A Gut Feeling: DNA Metabarcoding Gray Whale Diets

By Charles Nye, graduate student, OSU Department of Fisheries, Wildlife, & Conservation Sciences, Cetacean Conservation and Genomics Laboratory

Figure 1: An illustration (by me) of a feeding gray whale whose caudal end transitions into a DNA double helix.

Let’s consider how much stuff organisms shed daily. If you walk down a hallway, you’ll leave a microscopic trail of skin cells, evaporated sweat, and even more material if you so happen to sneeze or cough (as we’ve all learned). The residency of these bits and pieces in a given environment is on the order of days, give or take (Collins et al. 2018). These days, we can extract, amplify, and sequence DNA from leftover organismal material in environments (environmental DNA; eDNA), stomach contents (dietary DNA, dDNA), and other sources (Sousa et al. 2019; Chavez et al. 2021).

You might be familiar with genetic barcoding, where scientists are able to use documented and annotated pieces of a genome to identify a piece of DNA down to a species. Think of these as genetic fingerprints from a crime scene where all (described) species on Earth are prime suspects. With advancements in computing technology, we can barcode many species at the same time—a process known as metabarcoding. In short, you can now do an ecosystem-wide biodiversity survey without even needing to see your species of interest (Ficetola et al. 2008; Chavez et al. 2021).

(Before you ask: yes, people have tried sampling Loch Ness and came up with not a single strand of plesiosaur DNA (University of Otago, 2019).)

I received my crash course on metabarcoding when I was employed at the Monterey Bay Aquarium Research Institute (MBARI), right before grad school. There, I was employed to help refine eDNA survey field and laboratory methods (in addition to some cool robot stuff). Here at OSU, I use metabarcoding to research whale ecology, detection, and even a little bit of forensics  work. Cetacean species (or evidence thereof) I’ve worked on include North Atlantic right whales (Eubalaena glacialis), killer whales (Orcinus spp.), and gray whales (Eschrichtius robustus).

Long-time readers of the GEMM Lab Blog are probably quite knowledgeable about the summertime grays—the Pacific Coast Feeding Group (PCFG). All of us here at OSU’s Marine Mammal Institute (MMI) are keenly interested in understanding why these whales hang out in the Pacific Northwest during the summer months and what sets them apart from the rest of the Eastern North Pacific gray whale population. What interests me? Well, I want to double-check what they’re eating—genetically.

“What does my study species eat?” is a straightforward but underappreciated question. It’s also deceptively difficult to address. What if your species live somewhere remote or relatively inaccessible? You can imagine this is a common logistical issue for most research in marine sciences. How many observations do you need to make to account for seasonal or annual changes in prey availability? Do all individuals in your study population eat the same thing? I certainly like to mix and match my diet.

Gray whale foraging ecology has been studied comprehensively over the last several decades, including an in-depth stomach content evaluation by Mary Nerini in 1984 and GEMMer Lisa Hildebrand’s MSc research. PCFG whales seem to prefer shrimpy little creatures called mysids, along with Dungeness crab (Cancer magister) larvae, during their stay in the Pacific Northwest (PNW), most notably the mysid Neomysis rayii (Guerrero 1989; Hildebrand et al. 2021). Indeed, the average energetic values of common suspected prey species in PNW waters rival the caloric richness of Arctic amphipods (Hildebrand et al. 2021). However, despite our wealth of visual foraging observations, metabarcoding may add an additional layer of resolution. For example, the ocean sunfish (Mola mola) was believed to exclusively forage on gelatinous zooplankton, but a metabarcoding approach revealed a much higher diversity of prey items, including other bony fishes and arthropods (Sousa et al. 2016).

Given all this exposition, you may be wondering: “Charles—how do you intend on getting dDNA from gray whales? Are you going to cut them open?”

Figure 2: The battle station, a vacuum pump that I use to filter out all of the particulate matter from a gray whale dDNA sample. The filter is made of polycarbonate track etch material, which melts away in the DNA extraction process—quite handy, indeed!

No. I’m going to extract DNA from their poop.

Well, actually, I’ve been doing that for the last two years. My lab (Cetacean Conservation and Genomics Laboratory, CCGL) and GEMM Lab have been collaborating to make lemonade out of, er…whale poop. An archive of gray whale fecal samples (with ongoing collections every field season) originally collected for hormone analyses presented itself with new life—the genomics kind. In addition to community-level data, we are also able to recover informative DNA from the gray whales, including sex ID from “depositing” individuals, though the recovery rate isn’t perfect.

