Content uploaded by Dafni Anastasiadi
Author content
All content in this area was uploaded by Dafni Anastasiadi on May 25, 2021
Content may be subject to copyright.
A preview of the PDF is not available
A clockwork fish. Age‐prediction using DNA methylation‐based biomarkers in the European seabass
Abstract
Age‐related changes in DNA methylation do occur. Taking advantage of this, mammalian and avian epigenetic clocks have been constructed to predict age. In fish, studies on age‐related DNA methylation changes are scarce and no epigenetic clocks have been constructed. However, in fisheries and population dynamics studies there is a need for accurate estimation of age, something that is often impossible for some economically important species with the currently available methods. Here, we used the European sea bass, a marine fish where age can be known with accuracy, to construct a piscine epigenetic clock, the first one in a cold‐blooded vertebrate. We used targeted bisulfite sequencing to amplify 48 CpGs from four genes in muscle samples and applied penalized regressions to predict age. We, thus, developed an age predictor in fish that is highly accurate (0.824) and precise (2.149 years). In juvenile fish, accelerated growth due to elevated temperatures had no effect in age prediction, indicating that the clock is able to predict the chronological age independently of environmentally‐driven perturbations. An epigenetic clock developed using muscle samples accurately predicted age in samples of testis but not ovaries, possibly reflecting the reproductive biology of fish. In conclusion, we report the development of the first piscine epigenetic clock, paving the way for similar studies in other species. Piscine epigenetic clocks should be of great utility for fisheries management and conservation purposes, where age determination is of crucial importance.
Supplementary resource (1)
... Epigenetic clocks summarize age-associated increases (hypermethylation) or decreases (hypomethylation) in DNA methylation across a selected group of CpG sites throughout the genome, which can be used collectively to estimate chronological age . While the utility of epigenetic age estimation has been demonstrated in several fish species (European sea bass Dicentrarchus labrax, Anastasiadi and Piferrer 2020; zebrafish Danio rerio, Mayne et al. 2020; Australian lungfish Neoceratodus forsteri, Murray cod Maccullochella peelii, and Mary River cod Maccullochella mariensis, Mayne et al. 2021a; Japanese medaka Oryzias latipes, Bertucci et al. 2021; northern red snapper Lutjanus campechanus and red grouper Epinephelus morio, Weber et al. 2022; and golden perch Macquaria ambigua spp., Mayne et al. 2023), the application of this aging technique to a deepwater fish species has not yet been investigated. The development of epigenetic clocks in deepwater fishes would seem to be of great utility given the high error often associated with traditional aging techniques for deepwater fishes (Cailliet et al. 2001;Campana 2005) and their vulnerability to overfishing given their longevity and slow growth (Devine et al. 2006). ...
... The epigenetic clocks developed with length data also required fewer CpG sites to predict age, which is important to consider when generating assays for production aging (i.e., the efficient generation of age estimates for large numbers of individuals needed for fisheries assessment; Passerotti et al. 2020), such as via genotyping-in-thousands by sequencing (GT-seq; Campbell et al. 2015), because PCR assays are easier to design with fewer loci. While Anastasiadi and Piferrer (2020) reported that the inclusion of length data did not have an effect on epigenetic age estimation in the European sea bass (Dicentrarchus labrax), this difference may be due to the fact that length data were treated as a covariate in a regression of predicted age versus chronological age in that study, while length data were treated as predictor variables during epigenetic clock development in the present study. The accuracy and precision of the epigenetic clocks further improved when males and females were modeled separately, suggesting patterns of ageassociated DNA methylation are somewhat sex-specific. ...
Age estimates are essential for fisheries assessment and management, but deepwater (>200 m) fishes are often difficult to age using traditional techniques. Therefore, age-predictive epigenetic clocks were developed for a model deepwater reef fish, blackbelly rosefish Helicolenus dactylopterus, using two tissue types (fin clips and muscle; n = 61 individuals; 9−60 years) and Δ¹⁴C-validated consensus age estimates. The influence of biological information (length and sex) on epigenetic clock accuracy, and the potential for developing a multi-tissue clock, were also assessed. Bisulfite-converted restriction site-associated DNA sequencing (bsRADseq) was used to identify CpG sites (cytosines followed by guanines) exhibiting age-correlated DNA methylation, and epigenetic clocks showed strong agreement (R² > 0.98) between predicted and consensus ages. Including length and sex data enhanced accuracy and precision (R² > 0.99; mean absolute error < 1 year). Age-associated CpG sites were identified across tissues, but a multi-tissue clock performed poorly relative to single-tissue clocks. Overall, results demonstrate that accurate and precise epigenetic clocks can be developed for deepwater fishes, and the inclusion of biological information may enhance clock accuracy and precision.
