Shunsuke Takahashi

Bio: Shunsuke Takahashi is an academic researcher from Kyoto Prefectural University. The author has contributed to research in topics: Microbiome & Lactobacillus plantarum. The author has an hindex of 8, co-authored 9 publications receiving 881 citations.

Development of a prokaryotic universal primer for simultaneous analysis of Bacteria and Archaea using next-generation sequencing.

Abstract: For the analysis of microbial community structure based on 16S rDNA sequence diversity, sensitive and robust PCR amplification of 16S rDNA is a critical step. To obtain accurate microbial composition data, PCR amplification must be free of bias; however, amplifying all 16S rDNA species with equal efficiency from a sample containing a large variety of microorganisms remains challenging. Here, we designed a universal primer based on the V3-V4 hypervariable region of prokaryotic 16S rDNA for the simultaneous detection of Bacteria and Archaea in fecal samples from crossbred pigs (Landrace×Large white×Duroc) using an Illumina MiSeq next-generation sequencer. In-silico analysis showed that the newly designed universal prokaryotic primers matched approximately 98.0% of Bacteria and 94.6% of Archaea rRNA gene sequences in the Ribosomal Database Project database. For each sequencing reaction performed with the prokaryotic universal primer, an average of 69,330 (±20,482) reads were obtained, of which archaeal rRNA genes comprised approximately 1.2% to 3.2% of all prokaryotic reads. In addition, the detection frequency of Bacteria belonging to the phylum Verrucomicrobia, including members of the classes Verrucomicrobiae and Opitutae, was higher in the NGS analysis using the prokaryotic universal primer than that performed with the bacterial universal primer. Importantly, this new prokaryotic universal primer set had markedly lower bias than that of most previously designed universal primers. Our findings demonstrate that the prokaryotic universal primer set designed in the present study will permit the simultaneous detection of Bacteria and Archaea, and will therefore allow for a more comprehensive understanding of microbial community structures in environmental samples.

Alteration in the gastric microbiota and its restoration by probiotics in patients with functional dyspepsia.

Abstract: Objective The objective of this study was to comparatively analyse the gastric fluid (GF) microbiota between patients with functional dyspepsia (FD) and healthy controls (HC), and to assess the effect of probiotics on the microbiota. Design Twenty-four Japanese patients with FD who met the Rome III definition and 21 age-matched and gender-matched HC volunteers were enrolled. The patients with FD had been treated with LG21, a probiotic strain. The GF was sampled after an overnight fast using a nasogastric tube. The bile acids concentration was determined by ELISA. The V3-V4 region of 16S rRNA gene was amplified using bacterial DNA from the GF, and then about 30 000 high-quality amplicons per sample were grouped into operational taxonomic units for analyses. Results The ratio of GF samples in which the bile acids were detectable was significantly greater in the FD than in the HC groups. In the bacterial composition analysis at the phylum level, the GF microbiota had a Bacteroidetes > Proteobacteria abundance and an absence of Acidobacteria in the FD group, in contrast, the GF microbiota had a Bacteroidetes Proteobacteria abundance and the presence of Acidobacteria in the HC group. Probiotic therapy in patients with FD shifted the composition of the GF microbiota to that observed in the HC volunteers. Conclusions Alteration in the GF microbiota was found in patients with FD compared with HC volunteers. Reflux of the small intestinal contents, including bile acid and intestinal bacteria, to the stomach was suggested to induce a bacterial composition change and be involved in the pathophysiology underlying FD. Probiotics appear effective in the treatment of FD through the normalisation of gastric microbiota. Trial registration number UMINCTR 000022026; Results.

Hesperetin Modifies the Composition of Fecal Microbiota and Increases Cecal Levels of Short-Chain Fatty Acids in Rats

Abstract: There has been particular interest in the prebiotic-like effects of commonly consumed polyphenols. This study aimed to evaluate the effects of hesperidin (HD) and its aglycone hesperetin (HT), major flavonoids in citrus fruits, on the structure and activity of gut microbiota in rats. Rats ingested an assigned diet (a control diet, a 0.5% HT diet, or a 1.0% HD diet) for 3 weeks. Terminal restriction fragment length polymorphism analysis revealed that the proportion of Clostridium subcluster XIVa in the feces collected at the third week of feeding was significantly reduced by the HT diet: 19.8 ± 4.3% for the control diet versus 5.3 ± 1.5% for the HT diet (P < 0.01). There was a significant difference in the cecal pool of short-chain fatty acids (SCFA), the sum of acetic, propionic, and butyric acids, between the control diet (212 ± 71 μmol) and the HT diet (310 ± 51 μmol) (P < 0.05), whereas the HD diet exhibited no effects (245 ± 51 μmol). Interestingly, dietary HT resulted in a significant increase in the...

Changes in the gut microbiota composition and the plasma ghrelin level in patients with Helicobacter pylori-infected patients with eradication therapy.

Abstract: Objective To investigate the influence of antimicrobials on both the gut microbiota structure and the plasma ghrelin level using Helicobacter pylori-infected patients who underwent eradication therapy. Design Twenty H. pylori-infected patients (mean age 68.3 years old) who underwent eradication therapy participated in the study. For the therapy, patients had 1 week of triple therapy consisting of amoxicillin, clarithromycin and proton-pump inhibitors. Stool and blood samples were obtained before (S1), immediately after (S2) and/or 3 months after (S3) the therapies. The concentrations of ghrelin and leptin in the blood were assayed using an ELISA. The V3-V4 region of the 16S rRNA gene was amplified using bacterial DNA from the stool, and about 50 000 high-quality amplicons per sample were grouped into operational taxonomic units for bacteriological analyses. Results The Bacteroidetes:Firmicutes (B:F) ratio was significantly greater at S3 than S1 (P Conclusions Changes in the gut microbiota, such as the B:F ratio after treatment with antimicrobials, might cause a change in the plasma ghrelin level, as the direct and earliest target of antimicrobials would be the microbiota rather than the hormone-secreting system.

