The diversity and community structure of symbiotic cyanobacteria in hornworts inferred from long‐read amplicon sequencing

2021 ◽  
Vol 108 (9) ◽  
pp. 1731-1744 ◽  
Author(s):  
Jessica M. Nelson ◽  
Duncan A. Hauser ◽  
Fay‐Wei Li
Keyword(s):  
Community Structure ◽  
Amplicon Sequencing ◽  
Long Read

scholarly journals Ultra-accurate microbial amplicon sequencing with synthetic long reads

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Benjamin J. Callahan ◽  
Dmitry Grinevich ◽  
Siddhartha Thakur ◽  
Michael A. Balamotis ◽  
Tuval Ben Yehezkel
Keyword(s):  
Microbial Community ◽  
16S Rrna ◽  
Amplicon Sequencing ◽  
Species Level ◽  
Full Length ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Strain Identification ◽  
Long Reads ◽  
Long Read

Abstract Background Out of the many pathogenic bacterial species that are known, only a fraction are readily identifiable directly from a complex microbial community using standard next generation DNA sequencing. Long-read sequencing offers the potential to identify a wider range of species and to differentiate between strains within a species, but attaining sufficient accuracy in complex metagenomes remains a challenge. Methods Here, we describe and analytically validate LoopSeq, a commercially available synthetic long-read (SLR) sequencing technology that generates highly accurate long reads from standard short reads. Results LoopSeq reads are sufficiently long and accurate to identify microbial genes and species directly from complex samples. LoopSeq perfectly recovered the full diversity of 16S rRNA genes from known strains in a synthetic microbial community. Full-length LoopSeq reads had a per-base error rate of 0.005%, which exceeds the accuracy reported for other long-read sequencing technologies. 18S-ITS and genomic sequencing of fungal and bacterial isolates confirmed that LoopSeq sequencing maintains that accuracy for reads up to 6 kb in length. LoopSeq full-length 16S rRNA reads could accurately classify organisms down to the species level in rinsate from retail meat samples, and could differentiate strains within species identified by the CDC as potential foodborne pathogens. Conclusions The order-of-magnitude improvement in length and accuracy over standard Illumina amplicon sequencing achieved with LoopSeq enables accurate species-level and strain identification from complex- to low-biomass microbiome samples. The ability to generate accurate and long microbiome sequencing reads using standard short read sequencers will accelerate the building of quality microbial sequence databases and removes a significant hurdle on the path to precision microbial genomics.

scholarly journals Connecting structure to function with the recovery of over 1000 high-quality metagenome-assembled genomes from activated sludge using long-read sequencing

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Caitlin M. Singleton ◽  
Francesca Petriglieri ◽  
Jannie M. Kristensen ◽  
Rasmus H. Kirkegaard ◽  
Thomas Y. Michaelsen ◽  
...  
Keyword(s):  
16S Rrna ◽  
Wastewater Treatment Plants ◽  
In Situ Hybridisation ◽  
Amplicon Sequencing ◽  
Rrna Genes ◽  
Fluorescence In Situ Hybridisation ◽  
Sequencing Data ◽  
High Quality ◽  
16S Rrna Amplicon Sequencing ◽  
Long Read

AbstractMicroorganisms play crucial roles in water recycling, pollution removal and resource recovery in the wastewater industry. The structure of these microbial communities is increasingly understood based on 16S rRNA amplicon sequencing data. However, such data cannot be linked to functional potential in the absence of high-quality metagenome-assembled genomes (MAGs) for nearly all species. Here, we use long-read and short-read sequencing to recover 1083 high-quality MAGs, including 57 closed circular genomes, from 23 Danish full-scale wastewater treatment plants. The MAGs account for ~30% of the community based on relative abundance, and meet the stringent MIMAG high-quality draft requirements including full-length rRNA genes. We use the information provided by these MAGs in combination with >13 years of 16S rRNA amplicon sequencing data, as well as Raman microspectroscopy and fluorescence in situ hybridisation, to uncover abundant undescribed lineages belonging to important functional groups.

scholarly journals Microdiversity and phylogeographic diversification of bacterioplankton in pelagic freshwater systems revealed through long-read amplicon sequencing

