gray whale – Page 3 – Geospatial Ecology of Marine Megafauna Laboratory

Cold Fingers and Carabiners

By Hayleigh Middleton, GEMM Lab summer 2018 intern, entering OSU undergrad

Cold Fingers and Carabiners: that’s what most of the past three weeks have been about. We’ve progressively been getting up earlier—with many thanks to the coffee pot and multiple alarms— in order to be on the water collecting data before the wind and fog decide to kick in. Working on the ocean at 7 am with wet hands, metal equipment, a tight suit, and a “refreshing” breeze while trying to keep an eight-foot sit-on-top kayak from tipping over is challenging to say the least. Making sure the Theodolite is perfectly level on its tripod resting on sand-covered ground at the top of a cliff? Not much easier. The air is cold, the wind is cold, the equipment is cold, I’m cold, and now, everything is wet.

Rugged laptop on the cliff site. Photo by Hayleigh Middleton.

I absolutely love it.

Of all the ways I could have chosen to spend my summer before starting college at OSU, I’m so glad I took a chance and asked to spend it here. The official goals of our research project are to monitor and record the foraging habits of the Pacific Coast Feeding Group of gray whales, attempt to find out if specific individuals tend to have site fidelity and forage here year after year, and why or how they choose certain spots to feed over others. What does that mean for me? I get to kayak and take pictures of whales for six weeks! Of course, there’s a bunch of technical stuff and expensive equipment that took us two weeks to learn, but now we’re off to a great start and ready to learn more about these amazing creatures.

We have such a short amount of time to collect all this data to try and fill in the puzzle that is gray whale behavior, and we’re only a few weeks in, but I feel like I’ve already connected with this group of 60,000-pound mammals. That, in essence, is really what we’re doing here. We’re on top of a 33-meter-high cliff watching empty water for hours on the chance that we’ll be able to see a whale, identify it through photo-ID, track it with the theodolite to figure out its behavior, and use our kayak data to figure out its diet and feeding choices. Even though the whales forage up to two kilometers away from our tracking spot, it feels like they know we’re watching them. Sometimes it feels like they’re teasing us—we’ll see one, and once we get the sights fixed on it, it dives down and doesn’t come back up until we’ve turned our attention. One whale got into a very predictable pattern: three blows and then a deep dive, forage for five minutes, pop up half a viewfinder away, three more blows. We set our sights on the third blow and waited for her to resurface.

…and waited.

She swam away and didn’t show herself again.

Other times it’s like they conspire against us. Earlier this week, we spent most of the morning tracking the same whale. A couple hours into the track, another whale popped up right next to the first. Since we use a computerized tracking program, each whale is assigned a group number. That way, we can track each individual’s path and later match it to the photo identification database and sometimes a nickname. The two whales surfaced at just the right frequency and distance apart that deciding which number was currently up was guesswork for a good 15 minutes, but we gave them new track numbers and were able to sort it out later after reviewing our photos.

Searching for whales. Photo by Haley Kent.

On another day, we surveyed for whales until quitting time, which is 3:00 pm. About 2:30 pm, one was finally spotted. I named her Princess because she couldn’t be bothered to bring her body out of the water enough so we could mark her location or take a picture except for when her pectoral fin, the tip of which was “gloved” in white, came out and made a motion like a princess in a parade. When there are whales around, we can’t just say “oh look, 3:00 pm time to go” because this is important data to collect. So, we decided to wait until 3:30 pm to see if she surfaced again within visual range. 3:30 pm came and still no sign of her, so I packed up the theodolite and tripod. As soon as the box was closed, she blew, and another whale surfaced right in front of the cliff. We got some pictures of the closer one for a bit and decided that was enough. As the camera was being lowered into its case, another whale surfaced in the cove. It felt like the first went and told all the whales heading south “hey, these guys want to leave at 3, so show up right around then.” That day we got back to the lab around 5. Even though this meant being on the cliff for almost 10 hours that day, it was thrilling to have seen so many whales in one day.

Then there are times when the whales seem to beg for attention. On our third day on the cliff, we saw what we believe to be a juvenile come swimming into view. We assume that he was a juvenile because he was “small” and quite blank in terms of pigmentation and scarring. He was adorable. He stayed over at Mill Rocks for a while foraging, all of which we “fixed” into the tracking program via the Theodolite, and then he came toward us into the little kelp patch just in front of our cliff site. He would dive down, scoop up some zooplankton to eat, and resurface right in the middle of the kelp. The cutest part is that he would then proceed to roll around in the kelp and further drape himself in it.

Having such a young whale come and forage made us wonder if mothers who have site fidelity then teach their young “hey, you don’t have to go all the way north, there’s a ton of good food here in Port Orford.” Hopefully that’s one of the things we’ll be able to figure out with the data collected with this long–term study. But in the meantime, I still have three weeks of data to collect and a bunch more whales to meet.

Managing Oceans: the inner-workings of marine policy

By Alexa Kownacki, Ph.D. Student, OSU Department of Fisheries and Wildlife, Geospatial Ecology of Marine Megafauna Lab

When we hear “marine policy” we broadly lump it together with environmental policy. However, marine ecosystems differ greatly from their terrestrial counterparts. We wouldn’t manage a forest like an ocean, nor would we manage an ocean like a forest. Why not? The answer to this question is complex and involves everything from ecology to politics.

Oceans do not have borders; they are fluid and dynamic. Interestingly, by defining marine ecosystems we are applying some kind of borders. But water (and all its natural and unnatural content) flows between these ‘ecosystems’. Marine ecosystems are home to a variety of anthropogenic activities such as transportation and recreation, in addition to an abundance of species that represent the three major domains of biology: Archaea, Bacteria, and Eukarya. Humans are the only creatures who “recognize” the borders that policymakers and policy actors have instilled. A migrating gray whale does not have a passport stamped as it travels from its breeding grounds in Mexican waters to its feeding grounds in the Gulf of Alaska. In contrast, a large cargo ship—or even a small sailing vessel—that crosses those boundaries is subjected to a series of immigration checkpoints. Combining these human and the non-human facets makes marine policy complex and variable.

The eastern Pacific gray whale migration route includes waters off of Mexico, Canada, and the United States. Source: https://www.learner.org/jnorth/tm/gwhale/annual/map.html

Environmental policy of any kind can be challenging. Marine environmental policy adds many more convoluted layers in terms of unknowns; marine ecosystems are understudied relative to terrestrial ecosystems and therefore have less research conducted on how to best manage them. Additionally, there are more hands in the cookie jar, so to speak; more governments and more stakeholders with more opinions (Leslie and McLeod 2007). So, with fewer examples of successful ecosystem-based management in coastal and marine environments and more institutions with varied goals, marine ecosystems become challenging to manage and monitor.