... Anastasiadi et al. amplified 48 CpG sites from four genes in muscle samples of Dicentrarchus labrax, for which the age was accurately determined by targeting the sodium bisulfite sequences. They then applied penalized regression to predict age and constructed an epigenetic clock in fish [6]. Methylation is a significant epigenetic modification of eukaryotic genomic DNA, and it plays crucial roles in biological processes such as gene expression, embryonic development, cell differentiation, and gene imprinting [7,8]. ...
... (3) We then implemented the linking reaction system as shown in Table 2: the reaction conditions were a 4 • C connection for 6-8 h; (4) We then implemented the PCR reaction system and conditions as shown in Table 3; (6) After electrophoresis, SYBR Safe DNA dye was used for 3 min to observe the brightness of the target band (100 bp); (7) We then cut the desired strip and placed it into a 1.5 mL centrifuge tube, ground the glue with a grinding rod, added 30-40 µL of pure water, and let it stand at 4 • C 6-12 h; (8) We then introduced the barcode sequence and the PCR reaction system as shown in Table 4; (9) We then purified PCR products with the QIAquick PCR Purification Kit, eluted them with 15 µL of pure water, and then used QubitQuantity for determination. In general, the ideal concentration of purified products is 10-30 ng/µL; (10) If multiple libraries are built, libraries with different barcode numbers can be mixed according to the amount of sent measurement data. ...
Simple Summary
In this study, MethylRAD-Seq of the body wall tissues of Apostichopus japonicus at different ages was analyzed based on methylated RAD-Seq technology and, combined with GO and KEGG analyses, different genes related to the age of A. japonicus were screened, such as H2AX, Hsp90, Pepn, and CDC6. This provides reference significance for the identification of A. japonicus age.
Abstract
The A. japonicus industry has expanded significantly, but no research has focused on determining the age of A. japonicus during farming. Correctly estimating the age of A. japonicus can provide a decision-making basis for the breeding process and data for the protection of A. japonicus aquatic germplasm resources. DNA methylation levels in the body wall of Apostichopus japonicus at 4 months, 1 year, 2 years, and 3 years old were determined using MethylRAD-Seq, and differentially methylated genes were screened. A total of 441 and 966 differentially methylated genes were detected at the CCGG and CCWGG sites, respectively. Aspartate aminotransferase, succinate semialdehyde dehydrogenase, isocitrate dehydrogenase, the histone H2AX, heat shock protein Hsp90, aminopeptidase N, cell division cycle CDC6, Ras GTPase activating protein (RasGAP), slit guidance ligand slit1, integrin-linked kinase ILK, mechanistic target of rapamycin kinase Mtor, protein kinase A Pka, and autophagy-related 3 atg3 genes may play key roles in the growth and aging process of A. japonicus. This study provides valuable information regarding age-related genes for future research, and these candidate genes can be used to create an “epigenetic clock”.
... In any case, it is well accepted that aging is related, in mammals [67] and fish (killifish) used a model of vertebrate aging, with an overall hypomethylation that becomes first associated to a down-regulation of DNA methyltransferases. In the present study, this fits well with a global hypomethylation (58.5 %) among the discriminant DM regions in our age model of one-and three-year old fish, that at the same time would co-exist with a site-specific hypermethylation, as reported upon aging in both humans [69] and other animal models [67,70], including fish [28,71,72]. ...
