scholarly journals OGUs enable effective, phylogeny-aware analysis of even shallow metagenome community structures

2021 ◽  
Author(s):  
Qiyun Zhu ◽  
Shi Huang ◽  
Antonio Gonzalez ◽  
Imran McGrath ◽  
Daniel McDonald ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Human Microbiome ◽  
Human Microbiome Project ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Shotgun Metagenomics ◽  
Biologically Relevant ◽  
Metagenome Analysis ◽  
Host Sex

We introduce Operational Genomic Unit (OGU), a metagenome analysis strategy that directly exploits sequence alignment hits to individual reference genomes as the minimum unit for assessing the diversity of microbial communities and their relevance to environmental factors. This approach is independent from taxonomic classification, granting the possibility of maximal resolution of community composition, and organizes features into an accurate hierarchy using a phylogenomic tree. The outputs are suitable for contemporary analytical protocols for community ecology, differential abundance and supervised learning while supporting phylogenetic methods, such as UniFrac and phylofactorization, that are seldomly applied to shotgun metagenomics despite being prevalent in 16S rRNA gene amplicon studies. As demonstrated in one synthetic and two real-world case studies, the OGU method produces biologically meaningful patterns from microbiome datasets. Such patterns further remain detectable at very low metagenomic sequencing depths. Compared with taxonomic unit-based analyses implemented in currently adopted metagenomics tools, and the analysis of 16S rRNA gene amplicon sequence variants, this method shows superiority in informing biologically relevant insights, including stronger correlation with body environment and host sex on the Human Microbiome Project dataset, and more accurate prediction of human age by the gut microbiomes in the Finnish population. We provide Woltka, a bioinformatics tool to implement this method, with full integration with the QIIME 2 package and the Qiita web platform, to facilitate OGU adoption in future metagenomics studies.

scholarly journals Molecular profiling of microbiota in human oral cavity, stomach and intestine

2018 ◽  
Author(s):  
Qianqian Liu ◽  
Feizhou Zhu ◽  
Liyu Chen ◽  
Meihua Xu ◽  
Jianwei Chen ◽  
...  
Keyword(s):  
16S Rrna ◽  
Oral Cavity ◽  
16S Rrna Gene ◽  
Gastric Juice ◽  
Human Microbiome ◽  
Human Microbiome Project ◽  
Rrna Gene ◽  
Human Oral Cavity ◽  
Stool Microbiota ◽  
The 16S Rrna Gene

The microbiota in the human gut is not only a complicated microecological system but also plays important roles in both health and disease. In order to understand the roles of these gut bacteria, we determined the distribution of microbiota in different regions of the gut by sequencing the 16S rRNA gene V4 region of the bacteria in the saliva, gastric juice, and stool of healthy individuals. The 16S rRNA gene V3-V5 region sequences of saliva and stool microbiota were obtained from Human Microbiome Project (HMP) and the V4 sequence was obtained from the V3-V5 sequences by a program designed by Perl language. We found that the microbiota of the gastric juice is more similar to those in the saliva rather than that in the stool. The frequency of some taxa was significantly different among the three groups with the Streptococcus, Veillonella, Oribacterium, Selenomonas, Actinomyces, and Granulicatella most abundant in the saliva; the Prevotella, Neisseria, Actinobacillus, Treponema, and Helicobacter most abundant in the gastric juice; and the Bacteroides, Parabacteroides, Faecalibacterium, Sutterella, Ruminococcus, Oscillospira and Phascolarctobacterium most abundant in the stool. In addition, results from PICRUSt analyses suggest that the functions of microbiota in the gastric juice are more similar as those in the saliva than in the stool. Moreover, we also found that the membrane transport of the microbiota in the saliva is higher than that in the stool and gastric juice. To our knowledge, this is the first comprehensive comparison of microbiota in the human oral cavity, stomach, and intestine.

scholarly journals Molecular profiling of microbiota in human oral cavity, stomach and intestine

2018 ◽  
Author(s):  
Qianqian Liu ◽  
Feizhou Zhu ◽  
Liyu Chen ◽  
Meihua Xu ◽  
Jianwei Chen ◽  
...  
Keyword(s):  
16S Rrna ◽  
Oral Cavity ◽  
16S Rrna Gene ◽  
Gastric Juice ◽  
Human Microbiome ◽  
Human Microbiome Project ◽  
Rrna Gene ◽  
Human Oral Cavity ◽  
Stool Microbiota ◽  
The 16S Rrna Gene

The microbiota in the human gut is not only a complicated microecological system but also plays important roles in both health and disease. In order to understand the roles of these gut bacteria, we determined the distribution of microbiota in different regions of the gut by sequencing the 16S rRNA gene V4 region of the bacteria in the saliva, gastric juice, and stool of healthy individuals. The 16S rRNA gene V3-V5 region sequences of saliva and stool microbiota were obtained from Human Microbiome Project (HMP) and the V4 sequence was obtained from the V3-V5 sequences by a program designed by Perl language. We found that the microbiota of the gastric juice is more similar to those in the saliva rather than that in the stool. The frequency of some taxa was significantly different among the three groups with the Streptococcus, Veillonella, Oribacterium, Selenomonas, Actinomyces, and Granulicatella most abundant in the saliva; the Prevotella, Neisseria, Actinobacillus, Treponema, and Helicobacter most abundant in the gastric juice; and the Bacteroides, Parabacteroides, Faecalibacterium, Sutterella, Ruminococcus, Oscillospira and Phascolarctobacterium most abundant in the stool. In addition, results from PICRUSt analyses suggest that the functions of microbiota in the gastric juice are more similar as those in the saliva than in the stool. Moreover, we also found that the membrane transport of the microbiota in the saliva is higher than that in the stool and gastric juice. To our knowledge, this is the first comprehensive comparison of microbiota in the human oral cavity, stomach, and intestine.

