A dominant language for scientific communication can streamline the process of science, but it also can create barriers and inequality

Dr. Alejandro A. Fernández Ajó, Postdoctoral Scholar, Marine Mammal Institute – OSU Department of Fisheries, Wildlife, & Conservation Sciences, Geospatial Ecology of Marine Megafauna (GEMM) Lab.

The English language is recognized as the international language of science (Gordin, 2015); I believe this is a useful convention that allows scientists to communicate ideas and gain access to global scientific literature regardless of their origin or native tongue. However, this avenue for sharing knowledge is open only for those proficient in English, and many scientists and users of scientific information, such as policy makers and conservationists, communicate on a daily basis in languages other than English. This inevitably creates barriers to the transfer of knowledge between communities, potentially impacting conservation and management because scientific knowledge is often unavailable in local languages.

Although in non-English speaking countries, local journals are receptive to publishing scientific research in languages other than English (i.e., their local language), oftentimes these local journals are perceived as low-quality and have a relatively low impact factor, making publishing in such journals less attractive to scientists. Therefore, readers with language barriers only have access to limited studies and are often unaware of the most significant research, even when the research is conducted in their region. This situation can result in a void of information relevant for environmental policies and conservation strategies. Ensuring that research findings are available in the local language of the region in which the research is conducted is an important step in science communication, but one that is often neglected.

In addition, scientists with English as a Foreign Language (EFL) confront the added challenge of navigating a second language while writing manuscripts, preparing and presenting oral presentations, and developing outreach communications (Ramirez-Castaneda, 2020). For example, EFL researchers have reported that one of the primary targets of criticism for their manuscripts under review is often the quality of their English rather than the science itself (Drubin and Kellogg, 2012). In academia, most job interviews and PhD applications are conducted in English; and grant and project proposals are often required to be written in English, which can be particularly challenging and can impact the allocation of resources for research and conservation in non-English speaking regions.

I am from Argentina, and I am a native Spanish speaker. I am fortunate to have started learning English at an early age and continue practicing with international collaborations and traveling abroad. Being able to communicate in English has opened many doors for me, but I recognize that I am in a privileged position with respect to many Argentinians and South Americans in general, where the majority of the population receives minimal training in English and bilingualism with English is very low. Thus, socioeconomic status can influence English proficiency, which then determines scientific success and access to knowledge. I believe that the scientific community should be aware of these issues and work towards improving equality in the process of research collaborations. Providing opportunities for students, and enhancing the availability of scientific knowledge for non-English speaking communities, particularly when the research is relevant for such communities.

In this picture I am with an international group of Fulbright scholars during the Spring International Language Program at the University of Arkansas. This is on of many activities organized by the Fulbright program to create bridges across cultures and languages.

Fortunately, there are several examples pointing towards improving equality in the scientific process, access to knowledge, and opportunities for EFL communities in STEM careers. Several journals are now accepting, or considering to accept the publication of papers in multiple languages. One example of this is the journal Integrative Organismal Biology, which provides the option for publishing the paper abstract in multiple languages. In our recent publication, “Male Bowhead Whale Reproductive Histories Inferred from Baleen Testosterone and Stable Isotopes,” we provided an abstract in five different languages, including Inuktitut, one of primary languages of indigenous groups in the area. And, international exchange programs like the Fulbright Foreign Student Program, of which I was a beneficiary between 2018-2020, enable graduate students and young professionals from abroad to study and conduct research in the United States.

In an effort to contribute to addressing these problems, I am working with a group of colleagues from Argentina (María Constanza (Kata) Marchesi and Tomas Marina) to develop graduate level coursework that will be offered at the Universidad Nacional de la Patagonia in Puerto Madryn, Argentina, with the objective to enable students to learn effective communication using English in the scientific environment. Unfortunately, these types of programs focused on EFL proficiency for STEM students are currently rare in Argentina, but my hope is that our work can spur the creation of additional programs for EFL students in STEM across the region.

I want to finish this post with the acknowledgement of the huge support I have form the GEMM Lab, which welcomes diversity, equity, and inclusivity, and promotes a culture of anti-racism, transparency, and acceptance (See the GEMM Lab DEI statement here).

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References and Additional Readings

Gordin, M. D. (2015). Scientific Babel : How Science Was Done Before and After Global English. Chicago, IL: University of Chicago Press.

