Sorting states of environmental DNA: Effects of isolation method and water matrix on the recovery of membrane‐bound, dissolved, and adsorbed states of eDNA

Abstract

Environmental DNA (eDNA) once shed can exist in numerous states with varying behaviors including degradation rates and transport potential. In this study, we consider three states of eDNA: (1) a membrane-bound state referring to DNA enveloped in a cellular or organellar membrane, (2) a dissolved state defined as the extracellular DNA molecule in the environment without any interaction with other particles, and(3) an adsorbed state defined as extracellular DNA adsorbed to a particle surface in the environment. Capturing, isolating, and analyzing a target state of eDNA provides utility for better interpretation of eDNA degradation rates and transport potential.While methods for separating different states of DNA have been developed, they remain poorly evaluated due to the lack of state-controlled experimentation. We evaluated the methods for separating states of eDNA from a single sample by spiking DNA from three different species to represent the three states of eDNA as state-specific controls. We used chicken DNA to represent the dissolved state, cultured mouse cells for the membrane-bound state, and salmon DNA adsorbed to clay particles as the adsorbed state. We performed the separation in three water matrices, two environmental and one synthetic, spiked with the three eDNA states. The membrane-bound state was the only state that was isolated with minimal contamination from nontarget states. The membrane-bound state also had the highest recovery (54.11 ± 19.24%), followed by the adsorbed state (5.08 ± 2.28%), and the dissolved state had the lowest total recovery (2.21 ± 2.36%). This study highlights the potential to sort the states of eDNA from a single sample and independently analyze them for more informed biodiversity assessments. However, further method development is needed to improve recovery and reduce cross-contamination.

eDNA Adsorption onto Microplastics: Impacts of Water Chemistry and Polymer Physiochemical Properties

“…However, these soil components may have some different adsorbent behavior compared to MPs. In contrast to MPs, which showed no adsorption in soft synthetic freshwater, DNA readily adsorbed to mineral clays and activated carbon surfaces in molecular grade water. , They also exhibited significantly higher DNA absorption, with equilibrium concentrations exceeding 1000 ng DNA mg –1 for activated carbon, substantially larger than the highest MP equilibrium concentration observed in this study (421 ng DNA mg –1 63–125 μm PE particles incubated with 30 ng μL –1 DNA for 24 h) …”

“…Understanding adsorption mechanisms is crucial in eDNA passive sampling, as it guides efficient methods for desorption to isolate DNA for downstream molecular analyses. For example, employing phosphate buffers at high pH levels effectively reduces binding forces between clay surfaces and DNA phosphate backbones. , Given that the adsorption mechanism is governed by characteristics of the adsorbent material and water chemistry, future studies should consider whether the protective quality of the adsorbed DNA is impacted by the adsorption mechanism. Furthermore, it is known that phosphate-containing molecules can replace the adsorbed DNA from adsorbent surfaces, , so it is also important to know whether once adsorbed if the DNA remains adsorbed or can be replaced by other DNA molecules.…”

Suspended Materials Affect Particle Size Distribution and Removal of Environmental DNA in Flowing Waters

“…These are more resilient to inhibitors and particles but may have a lower DNA recovery rate and higher cost per sample. Nonetheless, our findings indicate that the use of these kits, or the introduction of desorption steps, may be essential to optimize eDNA capture in areas with high sedimentation, as eDNA adsorbed to particles will be a major component of the eDNA pool in these areas.…”

Detection of fish sedimentary DNA in aquatic systems: A review of methodological challenges and future opportunities

“…The persistence of fish DNA in the water column depends on whether the eDNA is intra‐ or extracellular, as well as the ecosystem's biotic and abiotic factors, such as water temperature, sunlight, pH, and microbial activity (Barnes et al, 2014; Barnes & Turner, 2016; Rourke et al, 2022). Environmental DNA can be found in a variety of states (such as extra‐organismal [dissolved or particle adsorbed], intracellular, and even held within an organelle), with environmental factors differentially affecting the recovery of each state (Brandão‐Dias et al, 2023; Kirtane et al, 2023; Mauvisseau et al, 2022; Nagler et al, 2022). Depending on the state of eDNA, it is anticipated that microbial activity and abundance, as well as temperature, pH, and oxygen concentrations, are crucial factors in the degradation of eDNA in the water column (Hofreiter et al, 2001; Seymour et al, 2018).…”

“…intracellular, and even held within an organelle), with environmental factors differentially affecting the recovery of each state (Brandão-Dias et al, 2023;Kirtane et al, 2023;Mauvisseau et al, 2022;Nagler et al, 2022). Depending on the state of eDNA, it is anticipated that microbial activity and abundance, as well as temperature, pH, and oxygen concentrations, are crucial factors in the degradation of eDNA in the water column (Hofreiter et al, 2001;Seymour et al, 2018).…”