A visual representation of what can happen when there are many groups with different goals: no one can easily get what they want. Image Source: The Brew Monks

With this in mind, it is understandable that there is no official manual on policy development. There is, however, a broadly standardized process of how to develop, implement, and evaluate environmental policies: 1) recognize a problem 2) propose a solution 3) choose a solution 4) put the solution into effect and 4) monitor the results (Zacharias pp. 16-21). For a policy to be deemed successful, specific criteria must be met, which means that a common policy is necessary for implementation and enforcement. Within the United States, there are a multiple governing bodies that protect the ocean, including the National Oceanic and Atmospheric Administration (NOAA), Environmental Protection Agency (EPA), Fish and Wildlife Service (USFWS), and the Department of Defense (DoD)—all of which have different mission statements, budgets, and proposals. To create effective environmental policies, collaboration between various groups is imperative. Nevertheless, bringing these groups together, even those within the same nation, requires time, money, and flexibility.

This is not to say that environmental policy for terrestrial systems, but there are fewer moving parts to manage. For example, a forest in the United States would likely not be an international jurisdiction case because the borders are permanent lines and national management does not overlap. However, at a state level, jurisdiction may overlap with potentially conflicting agendas. A critical difference in management strategies is preservation versus conservation. Preservation focuses on protecting nature from use and discourages altering the environment. Conservation, centers on wise-use practices that allow for proper human use of environments such as resource use for economic groups. One environmental group may believe in preservation, while one government agency may believe in conservation, creating friction amongst how the land should be used: timber harvest, public use, private purchasing, etc.

Linear representation of preservation versus conservation versus exploitation. Image Source: Raoof Mostafazadeh

Furthermore, a terrestrial forest has distinct edges with measurable and observable qualities; it possesses intrinsic and extrinsic values that are broadly recognized because humans have been utilizing them for centuries. Intrinsic values are things that people can monetize, such as commercial fisheries or timber harvests whereas extrinsic values are things that are challenging to put an actual price on in terms of biological diversity, such as the enjoyment of nature or the role of species in pest management; extrinsic values generally have a high level of human subjectivity because the context of that “resource” in question varies upon circumstances (White 2013). Humans are more likely to align positively with conservation policies if there are extrinsic benefits to them; therefore, anthropocentric values associated with the resources are protected (Rode et al. 2015). Hence, when creating marine policy, monetary values are often placed on the resources, but marine environments are less well-studied due to lack of accessibility and funding, making any valuation very challenging.

The differences between direct (intrinsic) versus indirect (extrinsic) values to biodiversity that factor into environmental policy. Image Source: Conservationscienceblog.wordpress.com

Assigning a cost or benefit to environmental services is subjective (Dearborn and Kark 2010). What is the benefit to a child seeing an endangered killer whale for the first time? One could argue priceless. In order for conservation measures to be implemented, values—intrinsic and extrinsic—are assigned to the goods and services that the marine environment provides—such as seafood and how the ocean functions as a carbon sink. Based off of the four main criteria used to evaluate policy, the true issue becomes assessing the merit and worth. There is an often-overlooked flaw with policy models: it assumes rational behavior (Zacharias 126). Policy involves relationships and opinions, not only the scientific facts that inform them; this is true in terrestrial and marine environments. People have their own agendas that influence, not only the policies themselves, but the speed at which they are proposed and implemented.

Tourists aboard a whale-watching vessel off of the San Juan Islands, enjoying orca in the wild. Image Source: Seattle Orca Whale Watching

One example of how marine policy evolves is through groups, such as the International Whaling Commission, that gather to discuss such policies while representing many different stakeholders. Some cultures value the whale for food, others for its contributions to the surrounding ecosystems—such as supporting healthy seafood populations. Valuing one over the other goes beyond a monetary value and delves deeper into the cultures, politics, economics, and ethics. Subjectivity is the name of the game in environmental policy, and, in marine environmental policy, there are many factors unaccounted for, that decision-making is incredibly challenging.

Efficacy in terms of the public policy for marine systems presents a challenge because policy happens slowly, as does research. There is no equation that fits all problems because the variables are different and dynamic; they change based on the situation and can be unpredictable. When comparing institutional versus impact effectiveness, they both are hard to measure without concrete goals (Leslie and McLeod 2007). Marine ecosystems are open environments which add an additional hurdle: setting measurable and achievable goals. Terrestrial environments contain resources that more people utilize, more frequently, and therefore have more set goals. Without a problem and potential solution there is no policy. Terrestrial systems have problems that humans recognize. Marine systems have problems that are not as visible to people on a daily basis. Therefore, terrestrial systems have more solutions presented to mitigate problems and more policies enacted.

As marine scientists, we don’t always immediately consider how marine policy impacts our research. In the case of my project, marine policy is something I constantly have to consider. Common bottlenose dolphins are protected under the Marine Mammal Protection Act (MMPA) and inhabit coastal of both the United States and Mexico, including within some Marine Protected Areas (MPA). In addition, some funding for the project comes from NOAA and the DoD. Even on the surface-level it is clear that policy is something we must consider as marine scientists—whether we want to or not. We may do our best to inform policymakers with results and education based on our research, but marine policy requires value-based judgements based on politics, economics, and human objectivity—all of which are challenging to harmonize into a succinct problem with a clear solution.

Two common bottlenose dolphins (coastal ecotype) traveling along the Santa Barbara, CA shoreline. Image Source: Alexa Kownacki

References:

Dearborn, D. C. and Kark, S. 2010. Motivations for Conserving Urban Biodiversity. Conservation Biology, 24: 432-440. doi:10.1111/j.1523-1739.2009.01328.x

Leslie, H. M. and McLeod, K. L. (2007), Confronting the challenges of implementing marine ecosystem‐based management. Frontiers in Ecology and the Environment, 5: 540-548. doi:10.1890/060093

Munguia, P., and A. F. Ojanguren. 2015. Bridging the gap in marine and terrestrial studies. Ecosphere 6(2):25. http://dx.doi.org/10.1890/ES14-00231.1

Rode, J., Gomez-Baggethun, E., Krause, M., 2015. Motivation crowding by economic payments in conservation policy: a review of the empirical evidence. Ecol. Econ. 117, 270–282 (in this issue).

White, P. S. (2013), Derivation of the Extrinsic Values of Biological Diversity from Its Intrinsic Value and of Both from the First Principles of Evolution. Conservation Biology, 27: 1279-1285. doi:10.1111/cobi.12125

Zacharias, M. 2014. Marine Policy. London: Routledge.

Coastal oceanography takes patience

Joe Haxel, Acoustician, Assistant Professor, CIMRS/OSU

Greetings GEMM Lab blog readers. My name is Joe Haxel and I’m a close collaborator with Leigh and other GEMM lab members on the gray whale ecology, physiology and noise project off the Oregon coast. Leigh invited me for a guest blog appearance to share some of the acoustics work we’ve been up to and as you’ve probably guessed by now, my specialty is in ocean acoustics. I’m a PI in NOAA’s Pacific Marine Environmental Laboratory’s Acoustics Program and OSU’s Cooperative Institute for Marine Resources Studies where I use underwater sound to study a variety of earth and ocean processes.