Ramírez-Castañeda V (2020) Disadvantages in preparing and publishing scientific papers caused by the dominance of the English language in science: The case of Colombian researchers in biological sciences. PLoS ONE 15(9): e0238372. https://doi.org/10.1371/journal.pone.0238372

Drubin, D. G., and Kellogg, D. R. (2012). English as the universal language of science: opportunities and challenges. Mol. Biol. Cell 23:1399. doi: 10.1091/mbc.E12-02-0108

Amano, T., González-Varo, J. P., & Sutherland, W. J. (2016). Languages are still a major barrier to global science. PLoS Biology, 14(12), e2000933. https://doi.org/10.1371/journal.pbio.2000933

Marden, E., Abbott, R. J., Austerlitz, F., Ortiz-Barrientos, D., Rieseberg, L. H. (2021). Sharing and reporting benefits from biodiversity research. Molecular Ecology, 30(5), 1103–1107. https://doi.org/10.1111/mec.15702

Márquez, M. C., & Porras, A. M. (2020). Science communication in multiple languages Is critical to Its effectiveness. Frontiers in Communication, 5(May). https://doi.org/10.3389/fcomm.2020.00031

Ramírez-Castañeda V (2020) Disadvantages in preparing and publishing scientific papers caused by the dominance of the English language in science: The case of Colombian researchers in biological sciences. PLoS ONE 15(9): e0238372. https://doi.org/10.1371/journal.pone.0238372

Trisos, C. H., Auerbach, J., & Katti, M. (2021). Decoloniality and anti-oppressive practices for a more ethical ecology. Nature Ecology and Evolution, 5(9), 1205–1212. https://doi.org/10.1038/s41559-021-01460-w

Woolston, C, & Osório, J. (2019). When English is not your mother tongue. Nature 570, 265-267. https://doi.org/10.1038/d41586-019-01797-0

Letter to the Editor of Marine Mammal Science: Enhancing the impact and inclusivity of research by embracing multi-lingual science communication (2022) DOI: 10.13140/RG.2.2.29934.08001 http://dx.doi.org/10.13140/RG.2.2.29934.08001

Leal, J. S., Soares, B., Franco, A. C. S., de Sá Ferreira Lima, R. G., Baker, K., & Griffiths, M. (2022). Decolonizing ecological research: a debate between global North geographers and global South field ecologists. https://doi.org/10.31235/osf.io/wbzh2

A pregnancy test for whales?! Why and how?

Dr. Alejandro A. Fernández Ajó, Postdoctoral Scholar, Marine Mammal Institute – OSU Department of Fisheries, Wildlife, & Conservation Sciences, Geospatial Ecology of Marine Megafauna (GEMM) Lab.

I often receive two reactions when asked what I am currently working on; one is “Wow! That is a very cool job, it must be amazing to work with such incredible animals!”, the other is “How do you do that and why is that important?”. So, today I decided to blog about some of the reasons why it is important to develop a pregnancy test for gray whales and how we are doing this.

In a previous blogpost, I described the many ways in which whales play critical roles in sustaining marine ecosystem. Briefly, whales can enhance marine productivity by vertically and horizontally mixing of ocean waters, promoting primary production, and mitigating climate change by sequestering carbon with their large biomass and long life-span (1-3). Even after they die, their carcasses can contribute to biodiversity creating new habitat on the seafloor (4). But, over several decades, the whaling industry drastically removed whales around the globe, with some species and populations depleted to near extinction (5). Consequently, these depleted whale populations now play a diminished role in ocean ecosystem processes and their recovery is currently challenged by an increasing number of modern anthropogenic impacts. Hence, working towards whale conservation is essential for keeping a healthy marine ecosystem.

Working and designing effective strategies for conservation biology often involves gaining knowledge regarding the reproductive parameters of individual animals in wild populations. This information is critical for understanding population trends and the underlaying mechanisms that affect animal welfare and their potential for recovery. However, getting such information from free-living whales can be challenging (see Hunt et al. 2013). While we know that whales typically have long life-spans, lengthy generation times, extended parental care, and high survival rates, detailed knowledge on the life history and general reproductive biology of free-ranging whales is limited for the majority of the whale populations. In fact, much of what we do know about whale reproduction is derived from whaling records. Only recently, conservation physiology approaches (see our previous post here) have contributed alternative and non-invasive methods for monitoring key physiological processes that can help monitor a whale’s reproductive biology and determine reproductive parameters such as sexual maturity and pregnancy (6-9).

In this clip you can see an example of a fecal sample collection from a gray whale off the Oregon coast. We can look at hormones in the fecal samples which are useful indicators for endocrine assessments of free-swimming whales. Fecal sample and footage filmed under NOAA/NMFS permit #16111.