Microbiome ◽  
2021 ◽  
Vol 9 (1) ◽  
Author(s):  
Yusuke Okazaki ◽  
Shohei Fujinaga ◽  
Michaela M. Salcher ◽  
Cristiana Callieri ◽  
Atsushi Tanaka ◽  
...  
Keyword(s):  
16S Rrna ◽  
Regional Scale ◽  
Scale Up ◽  
Amplicon Sequencing ◽  
Freshwater Ecosystems ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Metagenomic Sequencing ◽  
Long Read

Abstract Background Freshwater ecosystems are inhabited by members of cosmopolitan bacterioplankton lineages despite the disconnected nature of these habitats. The lineages are delineated based on > 97% 16S rRNA gene sequence similarity, but their intra-lineage microdiversity and phylogeography, which are key to understanding the eco-evolutional processes behind their ubiquity, remain unresolved. Here, we applied long-read amplicon sequencing targeting nearly full-length 16S rRNA genes and the adjacent ribosomal internal transcribed spacer sequences to reveal the intra-lineage diversities of pelagic bacterioplankton assemblages in 11 deep freshwater lakes in Japan and Europe. Results Our single nucleotide-resolved analysis, which was validated using shotgun metagenomic sequencing, uncovered 7–101 amplicon sequence variants for each of the 11 predominant bacterial lineages and demonstrated sympatric, allopatric, and temporal microdiversities that could not be resolved through conventional approaches. Clusters of samples with similar intra-lineage population compositions were identified, which consistently supported genetic isolation between Japan and Europe. At a regional scale (up to hundreds of kilometers), dispersal between lakes was unlikely to be a limiting factor, and environmental factors or genetic drift were potential determinants of population composition. The extent of microdiversification varied among lineages, suggesting that highly diversified lineages (e.g., Iluma-A2 and acI-A1) achieve their ubiquity by containing a consortium of genotypes specific to each habitat, while less diversified lineages (e.g., CL500-11) may be ubiquitous due to a small number of widespread genotypes. The lowest extent of intra-lineage diversification was observed among the dominant hypolimnion-specific lineage (CL500-11), suggesting that their dispersal among lakes is not limited despite the hypolimnion being a more isolated habitat than the epilimnion. Conclusions Our novel approach complemented the limited resolution of short-read amplicon sequencing and limited sensitivity of the metagenome assembly-based approach, and highlighted the complex ecological processes underlying the ubiquity of freshwater bacterioplankton lineages. To fully exploit the performance of the method, its relatively low read throughput is the major bottleneck to be overcome in the future.

scholarly journals Long-Read Amplicon Sequencing of Nitric Oxide Dismutase (nod) Genes Reveal Diverse Oxygenic Denitrifiers in Agricultural Soils and Lake Sediments

2020 ◽  
Vol 80 (1) ◽  
pp. 243-247 ◽  
Author(s):  
Baoli Zhu ◽  
Zhe Wang ◽  
Dheeraj Kanaparthi ◽  
Susanne Kublik ◽  
Tida Ge ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Lake Sediments ◽  
Agricultural Soils ◽  
Amplicon Sequencing ◽  
Nod Genes ◽  
Long Read

Impacts of switching tillage to no-tillage and vice versa on soil structure, enzyme activities, and prokaryotic community profiles in Argentinean semi-arid soils

2021 ◽  
Author(s):  
L A Gabbarini ◽  
E Figuerola ◽  
J P Frene ◽  
N B Robledo ◽  
F M Ibarbalz ◽  
...  
Keyword(s):  
Community Structure ◽  
Soil Structure ◽  
Management Practices ◽  
Soil Health ◽  
Amplicon Sequencing ◽  
16S Rrna Genes ◽  
Rrna Genes ◽  
Prokaryotic Community ◽  
C Cycle

