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Consumption of fermentable dietary fibers (DFs), which can induce growth and/or activity of specific beneficial populations, is suggested a promising strategy to modulate the gut microbiota and restore health in microbiota-linked diseases. Until today, inulin and fructo-oligosaccharides (FOS) are the best studied DFs, while little is known about the gut microbiota-modulating effects of β-glucan, α-galactooligosaccharide (α-GOS) and xylo-oligosaccharide (XOS). Here, we used three continuous in vitro fermentation PolyFermS model to study the modulating effect of these DFs on two distinct human adult proximal colon microbiota, independently from the host. Supplementation of DFs, equivalent to a 9 g daily intake, induced a consistent metabolic response depending on the donor microbiota. Irrespective to the DF supplemented, the Bacteroidaceae-Ruminococcaceae dominated microbiota produced more butyrate (up to 96%), while the Prevotellaceae-Ruminococcaceae dominated microbiota produced more propionate (up to 40%). Changes in abundance of specific bacterial taxa upon DF supplementation explained the observed changes in short-chain fatty acid profiles. Our data suggest that the metabolic profile of SCFA profile may be the most suitable and robust read-out to characterize microbiota-modulating effects of a DF and highlights importance to understand the inter-individual response to a prebiotic treatment for mechanistic understanding and human application.

Background Administering probiotic strains of Limosilactobacillus reuteri to poultry has been shown to improve poultry performance and health. Some strains of L. reuteri taxa can produce reuterin, a broad-spectrum antimicrobial compound from glycerol conversion, with high inhibitory activity against enterobacteria. However, little is known about the metabolism of glycerol in the complex chicken cecal microbiota nor the effect of glycerol, either alone or combined with L. reuteri on the microbiota. In this study, we investigated the effect of L. reuteri PTA5_F13, a high-reuterin-producing chicken strain and glycerol, alone or combined, on broiler chicken cecal microbiota composition and activity using the continuous PolyFermS model recently developed to mimic chicken cecal fermentation. Methods Three independent PolyFermS chicken cecal microbiota models were inoculated with immobilized cecal microbiota from different animals and operated continuously. The effects of two additional levels of glycerol (50 and 100 mM) with or without daily supplementation of chicken-derived L. reuteri PTA5_F13 (10 7  CFU/mL final concentration) were tested in parallel second-stage reactors continuously inoculated with the same microbiota. We analyzed the complex chicken gut microbiota structure and dynamics upon treatment using 16S rRNA metabarcoding and qPCR. Microbiota metabolites, short-chain and branched-chain fatty acids, and glycerol and reuterin products were analyzed by HPLC in effluent samples from stabilized reactors. Results Supplementation with 100 mM glycerol alone and combined with L. reuteri PTA5_F13 resulted in a reproducible increase in butyrate production in the three modelled microbiota (increases of 18 to 25%). Glycerol alone resulted also in a reduction of Enterobacteriaceae in two of the three microbiota, but no effect was detected for L. reuteri alone. When both treatments were combined, all microbiota quantitatively inhibited Enterobacteriaceae , including in the last model that had very high initial concentrations of Enterobacteriaceae . Furthermore, a significant 1,3-PDO accumulation was measured in the effluent of the combined treatment, confirming the conversion of glycerol via the reuterin pathway. Glycerol supplementation, independent of L. reuteri addition, did not affect the microbial community diversity. Conclusions Glycerol induced a stable and reproducible butyrogenic activity for all tested microbiota and induced an inhibitory effect against Enterobacteriaceae that was strengthened when reuterin-producing L. reuteri was spiked daily. Our in vitro study suggests that co-application of L. reuteri PTA5_F13 and glycerol could be a useful approach to promote chicken gut health by enhancing metabolism and protection against Enterobacteriaceae .

