet. Key words: carbohydrate, digestibility, in vitro model, pig, sugar beet pulp, wheat bran

The aim of this study was to develop sustained release microspheres of capsicum oleoresin as an alternative to in-feed additives. Two spray-cooling technologies, a fluidized air bed using a spray nozzle system and a vibrating nozzle system placed on top of a cooling tower, were used to microencapsulate 20% of capsicum oleoresin in a hydrogenated, rapeseed oil matrix. Microencapsulation was intended to reduce the irritating effect of capsicum oleoresin and to control its release kinetics during consumption by the animal. Particles produced by the fluidized air bed process (batch F1) ranged from 180 to 1,000 [micro]m in size. The impact of particle size on release of capsaicin, the main active compound of capsicum oleoresin, was studied after sieving batch F1 to obtain 4 formulations: F1a (180 to 250 [micro]m), F1b (250 to 500 [micro]m), F1c (500 to 710 [micro]m), and F1d (710 to 1,000 [micro]m). The vibrating nozzle system can produce a monodispersive particle size distribution. In this study, particles of 500 to 710 [micro]m were made (batch F2). The release kinetics of the formulations was estimated in a flow-through cell dissolution apparatus (CFC). The time to achieve a 90% dissolution value (T90%) of capsaicin for subbatches of F1 increased with the increase in particle size (P < 0.05), with the greatest value of 165.5 [+ or -] 13.2 min for F1d. The kinetics of dissolution of F2 was slower than all F1 subbatches, with a T90% of 422.7 [+ or -] 30.0 rain. Nevertheless, because CFC systems are ill suited for experiments with solid feed and thus limit their predictive values, follow-up studies were performed on F1c and F2 using an in vitro dynamic model that simulated more closely the digestive environment. For both formulations a lower quantity of capsaicin dialyzed was recorded under fed condition vs. fasting condition with 46.9% [+ or -] 1.0 vs. 74.7% [+ or -] 2.7 for F1c and 32.4% [+ or -] 1.4 vs. 44.2% [+ or -] 2.6 for F2, respectively. This suggests a possible interaction between capsaicin and the feed matrix. Moreover, 40.4 [+ or -] 3.9% of the total capsaicin intake in F2 form was dialyzed after 8 h of digestion when feed had been granulated vs. 32.4 [+ or -] 1.4% when feed had not been granulated, which suggests that the feed granulation process could lead to a partial degradation of the microspheres and to a limitation of the sustained release effect. This study demonstrates the potential and the limitations of spray-cooling technology to encapsulate feed additives. Key words: capsaicin, capsicum oleoresin, microencapsulation, spray cooling

The aim of this study was to develop sustained release plant extracts as a potential alternative to antibiotic growth promoters for growing pigs. Pellets with a core based on microcrystalline cellulose and 3 active compounds (eugenol, carvacrol, and thymol) were prepared using rotary fluidized-bed technology. Two particle sizes were produced that had a mean size of approximately 250 and 500 [micro]m. Results show the process was able to produce pellets with a spherical and homogenous form when 10% of the active compounds were incorporated into the core. When active compounds were increased to 20%, the pellet became stickier, and the yield decreased from 90 to 65%. Different amounts of coating in the form of an aqueous-based ethylcellulose (EC) dispersion (Surelease) were applied to the core to modify the release of active compounds. The efficacy of the coating was evaluated in vitro using a flow-through cell apparatus. The time to achieve 50 and 90% dissolution increased with the increase in particle size (P < 0.05) and the increase in EC-coating level from 10 to 20% (wt/wt; P < 0.05), indicating the ability of the process to slow release depending on particle size and the amount of polymer applied. Differences in the release of the active compounds were observed in the same formulation of pellets, except for the formulation with small 10%-EC-coated particles, in which the active compounds were rapidly dissolved (more than 85% in 15 min or less). For all other formulations, the dissolution time for eugenol was always faster than for thymol or carvacrol. The close monitoring of plant extract behavior in the gastrointestinal tract could become a key factor in the continued use of phyto-molecules as alternatives to antibiotic growth promoters and in optimizing the balance between cost and efficacy. Different microencapsulation technologies can be used, of which the rotary fluidized bed warrants consideration because of the quality of the products obtained. Key words: feed additive, growing pig, in vitro dissolution, pellet, plant extract, rotary fluidized bed

A Recombinant Saccharomyces cerevisiae strain expressing plant is used as a model to study the role of cytochrome P450 in the digestive environment of a rat. The approach can be further used to develop new drug delivery systems based on yeast catalyzing a bioconversion reaction in the digestive tract.