As a component of the gray whale noise project, during the field seasons of 2016 and 2017 we recorded some of the first measurements of ambient sound in the shallow coastal waters off Oregon between 7 and 20 meters depth. In the passive ocean acoustics world this is really shallow, and with that comes all kinds of instrument and logistical challenges, which is probably one of the main reasons there is little or no acoustic baseline information in this environment.

For instance, one of the significant challenges is rooted in the hydrodynamics surrounding mobile recording systems like the drifting hydrophone we used during the summer field season in 2016 (Fig 1). Decoupling motion of the surface buoy (e.g., caused by swell and waves) from the submerged hydrophone sensor is critical, and here’s why. Hydrophones convert pressure fluctuations at the sensor/ water interface to a calibrated voltage recorded by a logging system. Turbulence resulting from moving the sensor up and down in the water column with surface waves introduces non-acoustic pressure changes that severely contaminate the data for noise level measurements. Vertical and horizontal wave motions are constantly acting on the float, so we needed to engineer compliance between the surface float and the suspended hydrophone sensor to decouple these accelerations. To overcome this, we employed a couple of concepts in our drifting hydrophone design. 1) A 10 cm diameter by 3 m long spar buoy provided floatation for the system. Spar buoys are less affected by wave motion accelerations compared to most other types of surface floatation with larger horizontal profiles and drag. 2) A dynamic shock cord that could stretch up to double its resting length to accommodate vertical motion of the spar buoy; 3) a heave plate that significantly reduced any vertical motion of the hydrophone suspended below it. This was a very effective design, and although somewhat cumbersome in transport with the RHIB between deployment sites, the acoustic data we collected over 40 different drifts around Newport and Port Orford in 2016 was clean, high quality and devoid of system induced contamination.

Figure 1. The drifting hydrophone system used for 40 different drifts recording ambient noise levels in 7-20 m depths in the Newport and Port Orford, OR coastal areas.

Spatial information from the project’s first year acoustic recordings using the drifting hydrophone system helped us choose sites for the fixed hydrophone stations in 2017. Now that we had some basic information on the spatial variability of noise within the study areas we could focus on the temporal objectives of characterizing the range of acoustic conditions experienced by gray whales over the course of the entire foraging season at these sites in Oregon. In 2017 we deployed “lander” style instrument frames, each equipped with a single, omni-directional hydrophone custom built by Haru Matsumoto at our NOAA/OSU Acoustics lab (Fig. 2). The four hydrophone stations were positioned near each of the ports (Yaquina Bay and Port Orford) and in partnership with the Oregon Department of Fish and Wildlife Marine Reserves program in the Otter Rock Marine Reserve and the Redfish Rocks Marine Reserve. The hydrophones were programmed on a 20% duty cycle, recording 12 minutes of every hour at 32 kHz sample rate, providing spectral information in the frequency band from 10 Hz up to a 13 kHz.

Figure 2. The hydrophone (black cylinder) on its lander frame ready for deployment.

Here’s where the story gets interesting. In my experience so far putting out gear off the Oregon coast, anything that has a surface expression and is left out for more than a couple of weeks is going to have issues. Due to funding constraints, I had to challenge that theory this year and deploy 2 of the units with a surface buoy. This is not typically what we do with our equipment since it usually stays out for up to 2 years at a time, is sensitive, and expensive. The 2 frames with a surface float were going to be deployed in Marine Reserves far enough from the traffic lanes of the ports and in areas with significantly less traffic and presumably no fishing pressure. The surface buoy consisted of an 18 inch diameter hard plastic float connected to an anchor that was offset from the instrument frame by a 150 foot weighted groundline. The gear was deployed off Newport in June and Port Orford in July. What could go wrong?

After monthly buoy checks by the project team, including GPS positions, and buoy cleanings my hopes were pretty high that the surface buoy systems might actually make it through the season with recoveries scheduled in mid-October. Had I gambled and won? Nope. The call came in September from Leigh that one of the whale watching outfits in Depoe Bay recovered a free floating buoy matching ours. Bummer. Alternative recovery plans initiated and this is where things began to get hairy. Fortunately, we had an ace in our back pocket. We have collaborators at the Oregon Coast Aquarium (OCA) who have a top-notch research diving team led by Jim Burke. In the last week of October, they performed a successful search dive on the missing unit near Gull Rock and attached a new set of floats directly to the instrument frame. The divers were in the water for a short 20 minutes thanks to the good series of marks recorded during the buoy checks throughout the summer (Fig. 3).

Figure 3. OCA divers, Jenna and Doug, heading out for a search dive to locate and mark the Gull Rock hydrophone lander.

We had surface marker floats on the frame, but there was a new problem. Video taken by Jenna and Doug from the OCA dive team revealed the landers were pretty sanded in from a couple of recent October storms (Fig. 4). Ugghhh!

Figure 4. Sanded in lander at Gull Rock. Notice the sand dollars and bull kelp wrapped on the frame.

Alternative recovery plan adjustment: we’re gonna need a diver assisted recovery with 2 boats. One to bring a dive team to air jet the sand out away from the legs of the frame and another larger vessel with pulling power to recover the freed lander. Enter the R/V Pacific Surveyor and Capt. Al Pazar. Al, Jim and I came up with a new recovery plan and only needed a decent weather window of a few hours to get the job done. Piece of cake in November off the Oregon coast, right?

The weather finally cooperated in early December in-line with the OCA dive team and R/V Pacific Surveyor’s availability. The 2 vessels and crew headed up to Gull Rock for the first recovery operation of the day. At first we couldn’t locate the surface floats. Oh no. It seemed the rough fall/ winter weather and high seas since late October were too much for the crab floats? As it turns out, we eventually found the floats eastward about 200 m but couldn’t initially see them in the glare and whitecapping conditions that morning. The lander frame had broken loose from its weakened anchor legs in the heavy weather (as it was designed to do through an Aluminum/ Stainless Steel galvanic reaction over time) and rolled or hopped eastward by about 200 m (Fig. 5). Oh dear!

Figure 5. A hydrophone lander after recovery. Notice all but 1 of the concrete anchor legs missing from the recovered lander and the amount of bio-fouling on the hydrophone (compared to Figure 2).

Thankfully, the hydrophone was well protected, and no air jetting was required. With OCA divers out of the water and clear, the Pacific Surveyor headed over to the floats and easily pulled the lander frame and hydrophone on board (Fig. 6). Yipee!

On to the next hydrophone station. This station, deployed ~ 800 m west of the south reef off of South Beach near the Yaquina Bay port entrance. It was deployed entirely subsurface and was outfitted with an acoustic release transponder that I could communicate with from the surface and command to release a pop-up messenger float and line for eventual recovery of the instrument frame. Once on station, communication with the release was established easily (a good start) and we began ranging and moving the OCA vessel Gracie Lynn in to a position within about 2 water depths of the unit (~40 m). I gave the command to the transponder and the submerged release confirmed it was free of its anchor and heading for the surface, but it never made it. Uh oh. Turns out this lander had also broke free of its anchored legs and rolled/ hopped 800 m eastward until it was pinned up against the boulder structure of the south reef. Amazingly, OCA divers Jenna and Doug located the messenger float ~ 5 m below the surface and the messenger line had been fouled by the rolling frame so it could not reach the surface. They dove down the messenger line and attached a new recovery line to the lander frame and the Pacific Surveyor hauled up the frame and hydrophone in-tact (Fig. 6). Double recovery success!