DNA-methylation clocks inform not only about chronological but also biological age, which brings a high resolution and precise understanding of age-related pathology and physiology. Attempts based on transcriptomic and epigenetic approaches arise as integrative biomarkers linking the quantification of stress response with a given fitness trait and may help to identify biological age markers, also considered welfare indicators. In gilthead sea bream, targeted gene expression and DNA-methylation analyses in white skeletal muscle proved sirt1 as a reliable marker of age-mediated changes of energy metabolism. To complete the list of welfare auditing biomarkers, wide-analyses of gene expression and DNA-methylation in one- and three-year old fish were combined. After discriminant analysis, 668 differentially expressed transcripts were matched with those containing differentially methylated (DM) regions (14,366), and 172 were overlapping. Through enrichment analyses and selection, two sets of genes were retained: 33 showing an opposite trend for DNA-methylation and expression, and 57 down-regulated and hypo-methylated. The first set displayed apparently a more reproducible and reliable pattern and 10 multifunctional genes with DM CpG in regulatory regions (sirt1, smad1, ramp1, psmd2 – up-regulated; col5a1, calcrl, bmp1, thrb, spred2, atp1a2 – down-regulated) were deemed candidate biological age markers for an improved welfare auditing in gilthead sea bream.
... The aging process can be accelerated by clockwork. 39 The mTOR is a serine/threonine kinase that is found in two functionally distinct signaling complexes in mammals. It functions as a vital core nutrient indicator. ...
Circadian rhythms are oscillations in physiology and behavior caused by the circadian regulator. Cryptochromes, Periods, and Bmal1 are circadian clock genes that have been linked to aging and cancer. Human pathologies alter circadian clock gene expression, and transgenic rats with clock gene defects progress to cancer and age prematurely. In the growth of age‐linked pathologies and carcinogenesis, cell proliferation and genome integrity play critical roles. The relationship concerning the cell cycle regulation and circadian clock is discussed in this article. The circadian clock controls the behavior and countenance of many main cell cycle and cell cycle check‐point proteins, and cell cycle‐associated proteins, in turn, control the activity and expression of circadian clock proteins. The circadian clock can be reset by DNA disruption, providing a molecular mechanism for mutual control amid the cell cycle and the clock. This circadian clock‐dependent regulation of cell proliferation, composed with other circadian clock‐dependent physiological functions including metabolism control, genotoxic and oxidative stress response, and DNA repair, unlocks new avenues for studying the processes of aging and carcinogenesis.
... The potential of eRNA in biomonitoring has been discussed in various studies, highlighting its ability to significantly improve the resolution of organism detection and offer valuable insights into ecological applications (Jo, 2023;Yates et al., 2021). Moreover, epigenetic modifications have been used to develop epigenetic clocks for non-model fish, which could contribute to high precision age estimation (Anastasiadi and Piferrer, 2020;Mayne et al., 2021). Although some variables may bias and mask the correlation between specific epigenetic signals and physiological traits (Schadewell and Adams, 2021;Sigsgaard et al., 2020), epigenetics still has the potential to be a powerful tool in this field. ...
Environmental DNA (eDNA) monitoring, a rapidly advancing technique for assessing biodiversity and ecosystem health, offers a noninvasive approach for detecting and quantifying species from various environmental samples. In this review, a comprehensive overview of current eDNA collection and detection technologies is provided, emphasizing the necessity for standardization and automation in aquatic ecological monitoring. Furthermore, the intricacies of water bodies, from streams to the deep sea, and the associated challenges they pose for eDNA capture and analysis are explored. The paper delineates three primary eDNA survey methods, namely, bringing back water, bringing back filters, and bringing back data, each with specific advantages and constraints in terms of labor, transport, and data acquisition. Additionally, innovations in eDNA sampling equipment, including autonomous drones, subsurface samplers, and in-situ filtration devices, and their applications in monitoring diverse taxa are discussed. Moreover, recent advancements in species-specific detection and eDNA metabarcoding are addressed, highlighting the integration of novel techniques such as CRISPR-Cas and nanopore sequencing that enable precise and rapid detection of biodiversity. The implications of environmental RNA and epigenetic modifications are considered for future applications in providing nuanced ecological data. Lastly, the review stresses the critical role of standardization and automation in enhancing data consistency and comparability for robust long-term biomonitoring. We propose that the amalgamation of these technologies represents a paradigm shift in ecological monitoring, aligning with the urgent call for biodiversity conservation and sustainable management of aquatic ecosystems.
