Assessing the performance and efficiency of environmental DNA/RNA capture methodologies under controlled experimental conditions

Growing interest and affordability of environmental DNA and RNA (eDNA and
eRNA) approaches for biodiversity assessments and monitoring of complex
ecosystems have led to the emergence of manifold protocols for nucleic
acids (NAs) isolation and processing. Although there is no consensus on a
standardized workflow, the common practice for water samples is to
concentrate NAs via filtration using varying pore size membranes. Using
the smallest pore is assumed to be most efficient for NAs capture from a
wide range of material (including sub-cellular particles), however a
trade-off must occur between detection of a meaningful molecular signal
and cost/time effort when processing samples using fine pore membranes.
Comparative studies involving formal efficiency assessments are lacking,
which restricts informed decision-making around an optimized sampling
approach for applications such as biosurveillance (i.e., detection and
monitoring of target taxa - nuisance organisms, endangered and indicator
taxa or other species of economic or cultural importance). Here, we
present an experimental study using an easily cultured microalgal species
(Alexandrium pacificum) to test different filter membranes for capturing
NAs in the context of cost/time effort and cell fractions encountered in
nature (whole cells, partially lysed, and naked NAs). The results showed
no statistically significant difference between membrane types for
capturing target eDNA signal from intact and partially lysed cell
treatments. In terms of time effort and volume processed, higher
efficiency ratings were obtained with the larger pore size (5 µm)
cellulose membranes. Positively charged nylon demonstrated enhanced
capture of naked NAs, and especially eRNA signal, across treatments. Our
findings support using coarse pore size filters for adequate capture of
target NA signal (from both eDNA and eRNA) with less processing time. The
framework presented here can provide a quick and robust feasibility check
and comparative assessment of new and existing NA processing technologies,
and allows sufficient control over multiple parameters, including
physical-chemical water properties, temporal scales, and concentration and
type of input material.