Figure 6. R/V Pacific Surveyor recovering hydrophone landers off Gull Rock and South Beach.

The hydrophone data from both systems looks outstanding and analysis is underway. This recovery effort took a huge amount of patience and the coordination of 3 busy groups (NOAA/OSU, OCA, Capt. Al). Thanks to these incredible collaborations and some heroic diving from Jim Burke and his OCA dive team, we now have a unique and unprecedented shallow water passive acoustic data set from the energetic waters off the Oregon coast.

So that’s some of the story from the 2016 and 2017 field season acoustic point of view. I’ll save the less exciting, but equally successful instrument recoveries from Port Orford for another time.

Finding the hot spot: incorporating thermal imagery into our whale research

By Leila Lemos and Leigh Torres

A couple weeks ago the GEMM Lab trialed something new in our gray whale research: the addition of a thermal imaging camera to our drone.

For those who do not know what a thermal imaging camera is, it is a device that uses infrared radiation to form an object, and operates in wavelengths as long as 14,000 nm (14 µm). A thermal camera uses a similar procedure as a normal camera, but responds to infrared radiation rather than visible light. It is also known as an infrared or thermographic camera.

All objects with a temperature above absolute zero emit infrared radiation, and thermography makes it possible to see with or without visible light. The amount of radiation emitted by an object intensifies with temperature, thus thermography allows for perception of temperature variations. Humans and other warm-blooded animals are easily detectable via infrared radiation, during the day or the night.

Infrared radiation was first discovered in 1800, by the astronomer Frederick William Herschel. He discovered infrared light by using a prism and a thermometer (Fig.1). He called it the infrared spectrum “dark heat”, which falls between the visible and microwave bands on the electromagnetic spectrum (Hitch 2016).

Figure 1: Astronomer Frederick William Herschel discovers infrared light by using a prism and a thermometer.
Source: NASA, 2012.

Around 30 years later it was possible to detect a person using infrared radiation within ten meters distance, and around 50 years later it was possible to detect radiation from a cow at 400 meters distance, as technology became gradually more sensitive (Langley, 1880).

Thermography nowadays is applied in research and development in a variety of different fields in industry (Vollmer and Möllmann 2017). Thermal imaging is currently applied in many applications, such as night vision, predictive maintenance, reducing energy costs of processes and buildings, building and roof inspection, moisture detection in walls and roofs, energy auditing, refrigerant leaks and detection of gas, law enforcement and anti-terrorism, medicinal and veterinary thermal imaging, astronomy, chemical imaging, pollution effluent detection, archaeology, paranormal investigation, and meteorology.

Some of the most interesting examples of its application are:

Detection of the presence of icebergs, increasing safety for navigators.
Detection of bombs
Non-invasive detection of breast cancer (Fig.2)
Detection of fire, and detection of fire victims in smoke-filled rooms or hidden under plywood, by the fire departments (Fig.3)

Figure 2: Thermography approved in 1982 to detect breast cancer. Method is able to detect 95% of early stages cancers.
Source: Hitch, 2016.

Figure 3: The use of thermal imaging cameras by the fire departments.
Source: MASC, 2017.

In environmental research, the thermal imaging camera is an interesting tool used to detect wildlife presence (especially for nocturnal species), to monitor wildlife and detect disease (Fig.4), and to better understand thermal patterns in animals (Fig.5), among others.

Figure 4: Wildlife monitoring: detection of mange infection in wolves of Yellowstone National Park. During winter, wolves infected with mange can suffer a substantial amount of heat loss compared to those without the disease, according to a study by the U.S. Geological Survey and its partners.
Source: Wildlife Research News 2012; USGS 2016.

Figure 5: Study on thermal patterns and thermoregulation abilities of emperor penguins in Antarctica.
Source: BBC 2013.

Now that thermal cameras are small enough for attachment to drones, we are eager to monitor whales with this device to potentially identify injuries and infections. This non-invasive method could contribute another aspect to our on-going blue and gray whale health assessment work. However, dealing with new technology is never easy and we are working to optimize settings to collect the data needed. Our test flights with the thermal camera were successful – we captured images and retrieved the expensive camera (always a good thing!) – but the whale images were less clear than desired. The camera was able to detect thermal variation between our research vessel and the ocean (Fig. 6: boat and people are displayed as hot coloration (yellow, orange and red tones), while the ocean exhibited a cold coloration (purple). Yet, the camera’s ability to differentiate thermal content of the whale while surfacing from the ocean was less evident (Fig. 7). We believe this problem is due to automatic gain control settings by the camera that essentially continually shifts the baseline temperature in the image so that thermal contrast between the whale and ocean was not very strong, except for those hot blow holes shinning like devil eyes (Fig. 7). We are working to adjust these gain settings so that our next trial will be more successful, and next time we will see our whales in all their colorful thermal glory.

Figure 6: Thermal image of the R/V Ruby captured by a thermal camera flown on a drone by the GEMM Lab on September 09th, 2017.
Source: GEMMLab 2017.

Figure 7. Thermal image of a gray whale captured by a thermal camera flown on a drone by the GEMM Lab on September 09th, 2017. Notice the ‘hot’ color (yellow-orange) of the blow holes indicating the heat within the whale’s body. (Image captured under NOAA/NMFS permit #16111).

References

BBC. 2013. In pictures: Emperor penguins’ ‘cold coat’ discovered. Available at: http://www.bbc.co.uk/nature/21669963

Hitch J. 2016. A Brief History of Thermal Cameras. Available at: http://www.newequipment.com/technology-innovations/brief-history-thermal-cameras /gallery?slide=1

Langley SP. 1880. The bolometer. Vallegheny Observatory, The Society Gregory, New York, NY, USA.

MASC. 2017. Thermal Imaging Camera. Available at: https://duckduckgo.com/ ?q=detection+of+victim+fire+department+thermal+camera&atb=v76-7_u&iax=1&ia= images&iai=http%3A%2F%2Fwww.masc.sc%2FSiteCollectionImages%2Fuptown%2F Super_Red_Hot.jpg

NASA. 2012. Beyond the Visible Light. Available at: https://www.nasa.gov/topics/ technology/features/webb-beyond-vis.html

USGS. 2016. Study Shows Cold and Windy Nights Physically Drain Mangy Wolves. Available at: https://www.usgs.gov/news/study-shows-cold-and-windy-nights-physically-drain-mangy-wolves

Vollmer M. and Möllmann KP. 2018. Infrared Thermal Imaging: Fundamentals, research and Applications. Second Edition. Wiley-VCH: Weinheim, Germany.