Because the GEMM Lab/MMI can non-invasively collect multiple samples from the same individuals over time, dDNA metabarcoding is a great way to repeatedly evaluate the diets of the PCFG, just shy of being at the right place at the right time with a GoPro or drone to witness a feeding event.  While we can get stomach contents and even usable dDNA from a naturally deceased whale, those data may not be ideal. How representative a stranded whale is of the population is dependent on the cause of death; an emaciated or critically injured individual, for example, is a strong outlier.

Figure 3: Presence/absence of the top 10 most-common taxonomic Families observed in the PCFG gray whale dDNA dataset (n = 20, randomly selected). Filled-in dots indicate at least one genetic read associated with that Family, and empty dots indicate none. Note the prey taxa: mysids (Mysidae), krill (Euphausiidae), and olive snails (Olividae).

Here’s a snapshot of progress to date for this dDNA metabarcoding project. I pulled out twenty random samples from my much larger working dataset (n = 82) for illustrative purposes (and legibility). After some bioinformatic wizardry, we can use a presence/absence approach to get an empirical glimpse at what passes through a PCFG gray whale. While I am able to recover species-level information, using higher-level taxonomic rankings summarizes the dataset in a cleaner fashion (and also, not every identifiable sequence resolves to species).

The title of most commonly observed prey taxa belongs to our friends, the mysids (Mysidae). Surprisingly, crabs and amphipods are not as common in this dataset, instead losing to krill (Euphausiidae) and olive snails (Olividae). The latter has been found in association with gray whale foraging grounds but not documented in a prey study (Jenkinson 2001). We also get an appreciable amount of interference from non-prey taxa, most notably barnacles (Balanidae), with an honorable mention to hydrozoans (Clytiidae, Corynidae). While easy to dismiss as background environmental DNA, as gray whales do forage at the benthos, these taxa were physically present and identifiable in Nerini’s (1984) gray whale stomach content evaluation.

So—can we conclude that barnacles and hydrozoans are an important part of a gray whale’s diet, as much as mysids? From decades of previous observations, we might say…probably not. Gray whales are actively targeting patches of crabby, shrimpy zooplankton things, and even employ novel foraging strategies to do so (Newell & Cowles 2006; Torres et al. 2018). However, the sheer diversity of consumed species does present additional dimensionality to our understanding of gray whale ecology.

The whales are eating these ancillary organisms, whether they intend to or not, and this probably does influence population dynamics, recruitment, and succession in these nearshore benthic habitats. After all, the shallow pits that gray whales leave behind post-feeding provide a commensal trophic link with other predatory taxa, including seabirds and groundfish (Oliver & Slattery 1985). Perhaps the consumption of these collateral species affects gray whale energetics and reflects on their “performance”?

I hope to address all of this and more in some capacity with my published work and graduate chapters. I’m confident to declare that we can document diet composition of PCFG whales using dDNA metabarcoding, but what comes next is where one can get lost in the sea(weeds). How does the diet of individuals compare to one another? What about at differing time points? Age groups? How many calories are in a barnacle? No need to fret—this is where the fun begins!


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https://www.otago.ac.nz/news/news/otago717609.html. [accessed 2023 Apr 25]

The learning curve never stops as the GRANITE project begins its seventh field season

Clara Bird, PhD Student, OSU Department of Fisheries, Wildlife, and Conservation Sciences, Geospatial Ecology of Marine Megafauna Lab

When I thought about what doing fieldwork would be like, before having done it myself, I imagined that it would be a challenging, but rewarding and fun experience (which it is). However, I underestimated both ends of the spectrum. I simultaneously did not expect just how hard it would be and could not imagine the thrill of working so close to whales in a beautiful place. One part that I really did not consider was the pre-season phase. Before we actually get out on the boats, we spend months preparing for the work. This prep work involves buying gear, revising and developing protocols, hiring new people, equipment maintenance and testing, and training new skills. Regardless of how many successful seasons came before a project, there are always new tasks and challenges in the preparation phase.

