Assessing reproductive hormones in adult female blue whales (Balaenoptera musculus) by analyzing historic baleen samples from the 1940’s

Monitoring reproductive parameters such as gestation period and calving intervals in blue whale (Balaenoptera musculus) populations is difficult due to limitations in hormone data collection methods for large whales. The most significant hindrance is that it is impossible to collect blood samples from these animals due to their size; therefore, alternative methods for hormone analysis must be investigated. Recent studies have shown that hormones like progesterone can be detected in baleen powder of other mysticete whale species like bowhead (Balaena mysticetus) and North Atlantic right (Eubalaena glacialis) whales. I hypothesized that baleen from female blue whales would contain regions of high progesterone indicating prior pregnancies. To test this, I ran enzyme immunoassays to quantify progesterone from serial samples taken along the length of historic baleen plates of four female blue whales, two of which were known to be pregnant upon capture. All of the specimens in this study were originally captured by post World War II era Japanese commercial whaling vessels and are archived at the Smithsonian National Museum of Natural History. This time period overlaps a pause in commercial whaling globally, and predates significant impacts of climate change, implying a relatively low stress environment for this population compared to recent years. Most baleen plates had several broad regions of high progesterone, as predicted, with patterns suggesting a 2-year calving interval for all but one female. A possible reason for this outlier could be due to the age of the individual, as she was the smallest individual in the sample size and may have not yet been reproducing. Two individuals known to be pregnant at death had high progesterone in the most recently grown baleen, as predicted. These findings may clarify historic norms of reproduction in blue whales, and could be helpful for comparisons to modern populations.

Hi, everyone! My name is Nadia Gray. I’m an Environmental and Sustainability Studies student, and for this semester I’ve been working with Dr. Kathleen Hunt of the Biology department here at George Mason, and my project is assessing reproductive hormones in adult female blue whales by analyzing historic baleen samples from the 1940’s. So let’s talk about the blue whale. So this is a marine mammal, and it is the largest animal ever to have lived. As far as we know. it is about as big as 3 school buses lined up bumper to bumper, and as heavy as 25 African elephants in one room and on average, they can live anywhere between 80- 90 years. And the reason why we’re talking about the blue whale is because, despite the fact that commercial whaling has been banned since 1986. This animal is still considered endangered by the IUCN Red List. This is due to other stressors, such as climate change, and although their population is increasing, we still have yet to see stable numbers. And that being said, we also don’t know that much about blue whale reproduction, and that is because these animals are too large to collect blood samples, from which is how hormone data is generally generated. There have been some studies trying to use other methods, such as fecal samples, respiratory vapor, and blubber collections. However, just do the nature of these animals. It is a very difficult method. For this project we’re going to be looking into using baleen as a method. and balen is the keratin-based apparatus you see growing inside of a whale’s mouth, and it is used to filter feed out krill. And for this study we’re also going to be looking at the steroid hormone progesterone, and that is because in high amounts we believe that progesterone can indicate pregnancy. For our research questions we’re going to be looking at is progesterone on detectable in historic baleen. How long is the gestation period of a female blue whale? And what was the number of calving intervals during the mid to late 1940’s. The sample size that we looked at in this project consists of 4 adult females, 2 of which were pregnant upon capture, and 3 of which had an intact root. And that is an important piece of data, because baleen grows similarly to how teeth grow where it has a root that anchors into the gums, but isn’t necessarily visible. So if you were to cut at the gum line, you’d be losing out that little last piece of data. The first step is to collect your samples. So this was done by drilling along the length of each baleen plate and collecting powder samples every centimeter. next up is to weigh out the powder of each centimeter of the baleen plate. We measured out around 20 grams for each specimen. And then you can move on to the extraction process, which is where you are, adding methanol to the powder, and that is what pulls out the hormones. And then the next step is, you are going to remove that methanol that has been sitting in the powder and separate it from the powder pellet that you see there. Next, you’re going to move on to the reconstitution phase which this is after you have dried down that previous methanol sample, and you are essentially rehydrating it so you can create samples with it, and then you’re going to have to go ahead and dilute those samples, because the progesterone, and sometimes is too high in specimens to be able to actually read it. And for my specimens. All of them were one to 4 dilutions, except for one that was one to 40. And then finally, you can move on to the assay portion. So each sample that you have created is going to go into each of these wells, and once you’re done with that, it’ll look something like this and then you can place the assay tray into what is called a spectrophotometer. And this brings us to our results. So this is what the raw data looks like. Everywhere you see light colors. That is what is considered to be high progesterone, and everywhere you see is dark. That is what we consider to be low progesterone, and the unit that is used in the spectrophotometer is called ocular density. And then this is what the final data ends up looking like. These graphs are made through a program called Prism, and if you are interested in a more in depth description of what is going on with each of these graphs. You are going to have to look at my poster of the same name, but essentially anywhere you see, a yellow bar is what we consider to be high progesterone. and on average, we found that these whales were experiencing, on average, of a 2 year calving interval, with the exception of
Figure D, which that one only had 2 high progesterone points, and prior to that was all low progesterone, which we later found out was the smallest world population, so this could potentially indicate that she had not yet been reproducing, because she might have been younger than the other specimens in this sample. And then the next steps: The purpose of this research is to generate baseline data for future hormone studies in blue whales. So essentially we are creating a control group and by testing whales from other time periods we can use this and determine what is considered normal reproduction. This knowledge can also be used to apply towards future management plans of the species. And this brings us to the end of the presentation. I would like to thank Dr. Michael Mcgowan and John Sosky from the Smithsonian National Museum of Natural history. I’d like to thank Allie Case and Dr. Kathleen Hunt over at side Tech at George Mason University, as well as Dr. Alyson, Fleming and Malia Smith, of the University of North Carolina, Wilmington.