Author: fernaale

Scouting mission to Kodiak: Reconnaissance of potential gray whale research in Kodiak, Alaska.

Dr. KC Bierlich, Dr. Alejandro Fernández Ajó, and Dr. Leigh Torres, OSU Department of Fisheries, Wildlife, & Conservation Sciences, Geospatial Ecology of Marine Megafauna Lab

Eastern North Pacific (ENP) gray whales (Eschrichtius robustus) undertake one of the longest annual migrations of any mammal, traveling from their winter breeding grounds in the warm waters of Baja California, Mexico to their summer feeding grounds in the icy waters of the Bering and Chukchi Seas1,2. Yet, a distinct subgroup of this population, called the Pacific Coast Feeding Group (PCFG), instead shorten their migration farther south to spend the summer foraging along waters from northern California, USA to northern British Columbia, Canada1 (Figure 1). On these summer feeding grounds gray whales will forage almost continuously to increase their energy reserves to support migration and reproduction during the rest of the year.

The GEMM Lab has been studying the ecology and physiology of the PCFG gray whales in Oregon waters since 2015, combining traditional photo-ID and behavioral observation methods with fecal sample collection, drone flights, and prey assessment to integrate data on individual whale behavior, nutritional status, prey consumption, and hormone variation. These multidisciplinary methods have proven effective to obtain an improved understanding of PCFG gray whale body condition and hormone variation by demographic unit and over time3,4,5, as well as prey energetics and foraging ecology6.

Figure 1. Left: ENP Gray whale´s range from the breeding grounds in Baja California, Mexico to the northernmost feeding grounds in the Arctic. Right: Overview of Kodiak Island; the red square shows a zoom in image of the study area, including the shore – and boat-based data collection sites in yellow.

Since the PCFG remains a small proportion (~230 individuals) of the larger eastern ENP population (~20,000 individuals), the GEMM Lab and multiple collaborators are interested in extending the research design implemented by the GEMM Lab in Oregon to study gray whale ecology and physiology of whales feeding on the more northern foraging grounds. The goal would be to fill some of the many critical knowledge gaps including gray whale resilience and response to climate change, connectivity between foraging grounds, population dynamics of the PCFG and ENP, and physiological variation (body condition, hormones) as a function of habitat, prey, demography, and time of year.

Kodiak Island, Alaska is a middle distance between PCFG foraging grounds in Newport, Oregon and the traditional ENP foraging grounds in Chukchi and Beaufort Seas (Figure 1). Two studies documented high gray whale encounter rates in Ugak Bay in Kodiak Island, including during summer months when foraging behavior was observed7,8. Evidence from photo-ID matches in these studies indicated that some PCFG whales might also extends their feeding grounds further north to Kodiak Island7,8.

During August-September of this year, GEMM Lab postdocs KC Bierlich and Alejandro Fernández Ajó traveled to Kodiak Island to assess opportunities for researching gray whales in the area. The mission objectives included determining gray whale presence, assessing behavioral states and foraging areas, determining feasibility of drone operations and fecal sample collection, collecting photo ID images, assessing feasibility of boat and shore-based operations in Ugak Bay (Figure 1), and connecting with local scientists and stakeholders interested in collaborating.

We landed in Kodiak the evening of August 28 (Figure 2), with a beautiful sunset. The next morning, we met our captain, Alexus Kwachka, over breakfast to discuss a plan for going offshore to look for gray whales later in the week. Alexus is a local fisherman in Kodiak with over 30 years of experience fishing in Alaska and incredible knowledge on local wildlife and navigating the rough Alaskan seas. It was particularly interesting to hear his stories on the local changes he has noticed over the years, not just in weather and fishing, but also in the seals, birds, and whales.

Figure 2. Arriving to Kodiak after a long day of travel.

