Single Fragment or Bulk Soil DNA Metabarcoding: Which is Better for Characterizing Biological Taxa Found in Surface Soils for Sample Separation?

Abstract

In forensic geology casework, sample size typically limits routine characterization of material using bulk approaches. To address this, DNA-based characterization of biological taxa has received attention, as the taxa present can be useful for sample-to-sample comparisons and source attribution. In our initial work, low biodiversity was captured when DNA barcodes were Sanger-sequenced from plant and insect fragments isolated from 10 forensic-type surface soils. Considering some forensic laboratories now have access to massively parallel sequencing platforms, we assessed whether biological taxa present in the same surface soils could be better characterized using DNA metabarcoding. To achieve this, plant and animal barcodes were amplified and sequenced on an Illumina® MiniSeq for three different DNA sample types (n = 50): individual fragments used in our initial study, and 250 and 100 mg of bulk soil (from the 10 sites used in the initial study). A total of 572 unique target barcode sequences passed quality filtering and were used in downstream statistical analyses: 54, 321, and 285 for individual fragments, 100 mg, and 250 mg bulk soil samples, respectively. Plant barcodes permitted some spatial separation of sample sites in non-metric multidimensional scaling plots; better separation was obtained for samples prepared from bulk soil. This study confirmed that bulk soil DNA metabarcoding is a better approach for characterizing biological taxa present in surface soils, which could supplement traditional geologic examinations.

Exploring plant diversity through soil DNA in Thai national parks for influencing land reform and agriculture planning

“…Inability to detect this species in some replicates might be due to the effects of PCR and variation in the soil material used for DNA extractions. Similar results have been previously published and thus sensitivity and reproducibility of the eDNA metabarcoding results have been a challenging issue ( Boggs, Scheible & Meiklejohn, 2019 ). To avoid replicate variation, it has been suggested that performing technical replicates during PCR and pooling them before downstream processing could be effective ( Lauber et al, 2013 ; Young, Weyrich & Cooper, 2016 ; Demanèche et al, 2017 ).…”

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

“…Raw sequence data was processed and analyzed following the pipeline described in [20]. Briefly, raw reads were filtered and trimmed using the default parameters of the DADA2 pipeline [21].…”

Assessing three soil removal methods for environmental DNA analysis of mock forensic geology evidence

“…DNA metabarcoding is an approach that allows for the amplification of small, yet, informative regions of the genome to characterize biological communities. Research assessing the bacterial [14,15], plant [14,16,17], insect [16,17], fungal [17,18], and eukaryotic [14,17] communities within mock soil core evidence has highlighted promising outcomes for use in forensic geology casework. Other studies have incorporated realistic crime scene scenarios by including mock evidence in their characterization of the bacterial [19,20] and eukaryotic [21] communities.…”