CoMiniGut—a small volumein vitrocolon model for the screening of gut microbial fermentation processes

PeerJ ◽

10.7717/peerj.4268 ◽

2018 ◽

Vol 6 ◽

pp. e4268 ◽

Cited By ~ 19

Author(s):

Maria Wiese ◽

Bekzod Khakimov ◽

Sebastian Nielsen ◽

Helena Sørensen ◽

Frans van den Berg ◽

...

Keyword(s):

Relative Abundance ◽

High Throughput Sequencing ◽

Microbial Community Composition ◽

Short Chain Fatty Acids ◽

Rrna Gene ◽

Microbial Fermentation ◽

Data Sets ◽

Microbial Composition ◽

Working Volume

Driven by the growing recognition of the influence of the gut microbiota (GM) on human health and disease, there is a rapidly increasing interest in understanding how dietary components, pharmaceuticals and pre- and probiotics influence GM.In vitrocolon models represent an attractive tool for this purpose. With the dual objective of facilitating the investigation of rare and expensive compounds, as well as an increased throughput, we have developed a prototypein vitroparallel gut microbial fermentation screening tool with a working volume of only 5 ml consisting of five parallel reactor units that can be expanded with multiples of five to increase throughput. This allows e.g., the investigation of interpersonal variations in gut microbial dynamics and the acquisition of larger data sets with enhanced statistical inference. The functionality of thein vitrocolon model, Copenhagen MiniGut (CoMiniGut) was first demonstrated in experiments with two common prebiotics using the oligosaccharide inulin and the disaccharide lactulose at 1% (w/v). We then investigated fermentation of the scarce and expensive human milk oligosaccharides (HMOs) 3-Fucosyllactose, 3-Sialyllactose, 6-Sialyllactose and the more common Fructooligosaccharide in fermentations with infant gut microbial communities. Investigations of microbial community composition dynamics in the CoMiniGut reactors by MiSeq-based 16S rRNA gene amplicon high throughput sequencing showed excellent experimental reproducibility and allowed us to extract significant differences in gut microbial composition after 24 h of fermentation for all investigated substrates and fecal donors. Furthermore, short chain fatty acids (SCFAs) were quantified for all treatments and donors. Fermentations with inulin and lactulose showed that inulin leads to a microbiota dominated by obligate anaerobes, with high relative abundance of Bacteroidetes, while the more easily fermented lactulose leads to higher relative abundance of Proteobacteria. The subsequent study on the influence of HMOs on two infant GM communities, revealed the strongest bifidogenic effect for 3′SL for both infants. Inter-individual differences of infant GM, especially with regards to the occurrence of Bacteroidetes and differences in bifidobacterial species composition, correlated with varying degrees of HMO utilization foremost of 6′SL and 3′FL, indicating species and strain related differences in HMO utilization which was also reflected in SCFAs concentrations, with 3′SL and 6′SL resulting in significantly higher butyrate production compared to 3′FL. In conclusion, the increased throughput of CoMiniGut strengthens experimental conclusions through elimination of statistical interferences originating from low number of repetitions. Its small working volume moreover allows the investigation of rare and expensive bioactives.

Production of conjugated linoleic acid (CLA): effect of inulin on microbial composition and CLA concentration in a human intestinal model

Proceedings of The Nutrition Society ◽

10.1017/s0029665120005777 ◽

2020 ◽

Vol 79 (OCE2) ◽

Author(s):

Ilaria Carafa ◽

Domenico Masuero ◽

Urska Vrhovsek ◽

Giovanni Bittante ◽

Elena Franciosi ◽

...

Keyword(s):

Linoleic Acid ◽

Gut Microbiota ◽

Relative Abundance ◽

Illumina Miseq ◽

Ultra Performance Liquid Chromatography ◽

Rrna Gene ◽

Human Gut ◽

Microbial Composition ◽

Human Gut Microbiota

AbstractConjugated linoleic acids (CLAs) show a number of putative health-promoting activities including anti-carcinogenic, anti-adipogenic, anti-diabetogenic, anti-inflammatory and antioxidant actions. CLAs are naturally produced by ruminal bacteria and several studies demonstrate that various lactobacilli and bifidobacteria are also able to produce CLAs in vitro from linoleic acid (LA). However, the ability of the human gut microbiota to produce CLA is less extensively studied. Our hypothesis is that the human gut microbiota is able to convert LA to CLA, and that the readily fermentable fiber inulin would positively modulate the growth of CLA-producing bacteria and, consequently increase the CLA content in the intestine.The capability of the faecal microbiota from five healthy donors to produce CLA was tested in anaerobic batch cultures for 48 hours at pH 5.5 and 6.5. Test treatments were linoleic acid (LA; 1 mg/mL) + bovine serum albumin (BSA; 0.2 mg/mL), and LA (1 mg/mL) + BSA (0.2 mg/mL) + inulin (1%, w/v) compared to a control BSA (0.2 mg/mL) fermentation. The microbial composition was analyzed 0, 24 and 48 hours after starting the fermentation by 16S rRNA gene Illumina MiSeq sequencing (V3-V4 region). CLAs were quantified by Ultra performance liquid chromatography - tandem mass spectrometer (UPLC-MS/MS) and bi-dimensional gas chromatography (GC x GC).The inclusion of LA + BSA + inulin at pH 5.5 significantly increased the relative abundance of Collinsella aerofaciens (p < 0.05), and tended to increase the relative abundance of bifidobacteria. LA + BSA + inulin at both pH 5.5 and 6.5 reduced the relative abundance of Parabacteroides, Bilophila, Clostridia and Enterobacteriaceae (p < 0.05). The concentration of CLA, in particular the isomer cis9,trans11 C18:2, was significantly higher in the LA + BSA + inulin group at pH 5.5 after 24 and 48 hours fermentation.The data show that the treatment LA + BSA + inulin at pH 5.5 induce substantial changes in microbiota composition, including bifidogenesis and CLA production in a human intestinal microbiota model. The changes of relative abundance detected are consistent with changes in gut bacteria previously linked to human health. Collinsella aerofaciens has been reported for reducing bloating, in particular in subjects suffering from irritable bowel syndrome, while Clostridia, Bilophila and Enterobacteriaceae causes human infections. In addition, the increase of bifidobacteria and LAB, which have previously been shown in vitro to produce CLA, may also be involved in CLA production under simulated cecal microbiome. These preclinical observations warrant confirmation in suitably designed animal and human mechanistic studies.

Phosphorus forms in the sediment of seagrass meadows affected mainly by fungi rather than bacteria: a preliminary study based on 31P-NMR and high-throughput sequencing

Oceanological and Hydrobiological Studies ◽

10.1515/ohs-2020-0036 ◽

2020 ◽

Vol 49 (4) ◽

pp. 408-420

Author(s):

Muqiu Zhao ◽

Hui Wang ◽

Shuai Wang ◽

Qiuying Han ◽

Yunfeng Shi

Keyword(s):

High Throughput ◽

Relative Abundance ◽

Tidal Flat ◽

High Throughput Sequencing ◽

Coastal Wetland ◽

Microbial Community Composition ◽

Rrna Gene ◽

Wetland Ecosystems ◽

Seagrass Meadows ◽

P Cycling

AbstractMicroorganisms play an important role in the circulation of phosphorus (P) in the sediment of coastal wetland ecosystems. In this study, solution 31P nuclear magnetic resonance (NMR) was used to determine different forms of P in the sediments of four different seagrass meadows and a bare tidal flat, while high-throughput 16S and ITS rRNA gene sequencing was used to determine the microbial community composition. The solution 31P-NMR spectra revealed six forms of the P compounds detected by the NaOH-EDTA extraction of sediments, where Ortho-P was the most dominant P compound, followed by Mono-P. The Po compounds were more varied in the seagrass meadow sediments and more abundant compared to the bare tidal flat. Bacterial communities in the sediments collected from E. acoroides and fungal communities in the bare tidal flat were relatively different from those at the other sites. The relative abundance of P-cycling-related fungi belonging to the phylum Ascomycota was 26.20% and was much higher than that of bacteria (only 0.29%) belonging to the class Bacilli. Mono-P was the major factor determining the distribution of P-cycling-related fungi and negatively correlated with the relative abundance of Aspergillus and Trichoderma. We believe that fungi can affect P forms in the sediment of seagrass meadows more than bacteria.

Xylitol enhances synthesis of propionate in the colon via cross-feeding of gut microbiota

Microbiome ◽

10.1186/s40168-021-01029-6 ◽

2021 ◽

Vol 9 (1) ◽

Author(s):

Shasha Xiang ◽

Kun Ye ◽

Mian Li ◽

Jian Ying ◽

Huanhuan Wang ◽

...

Keyword(s):

Gut Microbiome ◽

Lactobacillus Reuteri ◽

Carbon Sources ◽

Short Chain Fatty Acids ◽

Microbial Composition ◽

Key Enzymes ◽

Rrna Sequencing ◽

Phosphate Acetyltransferase

Abstract Background Xylitol, a white or transparent polyol or sugar alcohol, is digestible by colonic microorganisms and promotes the proliferation of beneficial bacteria and the production of short-chain fatty acids (SCFAs), but the mechanism underlying these effects remains unknown. We studied mice fed with 0%, 2% (2.17 g/kg/day), or 5% (5.42 g/kg/day) (weight/weight) xylitol in their chow for 3 months. In addition to the in vivo digestion experiments in mice, 3% (weight/volume) (0.27 g/kg/day for a human being) xylitol was added to a colon simulation system (CDMN) for 7 days. We performed 16S rRNA sequencing, beneficial metabolism biomarker quantification, metabolome, and metatranscriptome analyses to investigate the prebiotic mechanism of xylitol. The representative bacteria related to xylitol digestion were selected for single cultivation and co-culture of two and three bacteria to explore the microbial digestion and utilization of xylitol in media with glucose, xylitol, mixed carbon sources, or no-carbon sources. Besides, the mechanisms underlying the shift in the microbial composition and SCFAs were explored in molecular contexts. Results In both in vivo and in vitro experiments, we found that xylitol did not significantly influence the structure of the gut microbiome. However, it increased all SCFAs, especially propionate in the lumen and butyrate in the mucosa, with a shift in its corresponding bacteria in vitro. Cross-feeding, a relationship in which one organism consumes metabolites excreted by the other, was observed among Lactobacillus reuteri, Bacteroides fragilis, and Escherichia coli in the utilization of xylitol. At the molecular level, we revealed that xylitol dehydrogenase (EC 1.1.1.14), xylulokinase (EC 2.7.1.17), and xylulose phosphate isomerase (EC 5.1.3.1) were key enzymes in xylitol metabolism and were present in Bacteroides and Lachnospiraceae. Therefore, they are considered keystone bacteria in xylitol digestion. Also, xylitol affected the metabolic pathway of propionate, significantly promoting the transcription of phosphate acetyltransferase (EC 2.3.1.8) in Bifidobacterium and increasing the production of propionate. Conclusions Our results revealed that those key enzymes for xylitol digestion from different bacteria can together support the growth of micro-ecology, but they also enhanced the concentration of propionate, which lowered pH to restrict relative amounts of Escherichia and Staphylococcus. Based on the cross-feeding and competition among those bacteria, xylitol can dynamically balance proportions of the gut microbiome to promote enzymes related to xylitol metabolism and SCFAs.

Disentangling Responses of the Subsurface Microbiome to Wetland Status and Implications for Indicating Ecosystem Functions

Microorganisms ◽

10.3390/microorganisms9020211 ◽

2021 ◽

Vol 9 (2) ◽

pp. 211

Author(s):

Jie Gao ◽

Miao Liu ◽

Sixue Shi ◽

Ying Liu ◽

Yu Duan ◽

...

Keyword(s):

Microbial Community ◽

High Throughput Sequencing ◽

Carbon Fixation ◽

Microbial Community Composition ◽

Ecosystem Functions ◽

Microbial Composition ◽

Wetland Ecosystems ◽

Soil Microbial ◽

Geochemical Properties ◽

Microbial Groups

In this study, we analyzed microbial community composition and the functional capacities of degraded sites and restored/natural sites in two typical wetlands of Northeast China—the Phragmites marsh and the Carex marsh, respectively. The degradation of these wetlands, caused by grazing or land drainage for irrigation, alters microbial community components and functional structures, in addition to changing the aboveground vegetation and soil geochemical properties. Bacterial and fungal diversity at the degraded sites were significantly lower than those at restored/natural sites, indicating that soil microbial groups were sensitive to disturbances in wetland ecosystems. Further, a combined analysis using high-throughput sequencing and GeoChip arrays showed that the abundance of carbon fixation and degradation, and ~95% genes involved in nitrogen cycling were increased in abundance at grazed Phragmites sites, likely due to the stimulating impact of urine and dung deposition. In contrast, the abundance of genes involved in methane cycling was significantly increased in restored wetlands. Particularly, we found that microbial composition and activity gradually shifts according to the hierarchical marsh sites. Altogether, this study demonstrated that microbial communities as a whole could respond to wetland changes and revealed the functional potential of microbes in regulating biogeochemical cycles.