Next, we met with Sun’aq Tribe’s biologist Matthew Van Daele, who coordinates the marine mammal stranding network on Kodiak Island and has a deep knowledge of the locations to find whales. Matt showed us several great spots to scout for gray whales along the shore in the Pasagshak area (Figure 1), which overlooks Ugak Bay and is about 1 hour drive from Kodiak (Figure 3). Along the way, Matt discussed the high mortality rate of gray whales he has observed over the past two years and his concerns about some skinny whales in the area he recently observed during aerial surveys. Since 2019, an Unusual Mortality Event (UME) of gray whales along the whole North Pacific west coast (Mexico, USA, Canada) has impacted the ENP gray whales and while the exact cause(s) of these mortalities is largely unknown, evidence suggests reduced nutritional status may be a likely cause of death9. We learned from Matt that while gray whale strandings are decreasing compared to the previous two years, the numbers are still concerningly high. It was an absolute pleasure spending the day with Matt, as being born and raised in Kodiak he has such great knowledge of the area and the local wildlife. Together we saw Kodiak’s beautiful landscape with lots of different wildlife, which included some huge Kodiak brown bears a few hundred meters away from the road (Figure 4).

Figure 3. The views from Pasagshak Point that are good observation locations for gray whales. The gray arrows represent the view looking left (A) and right (C) from Pasagshak Point (B). A panorama of the view from left to right on the point is also shown (D). Photo: KC Bierlich.
Figure 4. Sighting of a Kodiak brown bear (Ursus arctos) off the roadside on our way to Pasagshak. The Kodiak brown bear is the largest recognized subspecies (or population) of the brown bear, and one of the largest bears alive today. Photo: Alejandro Fernández Ajó.

The next day, the weather was great, so we returned to the Pasagshak lookout points to spend the day looking for whales. We spotted several gray whales from the cliffs and shore. At Burton Beach, we spotted a gray whale very close to shore that first appeared to be traveling, but then changed direction and started moving closer inshore –less than 10 m from where we were standing on the beach! The whale then swam back and forth along the shore, providing an opportunity to collect photos of its right and left side to use for photo ID. KC flew the drone over the whale and recorded some amazing behavior of lateral swimming and great images for photogrammetry. Our excitement was sky high as within two days on the trip we had documented the presence of gray whales, recorded the best places to work from land, and even captured some interesting behavior, photo ID, and photogrammetry data from shore! (Figure 5).

Figure 5. Gray whale feeding off Burton Beach, Kodiak Island. This photo was taken from the shore, as the whale swam back and forth amazingly close to the shoreline. In this picture you can see the whale´s head from a ventral perspective. Photo: Alejandro Fernández Ajó / GEMM Lab. Photograph captured under NOAA/NMFS permit #21678.

The weather deteriorated over the next couple days, bringing foggy and rainy conditions. We used this time to process data and meet with some of the local researchers. When the weather conditions improved, we met back up with Alexus and boarded his fishing vessel, “No Point”, and headed off to Ugak Bay to look for gray whales. During transit we encountered a humpback whale mother-calf pair lunge feeding and breaching (Figure 6). As we approached Pasagshak we sighted a gray whale diving and benthic feeding in 60 m water depth, and then 2-3 other individual whales exhibiting the same behavior close by. We collected photo ID data, but high wind conditions hindered drone operations, so we continued surveying further into Ugak Bay and turned around following the coast towards Gull Point (Figure 7).

Figure 6. A breaching humpback whale on the way to Ugak Bay from Kodiak. Photo: Alejandro Fernández Ajó / GEMM Lab. Photograph captured under NOAA/NMFS permit #21678.
Figure 7. Track line (shown in blue) of boat-based operations. White circles represent the locations for sightings of gray whales.

During our survey effort we spotted a gray whale foraging on a shallow rocky, kelp reef (12 m depth) along the northwest point of Ugak Bay. This sighting was similar to behavior we often observe in Oregon, with whales feeding in near shore shallow, reef habitats. Conditions for flying the drone were still too windy, but we observed the whale defecate and collected a fecal sample! For us, fecal samples are like “biological gold”, as we can study hormones (which include assessments of their reproductive status, nutritional condition, sex determination, and stress levels), genetics, prey, and much more! We were so excited to collect this sample because it provides the chance to start looking at the physiological parameters of these Alaskan whales and compare findings to what we observe in samples collected from whales in Oregon (Figure 8).