Wildlife Research News, 2012. Tool: Infrared Monitoring. Available at: https://wildliferesearchnews.wordpress.com/2012/04/24/tool-infrared-monitoring/

New steps towards community engagement: introducing high schoolers to the field

By Florence Sullivan, MSc, GEMM Lab Research Assistant

This summer, I had the pleasure of returning to Port Orford to lead another field season of the GEMM Lab’s gray whale foraging ecology research project. While our goal this summer was to continue gathering data on gray whale habitat use and zooplankton community structure in the Port Orford region, we added in a new and exciting community engagement component: We integrated local high school students into our research efforts in order to engage with the local community to promote interest in the OSU field station and the research taking place in their community. Frequent blog readers will have seen the posts written by this year’s interns (Maggie O’Rourke Liggett, Nathan Malamud, and Quince Nye) as they described how they became interns, their experience doing fieldwork, and some lessons they’ve learned from the project. I am very impressed with the hard work and effort that all three of them put into making this field season a success. (Getting out of a warm bed, and showing up at the field station at 6am sharp for five weeks straight is no easy feat for high-schoolers or an undergrad student during summer break!)

Quince hard at work scanning the horizon for whale spouts. photo credit: Alexa Kownacki

During the month of August, our team collected the following data on whale distribution and behavior:

Spent 108 hours on the cliff looking for whales
Spent 11 hours actively tracking whales with the theodolite
Collected 19 whale tracklines
Identified 15 individual whales using photo-ID – Two of those whales came back 3 times each, and one of them was a whale nick-named “Buttons” who we had tracked in 2016 as well.

We also collected data on zooplankton – gray whale prey – in the area:

Collected 134 GoPro videos of the water column at the 12 kayak sample sites
Did approximately 147 zooplankton net tows
Collected 64 samples for community analysis to see what species of zooplankton were present
Collected 115 samples for energetic analysis to determine how many calories can be derived from each zooplankton

The 2017 field team. From left to right: Tom Calvanese (Field Station Manager), Florence Sullivan (Project Lead), Quince Nye, Maggie O’Rourke-Liggett, and Nathan Malamud. Photo credit: Alexa Kownacki

Since I began this project in 2015, I have been privileged to work with some truly fantastic interns. Each year, I learned new lessons about how to be an effective mentor, and how to communicate our research goals and project needs more clearly. This year was no exception, and I worked hard to bring some of the things I’ve learned into my project planning. As the team can tell you, science communication, and the benefits of building good will and strong community relationships were heavily emphasized over the course of the internship. Everyone was encouraged to use every opportunity to engage with the public, explain our work, and pass on new things they had learned. Whenever the team encountered other kayakers out on the water, we took the time to share any cool zooplankton samples we gathered that day, and explain the goals of our research. Maggie and I also took the opportunity to give a pair of evening lectures at Humbug Mountain State Park, which were both well attended by curious campers.

Florence and Maggie give evening lectures at Humbug Mountain State Park

In addition, the team held a successful final community presentation on September 1 at the Port Orford Field Station that 45 people attended! In the week leading up to the presentation, Quince and Nathan spent many long hours working diligently on the powerpoint presentation, while Maggie put together a video presentation of “the intern experience” (Click here for the video showcased on last week’s blog). I am incredibly proud of Nathan and Quince, and the clear and confident manner in which they presented their experience to the audience who showed up to support them. They easily fielded the following questions:

Q: “How do you tell the difference between a whale that is searching or foraging?”

A: When we look at the boundaries of our study site, a foraging whale consistently comes up to breathe in the same spot, while a searching whale covers a lot of distance going back and forth without leaving the general area.

Q: “How do we make sure that this program continues?”

A: Stay curious and support your students as they take on internships, support the field station as it seeks to provide resources, and if possible, donate to funds that raise money for research efforts.

Nathan talks about the plankton results during the final community presentation. photo credit: Alexa Kownacki

The audience during the final community presntation. photo credit: Alexa Kownacki

Quince and Nathan answer questions at the end of the community presentation. photo credit: Alexa Kownacki

When communicating science, it is important to results into context. In addition to showcasing the possibilities of excellent research with positive community support, and just how much a trio of young people can grow over the course of 6 weeks, this summer has highlighted the value of long term monitoring studies, particularly when studying long-lived animals such as whales. We saw far fewer whales this summer than compared to the two previous years, and the whales spent much less time in the Port Orford area (Table 1). As a scientist, knowing where whales are not (absence data) is just as important as knowing where whales are (presence data), and these marked differences drive our hypotheses! What has changed in the system? What can explain the differences in whale behavior between years? Does it have to do with food quality or availability? (This is why we have been gathering all those zooplankton samples.) Does it have to do with other oceanographic factors or human activities?

Table 1. Summary of whale tracking efforts for the three seasons of field work in Port Orford. Notice how in 2017 we only collected 194 whale location points (theodolite marks). This is about 92% less than in the previous years.

	2015	2016	2017
Hours spent watching	72:49	148:30	108
Hours spent tracking	80:39*	82:30	11
Number of individuals	43	50	15
Number of theodolite marks	2483	2414	194

*we often tracked more than one individual simultaneously in 2015

Long term monitoring projects give us a chance to notice differences between years, and ask questions about what are normal fluctuations in the system, and what are abnormal. On top of that, projects like this create the opportunity for additional internships, and to mentor more students in the scientific method of investigation. There is so much still to be explored in the Port Orford ecosystem, and I truly hope this program is able to continue. If you are interested in making a monetary contribution to sustain this research and internship program, donations can be accepted here (gemm lab fund) and here (field station fund).

Quince records zooplankon sample weights in the wet lab.

Quince sorts through a zooplankton sample in the wet lab.

Nathan stores zooplankton community analysis samples

Maggie, Florence and Quince enjoy the eclipse!

Quince and Maggie bundle up on the cliff as they watch for whales.

Nathan and Quince organize data on the computer at the end of the day.

Quince and Nathan build sand castles as we wait for the fog to clear before launching the research kayak

This research and student internships would not have been possible without the generous support from Oregon Sea Grant, the Oregon Coast STEM hub, the Port Orford Field Station, South Coast Tours, partnerships with the Bernard and Chapman labs, the OSU Marine Mammal Institute, and the Geospatial Ecology of Marine Megafauna Lab.

Diving Deeper

By Taylor Mock, GEMM Lab intern

Greetings, all!

My name is Taylor Mock. Since February I have been volunteering in the GEMM Lab and am ecstatic to make my online debut as part of the team!

For many years, I had a shallow relationship with Hatfield Marine Science Center. As a Newport native, I would spend mornings and evenings glancing over at the Hatfield buildings while driving over the bridge to and from school. I was always intrigued. Sure, I would hear snippets of research from my peers about what projects their parents were involved in, but the inner workings of the complex mystified me.

Toward the end of my Freshman year in 2012 at Westmont College in Santa Barbara, California, my mom asked me what my summer plans were. I replied with the typical “I don’t know… Get a job?” She insisted that instead of a job I think about getting an internship; experience that will last more than a summer. I inquired through a family friend (because every person in this little community is woven together some way or another) if any internships or volunteer opportunities were available at Hatfield. She pointed me in the direction of the Environmental Protection Agency and thus began my Hatfield volunteering saga. I worked that summer, and the next, at the EPA under the direction of Ted DeWitt and Jody Stecher on denitrification studies in estuarine marshes. That summer provided me a glorious front row seat to field research and a greater understanding of my potential as a person and as a scientist. Now, this experience was marvelous, but I knew shortly after starting that my heart was elsewhere.

It was during my study abroad semester in Belize as part of my internship at the Toledo Institute for Development and Environment (TIDE) that I realized I wanted to work with marine macroorganisms. At TIDE, I engaged in radio telemetry conservation efforts tracking Hicatee (Dermatemys mawii) aquatic turtles. We would spend days on a small boat floating through canals and setting nets in hopes of capturing individuals of this small population to outfit them with radio tracking devices. These would be later used to track foraging, mating, and travel patterns in the region. It was an amazing time, to say the least. I remember waking up on my 21st birthday from my camping hammock and staring up at the lush rainforest above my head with a warm breeze across my face, followed by spending the day in the presence of these glorious creatures. It was heaven. I returned to Westmont the following term and took a Marine Mammal Eco-Physiology course and absolutely fell in love with Cetacea. Yes, I had always been captivated by this clade of beings (and truthfully when I was eight years old had a book on “How to Become a Marine Mammal Trainer”), but this was deeper. Of course, pinnipeds and otters and polar bears and manatees were enjoyable to learn about. There was something about the Cetacea though and how they migrated up and down the coast (just like me!) that I really connected with. My time learning about these animals created an intimate understanding of another group of species that developed into a rich, indescribable empathetic connection. I had to take a couple years away from scholastics and away from biology for health and wellness reasons. One day, though, a couple years after graduating and returning to Newport I rekindled with Jody from the EPA. He asked me if I would like to volunteer under Leigh Torres in the Marine Mammal Institute at HMSC. I do not think I could have possibly said no. I have been enjoying my time in the GEMM Lab ever since!

Though I am available to help anyone with any task they need, the work I do mostly centers around photogrammetry.

Using photogrammetry skills to measure gray whales in the GEMM Lab.

Photogrammetry, essentially, means geo-spatially measuring objects using photographs. What that looks like for me is taking an aerial photograph (extracted from overhead drone video footage) of a whale, running the image through a computer program called “Matlab”, taking a series of measurements from the whale (e.g., tip of the mandible to the notch of the fluke, distance between each tip of the fluke, and several measurements across the midsection of the whale). Several images of individuals are processed in order to find an average set of measurements for each whale.

Final result of the photogrammetry method on a gray whale

You might be wondering, “How can one measure the distance accurately from just a photograph?” I am glad you asked! The drones are outfitted with a barometer to measure the atmospheric pressure and, in turn, altitude. The changing altitudes are recorded in a separate program that is run simultaneously with the video footage. Thus, we have the altitudinal measurements for every millisecond of the drone’s flight. To monitor the accuracy and functionality of the barometer, calibrations are completed upon deployment and retrieval of each drone flight. To calibrate: the initial takeoff height is measured, a board of known length is thrown into the water, the drone will then rise or lower slowly above the board between 10 and 40 m, photographs of the board are then taken from varying altitudes, and are processed in Matlab.

During my time in the GEMM Lab, I have had the pleasure of completing photogrammetry assignments for both Leila on the Oregon Coast gray whale and for Dawn on the New Zealand blue whale projects. These ladies, and the other members of the GEMM Lab, have been so patient and gracious in educating me on the workings of Matlab and the video processing systems. It is a distinct honor working with them and to delight in the astounding nature of these creatures together. Each day I am struck in sheer awe of how beautiful and powerful these whales truly are. Their graceful presence and movement through the water rivals even the most skillful dancer.

Over the last 6 years, I am delighted to say that my relationship with Hatfield has become much deeper. The people and the experiences I have encountered during my time here, especially in the GEMM Lab, have been nothing short of incredible. I am sincerely grateful for this continued opportunity. It fills my soul with joy to engage in work that contributes to the well being of the ocean and its inhabitants.

Thank you, Leigh and all of the GEMM Lab members. I hope to continue volunteering with you for as long as you will have me.

Curiosity and Community, new ways of exploring our environment.

By Nathan Malamud, GEMM Lab summer intern, Pacific High School senior

I am someone who has lived in a small town for all his life. Pretty much everyone knows each other by their first name and my graduating class only has around 20 people. Everywhere you look you will find a farm, ranch, or cranberry bog (even our school has two bogs of their own!). Because of my small town life, I have a strong sense of community. However, I have also developed a curiosity about natural and global phenomena. I try to connect these two virtues by participating in scientific efforts that help my community. When I heard that the OSU Port Orford Field Station was offering internships, I knew right away that it would definitely be a great experience for me.

The view from our field site at Fort Point in Port Orford

Port Orford, on Oregon’s southern coast, is a town that is closely tied to the ocean. So naturally, it’s important to understand and monitor our surroundings so that our town can thrive. Last year, my Marine Science class helped me further understand the complexity of the ocean. Our first semester taught us all about marine biology, zoology, and ecology. Our second semester immersed us into oceanography, ocean geology, and ocean chemistry. During the second semester, we also took trips to our town’s marine science center and to the marine reserve near Rocky Point. I loved this course and decided to try to expand my knowledge about the subject by going to the OSU Field Station.

Our safety instructor teaches takes us through basic paddling techniques

As an intern, I am currently working with three teammates to understand the feeding behavior of gray whales – what places they like to eat zooplankton the most and why they like to eat there. This whale project helps our community by Port Orford enabling high school students to perform college-level scientific research and inquiry, as well as allowing us to learn valuable skills such as CPR, surveying using a theodolite, working with chemicals in a lab, and data processing.

We had to learn how to rescue ourselves just in case we have an accident in the boat.

This internship with OSU’s GEMM Lab has taught me many new skills and given me new experiences that I have never had before. Before this internship, I had never been in a kayak. Now, I go out on the water nearly every other day! When on the water, I always try to sharpen my navigating skills. I use a GPS to pinpoint the locations of our sampling stations, and I communicate to my partner where we need to go and how we will get there.

Its very important to stretch before kayaking every morning.

Once we are there, it is my job to keep the boat close to the station location so that my partner can get accurate samples. This part is a very tricky task, because not only do I have to pay attention to the GPS to make sure we are within 10 meters of the spot, but I also have to pay attention to my surroundings. I have to look at the ocean, and figure out what direction the waves are coming from. I have to watch how external forces, like wind and currents, can cause the boat to drift far from station, and I have to correct drifting with gentle paddle strokes. This is hard, especially since the kayak is so light and easy to get pushed around by the wind. However, despite the difficulty, I have learned that it is crucial not to panic. Frustration only makes things worse. The key is to maintain a harmonic balance of concentration and zen.

I have also learned that when collecting data in the field, it’s important to observe and document as much as possible. When we are in the kayak, we have 12 stations that we try to visit every day (as long as the weather cooperates). At each station, we first use a secchi disk to test the water clarity, then lower the GoPro to film the water column and see where the zooplankton are. Sometimes we catch other interesting things on the video too, such as siphonophores (my personal favorites are jellies and salps) and rockfish.