Research has been conducted on model recombinant Saccharomyces cerevisiae which expresses heterologous CA4H. The survival rate and bioconversion activity of this recombinant have been investigated via the use of recombinant yeasts as convenient hosts for biodrug development, and the details are reported.

During the last decade, there has been a growing interest in understanding the fate of food during digestion in the gastrointestinal tract in order to strengthen the possible effects of food on human health. Ideally, food digestion should be studied in vivo on humans but this is not always ethically and financially possible. Therefore simple static in vitro digestion models mimicking the gastrointestinal tract have been proposed as alternatives to in vivo experiments but these models are quite basic and hardly recreate the complexity of the digestive tract. In contrast, dynamic models that allow pH regulation, flow of the food and injection in real time of digestive enzymes in the different compartments of the gastrointestinal tract are more promising to accurately mimic the digestive process. Most of the systems developed so far have been compared for their performances to in vivo data obtained on animals and/or humans. The objective of this article is to review the validation towards in vivo data of some of the dynamic digestion systems currently available in order to determine what aspects of food digestion they are able to mimic. Eight dynamic digestion systems are presented as well as their validation towards in vivo data. Advantages and limits of each simulator is discussed. This is the result of a cooperative international effort made by some of the scientists involved in Infogest, an international network on food digestion. [ABSTRACT FROM AUTHOR]

This paper introduces a new type of system to simulate conditions in the large intestine. This system combines removal of metabolites and water with peristaltic mixing to obtain and handle physiological concentrations of microorganisms, dry matter and microbial metabolites. The system has been designed to be complementary to the dynamic multi-compartmental system that simulates conditions in the stomach and small intestine described by Minekus et al. [Minekus M, Marteau P, Havenaar R, Huis in't Veld JHJ (1995) ATLA 23:197–209]. High densities of microorganisms, comparable to those found in the colon in vivo, were achieved by absorption of water and dialysis of metabolites through hollow-fibre membranes inside the reactor compartments. The dense chyme was mixed and transported by peristaltic movements. The potential of the system as a tool to study fermentation was demonstrated in experiments with pectin, fructo-oligosaccharide, lactulose and lactitol as substrates. Parameters such as total acid production and short-chain fatty acid (SCFA) patterns were determined with time to characterize the fermentation. The stability of the microflora in the system was tested after inoculation with fresh fecal samples and after inoculation with a microflora that was main-tained in a fermenter. Both approaches resulted in total anaerobic bacterial counts higher than 1010 colony-forming units/ml with physiological levels of Bifidobacterium, Lactobacillus, Enterobacteriaceae and Clostridium. The dry matter content was approximately 10%, while the total SCFA concentration was maintained at physiological concentrations with similar molar ratios for acetic acid, propionic acid and butyric acid as measured in vivo. [ABSTRACT FROM AUTHOR]

The methanogenic strain Mx-05 T was isolated from the human fecal microbiome. A phylogenetic analysis based on the 16S rRNA gene and protein marker genes indicated that the strain is affiliated with the order Methanomassiliicoccales . It shares 86.9% 16S rRNA gene sequence identity with Methanomassiliicoccus luminyensis , the only member of this order previously isolated. The cells of Mx-05 T were non-motile cocci, with a diameter range of 0.4-0.7 μm. They grew anaerobically and reduced methanol, monomethylamine, dimethylamine, and trimethylamine into methane, using H 2 as an electron donor. H 2 /CO 2 , formate, ethanol, and acetate were not used as energy sources. The growth of Mx-05 T required an unknown medium factor(s) provided by Eggerthella lenta and present in rumen fluid. Mx-05 T grew between 30 °C and 40 °C (optimum 37 °C), over a pH range of 6.9-8.3 (optimum pH 7.5), and between 0.02 and 0.34 mol.L -1 NaCl (optimum 0.12 mol.L -1 NaCl). The genome is 1.67 Mbp with a G+C content of 55.5 mol%. Genome sequence annotation confirmed the absence of the methyl branch of the H 4 MPT Wood-Ljungdahl pathway, as described for other Methanomassiliicoccales members. Based on an average nucleotide identity analysis, we propose strain Mx-05 T as being a novel representative of the order Methanomassiliicoccales , within the novel family Methanomethylophilaceae , for which the name Methanomethylophilus alvi gen. nov, sp. nov. is proposed. The type strain is Mx-05 T (JCM 31474T).