Gray whales (Eschrichtius robustus) in the Eastern North Pacific (ENP) typically undertake annual migrations between their lower latitude breeding grounds in the coastal waters of the Baja California Peninsula, Mexico, and the foraging grounds located on the Bering and Chukchi Seas (10). However, among the ENP whales a distinct subgroup of about 230 whales shorten their migration to feed in the coastal waters of Northern California, Oregon, and southeastern Alaska (11). This group of whales is known as the gray whale Pacific Coast Feeding Group (PCFG).

Since 2016, the GEMM Lab has monitored individual gray whales within the PCFG off the Oregon coast (check the GRANITE project). Gray whales have a distinct mottled skin; and each individual whale presents a unique pigmentation pattern that allows for the individual identification of whales. We can identify who is who among the whales who visit the Oregon coast. In this way, we can keep a detailed record of re-sightings of known individuals (visit our new web site to know more about the lives of individual whales that visit the Oregon coast).  We have high individual re-sighting rates, so this unique opportunity helps us keep a long-term data series for individual whales to monitor their health, body condition, and reproductive status over time, and thus further develop and advance our non-invasive study methods.

We are combining behavioral and feeding ecology with drone photogrammetry and endocrinology of the same individual whales to help us understand the relationships between natural and anthropogenic drivers with biological parameters. In this way, following individual whales, we are developing sensitive biomarkers to monitor and infer about the population health, population trends, and identify stressors that impact their recovery and welfare. In particular, we are now working to develop a noninvasive approach to detect pregnancy in gray whales based on fecal hormone analyses.

In this picture you can see “Rose”, a gray whale calf, on top of her mother “Scarlett”. Scarlett is one of the most recognizable whales from the PCFG, due to a large scar on the right side of her back (not visible in this picture). She has been observed along the Pacific NW coast since 1996, so she is at least 26 years old today. We know 3 of her calves. Following individual whales like Scarlett is helping us to better understand the gray whale reproductive biology. Photo by Alejandro Fernandez Ajo taken under NOAA/NMFS permit #21678.

In marine mammals, the progesterone hormone is secreted in the ovaries during the estrous cycle and gestation, and is the predominant hormone responsible for sustaining pregnancy (12). As the hormones are cleared from the blood into the gut, they are metabolized and eventually excreted in feces; fecal samples represent a cumulative and integrated concentration of hormone metabolites (13-14), which are useful indicators for endocrine assessments of free-swimming whales. Several studies show that changes in hormone concentration correlate in meaningful ways with exposure to stressors (15-16) and changes in reproductive status (17-19). We are using our long data series of fecal hormones and individual life histories to advance our understanding on the gray whales’ reproductive biology. We are close to developing a technique that will allow us to detect pregnancy in whales based in fecal hormones analyses and photogrammetry. Stay tuned for results from this pregnancy test!

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References:

1- Pershing AJ, Christensen LB, Record NR, Sherwood GD, Stetson PB (2010) The impact of whaling on the ocean carbon cycle: Why bigger was better. PLoS ONE 5(8): e12444.

2- Roman J and McCarthy JJ. 2010. The whale pump: marine mammals enhance primary productivity in a coastal basin. PLoS ONE. 5(10): e13255.

3- Morissette L, Kaschner K, and Gerber LR. 2010. “Whales eat fish”? Demystifying the myth in the Caribbean marine ecosystem. Fish Fish 11: 388–404.

4- Smith CR, Roman J, Nation JB. A metapopulation model for whale-fall specialists: The largest whales are essential to prevent species extinctions. J. Mar. Res. 77, 283–302 (2019).

5- Branch TA, Williams TM. Legacy of industrial whaling. Whales. Whal. Ocean Ecosyst. 2006, 262–278 (2006).

6- Kellar NM, Keliher J, Trego ML, Catelani KN, Hanns C, George JC, et al. Variation of bowhead whale progesterone concentrations across demographic groups and sample matrices. Endanger Species Res 2013; 22:61–72. https://doi.org/10.3354/esr00537.

7- Pallin L, Robbins J, Kellar N, Berube M, Friedlaender A. Validation of a blubber-based endocrine pregnancy test for humpback whales. Conserv Physiol 2018;6:1 11. https://doi.org/10.1093/conphys/coy031PMID:29942518.

8-Hunt KE, Robbins J, Buck CL, Bérubé M, Rolland RM (2019) Evaluation of fecal hormones for noninvasive research on reproduction and stress in humpback whales (Megaptera novaeangliae). Gen Comp Endocrinol 280: 24–34.

9-Melica, V., Atkinson, S., Calambokidis, J., Lang, A., Scordino, J., & Mueter, F. (2021). Application of endocrine biomarkers to update information on reproductive physiology in gray whale (Eschrichtius robustus). Plos one, 16(8), e0255368.