Figure 8. A gray whale fecal sample right after being scooped from the water using nets attached long aluminum poles. Photo: KC Bierlich.

After a beautiful night anchored in a sheltered bay near Gull Point (Figure 9) we continued west to scan for whales. Back in Ugak Bay, we found six more gray whales diving and feeding in 50-60 m depth near the same location as the previous day off Pasagshak point. Weather conditions had finally improved, allowing us to fly the drone. We flew over four whales and collected video for behavior and photogrammetry analysis, which allows us to measure the body condition of the whales to assess how healthy it is (Figure 10).

Figure 9. “Home sweet home” for the night where our vessel “No point” anchored in a sheltered bay. Photo: Alejandro Fernández Ajó.
Figure 10. Drone image of two gray whales feeding near each other. Note the trailing sediment plume from the whale’s mouth and body indicating it was bottom feeding in a muddy benthic habitat. Photo: KC Bierlich. Photograph captured under NOAA/NMFS permit #21678.

Another highlight of our field work was the collection of a benthic prey sample using a Ponar grab sampler at this location in Pasagshak Bay where the whales were foraging. The bottom was muddy and rich with invertebrates; the sample literally looked like it was boiling from the amount of prey in it (Figure 11). From this sample, we will determine the invertebrate species and caloric content of these prey for comparison to the prey found in Oregon waters.

Figure 11. The Ponar bottom grab sample, full of invertebrate prey, taken near whales feeding in ~50-60 m depth. Photo: CK Bierlich.

Overall, this scouting mission to Kodiak was a great success! Through boat surveys, shore-based observations, and the conversations with locals, we determined the best areas and timing to effectively work from boats and shore to expand our gray whale research to Kodiak. Moreover, our scouting mission resulted in the collection of relevant pilot data including fecal samples for hormonal analyses, drone images for body condition and behavioral assessments, prey samples, and photo-ID images. This scouting mission identified several knowledge gaps regarding gray whale ecology, physiology, and population connectivity that can be feasibly addressed through expansion of GEMM Lab research efforts to the Alaskan region. Importantly, the trip facilitated important networking with locals to establish potential collaborations for future work. We are optimistic and excited to grow our collaborative research in Kodiak.

This pilot project was funded by sales and renewals of the special Oregon whale license plate, which benefits MMI. We gratefully thank all the gray whale license plate holders, who made this scouting trip possible.

References:

1 Calambokidis J, Darling JD, Deecke V, Gearin P, Gosho M, Megill W, et al. Abundance, range and movements of a feeding aggregation of gray whales (Eschrichtius robustus) from California to south- eastern Alaska in 1998. J Cetacean Res Manag 2002; 4:267–76.

2Stewart JD, Weller DW. Abundance of eastern North Pacific gray whales 2019/2020. 2021. U.S. Department of Commerce, NOAA Technical Memorandum NMFS-SWFSC-639. https://doi.org/10. 25923/bmam-pemorandum NMFS-SWFSC-639. https://doi.org/10. 25923/bmam-pe91.

3Lemos, L. S. et al. Assessment of fecal steroid and thyroid hormone metabolites in eastern North Pacific gray whales. Conserv. Physiol. 8, (2020).

4Lemos, L. S. et al. Stressed and slim or relaxed and chubby? A simultaneous assessment of gray whale body condition and hormone variability. Mar. Mammal Sci. 1–11 (2021). doi:10.1111/mms.12877

5Soledade Lemos, L., Burnett, J. D., Chandler, T. E., Sumich, J. L. & Torres, L. G. Intra‐ and inter‐annual variation in gray whale body condition on a foraging ground. Ecosphere 11, (2020).

6Hildebrand, L., Bernard, K. S. & Torres, L. G. (2021). Do Gray Whales Count Calories? Comparing Energetic Values of Gray Whale Prey Across Two Different Feeding Grounds in the Eastern North Pacific. Frontiers in Marine Science, 8(July), 1–13. https://doi.org/10.3389/fmars.2021.683634

7Gosho Merrill, Patrick Gearin, Ryan Jenkinson, Jeff Laake, Lori Mazzuca, David Kubiak, John Calambokidis, Will Megill, Brian Gisborne, Dawn Goley, Christina Tombach, James Darling, V. D. gosho_et_al._2011_-_sc-m11-awmp2.pdf. (2011).

8Moore, S. E., Wynne, K. M., Kinney, J. C. & Grebmeier, J. M. GRAY WHALE OCCURRENCE AND FORAGE SOUTHEAST OF KODIAK, ISLAND, ALASKA. Mar. Mammal Sci. 23, 419–428 (2007).

9Christiansen F, Rodríguez-González F, Martínez-Aguilar S, Urbán J and others (2021) Poor body condition associated with an unusual mortality event in gray whales. Mar Ecol Prog Ser 658:237-252. https://doi.org/10.3354/meps13585

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Optimizing hormone extraction protocols for whale baleen

By Alejandro Fernández Ajó, Postdoc, OSU Department of Fisheries, Wildlife, and Conservation Science, Geospatial Ecology of Marine Megafauna Lab

Large whale conservation is challenged by our limited understanding of the impacts of natural and anthropogenic disturbances on the whale´s health and its population level consequences. To better mitigate human-wildlife conflicts, we need to improve our ability to predict multi-scale responses of whales to disturbances, describe and identify disease dynamics, and understand the reproductive biology of whales (Madliger, 2020; McCormick and Romero, 2017). Conservation physiology and conservation endocrinology can provide tools to illuminate the underlying physiological mechanisms whales use to cope with changing environments and different stressors, thus filling information gaps to guide management and conservation actions.

In brief, conservation physiology is a multidisciplinary field wherein a broad suite of tools and concepts are used to understand how organisms and ecosystems respond to both environmental and anthropogenic change and stressors (Madliger et al., 2020). Conservation endocrinology is a subdiscipline within conservation physiology, which relies on endocrine measurements (hormone quantifications). However, monitoring the physiology of free ranging animals in wild populations presents many technical challenges and it is particularly difficult when studying whales. Traditionally, conservation endocrinology relied on laboratory analyses of plasma samples (derived from blood). Yet implementing this techniques for monitoring the physiology of mysticetes (baleen whales) is currently impossible, as there are no feasible, non- (or minimally) invasive, methods to obtain a blood sample from living large whales (Hunt et al., 2013).

Therefore, we are interested in the development and further validation of alternative sample types from whales to obtain endocrine data. During my Ph.D. dissertation I worked to develop and ground truth the endocrine analyses of whale baleen as a novel sample type that can be used for retrospective assessments of the whale´s physiology. Baleen, the filter-feeding apparatus of the mysticete whales (Figure 1), consists of long fringed plates of stratified, keratinized tissue that grow continuously and slowly downward from the whale´s upper jaw (Hunt et al., 2014). Baleen plates are readily accessible at necropsy and routinely collected from carcasses of stranded whales.

Figure 1: Southern Right Whale skim feeding at the surface. Baleen plates hanging downwards are visible in the image. Photo: Alejandro Fernández Ajó / Instituto de Conservación de Ballenas de Argentina.

Like hair, nails, feathers, spines, or horns of other animals, baleen is a keratinized tissue that can store steroid and thyroid hormones in detectable and relevant concentrations to provide an integrated measure of hormonal plasma levels over the period that the structure was growing. Thus, baleen contains a progressive time-series that captures months and often years of an individual’s endocrine history with sufficient temporal resolution to determine seasonal endocrine patterns allowing to explore questions that have historically been difficult to address in large whales, including pregnancy and inter-calving interval, age of sexual maturation, timing and duration of seasonal reproductive cycles, adrenal physiology, and metabolic rate. Additionally, their robust and stable keratin matrix allows baleen samples to be stored for years to decades, enabling the analysis and comparison of endocrine patterns from past and modern populations. Therefore, keratinized sample matrices are valuable tools to investigate reproductive and stress physiology in whales and other vertebrates.