We aimed to assess if casein structure affects its digestion and its subsequent amino acid delivery kinetic. Higher nitrogen levels were recovered in dialysates after in vitro digestions of sodium caseinate (SC, formed of small aggregates) compared to micellar casein (MC, native form of casein) and calcium caseinate (CC, intermediate structure). Likewise, plasma indispensable amino-acid concentration peak was higher after SC compared to MC or CC ingestion in healthy volunteers in a randomized, double blind, cross-over study. In pigs, gamma-scintigraphy using labelled meals revealed that SC was mainly localized in the proximal part of the stomach whereas MC was distributed in the whole gastric cavity. Caseins were found in both solid and liquid phases and partly hydrolyzed casein in the solid phase shortly after SC drink ingestion. These data support the concept of slow (MC) and rapid (SC) casein depending of casein structure, likely due to their intra-gastric clotting properties., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Gaelle BOUDRY reports financial support and equipment, drugs, or supplies were provided by Ingredia SA. Audrey BOULIER reports a relationship with Ingredia SA that includes: employment. Alain BANIEL reports a relationship with Ingredia SA that includes: employment. Gaelle BOUDRY reports a relationship with Ingredia SA that includes: travel reimbursement., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Metagenome analyses of the human microbiome suggest that horizontal gene transfer (HGT) is frequent in these rich and complex microbial communities. However, so far, only a few HGT studies have been conducted in vivo . In this work, three different systems mimicking the physiological conditions encountered in the human digestive tract were tested, including (i) the TNO gastro-Intestinal tract Model 1 (TIM-1) system (for the upper part of the intestine), (ii) the ARtificial COLon (ARCOL) system (to mimic the colon), and (iii) a mouse model. To increase the likelihood of transfer by conjugation of the integrative and conjugative element studied in the artificial digestive systems, bacteria were entrapped in alginate, agar, and chitosan beads before being placed in the different gut compartments. The number of transconjugants detected decreased, while the complexity of the ecosystem increased (many clones in TIM-1 but only one clone in ARCOL). No clone was obtained in a natural digestive environment (germfree mouse model). In the human gut, the richness and diversity of the bacterial community would offer more opportunities for HGT events to occur. In addition, several factors (SOS-inducing agents, microbiota-derived factors) that potentially increase in vivo HGT efficiency were not tested here. Even if HGT events are rare, expansion of the transconjugant clones can happen if ecological success is fostered by selecting conditions or by events that destabilize the microbial community. IMPORTANCE The human gut microbiota plays a key role in maintaining normal host physiology and health, but its homeostasis is fragile. During their transit in the gastrointestinal tract, bacteria conveyed by food can exchange genes with resident bacteria. New traits acquired by HGT (e.g., new catabolic properties, bacteriocins, antibiotic resistance) can impact the gut microbial composition and metabolic potential. We showed here that TIM-1, a system mimicking the upper digestive tract, is a useful tool to evaluate HGT events in conditions closer to the physiological ones. Another important fact pointed out in this work is that Enterococcus faecalis is a good candidate for foreign gene acquisition. Due to its high ability to colonize the gut and acquire mobile genetic elements, this commensal bacterium could serve as an intermediate for HGT in the human gut., Competing Interests: The authors declare no conflict of interest.