... The epigenetic mark cytosine DNA methylation (DNAme) is widespread in eukaryotes [11] and has been robustly associated with the ageing process [12,13]. Age-related changes in DNAme have been observed in multiple vertebrate groups, including fishes [14], amphibians [15], birds [16] and mammals [17], suggesting that the association between ageing and this epigenetic mark is evolutionarily conserved. By contrast, to our knowledge, only one study explored the role of DNAme in ageing in invertebrates and reported tentative yet mixed evidence indicating that DNAme might play a role in lifespan regulation in the honeybee Apis mellifera [18]. ...
Epigenetic alterations are a primary hallmark of aging. In mammals, age-related epigenetic changes alter gene expression profiles, disrupt cellular homeostasis and physiological functions and, therefore, promote aging. It remains unclear whether aging is also driven by epigenetic mechanisms in invertebrates. Here, we used a pharmacological hypomethylating agent (RG108) to evaluate the effects of DNA methylation on lifespan in an insect - the bumble bee Bombus terrestris. RG108 extended mean lifespan by 43% and induced the differential methylation of genes involved in hallmarks of aging, including DNA damage repair and chromatin organisation. Furthermore, the longevity gene sirt1 was overexpressed following the treatment. Functional experiments demonstrated that SIRT1 protein activity was positively associated with lifespan. Overall, our study indicates that epigenetic mechanisms are conserved regulators of lifespan in both vertebrates and invertebrates and provides new insights into how DNA methylation is involved in the aging process in insects.
... Such analogy further supports the relevance of DNA methylation in the regulation of gene expression in fish and in our experimental model in particular [9]. Otherwise, like in humans and other animal models [66,67], a global DNA hypo-methylation and site-specific hyper-methylation could be associated with aging in marine fish [68,69]. We may then speculate that changes in the epigenetic clock of REF animals might occur, reflecting perhaps an aging phenotype with an overall impairment of performance that affects among other traits, growth, swimming behaviour and the incidence of skeletal deformities [49][50][51][52]. ...
Background
Broodstock nutritional programming improves the offspring utilization of plant-based diets in gilthead sea bream through changes in hepatic metabolism. Attention was initially focused on fatty acid desaturases, but it can involve a wide range of processes that remain largely unexplored. How all this can be driven by a different genetic background is hardly underlined, and the present study aimed to assess how broodstock nutrition affects differentially the transcriptome and genome-wide DNA methylome of reference and genetically selected fish within the PROGENSA® selection program.
Results
After the stimulus phase with a low fish oil diet, two offspring subsets of each genetic background received a control or a FUTURE-based diet. This highlighted a different hepatic transcriptome (RNA-seq) and genome-wide DNA methylation (MBD-seq) pattern depending on the genetic background. The number of differentially expressed transcripts following the challenge phase varied from 323 in reference fish to 2,009 in genetically selected fish. The number of discriminant transcripts, and associated enriched functions, were also markedly higher in selected fish. Moreover, correlation analysis depicted a hyper-methylated and down-regulated gene expression state in selected fish with the FUTURE diet, whereas the opposite pattern appeared in reference fish. After filtering for highly represented functions in selected fish, 115 epigenetic markers were retrieved in this group. Among them, lipid metabolism genes (23) were the most reactive following ordering by fold-change in expression, rendering a final list of 10 top markers with a key role on hepatic lipogenesis and fatty acid metabolism (cd36, pitpna, cidea, fasn, g6pd, lipt1, scd1a, acsbg2, acsl14, acsbg2).
Conclusions
Gene expression profiles and methylation signatures were dependent on genetic background in our experimental model. Such assumption affected the magnitude, but also the type and direction of change. Thus, the resulting epigenetic clock of reference fish might depict an older phenotype with a lower methylation for the epigenetically responsive genes with a negative methylation-expression pattern. Therefore, epigenetic markers will be specific of each genetic lineage, serving the broodstock programming in our selected fish to prevent and mitigate later in life the risk of hepatic steatosis through changes in hepatic lipogenesis and fatty acid metabolism.
