Image Courtesy of Catherine Kwon.
The Galapagos Islands have been an archetypal example of evolution and natural selection since 1835, when famed naturalist Charles Darwin set foot on their remote shores. Upon arriving on the islands, Darwin discovered animals that were similar in many ways but differed in their adaptations to the unique environment of their respective home islands. These observations led Darwin to develop his revolutionary theory of natural selection, detailing how populations of organisms change and adapt to succeed in their specific environments.
One of the most iconic residents of the Galapagos Islands is the Galapagos giant tortoise. After arriving at the Galapagos Islands from mainland South America millions of years ago, the tortoises slowly evolved into fifteen unique species across ten of the islands in the Galapagos, with twelve of these species surviving to this day. These species are all specifically adapted to succeed in the isolated environments they inhabit. For decades, researchers believed that the tortoises of San Cristóbal Island—one of the oldest islands in the Galapagos archipelago—belonged to a single lineage. However, Adalgisa Caccone, a Senior Research Scientist in Yale’s Department of Ecology and Evolutionary Biology, along with a team of eleven colleagues, recently found evidence of a second, long-extinct lineage of tortoises on San Cristóbal Island. In her group’s new study, published in Heredity in February, Caccone and her team sequenced DNA from museum samples of tortoises more than a century old, suggesting the existence of a previously unknown lineage of tortoises.
A Long Time Coming
The story of this groundbreaking discovery began nearly three decades ago when Caccone and her husband, Professor Jeffrey Powell of the Department of Ecology and Evolutionary Biology, first became interested in studying the giant tortoises to save them from extinction. This threat has haunted the tortoise population since the arrival of humans to the Galapagos. To early fisherman and whalers, the turtles were a meal, not an evolutionary miracle. “They were a good source of fresh food because you can keep them on the boat without food or water for up to six months,” Caccone said. “It’s been estimated that 250,000 tortoises were taken across the islands.”
Caccone was confident that the key to managing endangered tortoise populations lay in piecing together the puzzle of their evolutionary origins. In collaboration with the Galapagos National Park and the Charles Darwin Foundation, she began collecting samples from the surviving populations of Galapagos tortoises.
The next step in uncovering the story of the tortoises’ evolution was interrogating their DNA. “We started sequencing small fragments of genes, like the mitochondrial DNA, some nuclear genes, and genotyping microsatellite loci,” Caccone said. “We started building the story.” By comparing the DNA data of the living populations, Caccone could identify similarities and differences, generating new understandings of how the tortoises might have spread across the islands and adapted.
Her efforts in uncovering these patterns led her to San Cristóbal Island. “San Cristóbal is one of the oldest islands of the Galapagos Islands, so it likely plays an important role in understanding how the colonization of the islands started,” Caccone said.
Still, a piece of the evolutionary puzzle was missing. “We soon realized that there were some questions that we could not address because we needed the DNA of the extinct species,” Caccone said.
A Surprising Solution
Caccone and her team found their solution in unexpected places: historical samples from a single living tortoise and tortoise bones collected from a cave during an expedition to San Cristóbal in 1906, which had been stored away in museum collections more than a century ago. Caccone set to work sequencing DNA from these samples—a particularly challenging task due to the age of the samples.
Using micro-CT scans, which use x-rays to build a 3-D image of the interior of an object on a very small scale, Caccone and her team could determine the sections of the old bones most likely to contain preserved DNA. They sequenced small sections of DNA from each bone, overcoming the challenges of working with tortoise bones that were hundreds of years old and highly degraded. “What you want to get is a sample that has a good amount of DNA preserved in it,” Caccone said. “Usually, bones are much better than tissues, because the external mineralized portion of the bone protects the DNA.”
Caccone expected to find a variety of DNA sequences related to the single existing haplotype, or set of DNA variations, found in the living tortoises from San Cristóbal. Instead, she found something very surprising. “We found that unlike in the existing population, there was variation, and that the variation was not related to the existing haplotype,” Caccone said. “It was something different. That’s when we said, okay, there’s something interesting here. Let’s get to work.”
And work they did—Caccone and her team convinced the Galapagos National Park, with which they are working in close collaboration, to organize a field expedition over San Cristóbal Island to check if there were any tortoises related to the lineage from the cave samples. The team collected hundreds of samples across the island, searching in remote places never previously sampled before. Caccone and her team selected a random collection of 129 blood samples from living tortoises to test if any showed signs of relatedness with the samples collected in the cave.
The genome-wide analyses of samples from living tortoises did not reveal any intermixing between the two lineages, suggesting the existence of an entirely different, long-dead lineage of tortoises on San Cristóbal Island. “The key finding is that the mitochondrial haplotypes for these cave specimens are highly distinct from the haplotype that’s in the living population,” Evelyn Jensen, first author on the Heredity paper and lecturer at Newcastle University, said in a podcast with Heredity. “The common ancestor of the cave and the living lineages diverged probably about 700,000 years ago. So, the tortoises that died falling into the cave are of a totally different lineage from the tortoises that live on the island today.”
New Ideas from Old Bones
Jensen and Caccone’s discovery has brought scientists closer to completing the puzzle of giant tortoise evolution and colonization in the Galapagos Islands. It has also powerfully demonstrated the importance of analyzing both historical and contemporary samples when studying these tortoises. “If we’re just looking at the populations that have survived to the present day, we’re missing some important pieces of the puzzle for reconstructing evolutionary patterns,” Jensen said in the Heredity podcast. “Just when we think we have the story worked out, we look at some dusty old bones in the museum. Now we’re potentially having to do a major rethink of our understanding of how these species arose and colonized the Galapagos archipelago.”
Caccone and her team are already working on their next step—obtaining more comprehensive DNA samples from the historical samples. The DNA data from this study is limited to mitochondrial DNA, a form of DNA that is more abundant and easier to isolate. However, this subset of DNA lacks much of the broader genetic information of the tortoises, limiting options for genetic analysis. In the future, Caccone hopes to isolate and sequence nuclear DNA, or DNA from the tortoise’s chromosomes, which will provide a much more complete picture of the genetic differences between the cave tortoises and the existing population. The ability to compare full nuclear DNA sequences between extinct and living populations holds the potential to elucidate new knowledge about the relationship between these two tortoise lineages. Further sequencing might even confirm that this unique lineage is an entirely new tortoise species—the evolutionary secrets within these bones have yet to be fully revealed.
About the Author: Sophia Burick is a first-year Molecular Biophysics and Biochemistry major in Timothy Dwight College. In addition to writing for YSM, Sophia sings with the New Blue, Yale’s oldest women’s acapella group, and works as a research assistant at the Yale School of Medicine.
Acknowledgments: The author would like to thank Adalgisa Caccone for her time and enthusiasm for her research.
Extra reading:
Jensen, E. L., Quinzin, M. C., Miller, J. M., Russello, M. A., Garrick, R. C., Edwards, D. L., Glaberman, S., Chiari, Y., Poulakakis, N., Tapia, W., Gibbs, J. P., & Caccone, A. (2022). A new lineage of Galapagos giant tortoises identified from museum samples. Heredity. https://doi.org/10.1038/s41437-022-00510-8
Citations:
Jensen, E. L., Quinzin, M. C., Miller, J. M., Russello, M. A., Garrick, R. C., Edwards, D. L., Glaberman, S., Chiari, Y., Poulakakis, N., Tapia, W., Gibbs, J. P., & Caccone, A. (2022). A new lineage of Galapagos giant tortoises identified from museum samples. Heredity. https://doi.org/10.1038/s41437-022-00510-8
Burgon, J., & Jensen, E. (2022, March 16). Galápagos giants. Heredity Podcast.