Gender assessment of the gut microbiome in obese patients

The aim. To assess the relationship between body mass index (BMI) and gut bacteria in men and women with obesity.Materials and methods. The study included 56 overweight patients, divided into 2 groups. The first group consisted of 27 women (the average age was 62 ± 2.2 years), the second group — 29 men (the average age was 55 ± 9 years). The Quetelet index (kg / m2) was calculated for all patients. To study the gut microbiome, the method of polymerase chain reaction in real time (RT-PCR) and metagenomic sequencing were used. DNA from feces was isolated using the Express-DNA-Bio DNA isolation kit (AlkorBio, Russia). To carry out RT-PCR, a set of reagents “Colonoflor-16” (“Alfalab”, Russia) was used. For microbiome sequencing, DNA libraries were prepared using the Illumina Nextera Sample Preparation Kit with DNA primers corresponding to the V3 — V4 regions of the 16S rRNA gene. The study of fecal samples was carried out using 16S rRNA gene sequencing on the Illumina platform (MiSeq sequencer).Results. It was revealed that a higher total number of bacteria, an increased content of Bacteroides fragilis group and Faeca-libacterium prausnitzii, is statistically significantly more common in women than in men. Strong negative correlations were found between BMI and total bacterial mass, between BMI and the number of Bacteroides fragilis group among women with grade I obesity. In overweight men, a correlation was found between BMI and the Bacteroides fragilis group / Faecalibacterium prausnitzii ratio.Conclusions. The total number of bacteria, the content of Bacteroides fragilis group and Faecalibacterium prausnitzii in the gut of patients have statistically significant associations with BMI, and probably can affect the formation of metabolic disorders to a greater extent in women than in men. To clarify the identified trends and patterns in this pilot study, further study of the microbiome with a large number of patients and additional analyzes of the metagenome (16S rRNA) and metabolome, a transcriptome, allowing to control the expression of key metabolic enzymes, largely associated with the compositional features of the gut microbiocenosis, is required.

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Deep Investigating the Changes of Gut Microbiome and Its Correlation With the Shifts of Host Serum Metabolome Around Parturition in Sows

Frontiers in Microbiology ◽

10.3389/fmicb.2021.729039 ◽

2021 ◽

Vol 12 ◽

Author(s):

Hao Fu ◽

Maozhang He ◽

Jinyuan Wu ◽

Yunyan Zhou ◽

Shanlin Ke ◽

...

Keyword(s):

16S Rrna ◽

Gut Microbiota ◽

16S Rrna Gene ◽

Gut Microbiome ◽

Bacterial Species ◽

Late Pregnancy ◽

Rrna Gene ◽

Metagenomic Sequencing ◽

Sequencing Data ◽

Functional Capacities

Parturition is a crucial event in the sow reproduction cycle, which accompanies by a series of physiological changes, including sex hormones, metabolism, and immunity. More and more studies have indicated the changes of the gut microbiota from pregnancy to parturition. However, what bacterial species and functional capacities of the gut microbiome are changed around parturition has been largely unknown, and the correlations between the changes of gut bacterial species and host metabolome were also uncovered. In this study, by combining 16S rRNA gene and shotgun metagenomic sequencing data, and the profiles of serum metabolome and fecal short-chain fatty acids (SCFAs), we investigated the changes of gut microbiome, serum metabolite features and fecal SCFAs from late pregnancy (LP) to postpartum (PO) stage. We found the significant changes of gut microbiota from LP to PO stage in both 16S rRNA gene sequencing and metagenomic sequencing analyses. The bacterial species from Lactobacillus, Streptococcus, and Clostridium were enriched at the LP stage, while the species from Bacteroides, Escherichia, and Campylobacter had higher abundances at the PO stage. Functional capacities of the gut microbiome were also significantly changed and associated with the shifts of gut bacteria. Untargeted metabolomic analyses revealed that the metabolite features related to taurine and hypotaurine metabolism, and arginine biosynthesis and metabolism were enriched at the LP stage, and positively associated with those bacterial species enriched at the LP stage, while the metabolite features associated with vitamin B6 and glycerophospholipid metabolism had higher abundances at the PO stage and were positively correlated with the bacteria enriched at the PO stage. Six kinds of SCFAs were measured in feces samples and showed higher concentrations at the LP stage. These results suggested that the changes of gut microbiome from LP to PO stage lead to the shifts of host lipid, amino acids and vitamin metabolism and SCFA production. The results from this study provided new insights for the changes of sow gut microbiome and host metabolism around parturition, and gave new knowledge for guiding the feeding and maternal care of sows from late pregnancy to lactation in the pig industry.

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A unique and reliable fecal DNA extraction method for 16S rRNA gene and shotgun metagenomic sequencing in the analysis of the human gut microbiome

10.21203/rs.3.rs-52279/v1 ◽

2020 ◽

Author(s):

Céline Elie ◽

Magali Perret ◽

Karen Louis ◽

Asmaà Fritah-Lafont ◽

Philippe Leissner ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Dna Extraction ◽

Gut Microbiome ◽

High Throughput Sequencing ◽

Rrna Gene ◽

Metagenomic Sequencing ◽

Human Gut ◽

Human Gut Microbiome ◽

Shotgun Metagenomic Sequencing

Abstract Background: The gut microbiome is widely analyzed using high-throughput sequencing, such as 16S rRNA gene amplicon sequencing and shotgun metagenomic sequencing (SMS). DNA extraction is known to have a large impact on the metagenomic analyses. The aim of this study was to select a unique and best performing DNA extraction protocol for both metagenomic sequencing methods. In that context, four commonly used DNA extraction methods were compared for the analysis of the gut microbiota. Commercial versions were evaluated against modified protocols using a stool preprocessing device (SPD, bioMérieux) in order to facilitate DNA extraction. Stool samples from nine healthy volunteers and nine patients with a Clostridium difficile infection were extracted with all protocols and sequenced with both metagenomic methods. Protocols were ranked using wet- and dry-lab criteria, including quality controls of the extracted genomic DNA, alpha-diversity, accuracy using a mock community of known composition and repeatability across technical replicates.Results: Independently of the sequencing methods used, SPD significantly improved efficiency of the four tested protocols compared with their commercial version, in terms of extracted DNA quality, accuracy of the predicted composition of the microbiota (notably for Gram-positive bacteria), sample alpha-diversity, and experimental repeatability. The best overall performance was obtained for the S-DQ protocol, SPD combined to the DNeasy PowerLyser PowerSoil protocol from QIAGEN.Conclusion: Based on this evaluation, we recommend to use the S-DQ protocol, to obtain standardized and high quality extracted DNA in the human gut microbiome studies.

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Faculty Opinions recommendation of Low Maternal Microbiota Sharing across Gut, Breast Milk and Vagina, as Revealed by 16S rRNA Gene and Reduced Metagenomic Sequencing.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.733207420.793548246 ◽

2018 ◽

Author(s):

Gregor Reid

Keyword(s):

Breast Milk ◽

16S Rrna ◽

16S Rrna Gene ◽

Rrna Gene ◽

Metagenomic Sequencing

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Comparison between 16S rRNA and shotgun sequencing data for the taxonomic characterization of the gut microbiota

Scientific Reports ◽

10.1038/s41598-021-82726-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Francesco Durazzi ◽

Claudia Sala ◽

Gastone Castellani ◽

Gerardo Manfreda ◽

Daniel Remondini ◽

...

Keyword(s):

16S Rrna ◽

Gut Microbiota ◽

16S Rrna Gene ◽

Gene Sequencing ◽

16S Rrna Gene Sequencing ◽

Shotgun Sequencing ◽

Rrna Gene ◽

Metagenomic Sequencing ◽

Experimental Conditions ◽

Rrna Gene Sequencing

AbstractIn this paper we compared taxonomic results obtained by metataxonomics (16S rRNA gene sequencing) and metagenomics (whole shotgun metagenomic sequencing) to investigate their reliability for bacteria profiling, studying the chicken gut as a model system. The experimental conditions included two compartments of gastrointestinal tracts and two sampling times. We compared the relative abundance distributions obtained with the two sequencing strategies and then tested their capability to distinguish the experimental conditions. The results showed that 16S rRNA gene sequencing detects only part of the gut microbiota community revealed by shotgun sequencing. Specifically, when a sufficient number of reads is available, Shotgun sequencing has more power to identify less abundant taxa than 16S sequencing. Finally, we showed that the less abundant genera detected only by shotgun sequencing are biologically meaningful, being able to discriminate between the experimental conditions as much as the more abundant genera detected by both sequencing strategies.

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Mucosa-Associated Faecalibacterium prausnitzii Phylotype Richness Is Reduced in Patients with Inflammatory Bowel Disease

Applied and Environmental Microbiology ◽

10.1128/aem.02006-15 ◽

2015 ◽

Vol 81 (21) ◽

pp. 7582-7592 ◽

Cited By ~ 49

Author(s):

Mireia Lopez-Siles ◽

Margarita Martinez-Medina ◽

Carles Abellà ◽

David Busquets ◽

Miriam Sabat-Mir ◽

...

Keyword(s):

Inflammatory Bowel Disease ◽

16S Rrna ◽

16S Rrna Gene ◽

Bowel Disease ◽

Denaturing Gradient Gel Electrophoresis ◽

Gastrointestinal Disease ◽

Rrna Gene ◽

Content Type ◽

Faecalibacterium Prausnitzii ◽

Inflammatory Bowel

ABSTRACTFaecalibacterium prausnitziidepletion in intestinal diseases has been extensively reported, but little is known about intraspecies variability. This work aims to determine if subjects with gastrointestinal disease host mucosa-associatedF. prausnitziipopulations different from those hosted by healthy individuals. A new species-specific PCR-denaturing gradient gel electrophoresis (PCR-DGGE) method targeting the 16S rRNA gene was developed to fingerprintF. prausnitziipopulations in biopsy specimens from 31 healthy control (H) subjects and 36 Crohn's disease (CD), 23 ulcerative colitis (UC), 6 irritable bowel syndrome (IBS), and 22 colorectal cancer (CRC) patients. The richness ofF. prausnitziisubtypes was lower in inflammatory bowel disease (IBD) patients than in H subjects. The most prevalent operational taxonomic units (OTUs) consisted of four phylotypes (OTUs with a 99% 16S rRNA gene sequence similarity [OTU99]), which were shared by all groups of patients. Their distribution and the presence of some disease-specificF. prausnitziiphylotypes allowed us to differentiate the populations in IBD and CRC patients from that in H subjects. At the level of a minimum similarity of 97% (OTU97), two phylogroups accounted for 98% of the sequences. Phylogroup I was found in 87% of H subjects but in under 50% of IBD patients (P= 0.003). In contrast, phylogroup II was detected in >75% of IBD patients and in only 52% of H subjects (P= 0.005). This study reveals that even though the main members of theF. prausnitziipopulation are present in both H subjects and individuals with gut diseases, richness is reduced in the latter and an altered phylotype distribution exists between diseases. This approach may serve as a basis for addressing the suitability ofF. prausnitziiphylotypes to be quantified as a putative biomarker of disease and depicting the importance of the loss of these subtypes in disease pathogenesis.

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Metagenome-validated Parallel Amplicon Sequencing and Text Mining-based Annotations for Simultaneous Profiling of Bacteria and Fungi: Vaginal Microbiome and Mycobiota in Healthy Women

10.21203/rs.3.rs-321778/v1 ◽

2021 ◽

Author(s):

Seppo Virtanen ◽

Schahzad Saqib ◽

Tinja Kanerva ◽

Pekka Nieminen ◽

Ilkka Kalliala ◽

...

Keyword(s):

16S Rrna ◽

16S Rrna Gene ◽

Cost Effective ◽

Amplicon Sequencing ◽

Rrna Gene ◽

Extensive Literature ◽

Metagenomic Sequencing ◽

Text Extraction ◽

Healthy Women ◽

Bacteria And Fungi

Abstract Background: Amplicon sequencing of kingdom-specific tags such as 16S rRNA gene for bacteria and internal transcribed spacer (ITS) region for fungi are widely used for investigating microbial populations. So far most human studies have focused on bacteria while studies on host-associated fungi in health and disease have only recently started to accumulate. To enable cost-effective parallel analysis of bacterial and fungal communities in human and environmental samples, we developed a method where 16S rRNA gene and ITS-1 amplicons were pooled together for a single Illumina MiSeq or HiSeq run and analysed after primer-based segregation. Taxonomic assignments were performed with Blast in combination with an iterative text-extraction based filtration approach, which uses extensive literature records from public databases to select the most probable hits that were further validated by shotgun metagenomic sequencing. Results: Using 50 vaginal samples, we show that the combined run provides comparable results on bacterial composition and diversity to conventional 16S rRNA gene amplicon sequencing. The text-extraction-based taxonomic assignment guided tool provided ecosystem specific annotations that were confirmed by Metagenomic Phylogenetic Analysis (MetaPhlAn). The metagenome analysis revealed distinct functional differences between the bacterial community types while fungi were undetected, despite being identified in all samples based on ITS amplicons. Co-abundance analysis of bacteria and fungi did not show strong between-kingdom correlations within the vaginal ecosystem of healthy women.Conclusion: Combined amplicon sequencing for bacteria and fungi provides a simple and cost-effective method for simultaneous analysis of microbiota and mycobiota within the same samples. Text extraction-based annotation tool facilitates the characterization and interpretation of defined microbial communities from rapidly accumulating sequencing and metadata readily available through public databases.

