Science & Tech

Sampling Procedures

sampling techniques
Credit
MLParker

Nautilus explores areas of the deep sea that have rarely -- and sometimes never -- been seen by human eyes. As such, it is crucial to have a clear record of the environmental conditions and species observed during expeditions. In addition to video imagery captured and observational data logged during dives, another important record of Nautilus’ explorations are the samples collected. The main types of physical samples collected by our Corps of Exploration are biological, geological, and water samples. Following a dive, samples are then brought to the surface for processing and preservation. 

Learn more about our Sampling Techniques and Processing Procedures below.

Sampling Techniques

Biological Sampling

A wide variety of biological samples are collected by Nautilus, and different techniques are needed depending on the size and fragility of the specimen. The most basic method is a “grab” with the manipulator arm located on ROV Hercules, which has two-fingered jaws on the end used for gripping. Different types of jaws, such as parallel and offset jaws, allow scientists to collect a variety of deep sea specimens.

These jaws can open and close to pick up larger samples, such as a clump of tubeworms or  invertebrate species on a rock. Collected specimens can then be placed into one of the bioboxes on Hercules, which are containers with watertight lids used to seal in the cooler waters of the deep sea as the ROV returns to the surface, where water temperatures are much warmer and could damage specimens. In all, Hercules is routinely equipped with eight biobox compartments: two large compartments on the front, as well as two large and four small compartments on the starboard (right) side.

For more delicate samples, specialized grippers replace the typical manipulator jaws to ensure the safe and effective collection of delicate specimens. Coral cutters have sharp blades capable of cutting a branch from a coral, and soft rubber above the blade to keep a tight hold on the sample without crushing it. An experimental specialized gripper nicknamed “squishy fingers”, uses low-pressure seawater to control the hydraulic movement of its three-fingered soft robotic hand. This allows our ROV pilots to pick up very fragile specimens, like a sea star or sea cucumber, without damaging their tissues. 

coral cutters and robotic hand
Credit
Ocean Exploration Trust
Coral cutters (left) have a special blade to cut samples. “Squishy fingers” (right) is a hydraulic robotic hand used to collect animals without harming them.

For some small, delicate, or flexible specimens, the “slurp” hose is used to ensure that the sample is collected and handled with the proper care. ROV pilots use the manipulator arm to pick up the handle of this vacuum-like hose, which then sucks up water, sediment, and small or delicate fauna into one of the slurp containers. There are eight chambers on Hercules, one of which is used to flush out the hose between samples, leaving seven discrete chambers for sample collection. Fine or coarse mesh can be placed at the chamber exits, preventing smaller organisms like microbes or particles from escaping. 

slurp
Credit
Ocean Exploration Trust/ONC
The slurp can be used to collect small fauna such as polychaete worms, limpets, snails, and sea spiders off a tubeworm bush.

A scoop is another way to  collect fauna off of the seafloor sediment that may otherwise be too delicate for the typical gripper jaws to handle. A scoop is picked up and held by the manipulator arm and can be made of many materials, like a large ice scoop or even a modified milk jug.

scoop
Credit
Ocean Exploration Trust
A scoop can also be used to collect sediment when layer preservation is not necessary.

Geological Sampling Techniques

Our team collects many types of geological samples, the most common being sediment and rocks. For medium to large rocks, a basic grab with the manipulator arm is usually most effective. However, it’s not always as easy as it sounds – sometimes a piece of rock has to be broken off of a much larger outcrop, or a small rock that appeared to be sitting on the surface of the sediment turns out to be massive and extends far below the seafloor! When rocks are “glued” together with a manganese coating, which is common in the deep sea, a crow bar or prying rod may be used to loosen or break off a rock sample.

For sediment sampling, the most frequently used technique is the push core. A push core is a clear tube with a handle for the manipulator arm to hold. As its name suggests, the core is pushed down into the sediment and then carefully pulled back out, complete with sediment. This method allows the horizontal layers of sediment and infauna to be preserved, for further analysis including for composition. The core is then pushed into a holder or “quiver” with a rubber stopper on the bottom to contain the sediment until the ROV is recovered at the surface. Our team will also use a scoop to collect sediment where layering is not important or use the slurp hose when collecting rubble. 

push core sampler
Credit
Ocean Exploration Trust
A push core sampler is deployed using the ROV manipulator arm (left). A rubber stopper keeps the sediment in place until the ROV is recovered.

Hydrological and Chemical Sampling Techniques

Water sampling on Nautilus is most frequently done using a “Niskin” bottle, which can capture certain water properties at different points during a dive. Niskin bottles are large grey cylinders on the port (left) side of Hercules. There are typically six niskins on the ROV, each of which has a wire trigger for the manipulator arm to pull in order to close the lid of a sample container. This traps 5 Liters (1.3 gallons) of ambient water inside the chamber and seals it off until the ROV returns to the surface. Typically, one Niskin bottle is reserved to record the background or control conditions, and this may be taken over a patch of bare sediment. The remaining Niskin bottles are taken around areas of interest, such as near high densities of deep-sea corals or sponges or near a hydrothermal vent or methane seep. Collected water can then be used for chemical analysis, detailing the composition within the area of interest, or for environmental DNA (eDNA) analyses conducted by our team aboard Nautilus

Niskin bottles
Credit
Ocean Exploration Trust
Six Niskin bottles are attached to the side of ROV Hercules to collect water samples at different depths during a dive.

