DOI: 10.5061/dryad.hx3ffbgm3
title: Epigenetic aging of Māui and Hector's dolphins

description: The age of an individual is an essential demographic parameter but is
 difficult to estimate without long-term monitoring or invasive sampling.
 Epigenetic approaches are increasingly used to age organisms, including
 non-model organisms such as cetaceans. Māui dolphins (Cephalorhynchus
 hectori maui) are a critically endangered subspecies endemic to Aotearoa
 New Zealand, and the age structure of this population is important for
 informing conservation. Here we present an epigenetic clock for aging Māui
 and Hector's dolphins (C. h. hectori) developed from methylation data
 using DNA from tooth aged individuals (n = 48). Based on this
 training dataset, the optimal model required only eight methylation sites,
 provided an age correlation of 0.95, and had a median absolute age error
 of 1.54 years. A leave-one-out cross-validation analysis with the same
 parameters resulted in an age correlation of 0.87 and median absolute age
 error of 2.09 years. To improve age estimate, we included previously
 published beluga whale (Delphinapterus leucas) data to develop a joint
 beluga/dolphin clock, resulting in a clock with comparable performance and
 improved estimation of older individuals. Application of the models to DNA
 from skin biopsy samples of living Māui dolphins revealed a shift in the
 median age of 8–9 years to a younger population aged 7–8 years 10 years
 later. These models could be applied to other dolphin species and
 demonstrate the ability to construct a clock even when the number of known
 age samples is limited, removing this impediment to estimating demographic
 parameters vital to the conservation of critically endangered species.
description: # Epigenetic aging of Māui and Hector's dolphins
 The files linked here
 accompany the article by Hernandez et al., "Using epigenetic clocks
 to investigate changes in the age structure of critically endangered Māui
 dolphins." Included here are the R code for the epigenetic clock
 construction and accompanying analyses and the files needed to replicate
 this work. Supplementary material is available from the journal website.
 ## Description of the data and file structure The methylation data is
 structured to where each row is a cytosine-guanine dinucleotide (CpG) site
 in the methylation array, and each column is an individual sample, as
 indicated by the variable Basename and ExternalSampleID. Metadata are
 structured to where each row is an individual sample, and the columns are
 associated metadata variables. Missing data are indicated by NAs.
 Individuals with a "no" in the CanBeUsedForAging variable or
 DOC_Cat "red" were excluded from epigenetic clock development as
 these individuals were technical outliers or considered unsuitable for
 aging. 1\. Revised_Ceph_Samples.csv Contains metadata and identifiers for
 the Māui and Hector's dolphin samples analyzed in the study relevant
 for R analyses. ### Variables * ExternalSampleID: In-house sample
 identifiers from Oregon State University * DOC_Code: an individual
 identifier assigned by the New Zealand Department of Conservation *
 Basename: An identifier for the sample when it was run on the methylation
 array by UCLA * Age: The initial tooth age estimate (in years) provided by
 Massey University * CanBeUsedForAging: Based on hierarchical clustering
 after the methylation array data were generated by UCLA, is the sample
 suitable for aging? Either yes, no or maybe. Samples with "no"
 were not used in clock calibration. * Female: a binary representation of
 sex, where 1 = female and 0 = male * SpeciesCommonName: The common name of
 the subspecies from which the sample originated * SpeciesLatinName: The
 binomial name of the subspecies from which the sample originated *
 DOC_cat: A categorical variable for samples that were analyzed for teeth
 aging. 'Green' denotes a confident age estimate.
 'Yellow' for some level of uncertainty in age estimate and to
 use with caution in clock calibration. 'Orange' for samples
 where only a minimum age estimate could be reached. 'Red' for
 samples where the tooth was unreadable so no age estimate was possible and
 it was recommended not to use the sample for clock calibration. NA for
 samples that did not have a tooth read for aging. Additional details about
 tooth aging are provided in Betty et al. 2022 (see citation in Hernandez
 et al. 2023 for full reference information). * ToothBatch: If a tooth was
 read for aging, the batch number for analysis. Either 1, 2, 3 or NA *
 SampleSource: Source for the tissue sample used for epigenetic aging,
 either 'Biopsy' for tissue from a biopsy dart, or
 'Necropsy' for post-mortem collection. 2\. HM_Clock_Clean.R The
 R code used to analyze the methylation data. Variables defined within.
 Data files are available through this repository or from NCBI GEO. ##
 Sharing/Access information The manifest for the methylation array
 (HorvathMammalMethylChip40) is available at the Gene Expression Omnibus
 (GPL28271: Illumina HorvathMammalianMethylChip40 Bead Chip). Beluga
 methylation data and some of the Māui and Hector's dolphin samples
 are archived in the NCBI GEO database, GSE164465, “Genome Methylation in
 Wild Beluga Whales.” Remaining Māui and Hector's dolphin methylation
 data are archived in the NCBI GEO database, GSE242072, “Epigenetic aging
 of Māui and Hector’s Dolphins” (Accession numbers GSM7748508 –
 GSM7748648). Beluga whale metadata are available through the supporting
 information for Bors et al. (2021) at
 ## Code/Software All analyses
 were run in R version 4.2.1 as implemented in RStudio. The necessary
 packages include the tidyverse suite, cowplot (for generating multi-panel
 figures) and glmnet (for clock models).