Abstract The effects of tillage on soil structure, physiology, and microbiota structure were studied in a long-term field experiment, with side-to-side plots, established to compare effects of conventional tillage (CT) vs. no-till (NT) agriculture. After 27 years, part of the field under CT was switched to NT and vice versa. Soil texture, soil enzymatic profiles, and the prokaryotic community structure (16S rRNA genes amplicon sequencing) were analysed at two soil depths (0–5, 5–10 cm) in samples taken 6, 18, and 30 months after switching tillage practices. Soil enzymatic activities were higher in NT than CT, and enzymatic profiles responded to the changes much earlier than the overall prokaryotic community structure. Beta diversity measurements of the prokaryotic community indicated that the levels of stratification observed in long-term NT soils were already recovered in the new NT soils thirty months after switching from CT to NT. Bacteria and Archaea OTUs, which responded to NT were associated with coarse soil fraction, SOC and C cycle enzymes while CT responders were related to fine soil fractions and S cycle enzymes. This study showed the potential of managing the soil prokaryotic community and soil health through changes in agricultural management practices.

scholarly journals High-throughput amplicon sequencing reveals the spatiotemporal effects, abiotic and biotic shaping factors for the microbial communities in tropical mangrove sediments in Sanya, China

2020 ◽  
Author(s):  
Wu Qu ◽  
Boliang Gao ◽  
Jie Wu ◽  
Min Jin ◽  
Jianxin Wang ◽  
...  
Keyword(s):  
Community Structure ◽  
Microbial Community ◽  
Microbial Communities ◽  
Fungal Community ◽  
Amplicon Sequencing ◽  
Biotic Factors ◽  
Community Structures ◽  
Mangrove Sediments ◽  
Element Cycling ◽  
Microbial Community Structures

Abstract Background Microbial roles in element cycling and nutrient providing are crucial for mangrove ecosystems and serve as important regulators for climate change in Earth ecosystem. However, some key information about the spatiotemporal influences and abiotic and biotic shaping factors for the microbial communities in mangrove sediments remains lacking. Methods In this work, 22 sediment samples were collected from multiple spatiotemporal dimensions, including three locations, two depths, and four seasons, and the bacterial, archaeal, and fungal community structures in these samples were studied using amplicon sequencing. Results The microbial community structures were varied in the samples from different depths and locations based on the results of LDA effect size analysis, principal coordinate analysis, the analysis of similarities, and permutational multivariate ANOVA. However, these microbial community structures were stable among the seasonal samples. Linear fitting models and Mantel test showed that among the 13 environmental factors measured in this study, the sediment particle size (PS) was the key abiotic shaping factor for the bacterial, archaeal, or fungal community structure. Besides PS, salinity and humidity were also significant impact factors according to the canonical correlation analysis (p ≤ 0.05). Co-occurrence networks demonstrated that the bacteria assigned into phyla Ignavibacteriae, Proteobacteria, Bacteroidetes, Chloroflexi, and Acidobacteria were the key biotic factors for shaping the bacterial community in mangrove sediments. Conclusions This work showed the variability on spatial dimensions and the stability on temporal dimension for the bacterial, archaeal, or fungal microbial community structure, indicating that the tropical mangrove sediments are versatile but stable environments. PS served as the key abiotic factor could indirectly participate in material circulation in mangroves by influencing microbial community structures, along with salinity and humidity. The bacteria as key biotic factors were found with the abilities of photosynthesis, polysaccharide degradation, or nitrogen fixation, which were potential indicators for monitoring mangrove health, as well as crucial participants in the storage of mangrove blue carbons and mitigation of climate warming. This study expanded the knowledge of mangroves for the spatiotemporal variation, distribution, and regulation of the microbial community structures, thus further elucidating the microbial roles in mangrove management and climate regulation.

scholarly journals Gut microbiota composition is associated with narcolepsy type 1

2020 ◽  
Vol 7 (6) ◽  
pp. e896
Author(s):  
Alexandre Lecomte ◽  
Lucie Barateau ◽  
Pedro Pereira ◽  
Lars Paulin ◽  
Petri Auvinen ◽  
...  
Keyword(s):  
Community Structure ◽  
Gut Microbiota ◽  
Bacterial Communities ◽  
Beta Diversity ◽  
Alpha Diversity ◽  
Amplicon Sequencing ◽  
Rrna Gene ◽  
Differential Abundance ◽  
Alpha And Beta Diversity