For example, as the GEMM Lab GRANITE project team geared up for its seventh field season, we had a few new components to prepare for. Just to remind you, the GRANITE (Gray whale Response to Ambient Noise Informed by Technology and Ecology) project’s field season typically takes place from June to mid-October of each year. Throughout this time period the field team goes out on a small RHIB (rigid hull inflatable boat), whenever the weather is good enough, to collect photo-ID data, fecal samples, and drone imagery of the Pacific Coast Feeding Group (PCFG) gray whales foraging near Newport, OR, USA. We use the data to assess the health, ecology and population dynamics of these whales, with our ultimate goal being to understand the effect of ambient noise on the population. As previous blogs have described, a typical field day involves long hours on the water looking for whales and collecting data. This year, one of our exciting new updates is that we are going out on two boats for the first part of the field season and starting our season 10 days early (our first day was May 20th). These updates are happening because a National Science Foundation funded seismic survey is being conducted within our study area starting in June. The aim of this survey is to assess geophysical structures but provides us with an opportunity to assess the effect of seismic noise on our study group by collecting data before, during, and after the survey. So, we started our season early in order to capture the “before seismic survey” data and we are using a two-boat approach to maximize our data collection ability.

While this is a cool opportunistic project, implementing the two-boat approach came with a new set of challenges. We had to find a second boat to use, buy a new set of gear for the second boat, figure out the best way to set up our gear on a boat we had not used before, and update our data processing protocols to include data collected from two boats on the same day. Using two boats also means that everyone on the core field team works every day. This core team includes Leigh (lab director/fearless leader), Todd (research assistant), Lisa (PhD student), Ale (new post-doc), and me (Clara, PhD student). Leigh and Todd are our experts in boat driving and working with whales, Todd is our experienced drone pilot, I am our newly certified drone pilot, and Lisa, Ale, and myself are boat drivers. Something I am particularly excited about this season is that Lisa, Ale, and I all have at least one field season under our belts, which means that we get to become more involved in the process. We are learning how to trailer and drive the boats, fly the drones, and handling more of the post-field work data processing. We are becoming more involved in every step of a field day from start to finish, and while it means taking on more responsibility, it feels really exciting. Throughout most of graduate school, we grow as researchers as we develop our analytical and writing skills. But it’s just as valuable to build our skillset for field work. The ocean conditions were not ideal on the first day of the field season, so we spent our first day practicing our field skills.

For our “dry run” of a field day, we went through the process of a typical day, which mostly involved a lot of learning from Leigh and Todd. Lisa practiced her trailering and launching of the boat (figure 1), Ale and Lisa practiced driving the boat, and I practiced flying the drone (figure 2). Even though we never left the bay or saw any whales, I thoroughly enjoyed our dry run. It was useful to run through our routine, without rushing, to get all the kinks out, and it also felt wonderful to be learning in a supportive environment. Practicing new skills is stressful to say the least, especially when there is expensive equipment involved, and no one wants to mess up when they’re being watched. But our group was full of support and appreciation for the challenges of learning. We cheered for successful boat launchings and dockings, and drone landings. I left that day feeling good about practicing and improving my drone piloting skills, full of gratitude for our team and excited for the season ahead.

Figure 1. Lisa (driving the truck) launching the boat.
Figure 2. Clara (seated, wearing a black jacket) landing the drone in Ale’s hands.

All the diligent prep work paid off on Saturday with a great first day (figure 3). We conducted five GoPro drops (figure 4), collected seven fecal samples from four different whales (figure 5), and flew four drone flights over three individuals including our star from last season, Sole. Combined, we collected two trifectas (photo-ID images, fecal samples, and drone footage)! Our goal is to get as many trifectas as possible because we use them to study the relationship between the drone data (body condition and behavior) and the fecal sample data (hormones). We were all exhausted after 10 hours on the water, but we were all very excited to kick-start our field season with a great day.

Figure 3. Lisa on the bow pulpit during our first sighting of the day.
Figure 4. Lisa doing a GoPro drop, she’s lowering the GoPro into the water using the line in her hands.
Figure 5. Clara and Ale collecting a fecal sample.

On Sunday, just one boat went out to collect more data from Sole after a rainy morning and I successfully flew over her from launching to landing! We have a long season ahead, but I am excited to learn and see what data we collect. Stay tuned for more updates from team GRANITE as our season progresses!

I love it when a plan comes together

By Dr. Leigh Torres


After four full-on days at sea covering 873 nautical miles, we are back in port as the winds begin to howl again and I now sip my coffee with a much appreciated still horizon. Our dedicated team worked the available weather windows hard and it paid off with more great absence data and excellent presence data too: blue whales, killer whales, common dolphins, and happily swimming pilot whales not headed to nearby Farewell Spit where a sad, massive stranding has occurred. It has been an exhausting, exhilarating, frustrating, exciting, and fulfilling time. As I reflect on all this work and reward, I can’t help but feel gratified for our persistent and focused planning that made it happen successfully. So, as we clean-up, organize data, process samples, and sit in port for a few days I would like to share some of our highlights over the past four days. I hope you enjoy them as much as we did.