However, due to its novelty, the extraction and analysis of hormones from baleen and other keratinized tissues requires both biological and analytical validations to ensure the method fulfills the requirements for its intended use. Baleen hormone analyses has already passed several essential assay validations, including parallelism and accuracy of immunoassays (Hunt et al., 2017b), and numerous biological validations, such as the study of animals with known physiological status (i.e., pregnancy, and known stress events such as entanglement in fishing gear or presence of lesions) to assess the degree to which the endocrine data reflect the physiology of the individual (Fernández Ajó et al., 2020, 2018; Hunt et al., 2018, 2017a; Lysiak et al., 2018; Palme, 2019). Yet, other questions essential for technical validation remain unknown, including choice and volume of extraction solvent, the effect of solvent-to-sample ratio (solvent:sample) on extraction yield, and the amount of sample (e.g., mg) needed for analysis to obtain reliable hormonal data.

In our recent contribution, Optimizing hormone extraction protocols for whale baleen: Tackling questions of solvent:sample ratio and variation, we aimed to tackle two of these important questions: “1) what is the minimum sample mass of baleen powder required to reliably quantify hormone content of baleen samples analyzed using commercially available enzyme immune assays (EIAs); and 2) what is the optimal ratio of solvent volume to sample mass for steroids extracted from baleen, i.e., the ratio that yields the maximum amount of hormone with high accuracy and low variability between replicates.”

We performed the extraction with methanol and tested a variety of sample masses with the objective to provide methodological guidance regarding optimizing sample mass and solvent volume for steroid hormone extraction from powdered baleen. Our results suggest that the optimal sample mass for methanol extraction of steroid hormones from baleen samples is 20 mg, and that larger sample masses did not produce either better yield or less variation in the apparent hormone per g of baleen sample (Figure 2). In addition, when the extraction was performed keeping the volume of solvent proportional to the sample mass (namely, a solvent:sample ratio of 80:1), masses as small as 10 mg yielded reliable hormone measurement (Figure 2).

Figure 2. Extraction experiment results for variable (solid circles) or constant (open circles) solvent:sample ratio (mean progesterone concentration ± standard error of the mean). Extractions for 5, 10, 20, 50, and 75 mg of baleen powder were performed in triplicate for each sample mass. Letters indicate significant differences between groups after Tukey’s post hoc test. The right-side panel presents a plot for 100 mg of baleen powder extracted with 8 mL of solvent performed in triplicate; error bars are too small to show in the figure. Source: Fernández Ajó et al., 2021.

Our results indicate how baleen hormone analytic techniques can be more widely employed on small sample masses from rare specimens (i.e., less sample than is currently employed, which is typically 75 mg or 100 mg in most studies to date), such as from natural history museums and stranding archives. Thus, we demonstrate that greater use of this valuable technique to reconstruct the endocrine and physiological history of individual whales over time can be achieved with reduced sample size (so reduced damage to the sample). I hope these findings encourage researchers to apply these methods more broadly to analyze historical archives of baleen plates that can date back to the era of commercial whaling, and modern archives of baleen collected from stranded animals to help continue further developing techniques that can make headway in gaining conservation-relevant physiological knowledge of this particularly challenging taxon.

