Aquatic eDNA samples consist of water that is passed through a filter to retain genetic material. There are a variety of methods for collecting aquatic DNA samples spanning a range in cost and complexity, from human-powered to automated.
Stage outcomes
At the end of this stage, you should know which sampling method you can use to meet your objective. You should be aware of what metadata to collect when going out into the field, and some key considerations around how to sample effectively.
Sampling Methods
There are a variety of methods that can collect samples at either the surface or deeper down in the water column. Collecting eDNA samples from the surface is generally the quickest and easiest method to collect a sample, and can be accomplished without the need for specialized equipment. However, it may be a poor indication or representation of the biodiversity in deeper waters.
The diversity of organisms detected using environmental DNA changes with depth in deeper, stratified water columns. To understand these changes in diversity or to target eDNA from specific subsurface habitats, it is necessary to collect eDNA samples below the surface.
Surface eDNA Sampling Methods
Syringe
Photo credit: Alan Cressler
The most basic way to collect an aquatic eDNA sample is to draw up sample water into a syringe and then push this through a filter, like a sterivex or a disc filter. Some of the major eDNA service providers employ this method in their aquatic eDNA sampling kits (see WilderLab or Jonah Ventures for examples).
Pole sampler
Many pole samplers are low-tech and can be built using supplies you can get at most hardware stores but can also be purchased from scientific suppliers. With these samplers, a bottle is placed at the sampling end and manually submerged to rinse, typically three times, and then filled with the sample.
eDNA sampling backpack
Photo credit: Smith-Root
In some cases, a pole sampler can be directly attached to a pump to draw up the water and directly pass it through a filter. An example of this is the Smith-Root eDNA sampler or eDNA Sampler Backpack combined with the single or trident Sampling Pole.
Shore-based pump-driven eDNA samplers
Photo credit: Ocean Diagnostics
Samples are directly drawn from the environment, passed through a filter, and the flow through is ideally collected into a container with graduations to keep track of the amount of water filtered. Some commercially available options include the Smith-Root eDNA Citizen Science Sampler and the Ocean Diagnostics eDNA surface sampler, while DIY versions have also been developed.
Subsurface eDNA Sampling Methods
Niskin bottle
Niskin bottles can be mounted on a rosette or connected directly to a line. The bottle is closed at the depth(s) of interest, brought back on board, and then drained into clean sample bottles for subsequent filtration.
Scuba
If samples need to be collected from a very specific habitat that is identified and verified visually, scuba divers can manually collect eDNA samples while on a dive survey in that habitat. This has the added advantage of being able to compare eDNA-derived biodiversity data with visually-obtained biodiversity data.
Towable eDNA sampling
eDNA can also be collected from a water body by towing a device fit with a filter membrane behind a vessel as a means to filter the water. Rather than the sample being collected from a discrete location, this collection approach yields a sample that integrates over a particular distance in a broader geographic location. An example can be found here.
When collecting and processing samples, it is important to collect information about the where, when, and how the samples are collected. This is called sample metadata. We will break the sample metadata down into two categories: field metadata, which describes sample collection, and lab metadata, which documents how samples are processed. This information is essential for analyzing and interpreting the genetic data.
Field metadata: This includes information about the location from which your sample(s) are collected including site name or ID, site latitude and longitude, information about the conditions (weather information, tide information, sea state information), members of the field team. It will also include information about the samples collected, such as unique sample ID, sample type (sample vs. control), sample depth, sample collection time, the time sample filtration began and ended, and volume of sample filtered.
Lab metadata: This includes information about how and when each sample is extracted, the volume and concentration of DNA after extraction, and additional information specific to the type of lab processing (e.g. metabarcoding, qPCR) that will be carried out for each sample. This information will be generated by the Service Provider when they process your samples.
The field and lab metadata are sometimes combined into a study metadata file.
Regardless of how a water sample is collected, the genetic material in that water must be collected onto a filter to be preserved, processed, and analyzed.When filtering your sample, there are a few key considerations to make around filter apparatus, filter pore size, filter membrane, filter type and size, to ensure you are capturing the correct genetic material.
Filtered aquatic samples will also need to be preserved properly to ensure the DNA material in the samples does not degrade before processing. There are a range of possibilities, including freezing, drying, and various chemical preservatives that can be used for long-term storage of DNA filters. Adding a preservative buffer to the filter is a common approach and there are a variety of buffers that can be added to a sample to preserve sample integrity. Some require a sample to be frozen after collection and filtration (No buffer, Ethanol, Sucrose Lysis Buffer, CTAB buffer) while others (Longmire’s Buffer, DNA/RNA Shield, RNALater) allow the sample to be kept at room or fridge temperatures and still remain viable.
A Service Provider may recommend a certain preservation method for the DNA extraction methods they use.
Resources
Resources associated with the Collection stage include:
Aquatic environmental DNA - a review of the macro-organismal biomonitoring revolution
Documentation
A paper describing the temporal trends in methods, major achievements, and knowledge gaps in the field of eDNA.
