Annotated bacterial chromosomes from frame-shift-corrected long read metagenomic data

Arumugam, Krithika; Bağcı, Caner; Bessarab, Irina; Beier, Sina; Buchfink, Benjamin; Górska, Anna; Qiu, Guofu; Huson, Daniel H.; Williams, Rohan B.H.

doi:10.1101/511683

Cited by 11 publications

(45 citation statements)

References 26 publications

(10 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…New long read analysis methods 13,14 and binning algorithms designed for long read metagenome data 15 have also appeared, anticipating the future expansion of metagenome data generated from these new instruments. More recent studies 12,[16][17][18][19][20][21][22][23] have collectively established that full length (or near-full length genomes) can be recovered from long read sequencing of complex communities, which sets the stage for further development of genome-resolved long read metagenomics.…”

Section: Introductionmentioning

confidence: 99%

Recovery of complete genomes and non-chromosomal replicons from activated sludge enrichment microbial communities with long read metagenome sequencing

Arumugam

Bessarab

Haryono

et al. 2021

npj Biofilms Microbiomes

Self Cite

View full text Add to dashboard Cite

New long read sequencing technologies offer huge potential for effective recovery of complete, closed genomes from complex microbial communities. Using long read data (ONT MinION) obtained from an ensemble of activated sludge enrichment bioreactors we recover 22 closed or complete genomes of community members, including several species known to play key functional roles in wastewater bioprocesses, specifically microbes known to exhibit the polyphosphate- and glycogen-accumulating organism phenotypes (namely Candidatus Accumulibacter and Dechloromonas, and Micropruina, Defluviicoccus and Candidatus Contendobacter, respectively), and filamentous bacteria (Thiothrix) associated with the formation and stability of activated sludge flocs. Additionally we demonstrate the recovery of close to 100 circularised plasmids, phages and small microbial genomes from these microbial communities using long read assembled sequence. We describe methods for validating long read assembled genomes using their counterpart short read metagenome-assembled genomes, and assess the influence of different correction procedures on genome quality and predicted gene quality. Our findings establish the feasibility of performing long read metagenome-assembled genome recovery for both chromosomal and non-chromosomal replicons, and demonstrate the value of parallel sampling of moderately complex enrichment communities to obtaining high quality reference genomes of key functional species relevant for wastewater bioprocesses.

show abstract

Section: Introductionmentioning

confidence: 99%

Recovery of complete genomes and non-chromosomal replicons from activated sludge enrichment microbial communities with long read metagenome sequencing

Arumugam

Bessarab

Haryono

et al. 2021

npj Biofilms Microbiomes

Self Cite

View full text Add to dashboard Cite

show abstract

“…In our case, we applied Guppy for basecalling, Canu for raw reads correction, and Medaka for polishing the assembled metagenomes. To ensure reducing the insertion and deletion error type, we further applied a frameshift-aware correction step [16] that improved the completeness and reduced the CDS number. Long-read metagenomics sequencing can be harnessed to build comprehensive and curated databases for in-depth characterization of novel bacterial diversity in the canine fecal microbiome or any other underexplored environment.…”

Section: Discussionmentioning

confidence: 99%

“…The next step for the HQ MAGs was to correct the frameshift errors, as described in [16], using Diamond 0.9.32 [28] and MEGAN-LR 6.19.1 [29]. We used ideel [30] to visualize the number of truncated openreading frames (ORF).…”

Section: Metagenomics Assembly and Polishingmentioning

confidence: 99%

“…This strategy was applied to assess the human GI microbiome [11], among others -such as mock communities [12], cow rumen [13], natural whey starter cultures [14], or wastewater [15]. Worthy of considering, some authors suggest that we may overcome the need for short reads to polish long-read data by either using correction software, such as frameshiftaware correction [16], or with ultra-deep coverage of the genomes [12].…”

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Long-read metagenomics retrieve complete single-contig bacterial genomes from canine feces