... By quantifying the methylation patterns of specific genomic regions, researchers can construct an "epigenetic clock" that correlates with chronological age [13,14]. This innovative approach offers a direct and accurate means of age determination, circumventing the limitations of traditional methods [15][16][17]. Furthermore, the epigenetic clock not only enhances our understanding of fish aging processes but also holds the potential to shed light on the effects of environmental factors on age-related changes, namely, temperature and food availability [13]. Integrating genomic information with traditional methods and modern imaging techniques presents an exciting multidisciplinary approach with which to advance age estimation accuracy. ...
Fish are the largest and most diverse group of vertebrates [...]
After three decades of working as a research scientist, I am stepping back to consider the events, questions, and principles that have guided my scientific journey. Important questions and research objectives have been how to implement the ecosystem approach to fisheries management in practice, the development of new data uses, the application of new observation methods and models, and estimating and accounting for uncertainty. Stakeholder engagement—why and how—is a topic that has increased in importance over time. While our observation methods did not change much over many decades, they are now changing rapidly due to new technological developments, but also societal and environmental changes.
Inferring the chronological and biological age of individuals is fundamental to population ecology and our understanding of ageing itself, its evolution, and the biological processes that affect or even cause ageing. Epigenetic clocks based on DNA methylation (DNAm) at specific CpG sites show a strong correlation with chronological age in humans, and discrepancies between inferred and actual chronological age predict morbidity and mortality. Recently, a growing number of epigenetic clocks have been developed in non-model animals and we here review these studies. We also conduct a meta-analysis to assess the effects of different aspects of experimental protocol on the performance of epigenetic clocks for non-model animals. Two measures of performance are usually reported, the R2 of the association between the predicted and chronological age, and the mean/median absolute deviation (MAD) of estimated age from chronological age, and we argue that only the MAD reflects accuracy. R2 for epigenetic clocks based on the HorvathMammalMethylChip4 was higher and the MAD scaled to age range lower, compared with other DNAm quantification approaches. Scaled MAD tended to be lower among individuals in captive populations, and decreased with an increasing number of CpG sites. We conclude that epigenetic clocks can predict chronological age with relatively high accuracy, suggesting great potential in ecological epigenetics. We discuss general aspects of epigenetic clocks in the hope of stimulating further DNAm-based research on ageing, and perhaps more importantly, other key traits.
Genetic selection is often implicated as the underlying cause of heritable phenotypic differences between hatchery and wild populations of steelhead trout (Oncorhynchus mykiss) that also differ in lifetime fitness. Developmental plasticity, which can also affect fitness, may be mediated by epigenetic mechanisms such as DNA methylation. Our previous study identified significant differences in DNA methylation between adult hatchery- and natural-origin steelhead from the same population that could not be distinguished by DNA sequence variation. In the current study, we tested whether hatchery-rearing conditions can influence patterns of DNA methylation in steelhead with known genetic backgrounds, and assessed the stability of these changes over time. Eyed-embryos from 22 families of Methow River steelhead were split across traditional hatchery tanks or a simulated stream-rearing environment for 8 months, followed by a second year in a common hatchery tank environment. Family assignments were made using a genetic parentage analysis to account for relatedness among individuals. DNA methylation patterns were examined in the liver, a relatively homogeneous organ that regulates metabolic processes and somatic growth, of juveniles at two time points: after eight months of rearing in either a tank or stream environment and after a subsequent year of rearing in a common tank environment. Further, we analyzed DNA methylation in the sperm of mature 2-year-old males from the earlier described treatments to assess the potential of environmentally-induced changes to be passed to offspring. Hepatic DNA methylation changes in response to hatchery versus stream-rearing in yearling fish were substantial, but few persisted after a second year in the tank environment. However, the early rearing environment appeared to affect how fish responded to developmental and environmental signals during the second year since novel DNA methylation differences were identified in the livers of hatchery versus stream-reared fish after a year of common tank rearing. Furthermore, we found profound differences in DNA methylation due to age, irrespective of rearing treatment. This could be due to smoltification associated changes in liver physiology after the second year of rearing. Although few rearing-treatment effects were observed in the sperm methylome, strong family effects were observed. These data suggest limited potential for intergenerational changes, but highlight the importance of understanding the effects of kinship among studied individuals in order to properly analyze and interpret DNA methylation data in natural populations. Our work is the first to study family effects and temporal dynamics of DNA methylation patterns in response to hatchery-rearing.