10-Swartz SL. Gray Whale. In: Wursig B, Thewissen JGM, Kovacs KM, editors. Encyclopedia of Marine Mammals (Third Edition). Elsevier;2018,p. 422–8.https://doi.org/10.1016/B978-0-12-804327-1.00140–0.

11-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.

12- Bronson, F. H. (1989). Mammalian reproductive biology. University of Chicago Press.

13-Wasser SK, Hunt KE, Brown JL, Cooper K, Crockett CM, Bechert U, Millspaugh JJ, Larson S, Monfort SL (2000) A generalized fecal glucocorticoid assay for use in a diverse array of nondomestic mammalian and avian species. Gen Comp Endocrinol120:260–275.

14- Hunt, K.E., Rolland, R.M., Kraus, S.D., Wasser, S.K., 2006. Analysis of fecal glucocorticoids in the North Atlantic right whale (Eubalaena glacialis). Gen. Comp. Endocrinol. 148, 260–272. https://doi.org/10.1016/j.ygcen.2006.03.01215.

15- Lemos, L.S., Olsen, A., Smith, A., Burnett, J.D., Chandler, T.E., Larson, S., Hunt, K.E., Torres, L.G., 2021. Stressed and slim or relaxed and chubby? A simultaneous assessment of gray whale body condition and hormone variability. Mar. Mammal Sci. 1–11. https://doi.org/10.1111/mms.12877

16- Rolland, R., McLellan, W., Moore, M., Harms, C., Burgess, E., Hunt, K., 2017. Fecal glucocorticoids and anthropogenic injury and mortality in North Atlantic right whales Eubalaena glacialis. Endanger. Species Res. 34, 417–429. https://doi.org/10.3354/esr00866.

17-Rolland, R.M., Hunt, K.E., Kraus, S.D., Wasser, S.K., 2005. Assessing reproductive status of right whales (Eubalaena glacialis) using fecal hormone metabolites. Gen. Comp. Endocrinol. 142, 308–317. https://doi.org/10.1016/j.ygcen.2005.02.002

18- Valenzuela Molina M, Atkinson S, Mashburn K, Gendron D, Brownell RL. Fecal steroid hormones reveal reproductive state in female blue whales sampled in the Gulf of California, Mexico. Gen Comp Endocrinol 2018;261:127–35.https://doi.org/10.1016/j.ygcen.2018.02.015 PMID:29476760.

19- Hunt, K. E., Robbins, J., Buck, C. L., Bérubé, M., & Rolland, R. M. (2019). Evaluation of fecal hormones for noninvasive research on reproduction and stress in humpback whales (Megaptera novaeangliae). General and Comparative Endocrinology, 280, 24-34.

Social turmoil due to the approval of an offshore oil exploration project off the coast of Argentina.

Dr. Alejandro A. Fernández Ajó, Postdoctoral Scholar, Marine Mammal Institute – OSU Department of Fisheries, Wildlife, & Conservation Sciences, Geospatial Ecology of Marine Megafauna (GEMM) Lab.

I just returned to my home country, Argentina, after over 2 years without leaving the USA due to COVID-19 travel restrictions. Being back with my family, my friends, my culture, and speaking my native language feels great and relaxing. However, I returned to a country struggling to rebound from the coronavirus pandemic. I am afraid this post pandemic scenario places Argentina in a vulnerable situation. The need for economic growth could result in decisions or policies that, in the long term, hurt the country, leaving environmental damage for potential economic growth.

Argentina holds extensive oil and gas deposits, including the world’s second largest gas formation, Vaca Muerta. Although offshore (i.e., in the ocean) oil exploration and exploitation are not yet extensively developed, the intention of offshore gas and oil drilling is on the agenda. In July 2021, a public hearing was held with the purpose to consider the environmental impact assessment for carrying out seismic exploration in the North Argentinian basin off the southern coast of the Buenos Aires province. Over 90% of the participants, including scientists, researchers, technicians from various institutions, non-governmental organizations and representatives of the fishing sector spoke against the project and highlighted the negative impacts that such activity can generate on marine life, and to other socioeconomic activities such as tourism and fishing, not only in Argentina but at the regional level.

Thousands of people marched along the beaches and the main coastal cities of Argentina to protest against the approval for a seismic explorations project in the Argentinian basin. Photo source: prensaobrera.com

Seismic prospections are usually done with the purpose for oil and gas exploitation and less frequently for research purposes. In seismic prospections, ships carry out explosions with airguns, whose sound waves reach the seabed, bounce back and are captured by receivers on the ships to map the petroleum deposits in seafloor and identify potential areas for hydrocarbon extractions. The sound emitted by the seismic airguns can reach extremely loud levels of sounds that travel for thousands of miles underwater. Such extreme high levels of sound can alter the behavior of many marine species, from the smallest planktonic species, to the largest marine mammals, masking their communication, causing physical and physiological stress, interfering with their vital functions, and reducing the local availability of prey (Di Iorio & Clark, 2010; Hildebrand, 2009; Weilgart, 2018).

Here you can listen to a short audio clip of a seismic airgun firing every ~8 seconds, a typical pattern. Close your eyes and imagine you are a whale living in this environment. Now, put the clip on loop and play it for three months straight. This would be the soundscape that whales living in a region of oil and gas exploration hear, as seismic surveys often last 1-4 months (see our previous post “Hearing is believing” for more details).

Despite the public rejection and the mounting evidence about the negative impacts and environmental risks associated with such activities, the government approved the initiation of the seismic prospection off the southern coast of Buenos Aires. In response, thousands of people marched along the beaches and the main coastal cities of Argentina to protest against the oil exploration project. The areas where the seismic surveys will be carried out overlap largely with the southern right whale’s migration routes and feeding areas during their spring and summer (Figure 1). Likewise, the area overlaps with highly productive areas in the ocean that hosts great biodiversity of species of ecological and commercial importance, including the feeding areas of seabirds, turtles and other marine mammals. Additionally, the seismic activity will endanger the health of the beaches of the coast of Buenos Aires and Uruguay where thousands of tourists spend the summer to escape from the large cities.

Figure 1. The map on the left is showing (light blue squares CAN_100, CAN_108, and CAN_114) the areas where seismic prospections are proposes. The map on the right is showing the individual satellite track lines for eleven individual southern right whales (SRW) during the feeding season. You can observe that the proposed area for seismic explorations overlaps with critical feeding habitat for the SRW. Source: Whale Conservation Institute of Argentina (ICB).

The impacts of these activities to marine wildlife are difficult to control and monitor (Elliott et al. 2019, Gordon et al, 2003), especially for large whales that are a very challenging taxa to study (Hunt et al. 2013). We know that the ability to perceive biologically important sounds is critical to marine mammals, and acoustic disturbance through human-generated noise can interfere with their natural functions. Sounds from seismic surveys are intense and have peak frequency bands overlapping those used by baleen whales (Di Lorio & Clark, 2010); however, evidence of interference with baleen whale acoustic communication, and the effects on their health and physiology are sparse. In this context, the GEMM Lab project GRANITE (Gray Whale Response to Ambient Noise Informed by Technology and Ecology), plans to generate information to fulfill these knowledge gaps and provide tools to aid conservation and management decisions in terms of allowable noise level in whale habitats. I am hopeful such information will reach decision makers and influence their decisions, however, sometimes it is frustrating to see how evidence-based information generated with high quality standards are often ignored.

The recent approval of the seismic exploration in Argentina is an example of my frustration. There is no way that the oil industry can guarantee a low-risk of impact on biodiversity and the environment. There are too many examples of environmental catastrophes related to the oil industries at sea that speak for themselves. Moreover, the promotion of such activities goes against the compromises assumed by the country to work to mitigate the effects of Climate Change, and to achieve the reductions of the greenhouse gas emissions to comply with the Paris Agreement. Decades of research help recognized the areas that would be impacted by these seismic activities as key habitat for the life cycle of whales, penguins, seals and more. But, apparently all these scientific data are ignored at the time of weighing the tradeoffs between “economic development” and environmental impacts. As a conservation biologist, I am questioning what can be done in order to be heard and significantly influence such decisions.

Did you enjoy this blog? Want to learn more about marine life, research, and conservation? Subscribe to our blog and get a weekly alert when we make a new post! Just add your name into the subscribe box on the left panel.

References:

  • Di Iorio, L., & Clark, C. W. (2010). Exposure to seismic survey alters blue whale acoustic communication. Biology Letters, 6(1), 51–54. https://doi.org/10.1098/rsbl.2009.0651
  • Weilgart, L. (2018). The impact of ocean noise pollution on fish and invertebrates. Report for OceanCare, Switzerland.
  • Elliott, B. W., Read, A. J., Godley, B. J., Nelms, S. E., & Nowacek, D. P. (2019). Critical information gaps remain in understanding impacts of industrial seismic surveys on marine vertebrates. In Endangered Species Research (Vol. 39, pp. 247–254). Inter-Research. https://doi.org/10.3354/esr00968
  • Gordon, J., Gillespie, D., Potter, J., Frantzis, A., Simmonds, M. P., Swift, R., & Thompson, D. (2003). A review of the effects of seismic surveys on marine mammals. Marine Technology Society Journal37(4), 16-34.
  • 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. Conservation Physiology, cot006–cot006. https://doi.org/10.1093/conphys/cot006

Harmful algal blooms expose southern right whales to domoic acid and can potentially cause endocrine alterations

Dr. Alejandro Fernández Ajó, OSU Department of Fisheries, Wildlife, & Conservation Sciences, Geospatial Ecology of Marine Megafauna Lab

Rises in ocean temperatures can lead to multiple alterations in marine ecosystems, including the increase and the frequency of Harmful Algal Blooms (HABs). HABs are characterized by the rapid growth of toxin-producing species of algae that can be harmful to people, animals, and the local ecology, even causing death in severe cases. Species of marine diatom within the genus Pseudo-nitzschia and Nitzschia can form HABs when they produce domoic acid (DA), a potent neurotoxin responsible for amnesic shellfish poisoning (D’Agostino et al., 2018, 2017).

Figure 1. Southern right whale (E. australis) mother and calf swimming at the gulfs of Peninsula Valdes, Argentina, during a phytoplankton bloom. Photo: Mariano Sironi / Instituto de Conservacion de Ballenas de Argentina.

During HABs, DA is transferred to higher organisms through the pelagic food web and is accumulated by intermediate vectors, such as copepods, euphausiids (i.e., krill), shellfish, and fish. As this neurotoxin affects top predators, DA poisoning poses a risk to the safety and health of humans and wildlife. This neurotoxin has caused mortality in many marine mammal species, including both pinnipeds and cetaceans (Gulland 1999; Lefebvre et al. 1999; Fire et al. 2010, 2021; Broadwater et al. 2018). In addition, the exposure to DA constitutes a stressor that may affect glucocorticoids (hormones involved in the stress response) concentrations.

The glucocorticoids (GCs; cortisol and corticosterone) are adrenal steroid hormones that maintain the essential functions of metabolism and energy balance in mammals. GCs can increase sharply in response to environmental stressors to elicit physiological and behavioral adaptations by individuals to support survival (Sapolsky et al. 2000; Bornier et al. 2009). However, with the chronic exposure to a stressor, this relationship can reverse, with GCs sometimes declining below its baseline levels (Dickens and Romero, 2013; Fernández Ajó et al., 2018). Moreover, DA can interfere with the stress response in mammals, and cause alterations in their physiological response. DA is an excitatory amino acid analog of glutamate (Pulido 2008), a well-known brain neurotransmitter that play an important role in the activation of the adrenal axis (which in turn regulate the production and secretion of the GCs) and regulate many of the pituitary hormones involved in the stress response (Brann and Mahesh 1994; Johnson et al. 2001). Hence, monitoring GC levels in marine mammals can be a potential useful metric for assessing the physiological impacts of exposure to DA.

Glucocorticoids are traditionally measured in plasma, but given that plasma sampling from free-ranging large whales is currently impossible, alternative sample types such as fecal samples, among others, can be utilized to quantify GCs in large whales (Ajó et al., 2021; Burgess et al., 2018, 2016; Fernández Ajó et al., 2020, 2018; Hunt et al., 2019, 2014, 2006; Rolland et al., 2017, 2005)(Figure 2). The analyses of fecal glucocorticoid metabolites (fGCm) is particularly useful for endocrine assessments of free-swimming whales, with several studies showing that fGCm correlate in meaningful ways with presumed stressors. For example, high levels of fGCm in North Atlantic right whales (NARW, Eubalaena glacialis) and in gray whales (Eschrichtius robustus) correlate with poor body condition (Hunt et al., 2006; Lemos et al., 2021), and fGCm increases were associated with whale entanglements and ship strikes (i.e., Lemos et al., 2020; Rolland et al., 2017).

Figure 2. Alternative samples types can be used to study hormones in large whales. 1-2-3 are sample types that can be obtained from free-living whales and provide a more instantaneous and acute measurement of the whales´ physiology. 4-5 can be obtained at necropsy when the whale is found dead at the beach and provide an integrated measure of the whale physiology that can expand through years or even the lifespan of an individual.