Background Folate (vitamin B9) occurs naturally mainly as tetrahydrofolate (THF), methyl-tetrahydrofolate (M-THF), and formyl-tetrahydrofolate (F-THF), and as dietary synthetic form (folic acid). While folate auxotrophy and prototrophy are known for several gut microbes, the specific folate forms produced by gut prototrophs and their impact on gut auxotrophs and microbiota remain unexplored. Methods Here, we quantified by UHPLC-FL/UV folate produced by six predicted gut prototrophs ( Marvinbryantia formatexigens DSM 14469 , Blautia hydrogenotrophica 10507  T , Blautia producta DSM 14466, Bacteroides caccae DSM 19024, Bacteroides ovatus DSM 1896 , and Bacteroides thetaiotaomicron DSM 2079  T ) and investigated the impact of different folate forms and doses (50 and 200 µg/l) on the growth and metabolism of the gut auxotroph Roseburia intestinalis in pure cultures and during fecal anaerobic batch fermentations (48 h, 37 °C) of five healthy adults. Results Our results confirmed the production of folate by all six gut strains, in the range from 15.3 ng/ml to 205.4 ng/ml. Different folate forms were detected, with THF ranging from 12.8 to 41.4 ng/ml and 5-MTHF ranging from 0.2 to 113.3 ng/ml, and being detected in all strains. Natural folate forms, in contrast to folic acid, promoted the growth and metabolism of the auxotroph R. intestinalis L1-82, with dose-dependent effects. During fecal batch fermentations, folate forms at both levels had no detectable effect on total bacteria concentration, on gut community composition and metabolic activity and on Roseburia spp. abundance, compared to the control without folate addition. Conclusions Our study demonstrates for the first time in vitro the production of different natural folate forms by predicted gut prototrophs and the stimulation on the growth of the folate auxotrophic butyrate-producing R. intestinalis L1-82. Surprisingly, folate did not impact fecal fermentations. Our data suggest that the dietary folate forms at the tested levels may only have limited effects, if any, on the human gut microbiota in vivo.

Reuterin is a broad-spectrum antimicrobial system produced by specific strains of Lactobacillus reuteri during anaerobic metabolism of glycerol. Acrolein is the main component responsible for its antimicrobial activity. Here, the sensitivity of Campylobacter jejuni (n = 51) and Campylobacter coli (n = 20) isolates from chicken meat and human stool samples to reuterin was investigated. The minimum inhibitory concentration (MIC) of C. jejuni and C. coli strains was measured between 1.5 and 3.0 µM of acrolein, below the MIC of the sensitive indicator strain Escherichia coli K12 (16.5 µM acrolein). The interaction of C. jejuni N16-1419 and the reuterin-producing L. reuteri PTA5_F13 was studied during 24 h co-cultures with or without glycerol. A high C. jejuni growth was observed in cultures without glycerol. In contrast, C. jejuni growth decreased from 7.3 ± 0.1 log CFU/mL to below detection limit (1 log CFU/mL) during co-cultures added with 28 mM glycerol. This bactericidal effect could be attributed to in situ reuterin production. The low MIC observed and the high sensitivity towards in situ produced reuterin suggests L. reuteri combined with glycerol, as a possible intervention option to reduce Campylobacter in the food chain.

The Sant'Elia nursery school in Como, designed by Giuseppe Terragni (1934-1937), is a landmark in 20th-century architecture and restoration. Its long-standing educational use frames the building's ‘use value’ in the discipline of modern heritage conservation. Through a critical reading of the construction documents and subsequent interventions – a ‘rehabilitation’ in the 1960s, ‘philological restoration’ in the 1980s, modern restoration techniques in 1998-2001 – the paper highlights how each intervention is a compromise between the cultural context in which it is carried out, the choices suggested by disciplinary developments, and the materiality of the building, constituting a representative case for modern school buildings.

