Journal: bioRxiv

Article Title: In-field metagenome and 16S rRNA gene amplicon nanopore sequencing robustly characterize glacier microbiota

doi: 10.1101/073965

Figure Lengend Snippet: In-field 16S rRNA gene amplicon sequencing of cryoconite bacterial communities showing (A) phylum and family level taxonomic distribution of (B) open and closed cryoconite holes sampled on Vestre Brøggerbreen. The top image shows a “closed” cryoconite hole where the snow and superimposed ice cover has been displaced to reveal the hole, while the middle image shows a seasonally-open cryoconite hole. The bottom image shows the arrangement of equipment for DNA extraction, 16S rRNA gene PCR and nanopore sequencing in a field lab.

Article Snippet: Samples were transferred to the NERC Arctic Research Station Ny Ålesund within three hours and DNA extracted and quantified as described above within the field lab. Bacterial 16S rRNA genes were amplified from 50 ng DNA per sample diluted to 10 µL nuclease free water in 50 µL with 1 × LongAmp Taq master mix (New England Biolabs, Inc), 1 µL 16S rRNA gene barcoded primer (ONT-SQK-RAB-201, Oxford Nanopore Technologies, Ltd) and 14 µL nuclease free water.

Techniques: Amplification, Sequencing, DNA Extraction, Nanopore Sequencing

Journal: bioRxiv

Article Title: In-field metagenome and 16S rRNA gene amplicon nanopore sequencing robustly characterize glacier microbiota

doi: 10.1101/073965

Figure Lengend Snippet: multivariate discrimination of cryoconite bacterial communities revealed by in-field 16S rRNA gene amplicon sequencing. Analyses are performed with data aggregated to phylum/proteobacterial class (A-B) or family-level taxa (C-D) with Hierarchical Cluster Analysis (HCA, subpanels A, C,) or Principal Cooordinates Analysis (PCoA, B,D,). Closed holes (VB1-3) and open holes (VB5-6) are ordinated by multivariate analysis of of fourth-root transformed Bray-Curtis distances of phylotype relative abundances.

Article Snippet: Samples were transferred to the NERC Arctic Research Station Ny Ålesund within three hours and DNA extracted and quantified as described above within the field lab. Bacterial 16S rRNA genes were amplified from 50 ng DNA per sample diluted to 10 µL nuclease free water in 50 µL with 1 × LongAmp Taq master mix (New England Biolabs, Inc), 1 µL 16S rRNA gene barcoded primer (ONT-SQK-RAB-201, Oxford Nanopore Technologies, Ltd) and 14 µL nuclease free water.

Techniques: Amplification, Sequencing, Transformation Assay

Journal: bioRxiv

Article Title: In-field metagenome and 16S rRNA gene amplicon nanopore sequencing robustly characterize glacier microbiota

doi: 10.1101/073965

Figure Lengend Snippet: Benchmarking nanopore 16S rRNA gene amplicon sequencing of cryoconite bacterial communities by comparison with laboratory-generated data. (A) Phylum and family level taxonomic distribution of cryoconite bacterial communities and (B) phylum and (C) family level taxon distributions used for principal coordinates analysis of fourth-root transformed Bray-Curtis distances of phylotype relative abundances discriminate between Arctic and alpine cryoconite communities. Arctic glaciers (blue, AB, ML,VB) and Alpine glaciers (red, GF, RM) are clearly ordinated.

Article Snippet: Samples were transferred to the NERC Arctic Research Station Ny Ålesund within three hours and DNA extracted and quantified as described above within the field lab. Bacterial 16S rRNA genes were amplified from 50 ng DNA per sample diluted to 10 µL nuclease free water in 50 µL with 1 × LongAmp Taq master mix (New England Biolabs, Inc), 1 µL 16S rRNA gene barcoded primer (ONT-SQK-RAB-201, Oxford Nanopore Technologies, Ltd) and 14 µL nuclease free water.

Techniques: Amplification, Sequencing, Comparison, Generated, Transformation Assay

Journal: bioRxiv

Article Title: In-field metagenome and 16S rRNA gene amplicon nanopore sequencing robustly characterize glacier microbiota

doi: 10.1101/073965

Figure Lengend Snippet: Benchmarking nanopore 16S rRNA gene amplicon sequencing of cryoconite bacterial communities by comparison with laboratory-generated data. Correlation of log relative abundances between (A) the ratio of Alphaproteobacteria and Betaproteobacteria (B) Cyanobacteria to Alphaproteobacteria:Betaproteobacteria and (C) key taxonomic groups revealed using nanopore and pyro-sequencing of 16S rRNA genes. Positive, significant or highly significant Pearson r correlations are observed for each taxon.