In Península Valdés, Argentina, southern right whales (SRW, E. australis) gather in large numbers to mate and nurse their calves during the austral winter months (Bastida and Rodríguez, 2009). SRWs are capital breeders, largely fasting during the breeding season and instead relying on stored blubber fuel reserves. However, they can occasionally feed on calanoid copepods (D’Agostino et al., 2018, 2016), particularly during the phytoplankton blooms that are dominated by diatoms of the genus Pseudo-nitzschia (Sastre et al. 2007; D’Agostino et al. 2015, 2018). Therefore, feeding SRWs in Península Valdés temporally overlap with these Pseudo-nitzschia blooms (D’Agostino et al. 2018, 2015) and represents a test case for assessing the relationship of DA exposure with GC levels (Figure 3).

Figure 3. Southern right whale (E. australis) skim feeding at the Peninsula Valdes breeding ground. Photo: Lucas Beltranino.

In our recent scientific publication (D’Agostino et al. 2021), we investigate SRW exposure to DA at their breeding ground in Peninsula Valdes and assessed its effects on fecal glucocorticoid concentrations. Although the sample size of this study is unavoidably small due to the difficulties of obtaining fecal samples from whales at their calving grounds where defecation is infrequent, we observed significantly lower fGCm in samples from whales exposed to DA (Figure 4). Our results agree with findings from a previous study in California sea lions (Zalophus californianus) exposed to DA, where these authors found a significant association of DA exposure with reduced serum cortisol (Gulland et al., 2009), which can be tentatively attributed to abnormal function of the adrenal axis due to the exposure.

Figure 4. Fecal glucocorticoid metabolite levels in southern right whales exposed (YES, solid triangles) and not-exposed (NO, open circles) to DA. Left panel: immunoreactive fecal corticosterone metabolites. Right panel: immunoreactive fecal cortisol metabolites. Hormone concentrations are expressed in ng of immunoreactive hormone per gram of dry fecal sample. Significant differences between groups are denoted with an asterisk (P<0.05). The black solid line indicates the mean for each group, and in parenthesis is the sample size for each group. Adapted from D’Agostino et al. 2021.

If ingestion of toxins produced by phytoplankton can result in long-term suppression of baseline GCs, whales and marine mammals in general, could suffer reduced ability to cope with additional stressors. The adrenal function is essential to maintain circulating blood glucose and other aspects of metabolism within normal bounds. Additionally, the ability to elevate GCs facilitates energy mobilization to physiologically cope with a stressful event and to initiate appropriate behavioral responses (i.e., flee from predators, heal wounds). Various toxicants have been shown to reduce adrenal function across taxa (Romero and Wingfield, 2016) and could have negative consequences on the ability of cetaceans to respond and adapt to ongoing environmental and anthropogenic changes. Compounding this problem, whales are exposed to an increasing number of stressors from multiple sources and with cumulative effects and they need to be able to physiologically respond to continue to reproduce and survive.

To our knowledge, this study provides the first quantification of fGCm levels in whales exposed to DA; and we hope this effort starts a growing dataset to which other researchers can add. Sampling and analysis of non-traditional matrices, such as feces, blubber, baleen and others, would likely increase sample sizes and thus our understanding of the interrelationships among DA exposure and age, sex, and reproductive status of cetaceans. Given that chronic exposure to DA could alter the capacity of animals to respond to stress, and indications that HABs are becoming more frequent and intense world-wide (Van Dolah 2000; Masó et al. 2006; Erdner et al. 2008), we believe that research evaluating the health status of marine mammal populations should include the assessment of stress physiology relative to natural and anthropogenic stressors including exposure to toxicants.

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References

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Bonier F, Moore IT, Martin PR, Robertson RJ (2009) The relationship between fitness and baseline glucocorticoids in a passerine bird. Gen Comp Endocrinol 163:208–213. https:// doi. org/ 10. 1016/j.ygcen. 2008. 12. 013.

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Broadwater MH, Van Dolah FM, Fire SE (2018) Vulnerabilities of marine mammals to harmful algal blooms. Harmful Algal Blooms 2:191–222.

Burgess, E.A., Hunt, K.E., Kraus, S.D., Rolland, R.M., 2016. Get the most out of blow hormones: Validation of sampling materials, field storage and extraction techniques for whale respiratory vapour samples. Conserv. Physiol. 4, cow024. https://doi.org/10.1093/conphys/cow024

Burgess, E.A., Hunt, K.E., Kraus, S.D., Rolland, R.M., 2018. Quantifying hormones in exhaled breath for physiological assessment of large whales at sea. Sci. Rep. 8, 10031. https://doi.org/10.1038/s41598-018-28200-8

D’Agostino VC, Hoffmeyer MS, Almandoz GO, Sastre V, Degrati M., 2015. Potentially toxic Pseudo-nitzschia species in plankton and fecal samples of Eubalaena australis from Península Valdés calv-ing ground, Argentina. J Sea Res 106:39–43. https:// doi. org/ 10.1016/j. seares.