Early life is a critical period where gut ecosystem and functions are being established with significant impact on health. For regulatory, technical, and cost reasons, in vitro gut models can be used as a relevant alternative to in vivo assays. An exhaustive literature review was conducted to adapt the Mucosal Artificial Colon (M-ARCOL) to specific physicochemical (pH, transit time, and nutritional composition of ileal effluents) and microbial parameters from toddlers in the age range of 6 months-3 years, resulting in the Tm-ARCOL. In vitro fermentations were performed to validate this newly developed colonic model compared to in vivo toddler data. Results were also compared to those obtained with the classical adult configuration. Fecal samples from 5 toddlers and 4 adults were used to inoculate bioreactors, and continuous fermentations were performed for 8 days. Gut microbiota structure (lumen and mucus-associated microbiota) and functions (gas and short-chain fatty acids) were monitored. Clearly distinct microbial signatures were obtained between the two in vitro conditions, with lower α-diversity indices and higher abundances of infant-related microbial populations (e.g., Bifidobacteriaceae, Enterobacteriaceae) in toddler versus adult conditions. In accordance with in vivo data, methane was found only in adult bioreactors, while higher percentage of acetate but lower proportions of propionate and butyrate was measured in toddlers compared to adults. This new in vitro model will provide a powerful platform for gut microbiome mechanistic studies in a pediatric context, both in nutritional- (e.g., nutrients, probiotics, prebiotics) and health-related (e.g., drugs, enteric pathogens) studies. KEY POINTS: • Development of a novel in vitro colonic model recapitulating the toddler environment. • Specific toddler versus adult digestive conditions are preserved in vitro. • The new model provides a powerful platform for microbiome mechanistic studies., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The human digestion is a multi-step and multi-compartment process essential for human health, at the heart of many issues raised by academics, the medical world and industrials from the food, nutrition and pharma fields. In the first years of life, major dietary changes occur and are concomitant with an evolution of the whole child digestive tract anatomy and physiology, including colonization of gut microbiota. All these phenomena are influenced by child exposure to environmental compounds, such as drugs (especially antibiotics) and food pollutants, but also childhood infections. Due to obvious ethical, regulatory and technical limitations, in vivo approaches in animal and human are more and more restricted to favor complementary in vitro approaches. This review summarizes current knowledge on the evolution of child gut physiology from birth to 3 years old regarding physicochemical, mechanical and microbial parameters. Then, all the available in vitro models of the child digestive tract are described, ranging from the simplest static mono-compartmental systems to the most sophisticated dynamic and multi-compartmental models, and mimicking from the oral phase to the colon compartment. Lastly, we detail the main applications of child gut models in nutritional, pharmaceutical and microbiological studies and discuss the limitations and challenges facing this field of research., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Natural mineral water (NMWs) intake has been traditionally used in the treatment of various gastrointestinal diseases. We investigated the effect of two French NMWs, one a calcium and magnesium sulphate, sodium chloride, carbonic, and ferruginous water (NMW1), the other a mainly bicarbonate water (NMW2) on the prevention of intestinal inflammation. Intestinal epithelial cells stimulated with heat inactivated Escherichia coli or H 2 O 2 were treated with NMWs to evaluate the anti-inflammatory effects. Moderate colitis was induced by 1% dextran sulfate sodium (DSS) in Balbc/J mice drinking NMW1, NWW2, or control water. General signs and histological features of colitis, fecal lipocalin-2 and pro-inflammatory KC cytokine levels, global mucosa-associated microbiota, were analyzed. We demonstrated that both NMW1 and NMW2 exhibited anti-inflammatory effects using intestinal cells. In induced-colitis mice, NMW1 was effective in dampening intestinal inflammation, with significant reductions in disease activity scores, fecal lipocalin-2 levels, pro-inflammatory KC cytokine release, and intestinal epithelial lesion sizes. Moreover, NMW1 was sufficient to prevent alterations in the mucosa-associated microbiota. These observations, through mechanisms involving modulation of the mucosa-associated microbiota, emphasize the need of investigation of the potential clinical efficiency of such NMWs to contribute, in human beings, to a state of low inflammation in inflammatory bowel disease.

In vitro gut models, such as the mucosal artificial colon (M-ARCOL), provide timely and cost-efficient alternatives to in vivo assays allowing mechanistic studies to better understand the role of human microbiome in health and disease. Using such models inoculated with human fecal samples may require a critical step of stool storage. The effects of preservation methods on microbial structure and function in in vitro gut models have been poorly investigated. This study aimed to assess the impact of three commonly used preserving methods, compared with fresh fecal samples used as a control, on the kinetics of lumen and mucus-associated microbiota colonization in the M-ARCOL model. Feces from two healthy donors were frozen 48 h at - 80 °C with or without cryoprotectant (10% glycerol) or lyophilized with maltodextrin and trehalose prior to inoculation of four parallel bioreactors (e.g., fresh stool, raw stool stored at - 80 °C, stool stored at - 80 °C with glycerol and lyophilized stool). Microbiota composition and diversity (qPCR and 16S metabarcoding) as well as metabolic activity (gases and short chain fatty acids) were monitored throughout the fermentation process (9 days). All the preservative treatments allowed the maintaining inside the M-ARCOL of a complex and functional microbiota, but considering stabilization time of microbial profiles and activities (and not technical constraints associated with the supply of frozen material), our results highlighted 48 h freezing at - 80 °C without cryoprotectant as the most efficient method. These results will help scientists to determine the most accurate method for fecal storage prior to inoculation of in vitro gut microbiome models. KEY POINTS: • In vitro ARCOL model reproduces luminal and mucosal human microbiome. • Short-term storage of fecal sample influences microbial stabilization and activity. • 48 h freezing at - 80°C: most efficient method to preserve microbial ecosystem. • Scientific and technical requirements: influencers of preservation method.