Article Snippet: Samples were transferred to the NERC Arctic Research Station Ny Ålesund within three hours and DNA extracted and quantified as described above within the field lab. Bacterial 16S rRNA genes were amplified from 50 ng DNA per sample diluted to 10 µL nuclease free water in 50 µL with 1 × LongAmp Taq master mix (New England Biolabs, Inc), 1 µL 16S rRNA gene barcoded primer (ONT-SQK-RAB-201, Oxford Nanopore Technologies, Ltd) and 14 µL nuclease free water.

Techniques: Amplification, Sequencing, Comparison, Generated

Journal: bioRxiv

Article Title: In-field metagenome and 16S rRNA gene amplicon nanopore sequencing robustly characterize glacier microbiota

doi: 10.1101/073965

Figure Lengend Snippet: Benchmarking nanopore16S rRNA gene amplicon sequencing by sequencing mock communities using in-field protocols. Panel (A) shows the expected taxonomic distribution of The ZymoBIOMICS Microbial Community Standard for each sample, spiked in triplicate with Micrococcus luteus NCTC2665 genomic DNA to afford theoretical relative abundances of 16S rRNA genes in the range of 0-1.2% of the total bacterial community. Panel (B) shows shows the observed data where all reads assigned to genus level (excepting the poorly resolvable Enterobacteriaceae while Panel (C) shows all reads at the family level. The expected and observed Micrococcus luteus spike is highlighted in yellow.

Article Snippet: Samples were transferred to the NERC Arctic Research Station Ny Ålesund within three hours and DNA extracted and quantified as described above within the field lab. Bacterial 16S rRNA genes were amplified from 50 ng DNA per sample diluted to 10 µL nuclease free water in 50 µL with 1 × LongAmp Taq master mix (New England Biolabs, Inc), 1 µL 16S rRNA gene barcoded primer (ONT-SQK-RAB-201, Oxford Nanopore Technologies, Ltd) and 14 µL nuclease free water.

Techniques: Amplification, Sequencing

Journal: Frontiers in Microbiology

Article Title: Marine vampires: Persistent, internal associations between bacteria and blood-feeding marine annelids and crustaceans

doi: 10.3389/fmicb.2022.1113237

Figure Lengend Snippet: Microbiome diversity analysis of obligate blood feeding (OBF) crustaceans and leeches, based on 16S rRNA gene sequence similarity, using Non-metric multidimensional scaling (NMDS) ordination plots based on Bray–Curtis similarity resemblance, at the (A,B) broad category level, including blood-feeding versus comparison samples of non-blood feeding (NonBF) taxa, biological surfaces, and seawater (SW) and at the (C,D) specific blood-feeding taxa level. (E,F) Relative abundance of bacterial community structure at the genus level, from marine blood-feeders collected primarily from southern California coastal waters, including isopods Elthusa and Nerocila , the copepod Lernanthropus (Lerna), and leeches Branchellion, Ostreobdella, and Pterobdella (specific specimens are listed in in the same order as shown in the bar charts). Assigned bacterial taxa are color-coded as shown below. Taxa in dark gray were only found in the non-blood feeder (NBF) or seawater (SW) samples. Taxa in light gray were minor taxa in all specimens. See for a full key.

Article Snippet: The raw Illumina 16S rRNA gene barcode sequences and metadata collected in this study are available from the NCBI Small Read Archive (BioProject # PRJNA910167).

Techniques: Sequencing, Comparison

Journal: Frontiers in Microbiology

Article Title: Marine vampires: Persistent, internal associations between bacteria and blood-feeding marine annelids and crustaceans

doi: 10.3389/fmicb.2022.1113237

Figure Lengend Snippet: Relative abundances of the 7 most prevalent Vibrio ribotypes (based on 99% 16S rRNA gene sequence similarity) from obligate marine blood-feeders including the isopods Elthusa and Nerocila (Nero), copepod Lernanthropus (Lerna), and leeches Branchellion, Ostreobdella, and Pterobdella, compared to non-blood-feeding crustaceans (Non-BF), swabs of fish skin, and seawater (SW). Bars are scaled according to percent abundance of each Vibrio ribotype as a function of the entire microbial community in that specimen (for reference, occasional numbers indicate the % abundance of that bar). A total of 135 Vibrionaceae ribotypes were recovered from all samples in total, but the 7 portrayed here accounted for 80% of the total Vibrionaceae diversity. See for the phylogenetic position of these ribotypes. Note the pooling of Vibrio ribotypes 19,850 with 213,579 (+) common only in the Nerocila specimens via amplicon sequencing. Occasional symbols note distinction within OBF taxa, including Pterobdella occidentalis recovered from the goby (#), and P. abditovesiculata from Hawaii (*), as well as Branchellion collected from rays other than the pacific ray (°), and Elthusa collected from killifish (+).