Bioconductor workflow for microbiome data analysis: from raw reads to community analyses

F1000Research ◽

10.12688/f1000research.8986.1 ◽

2016 ◽

Vol 5 ◽

pp. 1492 ◽

Cited By ~ 148

Author(s):

Ben J. Callahan ◽

Kris Sankaran ◽

Julia A. Fukuyama ◽

Paul J. McMurdie ◽

Susan P. Holmes

Keyword(s):

High Throughput Sequencing ◽

Linear Models ◽

Reference Database ◽

Rrna Gene ◽

Microbial Composition ◽

Nonparametric Testing ◽

Abundance Estimates ◽

Taxonomic Markers ◽

Microbiome Data ◽

Microbiome Data Analysis

High-throughput sequencing of PCR-amplified taxonomic markers (like the 16S rRNA gene) has enabled a new level of analysis of complex bacterial communities known as microbiomes. Many tools exist to quantify and compare abundance levels or microbial composition of communities in different conditions. The sequencing reads have to be denoised and assigned to the closest taxa from a reference database. Common approaches use a notion of 97% similarity and normalize the data by subsampling to equalize library sizes. In this paper, we show that statistical models allow more accurate abundance estimates. By providing a complete workflow in R, we enable the user to do sophisticated downstream statistical analyses, including both parameteric and nonparametric methods. We provide examples of using the R packages dada2, phyloseq, DESeq2, ggplot2 and vegan to filter, visualize and test microbiome data. We also provide examples of supervised analyses using random forests, partial least squares and linear models as well as nonparametric testing using community networks and the ggnetwork package.

Comparative Analysis of the Gut Microbiota of Adult Mosquitoes From Eight Locations in Hainan, China

Frontiers in Cellular and Infection Microbiology ◽

10.3389/fcimb.2020.596750 ◽

2020 ◽

Vol 10 ◽

Author(s):

Xun Kang ◽

Yanhong Wang ◽

Siping Li ◽

Xiaomei Sun ◽

Xiangyang Lu ◽

...

Keyword(s):

Gut Microbiota ◽

Disease Control ◽

Bacterial Communities ◽

Mosquito Species ◽

Microbial Community Composition ◽

Variable Region ◽

Rrna Gene ◽

Microbial Composition ◽

Quality Filtering ◽

And Function

The midgut microbial community composition, structure, and function of field-collected mosquitoes may provide a way to exploit microbial function for mosquito-borne disease control. However, it is unclear how adult mosquitoes acquire their microbiome, how the microbiome affects life history traits and how the microbiome influences community structure. We analyzed the composition of 501 midgut bacterial communities from field-collected adult female mosquitoes, including Aedes albopictus, Aedes galloisi, Culex pallidothorax, Culex pipiens, Culex gelidus, and Armigeres subalbatus, across eight habitats using the HiSeq 4000 system and the V3−V4 hyper-variable region of 16S rRNA gene. After quality filtering and rarefaction, a total of 1421 operational taxonomic units, belonging to 29 phyla, 44 families, and 43 genera were identified. Proteobacteria (75.67%) were the most common phylum, followed by Firmicutes (10.38%), Bacteroidetes (6.87%), Thermi (4.60%), and Actinobacteria (1.58%). The genera Rickettsiaceae (33.00%), Enterobacteriaceae (20.27%), Enterococcaceae (7.49%), Aeromonadaceae (7.00%), Thermaceae (4.52%), and Moraxellaceae (4.31%) were dominant in the samples analyzed and accounted for 76.59% of the total genera. We characterized the midgut bacterial communities of six mosquito species in Hainan province, China. The gut bacterial communities were different in composition and abundance, among locations, for all mosquito species. There were significant differences in the gut microbial composition between some species and substantial variation in the gut microbiota between individuals of the same mosquito species. There was a marked variation in different mosquito gut microbiota within the same location. These results might be useful in the identification of microbial communities that could be exploited for disease control.