ObjectiveTo test the hypothesis that narcolepsy type 1 (NT1) is related to the gut microbiota, we compared the microbiota bacterial communities of patients with NT1 and control subjects.MethodsThirty-five patients with NT1 (51.43% women, mean age 38.29 ± 19.98 years) and 41 controls (57.14% women, mean age 36.14 ± 12.68 years) were included. Stool samples were collected, and the fecal microbiota bacterial communities were compared between patients and controls using the well-standardized 16S rRNA gene amplicon sequencing approach. We studied alpha and beta diversity and differential abundance analysis between patients and controls, and between subgroups of patients with NT1.ResultsWe found no between-group differences for alpha diversity, but we discovered in NT1 a link with NT1 disease duration. We highlighted differences in the global bacterial community structure as assessed by beta diversity metrics even after adjustments for potential confounders as body mass index (BMI), often increased in NT1. Our results revealed differential abundance of several operational taxonomic units within Bacteroidetes, Bacteroides, and Flavonifractor between patients and controls, but not after adjusting for BMI.ConclusionWe provide evidence of gut microbial community structure alterations in NT1. However, further larger and longitudinal multiomics studies are required to replicate and elucidate the relationship between the gut microbiota, immunity dysregulation and NT1.

Using PacBio Long-Read High-Throughput Microbial Gene Amplicon Sequencing To Evaluate Infant Formula Safety

2016 ◽  
Vol 64 (37) ◽  
pp. 6993-7001 ◽  
Author(s):  
Yi Zheng ◽  
Xiaoxia Xi ◽  
Haiyan Xu ◽  
Qiangchuan Hou ◽  
Yanfei Bian ◽  
...  
Keyword(s):  
Infant Formula ◽  
High Throughput ◽  
Amplicon Sequencing ◽  
Long Read

Harnessing long-read amplicon sequencing to uncover NRPS and Type I PKS gene sequence diversity in polar desert soils

2019 ◽  
Vol 95 (4) ◽  
Author(s):  
Nicole Benaud ◽  
Eden Zhang ◽  
Josie van Dorst ◽  
Mark V Brown ◽  
John A Kalaitzis ◽  
...  
Keyword(s):  
Gene Sequence ◽  
Amplicon Sequencing ◽  
Sequence Diversity ◽  
Type I ◽  
Desert Soils ◽  
Polar Desert ◽  
Long Read

scholarly journals Rapid and real-time identification of fungi up to the species level with long amplicon Nanopore sequencing from clinical samples

2020 ◽  
Author(s):  
Sara D’Andreano ◽  
Anna Cuscó ◽  
Olga Francino
Keyword(s):  
Real Time ◽  
Clinical Signs ◽  
Amplicon Sequencing ◽  
Species Level ◽  
Clinical Samples ◽  
Microsporum Canis ◽  
Mock Community ◽  
Malassezia Pachydermatis ◽  
Long Read ◽  
Real Time Identification

ABSTRACTThe availability of long-read technologies, like Oxford Nanopore Technologies, provides the opportunity to sequence longer fragments of the fungal ribosomal operon, up to 6 Kb (18S-ITS1-5.8S-ITS2-28S), and to improve the taxonomy assignment of the communities up to the species level and in real-time. We assess the taxonomy skills of amplicons targeting a 3.5 Kb region (V3 18S-ITS1-5.8S-ITS2-28S D2) and a 6 Kb region (V1 18S-ITS1-5.8S-ITS2-28S D12) with the What’s in my pot (WIMP) classifier. We used the ZymoBIOMICS™ mock community and different microbiological fungal cultures as positive controls. Long amplicon sequencing correctly identified Saccharomyces cerevisiae and Cryptococcus neoformans from the mock community and Malassezia pachydermatis, Microsporum canis, and Aspergillus fumigatus from the microbiological cultures. Besides, we identified Rhodotorula graminis in a culture mislabeled as Candida spp.We applied the same approach to external otitis in dogs. Malassezia was the dominant fungal genus in dogs’ ear skin, whereas M. pachydermatis was the main species in the healthy sample. Conversely, we identified a higher representation of M. globosa and M. sympodialis in otitis affected samples. We demonstrate the suitability of long ribosomal amplicons to characterize the fungal community of complex samples, either healthy or with clinical signs of infection.

Sign in / Sign up

Export Citation Format

Share Document