Article Snippet: The raw Illumina 16S rRNA gene barcode sequences and metadata collected in this study are available from the NCBI Small Read Archive (BioProject # PRJNA910167).

Techniques: Sequencing, Amplification

Journal: Journal of Assisted Reproduction and Genetics

Article Title: Improving analysis of the vaginal microbiota of women undergoing assisted reproduction using nanopore sequencing

doi: 10.1007/s10815-022-02628-4

Figure Lengend Snippet: Comparison of classified species using short- or long-read sequencing approaches. The stacked bar plot of the fraction of reads shows the most abundant bacterial taxa classified from the 16S rRNA gene amplicons. The sample IDs are indicated on the top and the sequencing approaches are on the bottom x -axis. Illumina: Illumina short-read sequencing (V3F/V4R), Nanoporefull: nanopore long-read sequencing (27F-YM/1492R-Y), Nanoporev3: nanopore long-read sequencing (V3F/1492R-Y)

Article Snippet: After preliminary evaluating a subset of samples, the 16S rRNA gene Barcoding Kit does not recapitulate Illumina sequencing results by using the universal primer pair 27f/1429R (Supplementary Fig. ).

Techniques: Comparison, Sequencing

Journal: Frontiers in Microbiology

Article Title: Assembly and Succession of Iron Oxide Microbial Mat Communities in Acidic Geothermal Springs

doi: 10.3389/fmicb.2016.00025

Figure Lengend Snippet: The relative abundance of different phylotypes from “mature” Fe(III)-oxide mats of One Hundred Spring Plain and Beowulf Spring (YNP) determined using random DNA sequencing over multiple years (and sequencing technologies), or short-fragment 16S rRNA gene sequencing (Illumina iTag) .

Article Snippet: The abundance of phylotypes from mature Fe(III)-oxide mats were compared using Illumina 16S rRNA gene barcodes vs. Illumina random metagenome sequencing from matching samples.

Techniques: DNA Sequencing, Sequencing

Journal: Frontiers in Microbiology

Article Title: Assembly and Succession of Iron Oxide Microbial Mat Communities in Acidic Geothermal Springs

doi: 10.3389/fmicb.2016.00025

Figure Lengend Snippet: Relative abundance (16S rRNA gene iTags) of key lithoautotrophic microbial populations over the time course of slide incubations in One Hundred Spring Plain (A) and Beowulf (B) Springs ( Hydrogenobaculum spp. = open squares; Metallosphaera yellowstonensis = closed black circles) .

Article Snippet: The abundance of phylotypes from mature Fe(III)-oxide mats were compared using Illumina 16S rRNA gene barcodes vs. Illumina random metagenome sequencing from matching samples.

Techniques:

Journal: Frontiers in Microbiology

Article Title: Assembly and Succession of Iron Oxide Microbial Mat Communities in Acidic Geothermal Springs

doi: 10.3389/fmicb.2016.00025

Figure Lengend Snippet: Relative abundance (16S rRNA gene Illumina barcodes) of different phylotypes determined on slides incubated in One Hundred Spring Plain (A) and Beowulf (B) Springs from 4 to 70 days . The dendrograms on the y and x axes represent the Bray-Curtis dissimilarity matrix between taxon abundance at different time points and taxon abundances, respectively.

Article Snippet: The abundance of phylotypes from mature Fe(III)-oxide mats were compared using Illumina 16S rRNA gene barcodes vs. Illumina random metagenome sequencing from matching samples.

Techniques: Incubation

Journal: Frontiers in Microbiology

Article Title: Assembly and Succession of Iron Oxide Microbial Mat Communities in Acidic Geothermal Springs

doi: 10.3389/fmicb.2016.00025

Figure Lengend Snippet: Relative abundance of taxa (16S rRNA gene Illumina barcodes) as a function of depth in mature thermoacidic iron oxide mats from One Hundred Spring Plain (A) and Beowulf (B) Springs . The top position refers to the upper 1 mm oxygenated zone, the middle position represents depths of ~2–7 mm, and the bottom position represents hypoxic regions > 10 mm.

Article Snippet: The abundance of phylotypes from mature Fe(III)-oxide mats were compared using Illumina 16S rRNA gene barcodes vs. Illumina random metagenome sequencing from matching samples.

Techniques: