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Hydrodynamic chronoamperometry for probing kinetics of anaerobic microbial metabolism--case study of Faecalibacterium prausnitzii.

ABSTRACT: Monitoring in vitro the metabolic activity of microorganisms aids bioprocesses and enables better understanding of microbial metabolism. Redox mediators can be used for this purpose via different electrochemical techniques that are either complex or only provide non-continuous data. Hydrodynamic chronoamperometry using a rotating disc electrode (RDE) can alleviate these issues but was seldom used and is poorly characterized. The kinetics of Faecalibacterium prausnitzii A2-165, a beneficial gut microbe, were determined using a RDE with riboflavin as redox probe. This butyrate producer anaerobically ferments glucose and reduces riboflavin whose continuous monitoring on a RDE provided highly accurate kinetic measurements of its metabolism, even at low cell densities. The metabolic reaction rate increased linearly over a broad range of cell concentrations (9 × 10(4) to 5 × 10(7) cells.mL(-1)). Apparent Michaelis-Menten kinetics was observed with respect to riboflavin (KM = 6 μM; kcat = 5.3 × 10(5) s(-1), at 37 °C) and glucose (KM = 6 μM; kcat = 2.4 × 10(5) s(-1)). The short temporal resolution allows continuous monitoring of fast cellular events such as kinetics inhibition with butyrate. Furthermore, we detected for the first time riboflavin reduction by another potential probiotic, Butyricicoccus pullicaecorum. The ability of the RDE for fast, accurate, simple and continuous measurements makes it an ad hoc tool for assessing bioprocesses at high resolution.

SUBMITTER: Prevoteau A

PROVIDER: S-EPMC4486957 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Dietary intervention is effective in human health promotion through modulation of gut microbiota. Diet can cause single-nucleotide polymorphisms (SNPs) to occur in the gut microbiota, and some of these variations may lead to functional changes in human health. In this study, we performed a systematic SNP analysis based on metagenomic data collected from children with Prader-Willi syndrome (PWS, n = 17) and simple obese (SO) children (n = 19), who had better healthy conditions after receiving high-fiber diet intervention. We found that the intervention increased the SNP proportions of Faecalibacterium, Bifidobacterium, and Clostridium and decreased those of Bacteroides in all children. Besides, the PWS children had Collinsella increased and Ruminococcus decreased, whereas the SO had Blautia and Escherichia decreased. There were much more BiasSNPs in PWS than in SO (4,465 vs 303), and only 81 of them appeared in both groups, of which 78 were from Faecalibacterium prausnitzii, and 51 were nonsynonymous mutations. These nonsynonymous variations were mainly related to pathways of environmental adaptation and nutrition metabolism, particularly to carbohydrate and nucleotide metabolism. In addition, dominant strains carrying BiasSNPs in all children shifted from F. prausnitzii AF32-8AC and F. prausnitzii 942/30-2 to F. prausnitzii SSTS Bg7063 and F. prausnitzii JG BgPS064 after the dietary intervention. Furthermore, although the abundance of Bifidobacterium increased significantly by the intervention and became dominant strains responsible for nutrition metabolism, they had less BiasSNPs between the pre- and post-intervention group in comparison with Faecalibacterium. The finding of F. prausnitzii as important functional strains influenced by the intervention highlights the superiority of applying SNP analysis in studies of gut microbiota. This study provided evidence and support for the effect of dietary intervention on gut microbial SNPs, and gave some enlightenments for disease treatment.

Project description:The goal of our study was to isolate and characterize Faecalibacterium prausnitzii from fecal samples of healthy calves and piglets, in order to develop a novel probiotic for livestock animals. We identified 203 isolates of Faecalibacterium sp., which were clustered in 40 genetically distinct groups. One representative isolate from each cluster was selected for further characterization. The concentrations of the short chain fatty acids (SCFA) acetate, butyrate, propionate and isobutyrate in the culture media were measured by gas chromatography. We observed reduction in the concentration of acetate followed by concomitant increase in the concentration of butyrate, suggesting that the isolates were consuming acetate present in the media and producing butyrate. Butyrate production correlated positively with bacterial growth. Since butyrate has many benefits to the colonic epithelial cells, the selection of strains that produce higher amounts of butyrate is extremely important for the development of this potential probiotic. The effect of pH and concentration of bile salts on bacterial growth was also evaluated in order to mimic the conditions encountered by F. prausnitzii in vivo. The optimal pH for growth ranged between 5.5 and 6.7, while most isolates were inhibited by of the lowest concentration of bile salts tested (0.1%). Antimicrobial resistance profile showed that most isolates of Faecalibacterium sp. were resistant against ciprofloxacin and sulfamethoxazole-trimethoprim. More than 50% of the isolates were resistant to tetracycline, amikacin, cefepime and cefoxitin. A total of 19 different combinations of multidrug resistance were observed among the isolates. Our results provide new insights into the cultural and physiological characteristics of Faecalibacterium prausnitzii illustrating large variability in short chain fatty acid production, in vitro growth, sensitivity to bile salts, and antibiotic resistance and suggesting that future probiotic candidates should be carefully studied before elected for in vivo studies.

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Hydrodynamic chronoamperometry for probing kinetics of anaerobic microbial metabolism--case study of Faecalibacterium prausnitzii.

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