Bibliography:

Fernández Ajó, A., Hunt, K.E., Dillon, D., Uhart, M., Sironi, M., Rowntree, V., Loren Buck, C., 2021. Optimizing hormone extraction protocols for whale baleen: Tackling questions of solvent:sample ratio and variation. Gen. Comp. Endocrinol. 113828. https://doi.org/10.1016/j.ygcen.2021.113828

Fernández Ajó, A.A., Hunt, K.E., Giese, A.C., Sironi, M., Uhart, M., Rowntree, V.J., Marón, C.F., Dillon, D., DiMartino, M., Buck, C.L., 2020. Retrospective analysis of the lifetime endocrine response of southern right whale calves to gull wounding and harassment: A baleen hormone approach. Gen. Comp. Endocrinol. 296, 113536. https://doi.org/10.1016/j.ygcen.2020.113536

Fernández Ajó, A.A., Hunt, K.E., Uhart, M., Rowntree, V., Sironi, M., Marón, C.F., Di Martino, M., Buck, C.L., 2018. Lifetime glucocorticoid profiles in baleen of right whale calves: potential relationships to chronic stress of repeated wounding by Kelp Gulls. Conserv. Physiol. 6, 1–12. https://doi.org/10.1093/conphys/coy045

Hunt, K.E., Lysiak, N.S., Moore, M., Rolland, R.M., 2017a. Multi-year longitudinal profiles of cortisol and corticosterone recovered from baleen of North Atlantic right whales (Eubalaena glacialis). Gen. Comp. Endocrinol. 254, 50–59. https://doi.org/10.1016/j.ygcen.2017.09.009

Hunt, K.E., Lysiak, N.S., Robbins, J., Moore, M.J., Seton, R.E., Torres, L., Loren Buck, C., Buck, C.L., 2017b. Multiple steroid and thyroid hormones detected in baleen from eight whale species. Conserv. Physiol. 5. https://doi.org/10.1093/conphys/cox061

Hunt, K.E., Lysiak, N.S.J., Matthews, C.J.D., Lowe, C., Fernández Ajó, A., Dillon, D., Willing, C., Heide-Jørgensen, M.P., Ferguson, S.H., Moore, M.J., Buck, C.L., 2018. Multi-year patterns in testosterone, cortisol and corticosterone in baleen from adult males of three whale species. Conserv. Physiol. 6, 1–16. https://doi.org/10.1093/conphys/coy049

Hunt, K.E., Moore, M.J., Rolland, R.M., Kellar, N.M., Hall, A.J., Kershaw, J., Raverty, S.A., Davis, C.E., Yeates, L.C., Fauquier, D.A., Rowles, T.K., Kraus, S.D., 2013. Overcoming the challenges of studying conservation physiology in large whales: a review of available methods. Conserv. Physiol. 1, cot006–cot006. https://doi.org/10.1093/conphys/cot006

Hunt, K.E., Stimmelmayr, R., George, C., Hanns, C., Suydam, R., Brower, H., Rolland, R.M., 2014. Baleen hormones: a novel tool for retrospective assessment of stress and reproduction in bowhead whales (Balaena mysticetus). Conserv. Physiol. 2, cou030–cou030. https://doi.org/10.1093/conphys/cou030

Lysiak, N.S.J., Trumble, S.J., Knowlton, A.R., Moore, M.J., 2018. Characterizing the Duration and Severity of Fishing Gear Entanglement on a North Atlantic Right Whale (Eubalaena glacialis) Using Stable Isotopes, Steroid and Thyroid Hormones in Baleen. Front. Mar. Sci. 5, 1–13. https://doi.org/10.3389/fmars.2018.00168

Madliger, C. L., Franklin, C. E., Love, O. P., & Cooke, S.J. (Ed.), 2020. Conservation Physiology: Applications for Wildlife Conservation and Management., 1st ed. Oxford University Press. https://doi.org/10.1093/oso/ 978019883610.001.0001

McCormick, S.D., Romero, L.M., 2017. Conservation Endocrinology. Bioscience 67, 429–442. https://doi.org/10.1093/biosci/bix026

Palme, R., 2019. Non-invasive measurement of glucocorticoids: Advances and problems. Physiol. Behav. 199, 229–243. https://doi.org/10.1016/j.physbeh.2018.11.021