The team in action on the RV Star Keys. Callum Lilley (DOC) on the bow waiting for a biopsy opportunity, Dawn Barlow (OSU) on the radio communicating with the small boat, Kristin Hodge (Cornell) taking photos of whales, Captain James Dalzell (Western Work Boats) on the helm, and Chief Engineer Spock (Western Work Boats) keeping his eyes peeled for a blow. (Photo credit: L. Torres)


In the small boat off looking for whales in a lovely flat, calm sea with an oil rig in the background. (Photo credit: D. Barlow)


Small boat action with Todd Chandler (OSU) at the helm, Leigh Torres (OSU) on the camera getting photo-id images, and Callum Lilley (DOC) taking the biopsy shot, and the dart is visible flying toward the whale in the black circle. (Photo credit: D. Barlow)


The stars of the show: blue whales. A photograph captured from the small boat of one animal fluking up to dive down as another whale surfaces close by. (Photo credit: L. Torres)


Collecting oceanographic data: Spock and Jason (Western Work Boats) deploy the CTD from the Star Keys. The CTD is an instrument that measures temperature, salinity, fluorescence and depth continuously as it descends to the bottom and back up again. (Photo credit: L. Torres)


The recently manufactured transducer pole in the water off the RV Star Keys (left) deployed with the echosounder to collect prey availability data, including this image (right) of krill swarms near feeding blue whales. (Photo credit: L. Torres)


The small boat returns to the Star Keys loaded with data and samples, including a large fecal sample in the net: The pooper scooper Leigh Torres (OSU), the biopsy rifle expert Callum Lilley (DOC), and the boat operator Todd Chandler (OSU). (Photo credit: D. Barlow)


Drone operator and videographer, Todd Chandler (OSU) under the towel (crucial piece of gear) to minimize glare on the screen as he locates and records blue whales. (Photo credit: K. Hodge)


A still shot captured from the drone footage of two adult blue whales surfacing in close proximity. (Photo credit: T. Chandler)


The team in action looking for blue whales in ideal survey conditions with Mt. Taranaki in the background. Todd Chandler (OSU) enters survey data while Dawn Barlow (OSU) spies for whale blows. (Photo credit: L. Torres)


A late evening at-sea after a big day sees Callum Lilley (DOC) processing a blue whale biopsy sample for transport, storage and analysis. (Photo credit: K. Hodge)


And we can’t forget why so many have put time, money and effort into this project: These blue whales are feeding and living within a space exploited by humans for multiple purposes, so we must ensure minimal impacts to these whales and their sustained health. (Photo credit: D. Barlow)

The five senses of fieldwork

By Leila Lemos, PhD student


This summer was full of emotions for me: I finally started my first fieldwork season after almost a year of classes and saw my first gray whale (love at first sight!).

During the fieldwork we use a small research vessel (we call it “Red Rocket”) along the Oregon coast to collect data for my PhD project. We are collecting gray whale fecal samples to analyze hormone variations; acoustic data to assess ambient noise changes at different locations and also variations before, during and after events like the “Halibut opener”; GoPro recordings to evaluate prey availability; photographs in order to identify each individual whale and assess body and skin condition; and video recordings through UAS (aka “drone”) flights, so we can measure the whales and classify them as skinny/fat, calf/juvenile/adult and pregnant/non-pregnant.

However, in order to collect all of these data, we need to first find the whales. This is when we use our first sense: vision. We are always looking at the horizon searching for a blow to come up and once we see it, we safely approach the animal and start watching the individual’s behavior and taking photographs.

If the animal is surfacing regularly to allow a successful drone overflight, we stay with the whale and launch the UAS in order to collect photogrammetry and behavior data.

Each team member performs different functions on the boat, as seen in the figure below.

Figure 1: UAS image showing each team members’ functions in the boat at the moment just after the UAS launch.
Figure 1: UAS image showing each team members’ functions in the boat at the moment just after the UAS launch.


While one member pilots the boat, another operates the UAS. Another team member is responsible for taking photos of the whales so we can match individuals with the UAS videos. And the last team member puts the calibration board of known length in the water, so that we can later calculate the exact size of each pixel at various UAS altitudes, which allows us to accurately measure whale lengths. Team members also alternate between these and other functions.