Next we tow a zooplankton net through the water, and let it collect zooplankton of all shapes and sizes, from tiny mysids to skeleton shrimp. Then we proceed to the next station and repeat the process. We have to remember to label everything, and tell the GoPro camera what station we’re at so we can sort all the information correctly when we get back to the field station. At the end of the day, we log our data into a computer, and preserve half our plankton samples with ethanol, so that we can identify the species present. The other half gets frozen for caloric content analysis by our collaborator Dr. Kim Bernard to help us understand how much zooplankton a whale needs to eat to meet its energy needs each day.

By repeating this entire process every day, we are able to look at daily changes, which also helps us to better understand why whales spend time in certain areas and not others. Be sure to check out my teammate Maggie’s blog post about some of the tools and technologies we use to track the whales!

This whale project has been, and definitely still is, a great experience for me! I have learned a lot and have worked with some amazing people. I believe that I am learning many valuable skills, and that the skills I learn will allow me to help my community.

A new addition to the GEMM Lab

By Dr. Leigh Torres, GEMM Lab, OSU, Marine Mammal Institute

Prepping for fieldwork is always a complex mental and physical juggling act, especially for an equipment-rich, multi-disciplinary, collaborative project like our research project on the impacts of ocean noise on gray whale physiology. For me, the past couple months has consisted of remembering to coordinate equipment purchasing/testing/updating (cameras, drones, GoPros), obtaining all needed permits/licenses (NMFS, FAA, vessel), prepping data recording and management protocols (data sheets, dropbox), scheduling personnel (7 people over 5 months), organizing sampling gear (fecal nets, zooplankton traps), gathering all needed lab supplies (jars, filters, tubes), and hoping for good weather.

This list would normally be enough to overwhelm me, but this year we have also had the (fortunate) opportunity to outfit our own research vessel. The OSU Marine Mammal Institute (MMI) obtained a surplus 5.4 m coast guard RHIB (rigid inflatable haul boat) and generously handed it off to the GEMM Lab for our coastal Oregon research. Fantastic! But not perfect, of course. What the coast guard needs as a vessel, is not exactly what we need for whale research. When the vessel arrived it had a straddle seat occupying most of the limited interior space, which would make it very hard for three people to ride comfortably during a long day of survey effort or move around during whale sightings.

The RHIB in its original state, with the straddle seat taking up a majority of the interior space.

So, the boat needed a re-fit. And who better to do this re-fit than someone who has spent more than 15 years conducting whale research in a RHIB, is a certified ABYC marine electrician, and runs his own marine repair business? Who has such a qualified resume? My research technician (and husband), Todd Chandler.

Over the last two months Todd has meticulously rearranged the interior of the vessel to maximize the space, prioritize safety and comfort, balance the boat for stability, and allow for effective data collection. He removed the straddle seat, had a light-weight aluminum center console and leaning post built to just the right size and specs, installed and updated electronics (VHF, GPS chart plotter), re-ran the engine wiring (throttle, tilt, kill-switch), patched up a few (8!) leaks in the pontoons, ran new nav lights, installed new fuel tanks, and serviced the engine. Phew! He did an amazing job and really demonstrated his skills, handiwork, and knowledge of field research.

Todd, rightfully proud, with our newly designed RHIB.

The vessel now looks great, runs smoothly, and gives us the space needed for our work. But, she needed a name! So, on Saturday afternoon we hosted a GEMM Lab boat naming BBQ. Our research team and lab gathered in the sun to admire the vessel, eat good food, watch the kids run and play, and come up with boat names.

The gang gets a laugh at another good proposed name.

I was impressed by the appropriate, thoughtful, clever names put forth, like Adam’s rib, Cetacea, Oppo (re-arrange poop), and Whale Done. I was faced with a tough decision so I made everyone vote; three ticks each.

And the winner is…… Ruby: An appropriate name for a research vessel in the GEMM Lab. Perhaps someday we will have a fleet: Ruby, Emerald, Diamond… Ah, a girl can dream.

Now it’s time for the many hiccups, challenges, and rewards of a field season. So thanks to Todd, the MMI, the GEMM Lab, and our awesome team for getting us ready to go. Stay tuned for updates on the actual research (and how Ruby performs).

RV Ruby, ready to splash and find some whales.

Challenges of fecal hormone analyses (Round 2): finally in Seattle!

By Leila Lemos, Ph.D. Student, Department of Fisheries and Wildlife, OSU

In a previous blog of mine, you could read about the challenges I have been facing while I am learning to analyze the hormone content in fecal samples of gray whales (Eschrichtius robustus). New challenges appeared along the way over the last month, while I was doing my training at the Seattle Aquarium (Fig. 1).

My training lasted a week and I am truly grateful to the energy and time our collaborators Shawn Larson (research coordinator), Amy Green and Angela Smith (laboratory technicians) contributed. They accompanied me throughout my training to ensure I would be able to conduct hormonal analysis in the future, and to handle possible problems along the way.

The first step was weighing all of the fecal samples (Fig. 2A). Subsequently, the samples were transferred to appropriate glass tubes (Figs. 2B & 2C) for the next laboratorial step.

Figure 2: Analytical processes: (A) Sample weighing; (B) Transference of the sample to a glass tube; (C) Result from the performed steps.

The second conducted step was the hormone extraction. The extraction began with the addition of an organic solvent, called methanol (CH3OH), to the sample tubes (Fig. 3A & 3B). Hormones leach out from the samples and dissolve in the methanol, due to their affinity for this polar solvent.

Tubes were then placed on a plate shaker (Fig. 3C) for 30 minutes, which is used to mix the substances, in order extract the hormones from the fecal samples. The next step was to place the tubes in a centrifuge (Fig. 3D) for 20 minutes. The centrifuge uses the sedimentation principle, causing denser substances or particles to settle to the bottom of the tube, while the less dense substances rise to the top.

Figure 3: Analytical processes: (A) Methanol addition; (B) Sample + methanol; (C) Plate shaker; (D) Centrifuge.

After this process, the two different densities were separated: the high-density particles of the feces were in the bottom of the tube, while the methanol containing the extracted hormones was at the top. The top phase (methanol + hormones) was then pipetted into a different tube (Fig. 4A). The solvent was then evaporated, by using an air dryer apparatus (Fig. 4B), with only the hormones remaining in the tube.

The third performed step was dilution. A specific amount of water, measured in correlation with sample weight and to the amount of the methanol mixed with each sample, was added to each tube (Fig. 4C). Since the hormones were concentrated in the methanol, the readings would exceed the measurement limits of the equipment (plate reader). Thus, in order to prepare the extracts for the immunoassays, different dilutions were made.

Figure 4: Analytical processes: (A) Methanol transference; (B) Methanol drying; (C) Water addition.