Probiotic Supplementation in a Clostridium difficile-Infected Gastrointestinal Model Is Associated with Restoring Metabolic Function of Microbiota

Microorganisms ◽

10.3390/microorganisms8010060 ◽

2019 ◽

Vol 8 (1) ◽

pp. 60

Author(s):

Mohd Baasir Gaisawat ◽

Chad W. MacPherson ◽

Julien Tremblay ◽

Amanda Piano ◽

Michèle M. Iskandar ◽

...

Keyword(s):

Gut Microbiota ◽

Alpha Diversity ◽

Gastrointestinal Disorder ◽

Short Chain Fatty Acids ◽

Rrna Gene ◽

Metabolic Function ◽

Microbial Composition ◽

Single Strain ◽

Fecal Samples ◽

Metabolic Functions

Clostridium (C.) difficile-infection (CDI), a nosocomial gastrointestinal disorder, is of growing concern due to its rapid rise in recent years. Antibiotic therapy of CDI is associated with disrupted metabolic function and altered gut microbiota. The use of probiotics as an adjunct is being studied extensively due to their potential to modulate metabolic functions and the gut microbiota. In the present study, we assessed the ability of several single strain probiotics and a probiotic mixture to change the metabolic functions of normal and C. difficile-infected fecal samples. The production of short-chain fatty acids (SCFAs), hydrogen sulfide (H2S), and ammonia was measured, and changes in microbial composition were assessed by 16S rRNA gene amplicon sequencing. The C. difficile-infection in fecal samples resulted in a significant decrease (p < 0.05) in SCFA and H2S production, with a lower microbial alpha diversity. All probiotic treatments were associated with significantly increased (p < 0.05) levels of SCFAs and restored H2S levels. Probiotics showed no effect on microbial composition of either normal or C. difficile-infected fecal samples. These findings indicate that probiotics may be useful to improve the metabolic dysregulation associated with C. difficile infection.

Diet Breadth Affects Bacterial Identity but Not Diversity in the Pollen Provisions of Closely Related Polylectic and Oligolectic Bees

Insects ◽

10.3390/insects11090645 ◽

2020 ◽

Vol 11 (9) ◽

pp. 645

Author(s):

Jason A. Rothman ◽

Diana L. Cox-Foster ◽

Corey Andrikopoulos ◽

Quinn S. McFrederick

Keyword(s):

Host Species ◽

High Throughput Sequencing ◽

Microbial Community Composition ◽

Diet Breadth ◽

Solitary Bees ◽

Rrna Gene ◽

Osmia Lignaria ◽

Future Studies ◽

Functional Consequences ◽

Oligolectic Bees

Mounting evidence suggests that microbes found in the pollen provisions of wild and solitary bees are important drivers of larval development. As these microbes are also known to be transmitted via the environment, most likely from flowers, the diet breadth of a bee may affect the diversity and identity of the microbes that occur in its pollen provisions. Here, we tested the hypothesis that, due to the importance of floral transmission of microbes, diet breadth affects pollen provision microbial community composition. We collected pollen provisions at four sites from the polylectic bee Osmia lignaria and the oligolectic bee Osmia ribifloris. We used high-throughput sequencing of the bacterial 16S rRNA gene to characterize the bacteria found in these provisions. We found minimal overlap in the specific bacterial variants in pollen provisions across the host species, even when the bees were constrained to foraging from the same flowers in cages at one site. Similarly, there was minimal overlap in the specific bacterial variants across sites, even within the same host species. Together, these findings highlight the importance of environmental transmission and host specific sorting influenced by diet breadth for microbes found in pollen provisions. Future studies addressing the functional consequences of this filtering, along with tests for differences between more species of oligoletic and polylectic bees will provide rich insights into the microbial ecology of solitary bees.

In Vitro Fecal Fermentation Patterns of Arabinoxylan from Rice Bran on Gut Microbiota in Normal Weight and Overweight/Obese Subjects

Current Developments in Nutrition ◽

10.1093/cdn/nzaa062_017 ◽

2020 ◽

Vol 4 (Supplement_2) ◽

pp. 1560-1560

Author(s):

Inah Gu ◽

Wing Shun Lam ◽

Daya Marasini ◽

Cindi Brownmiller ◽

Brett Savary ◽

...