Background: Enterotoxigenic Escherichia coli (ETEC) substantially contributes to the burden of diarrheal illnesses in developing countries. With the use of complementary in vitro models of the human digestive environment, TNO gastrointestinal model (TIM-1), and Mucosal Simulator of the Human Intestinal Microbial Ecosystem (M-SHIME), we provided the first detailed report on the spatial-temporal modulation of ETEC H10407 survival, virulence, and its interplay with gut microbiota. These systems integrate the main physicochemical parameters of the human upper digestion (TIM-1) and simulate the ileum vs ascending colon microbial communities and luminal vs mucosal microenvironments, captured from six fecal donors (M-SHIME)., Results: A loss of ETEC viability was noticed upon gastric digestion, while a growth renewal was found at the end of jejunal and ileal digestion. The remarkable ETEC mucosal attachment helped to maintain luminal concentrations above 6 log 10  mL -1 in the ileum and ascending colon up to 5 days post-infection. Seven ETEC virulence genes were monitored. Most of them were switched on in the stomach and switched off in the TIM-1 ileal effluents and in a late post-infectious stage in the M-SHIME ascending colon. No heat-labile enterotoxin production was measured in the stomach in contrast to the ileum and ascending colon. Using 16S rRNA gene-based amplicon sequencing, ETEC infection modulated the microbial community structure of the ileum mucus and ascending colon lumen., Conclusions: This study provides a better understanding of the interplay between ETEC and gastrointestinal cues and may serve to complete knowledge on ETEC pathogenesis and inspire novel prophylactic strategies for diarrheal diseases.

The objective of this study was to evaluate the benefits of a new extraction process, the ipowder® technology, applied to Melissa officinalis L. Compared to M. officinalis ground dry leaves, the ipowder® had a similar phytochemical fingerprint but contained twice the concentration of rosmarinic acid (by HPTLC and HPLC) and had a two-fold greater antioxidant activity (DPPH* method). In vitro digestion experiments (TIM-1 model) showed better availability of rosmarinic acid for intestinal absorption with the ipowder® than with ground dry leaves, manifested by a three-fold reduction in the quantity of ingested product needed for delivery of the same amount of rosmarinic acid into the upper gastro-intestinal tract. This study shows that the ipowder® technology preserves all the original plant compounds intact while making some active ingredients more accessible and available to exert their effects. To obtain a given effect, the amount of ipowder® extract to ingest will therefore be lower; a reduction in the daily dosage will be more convenient for the patient and will improve patient compliance with supplementation.

Background : The availability of fresh stool samples is a prerequisite in most gut microbiota functional studies. Objective : Strategies for amplification and long-term gut microbiota preservation from fecal samples would favor sample sharing, help comparisons and reproducibility over time and between laboratories, and improve the safety and ethical issues surrounding fecal microbiota transplantations. Design : Taking advantage of in vitro gut-simulating systems, we amplified the microbial repertoire of a fresh fecal sample and assessed the viability and resuscitation of microbes after preservation with some common intracellular and extracellular acting cryoprotective agents (CPAs), alone and in different combinations. Preservation efficiencies were determined after 3 and 6 months and compared with the fresh initial microbiota diversity and metabolic activity, using the chemostat-based Environmental Control System for Intestinal Microbiota (ECSIM) in vitro model of the gut environment. Microbial populations were tested for fermentation gas, short-chain fatty acids, and composition of amplified and resuscitated microbiota, encompassing methanogenic archaea. Results : Amplification of the microbial repertoire from a fresh fecal sample was achieved with high fidelity. Dimethylsulfoxide, alone or mixed with other CPAs, showed the best efficiency for functional preservation, and the duration of preservation had little effect. Conclusions : The amplification and resuscitation of fecal microbiota can be performed using specialized in vitro gut models. Correct amplification of the initial microbes should ease the sharing of clinical samples and improve the safety of fecal microbiota transplantation. Abbreviations : CDI, Clostridium difficile infection; CPA, cryoprotective agent; D, DMSO, dimethylsulfoxide; FMT, fecal microbiota transplantation; G, glycerol; IBD, inflammatory bowel disease; P, PEG-4000, polyethylene glycol 4000 g.mol -1 ; SCFA, short-chain fatty acid; SNR, signal-to-noise ratio.

For ethical, technical, regulatory, and cost reasons, in vitro methods are increasingly used as an alternative to in vivo experimentations. The aim of the present study was to validate, according to in vivo data in living animals, a new in vitro model of the piglet colon, the PigutIVM, under both control conditions and antibiotic disturbance by the widely used colistin. The PigutIVM reproduces the main biotic and abiotic parameters of the piglet colon: temperature, pH, retention time, supply of ileal effluents, complex, and metabolically active microbiota and self-maintained anaerobiosis. Under both control and antibiotic-treated conditions, qPCR analyses showed that the main bacterial populations of piglet gut microbiota were similar in vitro and in vivo, with Pearson correlation coefficient higher than 0.9. During colistin administration, both in piglets and in the in vitro model, a significant decrease in Escherichia coli populations was observed together with changes in microbial composition of subdominant populations. SCFA concentrations were similar in vitro and in vivo and were not modified by colistin. Interestingly, the administration of the probiotic Saccharomyces cerevisiae var. boulardii CNCM I-1079 led in vitro to a decrease in E. coli levels, as previously observed when the antibiotic treatment was applied. This new in vitro model of the piglet colon provides a flexible, reproducible, and cost-effective tool for the screening of drugs or new dietary compounds, such as pre- or probiotics. It will be helpful for researchers, feed producers, or veterinarians when developing innovative non-antibiotic strategies.

Enterohemorrhagic Escherichia coli (EHEC) are major food-borne pathogens that constitute a serious public health threat. Currently, there is no specific treatment available for EHEC infections in human creating an urgent need for the development of alternative therapeutic strategies. Among them, one of the most promising approaches is the use of probiotic microorganisms. Even if many studies have shown the antagonistic effects of probiotic bacteria or yeast on EHEC survival, virulence, adhesion on intestinal epithelium or pathogen-induced inflammatory responses, mechanisms mediating their beneficial effects remain unclear. This review describes EHEC pathogenesis and novel therapeutic strategies, with a particular emphasis on probiotics. The interests and limits of a probiotic-based approach and the way it might be incorporated into global health strategies against EHEC infections will be discussed.

Background/aims: Human gut microbiota harbors numerous metabolic properties essential for the host's health. Increased intestinal transit time affects a part of the population and is notably observed with human aging, which also corresponds to modifications of the gut microbiota. Thus we tested the metabolic and compositional changes of a human gut microbiota induced by an increased transit time simulated in vitro., Methods: The in vitro system, Environmental Control System for Intestinal Microbiota, was used to simulate the environmental conditions of 3 different anatomical parts of the human colon in a continuous process. The retention times of the chemostat conditions were established to correspond to a typical transit time of 48 hours next increased to 96 hours. The bacterial communities, short chain fatty acids and metabolite fingerprints were determined., Results: Increase of transit time resulted in a decrease of biomass and of diversity in the more distal compartments. Short chain fatty acid analyses and metabolite fingerprinting revealed increased activity corresponding to carbohydrate fermentation in the proximal compartments while protein fermentations were increased in the lower parts., Conclusions: This study provides the evidence that the increase of transit time, independently of other factors, affects the composition and metabolism of the gut microbiota. The transit time is one of the factors that explain some of the modifications seen in the gut microbiota of the elderly, as well as patients with slow transit time.

Background: Enterohemorrhagic Escherichia coli (EHEC) are major foodborne pathogens that constitute a serious public health threat, mainly in young children. Shiga toxins (Stx) are the main virulence determinants of EHEC pathogenesis but adhesins like intimin (eae) and Long polar fimbriae (Lpf) also contribute to infection. The TNO GastroIntestinal Model (TIM) was used for a comparative study of EHEC O157:H7 survival and virulence under adult and child digestive conditions., Methods: Survival kinetics in the in vitro digestive tract were determined by plating while bacterial viability was assessed by flow cytometry analysis. Expression of stx, eae, and lpf genes was followed by reverse transcriptase-quantitative PCR (RT-qPCR) and Stx production was measured by ELISA (enzyme-linked immunosorbent assay)., Results: Upon gastrointestinal passage, a higher amount of viable cells was found in the simulated ileal effluents of children compared to that of adults (with 34 and 6% of viable cells, respectively). Expression levels of virulence genes were up to 125-fold higher in children. Stx was detected only in child ileal effluents., Conclusion: Differences in digestive physicochemical parameters may partially explain why children are more susceptible to EHEC infection than adults. Such data are essential for a full understanding of EHEC pathogenesis and would help in designing novel therapeutic approaches.