The fourth and final step was to finally conduct the assays. Each assay kit is specific to the hormone to be analyzed with specified instructions for each kit. Since we were analyzing four different hormones (cortisol, testosterone, progesterone, and triiodothyronine – T3) we followed four different processes accordingly.

First, a table was filled with the identification numbers of the samples to be analyzed in that specific kit (Fig. 5A). The kit (Fig. 5B) includes the plate reader and several solutions that are used in the process to prepare standard curves, to initiate or stop chemical reactions, among other functions.

A standard curve, also known as calibration curve, is a common procedure in laboratory analysis for determining the concentration of an element in an unknown sample. The concentration of the element is determined by comparison with a set of standard samples of known concentration.

The plate contains several wells (Fig. 5C & 5D), which are filled with the samples and/or these other solutions. When the plate is ready, (Fig.5D) it is carried to the microplate reader that measures the intensity of the color of each of the wells. The intensity of the color is inversely proportional to the concentration of the hormone in both the standards and the samples.

Figure 5: (A) Filling the assay table with the samples to be analyzed; (B) Assay kit to be used; (C) Preparation of the plate; (D) Plate ready to be read.

Since this is the first fecal hormone analysis being performed in gray whales, a validation process of the method is required. Two different tests (parallelism and accuracy) were performed with a pool of three different samples. Parallelism tests that the assay is measuring the antigen (hormone) of interest and also identifies the most appropriate dilution factor to be used for the samples. Accuracy tests that the assay measurement of hormone concentration corresponds to the true concentration of the sample (Brown et al. 2005).

This validation process only needs to be done once. Once good parallelism and accuracy results are obtained, and we have identified the correct dilution factor and approximate concentration of the samples, the samples are ready to be analyzed. Below you can see examples of a good parallelism test (parallel displacement; Fig. 6) and bad parallelism tests (Fig. 7) that indicate no displacement, low concentration or non-parallel displacement; and a good accuracy test (Fig. 8).

Figure 6: Example of a good parallelism test. The dark blue line indicates the standard curve; the pink line indicates a good parallelism test, showing a parallel displacement; and the ratios in black indicate the dilution factors.
Source: Brown et al. (2005)

Figure 7: Examples of bad parallelism tests. The dark blue line indicates the standard curve; the light blue line is an example of no displacement; the pink line is an example of low concentration of the sample; and the green line is an example of non-parallel displacement.
Source: Brown et al. (2005)

Figure 8: Example of a good accuracy test while analyzing hormone levels of pregnanediol glucuronide (Pdg) in elephant urine. The graph shows good linearity (R2 of 0.9986) and would allow for accurate concentration calculations.
Source: Brown et al. (2005)

After the validation tests returned reliable results, the samples were also analyzed. However, many complications were encountered during the assay preparations and important lessons were learned that I know will allow this work to proceed more smoothly and quickly in the future. For instance, I now know to try to buy assay kits of the same brand, and to be extremely careful while reading the manual of the process to be performed with the assay kit. With practice over the coming years, my goal is to master these assay preparations.

Now, the next step will be to analyze all of the results obtained in these analyses and start linking the multiple variables we have from each individual, such as age, sex and body condition. The results of this analysis will lead to a better understanding of how reproductive and stress hormones vary in gray whales, and also link these hormone variations to nutritional status and noise events, one of my PhD research goals.

Cited Literature:

Brown J, Walker S and Steinman K. 2005. Endocrine manual for reproductive assessment of domestic and non-domestic species. Smithsonian’s National Zoological Park, Conservation and Research Center, Virginia 1-69.

Challenges of fecal analyses (Round 1)

By Leila Lemos, Ph.D. Student, Department of Fisheries and Wildlife, OSU

Fieldwork is done for the year and lab analyses just started with some challenges. This is not unexpected since no previous hormonal analysis has been conducted with any gray whale tissue, and whale fecal sample analysis is a relatively new technique. So, I have been thinking, learning, consulting, and creating a methodology as I go along. I am grateful to the expert advice and help from many great collaborators:

Kathleen Hunt (Northern Arizona University, AZ, United States)
Shawn Larson (Seattle Aquarium, WA, United States)
Amy Green (Seattle Aquarium, WA, United States)
Rachel Ann Hauser-Davis (Fiocruz, RJ, Brazil)
Maziet Cheseby (Oregon State University, OR, United States)
Scott Klasek (Oregon State University, OR, United States)

I have learned that an important step before undertaking fecal a hormonal analysis is the desalting process of the samples since salts can interfere in hormonal determinations, leading to false results. In order to remove salt content, each sample was first filtered (Fig. 1A), to remove a majority of the salt water content (Fig. 1B) that is inevitably collected along with the fecal sample. Each sample was then re-suspended in ultra-pure water, to dilute the remaining salt content in a higher water volume (Fig. 1C).

Figure 1: Analytical processes: (A) Filtration of the samples; (B) Result from filtration; (C) Addition of pure water to the samples.

After these steps were completed for each sample, the samples were centrifuged (Fig. 2A) to precipitate the fecal matter and leave the lighter salt ions in the supernatant (the liquid lying above a solid residue; Fig. 2B). After finishing these two phases, the water was removed with aid of a plastic pippete (Fig. 2C), and I was left with only desalted fecal at the bottom of the tubes (Fig. 2D).

Figure 2: Analytical processes: (A) Samples centrifugation; (B) Result from the centrifugation; (C, D) Results from separating water and sample.

The fecal samples were then frozen at -80°C (Fig. 3A & 3B) and then freeze-dried on a lyophilizer for 2 days to remove all remaining water content (Fig. 3C). Finally, I have what I need: desalted, dry fecal samples, ready for hormone analysis (Fig. 3D).

Figure 3: Analytical processes: (A) Freezing process of the samples; (B) Frozen samples ready to go to the lyophilizer; (C) Samples in the lyophilizer; (D) Final result of the lyophilizing process.

Writing this now, this process seems simple, but it was laborious, and took time to find the equipment needed at the right times. The end product is crucial to get a good final result, so my time investment (and my own increased stress level!) was worth it. This type of analysis is very new for marine mammals and our research lab is still in the learning the best methods. Along the way we were unsure of some decisions, some mistakes were made, and we were afraid of losing precious fecal material. But, this is the fun and challenge of working with a new species and new type of sample and, importantly, we have developed a working protocol that should make the process more efficient and reduce our stress levels next time around.

At the end of this sample preparation process, our 53 samples look great and are ready to be analyzed during my training at the Seattle Aquarium. We are also planning to analyze the water that was removed from the samples (Fig. 2D) to see if any hormone leached out from the poop into the water.

Results from this process will aid in future whale fecal hormone studies. Perhaps only the centrifugation step is needed and we can discard the water without losing hormone content. Or, perhaps we need to analyze both portions of the sample and sum the hormones found in each. We shall know the answer when we get our hormone metabolite results. Just another protocol to be worked out as I move ahead with the hormone analysis of these fecal samples. And through all these challenges I keep the end goal of this work in my mind: to learn about the reproductive and stress hormonal variation in gray whales and to link these variations to nutritional status and noise events. Onward!

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