The Sant'Elia nursery school in Como, designed by Giuseppe Terragni (1934-1937), is a landmark in 20th-century architecture and restoration. Its long-standing educational use frames the building's ‘use value’ in the discipline of modern heritage conservation. Through a critical reading of the construction documents and subsequent interventions – a ‘rehabilitation’ in the 1960s, ‘philological restoration’ in the 1980s, modern restoration techniques in 1998-2001 – the paper highlights how each intervention is a compromise between the cultural context in which it is carried out, the choices suggested by disciplinary developments, and the materiality of the building, constituting a representative case for modern school buildings.

Fecal microbiota transplantation has been successfully applied in the treatment of recurrent Clostridium difficile infection and has been suggested as an alternative therapy for other intestinal disorders such as inflammatory bowel disease or metabolic syndrome. “Artificial” colonic microbiota delivered by PolyFermS continuous fermentation models can provide a controllable and reproducible alternative to fecal transplantation, but effective preservation strategies must be developed. In this study, we systematically investigated the response of sessile and planktonic artificial colonic microbiota to cryopreservation and lyophilization. We suggest that functional redundancy is an important factor in providing functional stability with respect to exposure to stress during processing and storage. Functional redundancy in compositionally reduced microbial systems may be considered when designing microbial products for therapy. Biofilm-associated, sessile communities represent the major bacterial lifestyle, whereas planktonic cells mainly appear during initial colonization of new surfaces. Previous research, mainly performed with pathogens, demonstrated increased environmental stress tolerance of biofilm-growing compared to planktonic bacteria. The lifestyle-specific stress response of colonic microbiota, both natural and fermentation produced, has not been addressed before. Planktonic and sessile “artificial” colonic microbiota delivered by PolyFermS continuous fermentation models can provide a controllable and reproducible alternative to fecal transplantation in treating gastrointestinal disorders. We therefore characterized planktonic and sessile microbiota produced in two PolyFermS models inoculated with immobilized fecal microbiota and comparatively tested their levels of tolerance of frozen storage (–80°C) and freeze-dried storage (4°C) for 9 months to mimic preservation strategies for therapeutic applications. Sessile microbiota harbored next to shared taxa a unique community distinguishable from planktonic microbiota. Synergistetes and Proteobacteria were highly represented in sessile microbiota, while Firmicutes were more abundant in planktonic microbiota. The community structure and metabolic activity of both microbiota, monitored during standardized reactivation batch fermentations, were better preserved after frozen storage than dried storage, indicated by higher Bray-Curtis similarity and enhanced recovery of metabolite production. For both lifestyles, reestablishment of Bacteroidaceae was impaired after frozen and dried storage along with reduced propionate formation. In contrast, butyrate production was maintained after reactivation despite compositional rearrangements within the butyrate-producing community. Unexpectedly, the rate of recovery of metabolite production was lower after preservation of sessile than planktonic microbiota. We speculate that higher functional dependencies between microbes might have led to the lower stress tolerance of sessile than planktonic microbiota. IMPORTANCE Fecal microbiota transplantation has been successfully applied in the treatment of recurrent Clostridium difficile infection and has been suggested as an alternative therapy for other intestinal disorders such as inflammatory bowel disease or metabolic syndrome. “Artificial” colonic microbiota delivered by PolyFermS continuous fermentation models can provide a controllable and reproducible alternative to fecal transplantation, but effective preservation strategies must be developed. In this study, we systematically investigated the response of sessile and planktonic artificial colonic microbiota to cryopreservation and lyophilization. We suggest that functional redundancy is an important factor in providing functional stability with respect to exposure to stress during processing and storage. Functional redundancy in compositionally reduced microbial systems may be considered when designing microbial products for therapy.

The Sant'Elia nursery school in Como, designed by Giuseppe Terragni (1934-1937), is a landmark in 20th-century architecture and restoration. Its long-standing educational use frames the building's ‘use value’ in the discipline of modern heritage conservation. Through a critical reading of the construction documents and subsequent interventions – a ‘rehabilitation’ in the 1960s, ‘philological restoration’ in the 1980s, modern restoration techniques in 1998-2001 – the paper highlights how each intervention is a compromise between the cultural context in which it is carried out, the choices suggested by disciplinary developments, and the materiality of the building, constituting a representative case for modern school buildings.