Chemical sampling can also be done using instruments that scientists have brought onboard for the duration of their expedition. These specialized chemical sampling methods can capture water or gases, and some instruments have the capability to maintain the atmospheric pressure from the depth at which the sample was collected. This allows for more accurate measurements of chemicals found at the seafloor. Instruments scientists bring can serve a wide variety of purposes, including measuring possible pollutants or contaminants in the deep sea.

funnel and sampler
Credit
Ocean Exploration Trust
Scientists bring a variety of sampling tools aboard Nautilus, such as this funnel that collects methane gas bubbles (left) and then draws the gas into the sampler (right) for extraction and analysis on Nautilus.

Other water measurements taken during each dive include temperature, dissolved oxygen, and salinity. Sometimes a Miniature Autonomous Plume Recorder (MAPR) is used on cruises visiting hydrothermal vents or methane seeps. The MAPR can be placed on Hercules or Argus, and operates for the duration of a dive, collecting data on temperature, pressure, turbidity (optical backscatter), and oxidation-reduction potential (ORP). Altogether these are the principle tracers for detecting active seafloor hydrothermal and chemosynthetic ecosystems.

Processing Procedures

Once a dive is completed, Hercules returns to Nautilus for recovery and all samples are taken off of the vehicle to be processed in the wet lab. Samples are carefully labeled throughout the process to ensure each can be traced from the start of its journey on the seafloor to its final destination on shore.  

Biological and geological samples from the bioboxes are placed into labeled bins before being transported into the wet lab refrigerator to await their turn in the processing line. Push cores and slurp containers are also removed from the vehicle and taken into the lab. Water samples are extracted from the grey Niskin cylinders and placed into bottles and taken into the wet lab fridge for processing or preservation.

Hercules bioboxes
Credit
Ocean Exploration Trust
Biological and geological samples are taken from bioboxes aboard Hercules and placed into plastic bins for future processing.

Biological samples are then placed into tubes or bags with ethanol, to preserve the DNA, and labeled with their sample ID (below). Some larger specimens, like corals and sponges, may have the bulk of the sample dried to preserve structure.  

Biological samples are then shipped to a repository, the Harvard Museum of Comparative Zoology, so that any scientist can request the sample for more detailed analyses. Biological samples are used for many purposes, and help inform many areas of science and policy. Genetic analyses show connections between populations of fish or invertebrates, which indicate how well a species in an area might bounce back if disrupted. Isotope analysis of tissue indicates food web connections, such as how much nutrient an organism received from chemosynthetic vs photosynthetic sources. Analyses of coral skeletons, especially long-lived species like bamboo corals, show past effects of climate, and help predict how changing climate may affect the future environment. Coral analyses also help track ocean acidification effects in the deep sea.

sample tags
Credit
Ocean Exploration Trust
Each sample is carefully photographed with a sample ID number and a scale bar.

Geological rock and rubble samples are dried and placed into labeled bags. Sometimes a rock saw or hammer is used to break open the rock to examine the inner composition, but frequently this analysis does not happen until shoreside scientists begin studying the sample. Push cores may be extruded from the tube into subsampled layers, or may have the top layer of water removed before being shipped intact. All geological samples are sent to a repository at the University of Rhode Island Graduate School of Oceanography. Analyses of these samples are used to examine things like changes in sedimentation over time, which can indicate ocean health and climate conditions or the types of infaunal organisms including microbes. Detailed examination of sediment mineral contents can help provide information about geohazards, like potential for slope collapse or history of volcanic eruptions. Geological samples are also often used for understanding formation history, age-dating and geochronology studies, and determining composition (e.g. types of minerals). 

processing notes
Credit
Erin Ranney
Notes are taken detailing features of the sample, such as size, color, method of preservation, and subsampled organisms.

One main use of water samples on Nautilus is environmental DNA (eDNA) extraction by filtering the water through a fine filter paper to capture all the DNA. eDNA is composed of small pieces of organismal DNA found within the environment, including shed skin cells, mucus, feces, or small pieces of tissue. This eDNA can show a wide variety of organisms that live or pass through an area, even if they are not observed during a dive. Research partners conduct analyses of eDNA to uncover coral, sponge, and fish species or seep and vent related species that occupy areas of interest. These types of information can be especially important in Marine Protected Areas and sanctuaries that contain essential fish habitat, to show which species are utilizing these resources, and which habitats would be most effective to preserve to keep fish stocks healthy for fisheries and the environment. This is a relatively new science technique that OET is aiding through the collection of samples partially ground-truthed with video data.

Other chemical and water analyses conducted by researchers can provide information about carbon, elemental, and nutrient cycling – crucial to the balanced health of our planet.  Some chemical studies examine the extent that pollution extends into the deep sea, such as levels of microplastics, heavy metal contamination, and remnant radioactivity. Most of these analyses take place back on shore, but occasionally, time-sensitive samples will degrade if not analyzed promptly – in which case, the scientist set up their analysis equipment in the wet lab on Nautilus.