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

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.

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

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.

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

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.

scholarly journals Microbiome and Metagenome Analyses of a Closed Habitat during Human Occupation

mSystems ◽  
2020 ◽  
Vol 5 (4) ◽  
Author(s):  
Ganesh Babu Malli Mohan ◽  
Ceth W. Parker ◽  
Camilla Urbaniak ◽  
Nitin K. Singh ◽  
Anthony Hood ◽  
...  
Keyword(s):  
16S Rrna ◽  
Metal Surfaces ◽  
Microbial Population ◽  
Microbial Populations ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Particle Board ◽  
Shotgun Metagenomics ◽  
Glass Surfaces ◽  
Glass Metal

ABSTRACT Microbial contamination during long-term confinements of space exploration presents potential risks for both crew members and spacecraft life support systems. A novel swab kit was used to sample various surfaces from a submerged, closed, analog habitat to characterize the microbial populations. Samples were collected from various locations across the habitat which were constructed from various surface materials (linoleum, dry wall, particle board, glass, and metal), and microbial populations were examined by culture, quantitative PCR (qPCR), microbiome 16S rRNA gene sequencing, and shotgun metagenomics. Propidium monoazide (PMA)-treated samples identified the viable/intact microbial population of the habitat. The cultivable microbial population ranged from below the detection limit to 106 CFU/sample, and their identity was characterized using Sanger sequencing. Both 16S rRNA amplicon and shotgun sequencing were used to characterize the microbial dynamics, community profiles, and functional attributes (metabolism, virulence, and antimicrobial resistance). The 16S rRNA amplicon sequencing revealed abundance of viable (after PMA treatment) Actinobacteria (Brevibacterium, Nesternkonia, Mycobacterium, Pseudonocardia, and Corynebacterium), Firmicutes (Virgibacillus, Staphylococcus, and Oceanobacillus), and Proteobacteria (especially Acinetobacter) on linoleum, dry wall, and particle board (LDP) surfaces, while members of Firmicutes (Leuconostocaceae) and Proteobacteria (Enterobacteriaceae) were high on the glass/metal surfaces. Nonmetric multidimensional scaling determined from both 16S rRNA and metagenomic analyses revealed differential microbial species on LDP surfaces and glass/metal surfaces. The shotgun metagenomic sequencing of samples after PMA treatment showed bacterial predominance of viable Brevibacterium (53.6%), Brachybacterium (7.8%), Pseudonocardia (9.9%), Mycobacterium (3.7%), and Staphylococcus (2.1%), while fungal analyses revealed Aspergillus and Penicillium dominance. IMPORTANCE This study provides the first assessment of monitoring cultivable and viable microorganisms on surfaces within a submerged, closed, analog habitat. The results of the analyses presented herein suggest that the surface material plays a role in microbial community structure, as the microbial populations differed between LDP and metal/glass surfaces. The metal/glass surfaces had less-complex community, lower bioburden, and more closely resembled the controls. These results indicated that material choice is crucial when building closed habitats, even if they are simply analogs. Finally, while a few species were associated with previously cultivated isolates from the International Space Station and MIR spacecraft, the majority of the microbial ecology of the submerged analog habitat differs greatly from that of previously studied analog habitats.

scholarly journals Low Maternal Microbiota Sharing across Gut, Breast Milk and Vagina, as Revealed by 16S rRNA Gene and Reduced Metagenomic Sequencing

Genes ◽  
2018 ◽  
Vol 9 (5) ◽  
pp. 231 ◽  
Author(s):  
Ekaterina Avershina ◽  
Inga Angell ◽  
Melanie Simpson ◽  
Ola Storrø ◽  
Torbjørn Øien ◽  
...  
Keyword(s):  
Breast Milk ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Rrna Gene ◽  
Metagenomic Sequencing

scholarly journals Intrinsic challenges in ancient microbiome reconstruction using 16S rRNA gene amplification

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Kirsten A. Ziesemer ◽  
Allison E. Mann ◽  
Krithivasan Sankaranarayanan ◽  
Hannes Schroeder ◽  
Andrew T. Ozga ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
Dna Amplification ◽  
Amplicon Sequencing ◽  
Universal Primers ◽  
Rrna Gene ◽  
Dental Calculus ◽  
Shotgun Metagenomics ◽  
Variable Regions ◽  
V3 Region

Abstract To date, characterization of ancient oral (dental calculus) and gut (coprolite) microbiota has been primarily accomplished through a metataxonomic approach involving targeted amplification of one or more variable regions in the 16S rRNA gene. Specifically, the V3 region (E. coli 341–534) of this gene has been suggested as an excellent candidate for ancient DNA amplification and microbial community reconstruction. However, in practice this metataxonomic approach often produces highly skewed taxonomic frequency data. In this study, we use non-targeted (shotgun metagenomics) sequencing methods to better understand skewed microbial profiles observed in four ancient dental calculus specimens previously analyzed by amplicon sequencing. Through comparisons of microbial taxonomic counts from paired amplicon (V3 U341F/534R) and shotgun sequencing datasets, we demonstrate that extensive length polymorphisms in the V3 region are a consistent and major cause of differential amplification leading to taxonomic bias in ancient microbiome reconstructions based on amplicon sequencing. We conclude that systematic amplification bias confounds attempts to accurately reconstruct microbiome taxonomic profiles from 16S rRNA V3 amplicon data generated using universal primers. Because in silico analysis indicates that alternative 16S rRNA hypervariable regions will present similar challenges, we advocate for the use of a shotgun metagenomics approach in ancient microbiome reconstructions.

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