Oral administration of Lactobacillus plantarum strain Lq80 to weaning piglets stimulates the growth of indigenous lactobacilli to modify the lactobacillal population.

Abstract: The composition and the number of fecal or intestinal lactobacilli were determined in weaned piglets before (pre-administration and day 0) and during (days 4, 7, and 14) the administration of Lactobacillus plantarum strain Lq80 (daily dose=1010 cells). Fecal or intestinal lactobacilli were isolated, and the isolates were grouped by RAPD-PCR and further identified by 16S rDNA partial sequences. During administration, the number of lactobacilli in feces and intestinal contents (log cfu/g) increased from day 0 to day 4 (7.96 vs. 10.07, p<0.05), to day 7 (7.96 vs. 10.18, p<0.05), and to day 14 (7.96 vs. 9.02, p=0.07) in the administered group, although L. plantarum was isolated from the feces of piglets in the administered group at 5.0×106 cfu/g. The level of culturable lactobacilli was significantly higher on day 7 in the administered group than in the control group (8.93 vs. 10.18, p=0.0002). Furthermore, the minor species in the lactobacillal population under the control condition become detectable with the administration of strain Lq80. The oral administration of L. plantarum strain Lq80 was effective to stimulate the development of the lactobacillal population in the intestine of post-weaning piglets.

Reduced Abundance of Butyrate-Producing Bacteria Species in the Fecal Microbial Community in Crohn's Disease.

Abstract: Background: The global alteration of the gut microbial community (dysbiosis) plays an important role in the pathogenesis of inflammatory bowel diseases (IBDs). Ho

Comparison of the gut microbiota composition between obese and non-obese individuals in a Japanese population, as analyzed by terminal restriction fragment length polymorphism and next-generation sequencing

Abstract: Obesity has become one of the most serious social problems in developed countries, including Japan. The relationship between the gut microbiota and obesity has recently attracted the attention of many researchers. Although the gut microbiota was long thought to contribute to obesity, the exact association remains largely unknown. We examined the human gut microbiota composition in a Japanese population in order to determine its relationship to obesity. Stool samples from 23 non-obese subjects (body mass index [BMI] <20 kg/m2) and 33 obese subjects (BMI ≥25 kg/m2) were collected and DNA was extracted prior to colonoscopy. After terminal restriction fragment length polymorphism (T-RFLP) analysis, samples from 10 subjects (4 non-obese and 6 obese) were selected and subjected to next-generation sequencing for species-level analysis. T-RFLP analysis showed significantly reduced numbers of Bacteroidetes and a higher Firmicutes to Bacteroidetes ratio in obese subjects compared with non-obese subjects. Bacterial diversity was significantly greater in obese subjects compared with non-obese subjects. Next-generation sequencing revealed that obese and non-obese subjects had different gut microbiota compositions and that certain bacterial species were significantly associated with each group (obese: Blautia hydrogenotorophica, Coprococcus catus, Eubacterium ventriosum, Ruminococcus bromii, Ruminococcus obeum; non-obese: Bacteroides faecichinchillae, Bacteroides thetaiotaomicron, Blautia wexlerae, Clostridium bolteae, Flavonifractor plautii). Gut microbial properties differ between obese and non-obese subjects in Japan, suggesting that gut microbiota composition is related to obesity.

Drought and host selection influence bacterial community dynamics in the grass root microbiome.

Abstract: Root endophytes have been shown to have important roles in determining host fitness under periods of drought stress, and yet the effect of drought on the broader root endosphere bacterial community remains largely uncharacterized. In this study, we present phylogenetic profiles of bacterial communities associated with drought-treated root and rhizosphere tissues of 18 species of plants with varying degrees of drought tolerance belonging to the Poaceae family, including important crop plants. Through 16S rRNA gene profiling across two distinct watering regimes and two developmental time points, we demonstrate that there is a strong correlation between host phylogenetic distance and the microbiome dissimilarity within root tissues, and that drought weakens this correlation by inducing conserved shifts in bacterial community composition. We identify a significant enrichment in a wide variety of Actinobacteria during drought within the roots of all hosts, and demonstrate that this enrichment is higher within the root than it is in the surrounding environments. Furthermore, we show that this observed enrichment is the result of an absolute increase in Actinobacterial abundance and that previously hypothesized mechanisms for observed enrichments in Actinobacteria in drought-treated soils are unlikely to fully account for the phenomena observed here within the plant root.

The role of probiotics, prebiotics and synbiotics in animal nutrition

Abstract: Along with the intensive development of methods of livestock breeding, breeders' expectations are growing concerning feed additives that would guarantee such results as accelerating growth rate, protection of health from pathogenic infections and improvement of other production parameters such as: absorption of feed and quality of meat, milk, eggs. The main reason for their application would be a strive to achieve some beneficial effects comparable to those of antibiotic-based growth stimulators, banned on 01 January 2006. High hopes are being associated with the use of probiotics, prebiotics and synbiotics. Used mainly for maintenance of the equilibrium of the intestinal microbiota of livestock, they turn out to be an effective method in fight against pathogens posing a threat for both animals and consumers. This paper discusses definitions of probiotics, prebiotics and synbiotics. Criteria that have to be met by those kinds of formulas are also presented. The paper offers a list of the most commonly used probiotics and prebiotics and some examples of their combinations in synbiotic formulas used in animal feeding. Examples of available study results on the effect of probiotics, prebiotics and synbiotics on animal health are also summarised.