EPIGENETIC CLOCKS: USING DNA TO ESTIMATE THE AGE OF DOLPHINS

You probably know your age and you might be good at estimating the ages of other people. However, have you ever tried to guess the age of an animal, for example a dolphin? Estimating the age of dolphins is really difficult, because their appearance does not change once they are fully grown. Fortunately, the shape of their DNA changes over time. This allowed us to develop a tool, called an epigenetic clock, to determine the age of dolphins based on their DNA. Using this clock, we can estimate the age of a dolphin with an accuracy of 2 years. Although it is not exact, this still helps us answer questions about the lives of dolphins, such as when they start their own families. Knowing dolphin ages can also tell us if a group of dolphins contains a healthy mix of older and younger individuals. Epigenetic clocks are now being developed for many other species, offering exciting new research opportunities.

You probably know your age and you might be good at estimating the ages of other people.However, have you ever tried to guess the age of an animal, for example a dolphin?Estimating the age of dolphins is really di cult, because their appearance does not change once they are fully grown.Fortunately, the shape of their DNA changes over time.This allowed us to develop a tool, called an epigenetic clock, to determine the age of dolphins based on their DNA.Using this clock, we can estimate the age of a dolphin with an accuracy of years.Although it is not exact, this still helps us answer questions about the lives of dolphins, such as when they start their own families.Knowing dolphin ages can also tell us if a group of dolphins contains a healthy mix of older and younger individuals.Epigenetic clocks are now being developed for many other species, o ering exciting new research opportunities.

HOW TO ESTIMATE THE AGE OF DOLPHINS
When studying animals, one of the most important pieces of information to know about individual animals is their age.Scientists must know how old an animal is to understand many parts of its life history and, for example, to estimate if a population of animals can survive.But estimating an animal's age is not easy when its date of birth is unknown.Across all animal species, there are no general signs for the aging process, such as the wrinkles or gray hair seen in humans.trunk of a tree.This way, teeth can help scientists find a dolphin's age quite accurately.However, this method is not pleasant for the dolphins and therefore di cult to do with live animals.For some species, such as Indo-Pacific bottlenose dolphins (Tursiops aduncus), a pattern of spots on their skin can tell us about their age, because they gain more spots as they grow older (Figure ).However, individuals tend to have di erent amounts of spots in general, and it is often quite di cult to see enough of a dolphin's body to clearly see and count the spots.So, trying to age an animal based on its spotting pattern will only give us a rough estimate.
Because of these di culties, most studies rely on data collected over a long time, in studies where animals have been regularly observed for many years since their birth, and thus their ages are known.For species that can live for many years, it can be di cult to gather enough data in a human's lifetime.

SHARK BAY DOLPHINS
One of the places where dolphins have been studied for many years is Shark Bay, Western Australia.There, scientists have observed a population of Indo-Pacific bottlenose dolphins for more than years [ ], using techniques called photo-identification and

PHOTO-IDENTIFICATION
A method of identifying individual dolphins using photos of their dorsal fins.The scars and marks on their fins make them unique, like a fingerprint in humans.
biopsy sampling.For photo-identification, researchers take photos

BIOPSY SAMPLING
A data collection technique where we take a small skin sample from a live dolphin using a little dart with a sharp hollow end.
of the dolphins' dorsal fins.The shape of the fin is like a human

DORSAL FIN
The fin on the back of a whale or dolphin.
fingerprint, and by comparing the fins of di erent animals, researchers can recognize individuals.Biopsy sampling means collecting a small piece of dolphin skin that can be analyzed in the lab to get the animals' DNA.Because we have been studying this dolphin population for a long time and we recognize the individual dolphins, we also know when one is born and, therefore, can calculate its age.Thanks to biopsy sampling we have small skin samples from all the dolphins we study.

WHAT IS EPIGENETICS?
Our dolphin DNA samples have opened doors to new methods of age estimation for our dolphin species, using epigenetics.Epigenetics

EPIGENETICS
The study of how our genes are a ected by our behaviors and the environment we are exposed to.
is the study of how the environment and an individual's behaviors can a ect the way their genes work.Epigenetic changes modify the shape of the DNA, which alters which genes are switched on or turned o .This shape change can happen by adding or removing small molecules to the DNA-a process called DNA methylation.As animals age, di erent sets of genes are switched on or "expressed", meaning that the DNA takes on di erent shapes throughout an animal's life (Figure ).levels of DNA methylation in dolphins of a known age, using a new tool called the HorvathMammalMethylChip [ ], which can measure DNA methylation on up to , sites on the DNA at once.We then built an epigenetic clock using machine learning, which we used to

MACHINE LEARNING
A type of artificial intelligence that learns from data.
estimate the age of other dolphins of the same species based on their methylation levels (Figure ).Our clock had an average accuracy of .years, meaning if we estimated an animal to be years old, it might actually be anywhere between -years old [ ].This is quite close and therefore a really good result!We also tested how well a di erent clock, developed using skin samples from a related species (common bottlenose dolphins [ ]), worked for our samples.Common bottlenose dolphins look almost the same as Indo-Pacific dolphins but are often a bit bigger.We found that the common bottlenose dolphin clock worked almost as well for our Indo-pacific bottlenose dolphins as for the species it was made for.

WHY IS THE EPIGENETIC CLOCK USEFUL?
Dolphins can live for more than years, and in the past it has been very di cult to gather information from many animals with known ages.By using the epigenetic clock, we can now estimate the age of individual dolphins just based on their DNA, which means we only need a small skin sample from each dolphin, and we are good to go!This makes our clock extremely useful for scientists that study Indo-Pacific bottlenose dolphins but who do not have access to long-term data.The fact that the common bottlenose dolphin clock also worked really well for our species means that even researchers studying other closely related species can use our clock.Because it is expensive to build an epigenetic clock, sharing a clock can save

WHAT ARE THE NEXT STEPS?
We now know the age for many dolphins in the Shark Bay population.
With this knowledge, we can ask further questions, such as at what time in their lives they produce the most calves or what influences their aging.We can also study whether males and females age di erently.
As so often in science, an answer leads to many more questions!This is an exciting time for us, and we cannot wait to get deeper into this work!

Figure
Figure Indo-pacific bottlenose dolphin with speckling pattern.Photo by Simon Allen.

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Figure dollars, which they can then use for other interesting research.
the highest levels of DNA methylation, while old individuals have the lowest.This change in methylation is also referred to as the epigenetic clock.Using DNA from skin samples, we measured the have