The butyrogenic capability of Lactobacillus ( L .) plantarum is highly dependent on the substrate type and so far not assigned to any specific metabolic pathway. Accordingly, we compared three genomes of L. plantarum that showed a strain-specific capability to produce butyric acid in human cells growth media. Based on the genomic analysis, butyric acid production was attributed to the complementary activities of a medium-chain thioesterase and the fatty acid synthase of type two (FASII). However, the genomic islands of discrepancy observed between butyrogenic L. plantarum strains (S2T10D, S11T3E) and the non-butyrogenic strain O2T60C do not encompass genes of FASII, but several cassettes of genes related to sugar metabolism, bacteriocins, prophages and surface proteins. Interestingly, single amino acid substitutions predicted from SNPs analysis have highlighted deleterious mutations in key genes of glutamine metabolism in L. plantarum O2T60C, which corroborated well with the metabolic deficiency suffered by O2T60C in high-glutamine growth media and its consequent incapability to produce butyrate. In parallel, the increase of glutamine content induced the production of butyric acid by L. plantarum S2T10D. The present study reveals a previously undescribed metabolic route for butyric acid production in L. plantarum , and a potential involvement of the glutamine uptake in its regulation.

ABSTRACT Biofilm-associated, sessile communities represent the major bacterial lifestyle, whereas planktonic cells mainly appear during initial colonization of new surfaces. Previous research, mainly performed with pathogens, demonstrated increased environmental stress tolerance of biofilm-growing compared to planktonic bacteria. The lifestyle-specific stress response of colonic microbiota, both natural and fermentation produced, has not been addressed before. Planktonic and sessile “artificial” colonic microbiota delivered by PolyFermS continuous fermentation models can provide a controllable and reproducible alternative to fecal transplantation in treating gastrointestinal disorders. We therefore characterized planktonic and sessile microbiota produced in two PolyFermS models inoculated with immobilized fecal microbiota and comparatively tested their levels of tolerance of frozen storage (–80°C) and freeze-dried storage (4°C) for 9 months to mimic preservation strategies for therapeutic applications. Sessile microbiota harbored next to shared taxa a unique community distinguishable from planktonic microbiota. Synergistetes and Proteobacteria were highly represented in sessile microbiota, while Firmicutes were more abundant in planktonic microbiota. The community structure and metabolic activity of both microbiota, monitored during standardized reactivation batch fermentations, were better preserved after frozen storage than dried storage, indicated by higher Bray-Curtis similarity and enhanced recovery of metabolite production. For both lifestyles, reestablishment of Bacteroidaceae was impaired after frozen and dried storage along with reduced propionate formation. In contrast, butyrate production was maintained after reactivation despite compositional rearrangements within the butyrate-producing community. Unexpectedly, the rate of recovery of metabolite production was lower after preservation of sessile than planktonic microbiota. We speculate that higher functional dependencies between microbes might have led to the lower stress tolerance of sessile than planktonic microbiota. IMPORTANCE Fecal microbiota transplantation has been successfully applied in the treatment of recurrent Clostridium difficile infection and has been suggested as an alternative therapy for other intestinal disorders such as inflammatory bowel disease or metabolic syndrome. “Artificial” colonic microbiota delivered by PolyFermS continuous fermentation models can provide a controllable and reproducible alternative to fecal transplantation, but effective preservation strategies must be developed. In this study, we systematically investigated the response of sessile and planktonic artificial colonic microbiota to cryopreservation and lyophilization. We suggest that functional redundancy is an important factor in providing functional stability with respect to exposure to stress during processing and storage. Functional redundancy in compositionally reduced microbial systems may be considered when designing microbial products for therapy.