Predicting provenance of forensic soil samples: Linking soil to ecological habitats by metabarcoding and supervised classification

Fløjgaard, Camilla; Frøslev, Tobias Guldberg; Brunbjerg, Ane Kirstine; Bruun, Hans Henrik; Moeslund, Jesper Erenskjold; Hansen, Anders J.; Ejrnæs, Rasmus

doi:10.1371/journal.pone.0202844

Cited by 41 publications

(31 citation statements)

References 39 publications

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“…The preparation of the fungal (ITS2) and eukaryote (18S) eDNA datasets have been published in Fløjgaard et al () and Frøslev et al () respectively. The arthropod DNA dataset was produced by extracting DNA from the ethanol from the bulk insect Malaise traps and metabarcoding with the arthropod specific COI primers ZBJ‐ArtF1c and ZBJ‐ArtR2c (Zeale et al ).…”

Section: Methodsmentioning

confidence: 99%

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Multi‐taxon inventory reveals highly consistent biodiversity responses to ecospace variation

Brunbjerg

Bruun

Dalby

et al. 2020

Oikos

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Amidst the global biodiversity crisis, identifying general principles for variation of biodiversity remains a key challenge. Scientific consensus is limited to a few macroecological rules, such as species richness increasing with area, which provide limited guidance for conservation. In fact, few agreed ecological principles apply at the scale of sites or reserve management, partly because most community‐level studies are restricted to single habitat types and species groups. We used the recently proposed ecospace framework and a comprehensive data set for aggregating environmental variation to predict multi‐taxon diversity. We studied richness of plants, fungi and arthropods in 130 sites representing the major terrestrial habitat types in Denmark. We found the abiotic environment (ecospace position) to be pivotal for the richness of primary producers (vascular plants, mosses and lichens) and, more surprisingly, little support for ecospace continuity as a driver. A peak in richness at intermediate productivity adds new empirical evidence to a long‐standing debate over biodiversity responses to productivity. Finally, we discovered a dominant and positive response of fungi and insect richness to organic matter accumulation and diversification (ecospace expansion). Two simple models of producer and consumer richness accounted for 77% of the variation in multi‐taxon species richness suggesting a significant potential for generalization beyond individual species responses. Our study widens the traditional conservation focus on vegetation and vertebrate populations unravelling the importance of diversification of carbon resources for diverse heterotrophs, such as fungi and insects.

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Section: Methodsmentioning

confidence: 99%

“…1.8) (Callahan et al ) to identify exact amplicon sequence variants (ESVs, see Callahan et al ) including removal of chimeras and post‐clustering curation using LULU (Frøslev et al ). Taxonomic assignment was done with a custom script (as in Fløjgaard et al ). OTUs not assigned to Arthropoda were discarded before further analyses.…”

Section: Methodsmentioning

confidence: 99%

Multi‐taxon inventory reveals highly consistent biodiversity responses to ecospace variation

Brunbjerg

Bruun

Dalby

et al. 2020

Oikos

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“…The preparation of the fungal and eukaryote eDNA datasets have been published in Frøslev, et al (44) and Fløjgaard, et al (45) respectively. The insect DNA dataset was produced by extracting DNA from the ethanol from the bulk insect Malaise traps and metabarcoding with an insect specific 16S primer.…”

Section: Methodsmentioning

confidence: 99%

“…The dried pellet was extracted with the Qiagen DNeasy blood and tissue kit (Qiagen, Germany) with minor modifications. The extracted DNA was normalized, amplified, sequenced and analyzed according to the overall procedures described for 16S insects in Fløjgaard, et al (45), but including curation of OTUs with LULU (26) and taxonomic assignment with a custom script, and exclusion of non-arthropod sequences. Data from the two different collecting events were handled separately and the sequences were then combined for each site.…”

Section: Methodsmentioning

confidence: 99%

Multi-taxon inventory reveals highly consistent biodiversity responses to ecospace variation