Cuscó¹,

Pérez

Viñes³

et al. 2021

Preprint

View full text Add to dashboard Cite

BackgroundLong-read sequencing in metagenomics facilitates the assembly of complete genomes out of complex microbial communities. These genomes include essential biologic information such as the ribosomal genes or the mobile genetic elements, which are usually missed with short-reads. We applied long-read metagenomics with Nanopore sequencing to retrieve high-quality metagenome-assembled genomes (HQ MAGs) from a dog fecal sample.ResultsWe used nanopore long-read metagenomics and frameshift aware correction on a canine fecal sample and retrieved eight single-contig HQ MAGs, which were > 90% complete with < 5% contamination, and contained most ribosomal genes and tRNAs. At the technical level, we demonstrated that a high-molecular-weight DNA extraction improved the metagenomics assembly contiguity, the recovery of the rRNA operons, and the retrieval of longer and circular contigs that are potential HQ MAGs. These HQ MAGs corresponded to Succinivibrio, Sutterella, Prevotellamassilia, Phascolarctobacterium, Catenibacterium, Blautia, and Enterococcus genera. Linking our results to previous gastrointestinal microbiome reports (metagenome or 16S rRNA-based), we found that some bacterial species on the gastrointestinal tract seem to be more canid-specific –Succinivibrio, Prevotellamassilia, Phascolarctobacterium, Blautia_A sp900541345–, whereas others are more broadly distributed among animal and human microbiomes –Sutterella, Catenibacterium, Enterococcus, and Blautia sp003287895. Sutterella HQ MAG is potentially the first reported genome assembly for Sutterella stercoricanis, as assigned by 16S rRNA gene similarity. Moreover, we show that long reads are essential to gain biological insights that are otherwise missed in short-read MAGs catalogs, as shown by the mobilome functions detected in the long-read HQ MAGs.ConclusionsWe recovered eight single-contig HQ MAGs from canine feces of a healthy dog with nanopore long-reads. We also retrieved relevant biological insights from these specific bacterial species previously missed in public databases, such as complete ribosomal operons and mobilome functions. The high-molecular-weight DNA extraction improved the assembly's contiguity, whereas the high-accuracy basecalling, the raw read error correction, the assembly polishing, and the frameshift correction reduced the insertions and deletion errors. Both experimental and analytical steps ensured the retrieval of complete bacterial genomes.

show abstract

“…For S. citri strain C5, contigs were assembled using Canu 1.8. An additional polishing step was performed by medaka v 1.0.3 (Oxford Nanopore Technologies) and frame-shift-corrected by DIAMOND v 0.9.28 [56] and MEGAN v 6.18.4 [57], following the pipeline described by Arumugam et al [58]. Approximately ~500 bp segments from each end of a contig were used for BLASTn search to check the contig singularity.…”

Section: De Novo Sequencing Assemblymentioning

confidence: 99%

Genome analysis of Spiroplasma citri strains from different host plants and its leafhopper vectors

Rattner

Thapa

Dang

et al. 2020

Preprint

View full text Add to dashboard Cite

Background Spiroplasma citri comprises a complex of bacteria that cause diseases in citrus, horseradish, carrot, sesame, and also infect a wide array of ornamental and weed species. S. citri is transmitted in a persistent propagative manner by the beet leafhopper, Neoaliturus tenellus in North America and Circulifer haematoceps in the Mediterranean region. Leafhopper transmission and the pathogen’s wide host range serve as drivers of genetic diversity. This diversity was examined by comparing the genome sequences of seven S. citri strains from the United States (BR12, CC2, C5, C189, LB319, BLH-13, and BLH-MB) collected from different hosts and times with other publicly available spiroplasmas.Results Phylogenetic analysis using 16S rRNA sequences from 39 spiroplasmas obtained from NCBI database showed that S. citri strains, along with S. kunkelii and S. phoeniceum, two other plant pathogenic spiroplasmas, formed a monophyletic group. To refine genetic relationships among S. citri strains, phylogenetic analyses with 863 core orthologous sequences were performed. Strains that clustered together were: CC-12 and C5; C189 and R8-A2; BR12, BLH-MB, BLH13 and LB 319. Strain GII3-3 remained in a separate branch. Sequence rearrangements were observed among S. citri strains, predominantly in the center of the chromosome. One to nine plasmids were identified in the seven S. citri strains analyzed in this study. Plasmids were most abundant in strains isolated from the beet leafhopper, followed by strains from carrot, Chinese cabbage, horseradish and citrus, respectively. All these S. citri strains contained one plasmid with high similarity to plasmid pSci6 from S. citri strain GII3-3X which is known to confer insect transmissibility. Additionally, 17 to 25 prophage-like elements were identified in these genomes, which may promote rearrangements and contribute to repetitive regions. Conclusions The genome of seven S. citri strains were found to contain a single circularized chromosome, ranging from 1.58 Mbp to 1.74 Mbp and 1,597-2,232 protein-coding genes. These strains possessed a plasmid similar to pSci6 from the GII3-3X strain associated with leafhopper transmission. Prophage sequences found in the S. citri genomes may contribute to the extension of its host range. These findings increase our understanding of S. citri genetic diversity.

show abstract

Annotated bacterial chromosomes from frame-shift-corrected long read metagenomic data

Cited by 11 publications

References 26 publications

Recovery of complete genomes and non-chromosomal replicons from activated sludge enrichment microbial communities with long read metagenome sequencing

Recovery of complete genomes and non-chromosomal replicons from activated sludge enrichment microbial communities with long read metagenome sequencing

Long-read metagenomics retrieve complete single-contig bacterial genomes from canine feces

Genome analysis of Spiroplasma citri strains from different host plants and its leafhopper vectors

Contact Info

Product

Resources

About