Sometimes we put the UAS in the air and no whales are at the surface, or we can’t find any. These animals only stay at the surface for a short period of time, so working with whales can be really challenging. UAS batteries only last for 15-20 minutes and we need to make the most of that time as we can. All of the members need to help the UAS pilot in finding whales, and that is when, besides vision, we need to use hearing too. The sound of the whale’s respiration (blow) can be very loud, especially when whales are closer. Once we find the whale, we give the location to the UAS pilot: “whale at 2 o’clock at 30 meters from the boat!” and the pilot finds the whale for an overflight.

The opposite – too many whales around – can also happen. While we are observing one individual or searching for it in one direction, we may hear a blow from another whale right behind us, and that’s the signal for us to look for other individuals too.

But now you might be asking yourself: “ok, I agree with vision and hearing, but what about the other three senses? Smell? Taste? Touch?” Believe it or not, this happens. Sometimes whales surface pretty close to the boat and blow. If the wind is in our direction – ARGHHHH – we smell it and even taste it (after the first time you learn to close your mouth!). Not a smell I recommend.

Fecal samples are responsible for the 5th sense: touch!

Once we identify that the whale pooped, we approach the fecal plume in order to collect as much fecal matter as possible (Fig.2).

Figure 2: A: the poop is identified; B: the boat approaches the feces that are floating at the surface (~30 seconds); C: one of the team members remains at the bow of the boat to indicate where the feces are; D: another team member collects it with a fine-mesh net. Filmed under NOAA/NMFS permit #16111 to John Calambokidis).
Figure 2: A: the poop is identified; B: the boat approaches the feces that are floating at the surface (~30 seconds); C: one of the team members remains at the bow of the boat to indicate where the feces are; D: another team member collects it with a fine-mesh net. Filmed under NOAA/NMFS permit #16111 to John Calambokidis).


After collecting the poop we transfer all of it from the net to a small jar that we then keep cool in an ice chest until we arrive back at the lab and put it in the freezer. So, how do we transfer the poop to the jar? By touching it! We put the jar inside the net and transfer each poop spot to the jar with the help of water pressure from a squeeze bottle full of ambient salt water.

Figure 3: Two gray whale individuals swimming around kelp forests. Filmed under NOAA/NMFS permit #16111 to John Calambokidis).
Figure 3: Two gray whale individuals swimming around kelp forests. Filmed under NOAA/NMFS permit #16111 to John Calambokidis).


That’s how we use our senses to study the whales, and we also use an underwater sensory system (a GoPro) to see what the whales were feeding on.

GoPro video of mysid swarms that we recorded near feeding gray whales in Port Orford in August 2016:

Our fieldwork is wrapping up this week, and I can already say that it has been a success. The challenging Oregon weather allowed us to work on 25 days: 6 days in Port Orford and 19 days in the Newport and Depoe Bay region, totaling 141 hours and 50 minutes of effort. We saw 195 whales during 97 different sightings and collected 49 fecal samples. We also performed 67 UAS flights, 34 drifter deployments (to collect acoustic data), and 34 GoPro deployments.

It is incredible to see how much data we obtained! Now starts the second part of the challenge: how to put all of this data together and find the results. My next steps are:

– photo-identification analysis;

– body and skin condition scoring of individuals;

– photogrammetry analysis;

– analysis of the GoPro videos to characterize prey;

– hormone analysis laboratory training in November at the Seattle Aquarium


For now, enjoy some pictures and a video we collected during the fieldwork this summer. It was hard to choose my favorite pictures from 11,061 photos and a video from 13 hours and 29 minutes of recording, but I finally did! Enjoy!

Figure 4: Gray whale breaching in Port Orford on August 27th. (Photo by Leila Lemos; Taken under NOAA/NMFS permit #16111 to John Calambokidis).
Figure 4: Gray whale breaching in Port Orford on August 27th. (Photo by Leila Lemos; Taken under NOAA/NMFS permit #16111 to John Calambokidis).


Figure 5: Rainbow formation through sunlight refraction on the water droplets of a gray whale individual's blow in Newport on September 15th. (Photo by Leila Lemos; Taken under NOAA/NMFS permit #16111 to John Calambokidis).
Figure 5: Rainbow formation through sunlight refraction on the water droplets of a gray whale individual’s blow in Newport on September 15th. (Photo by Leila Lemos; Taken under NOAA/NMFS permit #16111 to John Calambokidis).


Likely gray whale nursing behavior (Taken under NOAA/NMFS permit #16111 to John Calambokidis):