D’Agostino, V.C., Degrati, M., Santinelli, N., Sastre, V., Dans, S.L., Hoffmeyer, M.S., 2018. The seasonal dynamics of plankton communities relative to the foraging of the southern right whale (Eubalaena australis) in northern Patagonian gulfs, Península Valdés, Argentina. Cont. Shelf Res. 164, 45–57. https://doi.org/10.1016/j.csr.2018.06.003

D’Agostino, V.C., Degrati, M., Sastre, V., Santinelli, N., Krock, B., Krohn, T., Dans, S.L., Hoffmeyer, M.S., 2017. Domoic acid in a marine pelagic food web: Exposure of southern right whales Eubalaena australis to domoic acid on the Península Valdés calving ground, Argentina. Harmful Algae 68, 248–257. https://doi.org/10.1016/j.hal.2017.09.001

D’Agostino, V.C., Hoffmeyer, M.S., Degrati, M., 2016. Faecal analysis of southern right whales (Eubalaena australis) in Península Valdés calving ground, Argentina: Calanus australis, a key prey species. J. Mar. Biol. Assoc. United Kingdom 96, 859–868. https://doi.org/10.1017/S0025315415001897

Dickens, M.J., Romero, L.M., 2013. A consensus endocrine profile for chronically stressed wild animals does not exist. Gen. Comp. Endocrinol. 191, 177–189. https://doi.org/10.1016/j.ygcen.2013.06.014

Erdner DL, Dyble J, Parsons ML, Stevens RC, Hubbard KA, Wrabel ML, Moore SK, Lefebvre KA, Anderson DM, Bienfang P, Bidi-gare RR, Parker MS, Moeller P, Brand LE, Trainer VL (2008) Centers for Oceans and Human Health: a unified approach to the challenge of harmful algal blooms. Environ Health 7:S2. https://doi. org/ 10. 1186/ 1476- 069X-7- S2- S2.

Fernández Ajó, A., Hunt, K.E., Dillon, D., Uhart, M., Sironi, M., Rowntree, V., Buck, C.L., 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

Fire SE, Bogomolni A, DiGiovanni RA Jr, Early G, Leighfield TA, Matassa K, Miller GA, Moore KM, Moore M, Niemeyer M, Pugliares K (2021) An assessment of temporal, spatial and taxonomic trends in harmful algal toxin exposure in stranded marine mammals from the US New England coast. PLoS ONE 16(1):e0243570. https:// doi. org/ 10. 1371/ journ al. pone. 02435 70

Fire SE, Wang Z, Berman M, Langlois GW, Morton SL, Sekula-Wood E, Benitez-Nelson CR (2010) Trophic transfer of the harmful algal toxin domoic acid as a cause of death in a minke whale (Balaenoptera acutorostrata) stranding in southern California. Aquat Mamm 36(4):342–350. https:// doi. org/ 10. 1578/ AM. 36.4.2010. 342.

Gulland F.M., 1999. Domoic acid toxicity in California sea lions stranded along the central California Coast, May-October 1998. NOAA Tech. Memo. NMFS-OPR-8. USA National Marine Fisheries Service, US Department of Commerce.

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Hunt, K.E., Robbins, J., Buck, C.L., Bérubé, M., Rolland, R.M., 2019. Evaluation of fecal hormones for noninvasive research on reproduction and stress in humpback whales (Megaptera novaeangliae). Gen. Comp. Endocrinol. 280, 24–34. https://doi.org/10.1016/j.ygcen.2019.04.004

Hunt, K.E., Rolland, R.M., Kraus, S.D., Wasser, S.K., 2006. Analysis of fecal glucocorticoids in the North Atlantic right whale (Eubalaena glacialis). Gen. Comp. Endocrinol. 148, 260–272. https://doi.org/10.1016/j.ygcen.2006.03.012

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

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Lemos, L.S., Olsen, A., Smith, A., Burnett, J.D., Chandler, T.E., Larson, S., Hunt, K.E., Torres, L.G., 2021. Stressed and slim or relaxed and chubby? A simultaneous assessment of gray whale body condition and hormone variability. Mar. Mammal Sci. 1–11. https://doi.org/10.1111/mms.12877

Lemos, L.S., Olsen, A., Smith, A., Chandler, T.E., Larson, S., Hunt, K., Torres, L.G., 2020. Assessment of fecal steroid and thyroid hormone metabolites in eastern North Pacific gray whales. Conserv. Physiol. 8. https://doi.org/10.1093/conphys/coaa110

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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.

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).


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:

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