Age structure is a fundamental aspect of animal population biology. Age is strongly related to individual physiological condition, reproductive potential and mortality rate. Currently, there are no robust molecular methods for age estimation in birds. Instead, individuals must be ringed as chicks to establish known‐age populations, which is a labour intensive and expensive process. The estimation of chronological age using DNA methylation is emerging as a robust approach in mammals including humans, mice and some non‐model species. Here we quantified DNA methylation in whole blood samples from a total of 71 known‐age Short‐tailed shearwaters (Ardenna tenuirostris) using digital restriction enzyme analysis of methylation (DREAM). The DREAM method measures DNA methylation levels at thousands of CpG dinucleotides throughout the genome. We identified seven CpG sites with DNA methylation levels that correlated with age. A model based on these relationships estimated age with a mean difference of 2.8 years to known age, based on validation estimates from models created by repeated sampling of training and validation data subsets. Longitudinal observation of individuals re‐sampled over 1 or 2 years generally showed an increase in estimated age (6/7 cases). For the first time, we have shown that epigenetic changes with age can be detected in a wild bird. This approach should be of broad interest to researchers studying age biomarkers in non‐model species and will allow identification of markers that can be assessed using targeted techniques for accurate age estimation in large population studies.
This article is protected by copyright. All rights reserved.
ELife digest
Epigenetic marks are chemical modifications found throughout the genome – the DNA within cells. By influencing the activity of nearby genes, the marks govern developmental processes and help cells to adapt to changes in their surroundings. Some epigenetic marks can be gained or lost with age. A lot of aging research focuses on one type of mark, called “DNA methylation”. By measuring the presence or absence of specific methyl groups, scientists can estimate biological age – which may differ from calendar age.
Recent studies have developed computer models called epigenetic aging clocks to predict the biological age of mouse cells. These clocks use epigenetic data collected from the entire genomes of mice, and are useful for understanding how the aging process is affected by genetic parameters, diet, or other environmental factors. Yet, the genome sequencing methods used to construct most existing epigenetic clocks are expensive, labor-intensive, and cannot be easily applied to large groups of mice.
Han et al. have developed a new way to predict biological aging in mice that needs methylation information from just three particular sections of the genome. Even though this approach is much faster and less expensive than other epigenetic approaches to measuring aging, it has a similar level of accuracy to existing models. Han et al. use the new method to show that cells from different strains of laboratory mice age at different rates. Furthermore, in a strain that has a shorter life expectancy, aging seems to be accelerated.
The new approach developed by Han et al. will make it easier to study how aging in mice is affected by different interventions. Further studies will also be needed to better understand how epigenetic marks relate to biological aging.
The age profile of populations fundamentally affects their conservation status. Yet age is frequently difficult to assess in wild animals. Here, we assessed the use of DNA methylation of homologous genes to establish the age structure of a rare and elusive wild mammal: the Bechstein's bat (Myotis bechsteinii). We collected 62 wing punches from individuals whose ages were known as a result of a long‐term banding study. DNA methylation was measured at seven CpG sites from three genes which have previously shown age‐associated changes in humans and laboratory mice. All CpG sites from the tested genes showed a significant relationship between DNA methylation and age, both individually and in combination (multiple linear regression R²=0.58, p<0.001). Despite slight approximation around estimates, the approach is sufficiently precise to place animals into practically useful age cohorts. This method is of considerable practical benefit as it can reliably age individual bats. It is also much faster than traditional capture‐mark‐recapture techniques, with the potential to collect information on the age structure of an entire colony from a single sampling session to better inform conservation actions for Bechstein's bats. By identifying three genes where DNA methylation correlates with age across distantly related species, this study also suggests that the technique can potentially be applied across a wide range of mammals.
This article is protected by copyright. All rights reserved.
