Potential for cross-contamination of diatom DNA samples when using toothbrushes

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Abstract

The use of toothbrushes and similar devices for sampling diatoms from hard surfaces is a well-established approach. Toothbrushes are routinely cleaned and reused when sampling for analysis by light microscopy. This paper looks at the scale of contamination encountered when this technique is used to sample diatoms for metabarcoding analyses, as well as at the scale of contamination to be expected if stream, rather than distilled water, is used to wash diatoms from stones. Although some contamination attributable to toothbrushes was detected, read numbers were low and had no effect on index calculation or ecological status estimates. However, if the primary focus of a study is to thoroughly document diversity in a sample, then even this small level of contamination may be unacceptable and more stringent measures may be required.

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Environmental DNA metabarcoding of lake fish communities reflects long-term data from established survey methods.

Bernd Hänfling, Lori Lawson-Handley, Daniel S Read … (2016)

Organisms continuously release DNA into their environments via shed cells, excreta, gametes and decaying material. Analysis of this 'environmental DNA' (eDNA) is revolutionizing biodiversity monitoring. eDNA outperforms many established survey methods for targeted detection of single species, but few studies have investigated how well eDNA reflects whole communities of organisms in natural environments. We investigated whether eDNA can recover accurate qualitative and quantitative information about fish communities in large lakes, by comparison to the most comprehensive long-term gill-net data set available in the UK. Seventy-eight 2L water samples were collected along depth profile transects, gill-net sites and from the shoreline in three large, deep lakes (Windermere, Bassenthwaite Lake and Derwent Water) in the English Lake District. Water samples were assayed by eDNA metabarcoding of the mitochondrial 12S and cytochrome b regions. Fourteen of the 16 species historically recorded in Windermere were detected using eDNA, compared to four species in the most recent gill-net survey, demonstrating eDNA is extremely sensitive for detecting species. A key question for biodiversity monitoring is whether eDNA can accurately estimate abundance. To test this, we used the number of sequence reads per species and the proportion of sampling sites in which a species was detected with eDNA (i.e. site occupancy) as proxies for abundance. eDNA abundance data consistently correlated with rank abundance estimates from established surveys. These results demonstrate that eDNA metabarcoding can describe fish communities in large lakes, both qualitatively and quantitatively, and has great potential as a complementary tool to established monitoring methods.

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Annual time-series analysis of aqueous eDNA reveals ecologically relevant dynamics of lake ecosystem biodiversity

Iliana Bista, Gary Carvalho, Kerry Walsh … (2017)

The use of environmental DNA (eDNA) in biodiversity assessments offers a step-change in sensitivity, throughput and simultaneous measures of ecosystem diversity and function. There remains, however, a need to examine eDNA persistence in the wild through simultaneous temporal measures of eDNA and biota. Here, we use metabarcoding of two markers of different lengths, derived from an annual time series of aqueous lake eDNA to examine temporal shifts in ecosystem biodiversity and in an ecologically important group of macroinvertebrates (Diptera: Chironomidae). The analyses allow different levels of detection and validation of taxon richness and community composition (β-diversity) through time, with shorter eDNA fragments dominating the eDNA community. Comparisons between eDNA, community DNA, taxonomy and UK species abundance data further show significant relationships between diversity estimates derived across the disparate methodologies. Our results reveal the temporal dynamics of eDNA and validate the utility of eDNA metabarcoding for tracking seasonal diversity at the ecosystem scale.