Keyword(s):

Gut Microbiota ◽

Rice Bran ◽

Block Design ◽

16S Rrna Gene Sequencing ◽

Short Chain Fatty Acids ◽

Normal Weight ◽

Rrna Gene ◽

Gut Health ◽

The Impact

Abstract Objectives Arabinoxylan is a non-starch polysaccharide and rich in wheat, rice and many other cereal grains. Diets high in fiber help promoting gut health in obesity. The objective of this study was to investigate the impact of arabinoxylan from rice bran on the gut microbiota and short chain fatty acids (SCFA) in normal weight (NW) and overweight/obese (OO) subjects through in vitro fecal fermentation. Methods Arabinoxylan was extracted from rice bran fiber. For in vitro fecal fermentation, each fecal sample from NW (n = 6, 3 males and 3 females) and OO (n = 7, 3 males and 4 females) was diluted into anaerobic medium with three treatments: control (no substrates), fructooligosaccharides (FOS, a well-known prebiotic), and arabinoxylan. Samples were incubated at 37˚C and aliquots were taken at 0, 4, 8, 12 and 24 h. SCFA content from samples at all timepoints was analyzed using HPLC. Samples at 0 and 24 h were used for gut microbiota analysis through 16S rRNA gene sequencing. Statistical analyses were performed for the randomized complete block design, where the weight classes are confounded with blocks (subjects). Friedman test was used to determine the difference at 5% level of significance. Results As a result, arabinoxylan treatment significantly increased total SCFA concentration in both NW and OO subjects than control (P < 0.05), comparable to FOS treatment. Between weight classes under arabinoxylan treatment, OO group showed a significantly higher total SCFA content than NW group (P < 0.05). Arabinoxylan changed gut microbial population at the genus level, stimulating Bifidobacterium, Collinsella and Blautia and decreasing Clostridium XIVa and b, Dorea and Oscillibacter (P < 0.05). In addition, different microbiome population was shown in weight classes with three treatments, showing higher Bacteroides in NW and higher Prevotella in OO. Conclusions These results showed that arabinoxylan from rice bran modified gut microbiota in both weight classes, increasing total SCFA content. This study suggests that arabinoxylan from rice bran may have a potential impact on microbial gut health in obesity with prebiotic activities. Funding Sources University of Arkansas.

Assessing the Effects of Ginger Extract on Polyphenol Profiles and the Subsequent Impact on the Fecal Microbiota by Simulating Digestion and Fermentation In Vitro

Nutrients ◽

10.3390/nu12103194 ◽

2020 ◽

Vol 12 (10) ◽

pp. 3194

Author(s):

Jing Wang ◽

Yong Chen ◽

Xiaosong Hu ◽

Fengqin Feng ◽

Luyun Cai ◽

...

Keyword(s):

Gut Microbiota ◽

Ph Value ◽

Illumina Miseq ◽

Short Chain Fatty Acids ◽

Fecal Microbiota ◽

Rrna Gene ◽

Gut Health ◽

Ginger Extract ◽

Mixed Culture Fermentation

The beneficial effects of ginger polyphenols have been extensively reported. However, their metabolic characteristics and health effects on gut microbiota are poor understood. The purpose of this study was to investigate the digestion stability of ginger polyphenols and their prebiotic effects on gut microbiota by simulating digestion and fermentation in vitro. Following simulated digestion in vitro, 85% of the polyphenols were still detectable, and the main polyphenol constituents identified in ginger extract are 6-, 8-, and 10-gingerols and 6-shogaol in the digestive fluids. After batch fermentation, the changes in microbial populations were measured by 16S rRNA gene Illumina MiSeq sequencing. In mixed-culture fermentation with fecal inoculate, digested ginger extract (GE) significantly modulated the fecal microbiota structure and promoted the growth of some beneficial bacterial populations, such as Bifidobacterium and Enterococcus. Furthermore, incubation with GE could elevate the levels of short-chain fatty acids (SCFAs) accompanied by a decrease in the pH value. Additionally, the quantitative PCR results showed that 6-gingerol (6G), as the main polyphenol in GE, increased the abundance of Bifidobacterium significantly. Therefore, 6G is expected to be a potential prebiotic that improves human health by promoting gut health.