Contaminations in (meta)genome data: An open issue for the scientific community

G, De Simone; Pasquadibisceglie, Andrea; Proietto, Roberta; Polticelli, Fabio; Aime, Silvio; Camp, Huub J. M. Op den; Ascenzi, Paolo

doi:10.1002/iub.2216

Cited by 19 publications

(15 citation statements)

References 76 publications

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“…Importantly, this is a recurring problem which is completely ignored by most metagenomic analysis tools. We continually find new errors in all new metagenomic references, indicating that this problem is also growing (De Simone et al 2020).…”

Section: Resultsmentioning

confidence: 99%

Wochenende - modular and flexible alignment-based shotgun metagenome analysis

Rosenboom

Friedrich

Poertner

et al. 2022

Preprint

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Shotgun metagenome analysis provides a robust and verifiable method for comprehensive microbiome analysis of fungal, viral, archaeal and bacterial taxonomy, particularly with regard to visualization of read mapping location, normalization options, growth dynamics and functional gene repertoires. Current read classification tools use non-standard output formats, or do not fully show information on mapping location. As reference datasets are not perfect, portrayal of mapping information is critical for judging results effectively. Our alignment-based pipeline, Wochenende, incorporates flexible quality control, trimming, mapping, various filters and normalization. We observe stringent filtering of mismatches and use of mapping quality sharply reduces the number of false positives. Further modules allow genomic visualization, as well as integration and subsequent plotting of pipeline results. Our novel normalization approach additionally allows calculation of absolute abundance profiles by comparison with reads assigned to the human host genome. Wochenende has the ability to find and filter alignments to all kingdoms of life using both short and long reads, and requires only good quality reference genomes. Wochenende is available at https://github.com/MHH-RCUG/Wochenende.

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Section: Resultsmentioning

confidence: 99%

Wochenende - modular and flexible alignment-based shotgun metagenome analysis

Rosenboom

Friedrich

Poertner

et al. 2022

Preprint

View full text Add to dashboard Cite

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“…Furthermore, dark matter seems to be particularly common in eDNA as compared to bulk samples (Andújar et al 2018). However, it should be mentioned that the high number of prokaryotic sequences in COI metabarcoding data is also reflecting known issues with contamination (Kumar et al 2013;Dittami and Corre 2017;De Simone et al 2020), incorrectly labeled reference sequences (Steinegger and Salzberg 2020) and holobionts (Gilbert et al 2012;Salvucci 2016) in eukaryotic genomes.…”

Section: Discussionmentioning

confidence: 99%

The Dark mAtteR iNvestigator (DARN) tool: getting to know the known unknowns in COI amplicon data

Zafeiropoulos¹,

Gargan²,

Hintikka³

et al. 2021

MBMG

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The mitochondrial cytochrome C oxidase subunit I gene (COI) is commonly used in environmental DNA (eDNA) metabarcoding studies, especially for assessing metazoan diversity. Yet, a great number of COI operational taxonomic units (OTUs) or/and amplicon sequence variants (ASVs) retrieved from such studies do not get a taxonomic assignment with a reference sequence. To assess and investigate such sequences, we have developed the Dark mAtteR iNvestigator (DARN) software tool. For this purpose, a reference COI-oriented phylogenetic tree was built from 1,593 consensus sequences covering all the three domains of life. With respect to eukaryotes, consensus sequences at the family level were constructed from 183,330 sequences retrieved from the Midori reference 2 database, which represented 70% of the initial number of reference sequences. Similarly, sequences from 431 bacterial and 15 archaeal taxa at the family level (29% and 1% of the initial number of reference sequences respectively) were retrieved from the BOLD and the PFam databases. DARN makes use of this phylogenetic tree to investigate COI pre-processed sequences of amplicon samples to provide both a tabular and a graphical overview of their phylogenetic assignments. To evaluate DARN, both environmental and bulk metabarcoding samples from different aquatic environments using various primer sets were analysed. We demonstrate that a large proportion of non-target prokaryotic organisms, such as bacteria and archaea, are also amplified in eDNA samples and we suggest prokaryotic COI sequences to be included in the reference databases used for the taxonomy assignment to allow for further analyses of dark matter. DARN source code is available on GitHub at https://github.com/hariszaf/darn and as a Docker image at https://hub.docker.com/r/hariszaf/darn.

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“…However, as noted previously, these studies are once again hindered by the lack of diversity in modern microbiome studies [30,49]. With mostly modern, industrialized pets represented in studies, our definition of what constitutes a 'normal' canine microbiome is skewed, which limits our ability to successfully discriminate between ancient contaminants and ancient microbes of interest [59,60]. The dogs represented in these ancient studies are frequently human-associated, postdomestication; but their lifestyles, environment and diet mirror none of the dogs studied today [61][62][63][64][65][66].…”

Section: (B) Canine Microbiomes Over Timementioning

confidence: 99%

Geographically diverse canid sampling provides novel insights into pre-industrial microbiomes

et al. 2022

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Canine microbiome studies are often limited in the geographic and temporal scope of samples studied. This results in a paucity of data on the canine microbiome around the world, especially in contexts where dogs may not be pets or human associated. Here, we present the shotgun sequences of fecal microbiomes of pet dogs from South Africa, shelter and stray dogs from India, and stray village dogs in Laos. We additionally performed a dietary experiment with dogs housed in a veterinary medical school, attempting to replicate the diet of the sampled dogs from Laos. We analyse the taxonomic diversity in these populations and identify the underlying functional redundancy of these microbiomes. Our results show that diet alone is not sufficient to recapitulate the higher diversity seen in the microbiome of dogs from Laos. Comparisons to previous studies and ancient dog fecal microbiomes highlight the need for greater population diversity in studies of canine microbiomes, as modern analogues can provide better comparisons to ancient microbiomes. We identify trends in microbial diversity and industrialization in dogs that mirror results of human studies, suggesting future research can make use of these companion animals as substitutes for humans in studying the effects of industrialization on the microbiome.

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Contaminations in (meta)genome data: An open issue for the scientific community

Cited by 19 publications

References 76 publications

Wochenende - modular and flexible alignment-based shotgun metagenome analysis

Wochenende - modular and flexible alignment-based shotgun metagenome analysis

The Dark mAtteR iNvestigator (DARN) tool: getting to know the known unknowns in COI amplicon data

Geographically diverse canid sampling provides novel insights into pre-industrial microbiomes

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