In wild animal conservation, knowing the age of an individual animal is extremely beneficial. However, estimating the age is difficult for many species. Recently, epigenetics-based methods of estimating age have been reported. These studies were predominantly on humans with few reports on other animals, especially wild animals. In the present study, a chimpanzee (Pan troglodytes) age prediction model was developed based on the ELOVL2, CCDC102B, and ZNF423 genes that may also have application in human age prediction. Pyrosequencing was used to measure methylation in 20 chimpanzee blood samples and correlation between age and methylation status was calculated. Age and methylation of sites in ELOVL2 and CCDC102B were significantly correlated and an age prediction model was created using these genes. In the regression equation using only ELOVL2, the highest correlation coefficient was 0.741, with a mean absolute deviation (MAD) of 5.41, compared with the combination of ELOVL2 and CCDC102B, where the highest correlation coefficient was 0.742 and the MAD was 5.41. Although larger MADs were observed in chimpanzees than in humans based on these genes, the results indicate the feasibility of estimating chimpanzee age using DNA methylation, and can have implications in understanding the ecology of chimpanzees and chimpanzee conservation.
Several articles describe highly accurate age estimation methods based on human DNA-methylation data. It is not yet known whether similar epigenetic aging clocks can be developed based on blood methylation data from canids. Using Reduced Representation Bisulfite Sequencing, we assessed blood DNA-methylation data from 46 domesticated dogs (Canis familiaris) and 62 wild gray wolves (C. lupus). By regressing chronological dog age on the resulting CpGs, we defined highly accurate multivariate age estimators for dogs (based on 41 CpGs), wolves (67 CpGs), and both combined (115 CpGs). Age related DNA methylation changes in canids implicate similar gene ontology categories as those observed in humans suggesting an evolutionarily conserved mechanism underlying age-related DNA methylation in mammals.
Precocious sexual maturation affects approximately one‐third of farmed European sea bass males which, in turn, constitute most of the farmed fish of this species. To prevent sexual maturation, and to contribute to the production of monosex stocks, chromosome set manipulation procedures have been subject to intense investigation over the last 20 years. These include the induction of poliploidy (triploidy and tetraploidy) and the production of individuals with uniparental inheritance (mito‐ and meiogynogens and androgenetics). Several studies have examined different experimental conditions in order to establish optimized protocols based on the application of pressure and temperature shocks, to retain a full set of chromosomes, thus suppressing the extrusion of the second polar body, or the first cleavage in the zygote. The use of UV irradiation has also been evaluated to inactivate the DNA of exposed gametes for the application of induced gynogenesis and androgenesis.
Triploidy in the European sea bass results in gonadal sterility in both sexes, which can be of advantage for its production in aquaculture although, only in larger fish, it may represent a superior growth. On the other hand, the maintenance of gynogenetic and androgenetic clonal founders can contribute to a better understanding of the genetic basis of many complex traits of interest for fish farming. Although the performance of fish after ploidy manipulation, concerning the growth, reproductive activity, and proportion of sexes, has been well documented, further evaluations are still required before these fish can achieve societal acceptance and be considered for their applicability to the industry. Finally, benefits, considerations and future work are under discussion.
The integration of genomic and environmental influences into methylation patterns to bring about a phenotype is of central interest in developmental epigenetics, but many details are still unclear. The sex ratios of the species used here, the European sea bass, are determined by genetic and temperature influences. We created four families from parents known to produce offspring with different sex ratios, exposed larvae to masculinizing temperatures and examined, in juvenile gonads, the DNA methylation of seven genes related to sexual development by a targeted sequencing approach. The genes most affected by both genetics and environment were cyp19a1a and dmrt1, with contrasting sex-specific methylation and temperature responses. The relationship between cyp19a1a methylation and expression is relevant to the epigenetic regulation of sex, and we report the evidence of such relationship only below a methylation threshold, ~80%, and that it was sex-specific: negatively correlated in females but positively correlated in males. From parents to offspring, the methylation in gonads was midway between oocytes and sperm, with bias towards oocytes for amh-r2, er-β2, fsh-r and cyp19a1a. In contrast, dmrt1 levels resembled those of sperm. The methylation of individual CpGs from foxl2, er-β2 and nr3c1 were conserved from parents to offspring, whereas those of cyp19a1a, dmrt1 and amh-r2 were affected by temperature. Utilizing a machine-learning procedure based on the methylation levels of a selected set of CpGs, we present the first, to our knowledge, system based on epigenetic marks capable of predicting sex in an animal with ~90% accuracy and discuss possible applications.