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Alone Yet Not Alone: Frankia Lives Under the Same Roof With Other Bacteria in Actinorhizal Nodules

Frontiers in Microbiology ◽

10.3389/fmicb.2021.749760 ◽

2021 ◽

Vol 12 ◽

Author(s):

Faten Ghodhbane-Gtari ◽

Timothy D’Angelo ◽

Abdellatif Gueddou ◽

Sabrine Ghazouani ◽

Maher Gtari ◽

...

Keyword(s):

16S Rrna ◽

Plant Growth ◽

16S Rrna Gene ◽

Root Nodules ◽

Actinorhizal Plants ◽

Rrna Gene ◽

Plant Growth Promoting Bacteria ◽

Metagenomic Sequencing ◽

Plant Growth Promoting ◽

Growth Promoting

Actinorhizal plants host mutualistic symbionts of the nitrogen-fixing actinobacterial genus Frankia within nodule structures formed on their roots. Several plant-growth-promoting bacteria have also been isolated from actinorhizal root nodules, but little is known about them. We were interested investigating the in planta microbial community composition of actinorhizal root nodules using culture-independent techniques. To address this knowledge gap, 16S rRNA gene amplicon and shotgun metagenomic sequencing was performed on DNA from the nodules of Casuarina glauca. DNA was extracted from C. glauca nodules collected in three different sampling sites in Tunisia, along a gradient of aridity ranging from humid to arid. Sequencing libraries were prepared using Illumina NextEra technology and the Illumina HiSeq 2500 platform. Genome bins extracted from the metagenome were taxonomically and functionally profiled. Community structure based off preliminary 16S rRNA gene amplicon data was analyzed via the QIIME pipeline. Reconstructed genomes were comprised of members of Frankia, Micromonospora, Bacillus, Paenibacillus, Phyllobacterium, and Afipia. Frankia dominated the nodule community at the humid sampling site, while the absolute and relative prevalence of Frankia decreased at the semi-arid and arid sampling locations. Actinorhizal plants harbor similar non-Frankia plant-growth-promoting-bacteria as legumes and other plants. The data suggests that the prevalence of Frankia in the nodule community is influenced by environmental factors, with being less abundant under more arid environments.

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Yoghurt Consumption is Associated With Transient Changes in the Composition of the Human Gut Microbiome

10.21203/rs.3.rs-99718/v1 ◽

2020 ◽

Author(s):

Caroline Ivanne Le Roy ◽

Alexander Kurilshikov ◽

Emily Leeming ◽

Alessia Visconti ◽

Ruth Bowyer ◽

...

Keyword(s):

16S Rrna ◽

Gut Microbiota ◽

Visceral Fat ◽

Gut Microbiome ◽

Body Weight Gain ◽

Healthy Eating Index ◽

Rrna Gene ◽

Metagenomic Sequencing ◽

Sequencing Data ◽

Shotgun Metagenomic Sequencing

Abstract Background: Yoghurt contains live bacteria that could contribute via modulation of the gut microbiota to its reported beneficial effects such as reduced body weight gain and lower incidence of type 2 diabetes. To date, the association between yoghurt consumption and the composition of the gut microbiota is underexplored. Here we used clinical variables, metabolomics, 16S rRNA and shotgun metagenomic sequencing data collected on over 1000 predominantly female UK twins to define the link between the gut microbiota and yoghurt-associated health benefits. Results: According to food frequency questionnaires (FFQ), 73% of subjects consumed yoghurt. Consumers presented a healthier diet pattern (healthy eating index: beta = 2.17±0.34; P = 2.72x10-10) and improved metabolic health characterised by reduced visceral fat (beta = -28.18±11.71 g; P = 0.01). According to 16S rRNA gene analyses and whole shotgun metagenomic sequencing approach consistent taxonomic variations were observed with yoghurt consumption. More specifically, we identified higher abundance of species used as yoghurt starters Streptococcus thermophilus (beta = 0.41±0.051; P = 6.14x10-12) and sometimes added Bifidobacterium animalis subsp. lactis (beta = 0.30±0.052; P = 1.49x10-8) in the gut of yoghurt consumers. Replication in 1103 volunteers from the LifeLines-DEEP cohort confirmed the increase of S. thermophilus among yoghurt consumers. Using food records collected the day prior to faecal sampling we showed that increase in these two yoghurt bacteria could be transient. Metabolomics analysis revealed that B. animalis subsp. lactis was associated with 13 faecal metabolites including a 3-hydroxyoctanoic acid, known to be involved in the regulation of gut inflammation.Conclusions: Yoghurt consumption is associated with reduced visceral fat mass and changes in gut microbiome including transient increase of yoghurt-contained species (i.e. S. thermophilus and B. lactis).

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