Brunbjerg

Bruun

Dalby

et al. 2019

Preprint

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31Amidst the global biodiversity crisis, identifying drivers of biodiversity variation remains a key 32 challenge. Scientific consensus is limited to a few macroecological rules, such as species richness 33 increasing with area, which provide limited guidance for conservation. In fact, few agreed ecological 34 principles apply at the scale of sites or reserve management, partly because most community-level 35 studies are restricted to single habitat types and species groups. We used the recently proposed 36 ecospace framework and a comprehensive data set for aggregating environmental variation to 37 predict multi-taxon diversity. We studied richness of plants, fungi, and arthropods in 130 sites 38 representing the major terrestrial habitat types in Denmark. We found the abiotic environment 39 (ecospace position) to be pivotal for the richness of primary producers (vascular plants, mosses, and 40 lichens) and, more surprisingly, little support for ecospace continuity as a driver. A peak in richness 41 at intermediate productivity adds new empirical evidence to a long-standing debate over 42 biodiversity responses to productivity. Finally, we discovered a dominant and positive response of 43 fungi and insect richness to organic matter accumulation and diversification (ecospace expansion). 44Two simple models of producer and consumer richness accounted for 77 % of the variation in multi-45 taxon species richness suggesting a significant potential for generalization beyond individual species 46 responses. Our study widens the traditional conservation focus on vegetation and vertebrate 47 populations unravelling the importance of diversification of carbon resources for diverse 48 heterotrophs, such as fungi and insects. 49 50

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“…This result indicates that the number of unique sequences output from DADA2 cannot be used to infer experimental success, with respect to sequence quality or target specificity. In the limited previous DNA metabarcoding studies that targeted ITS2 for plant characterization from soil, larger numbers of target sequences per sample were recovered (e.g.,~10,000 sequences/sample) [4,23]. Two main differences in methodology can explain the increased recovery in those studies: 1) soil was stored at -80˚C prior to subsampling to preserve the biological material [4], and 2) either multiple DNA extractions were completed per sample (with the resulting DNA pooled to increase input for PCR; [4]), or a single extraction was completed using a large quantity of soil as input (4 gm; [23]).…”

Section: Barcode Sequence and Taxonomic Group Recoverymentioning

confidence: 99%

Optimization of the second internal transcribed spacer (ITS2) for characterizing land plants from soil

2020

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Molecular-based taxonomy, specifically DNA barcoding, has streamlined organism identification. For land plants, the recommended 2-locus barcode of rbcL and matK is not suitable for all groups, thus the second subunit of the nuclear internal transcribed spacer (ITS2) has received attention as a possible alternative. To date, evaluations of ITS2 have mostly been limited in scope to specific plant orders/families and single source material. Prior to using ITS2 to routinely characterize land plants present in environmental samples (i.e., DNA metabarcoding), a wet lab protocol optimized for bulk sample types is needed. To address this gap, in this study we determined the broad recoverability across land plants when using published ITS2 primer pairs, and subsequently optimized the PCR reaction constituents and cycling conditions for the best two performing primer pairs (ITS2F/ITSp4 and ITSp3/ITSu4). Using these conditions, both primer pairs were used to characterize land plants present in 17 diverse soils collected from across the US. The resulting PCR amplicons were prepared into libraries and pooled for sequencing on an Illumina ® MiniSeq. Our existing bioinformatics workflow was used to process raw sequencing data and taxonomically assign unique ITS2 plant sequences by comparison to GenBank. Given strict quality criteria were imposed on sequences for inclusion in data analysis, only 43.6% and 7.5% of sequences from ITS2F/ITSp4 and ITSp3/ITSu4 respectively remained for taxonomic comparisons;~7-11% of sequences originated from fungal co-amplification. The number of orders and families recovered did differ between primer pairs, with ITS2F/ITSp4 consistently outperforming ITSp3/ITSu4 by >15%. Primer pair bias was observed in the recovery of certain taxonomic groups; ITS2F/ITSp4 preferentially recovered flowering plants and grasses, whereas ITSp3/ITSu4 recovered more moss taxa. To maximize data recovery and reduce potential bias, we advocate that studies using ITS2 to characterize land plants from environmental samples such as soil use a multiple primer pair approach.

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Predicting provenance of forensic soil samples: Linking soil to ecological habitats by metabarcoding and supervised classification

Cited by 41 publications

References 39 publications

Multi‐taxon inventory reveals highly consistent biodiversity responses to ecospace variation

Multi‐taxon inventory reveals highly consistent biodiversity responses to ecospace variation

Multi-taxon inventory reveals highly consistent biodiversity responses to ecospace variation

Optimization of the second internal transcribed spacer (ITS2) for characterizing land plants from soil

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