First Atlantic satellite tracks of 'lost years' green turtles support the importance of the Sargasso Sea as a sea turtle nursery.

In-water behaviour and long-term movements of oceanic-stage juvenile sea turtles are not well described or quantified. This is owing to technological or logistical limitations of tracking small, fast-growing animals across long distances and time periods within marine habitats. Here, we present, to our knowledge, the first long-term offshore tracks of oceanic green turtles (Chelonia mydas) in western North Atlantic waters. Using a tag attachment technique developed specifically for young (less than 1 year old) green turtles, we satellite-tracked 21 oceanic-stage green turtles (less than 19 cm straight carapace length) up to 152 days using small, solar-powered transmitters. We verify that oceanic-stage green turtles: (i) travel to and remain within oceanic waters; (ii) often depart the Gulf Stream and North Atlantic Subtropical Gyre currents, orienting towards waters associated with the Sargasso Sea; (iii) remain at the sea surface, using thermally beneficial habitats that promote growth and survival of young turtles; and (iv) green turtles orient differently compared to same stage loggerhead turtles (Caretta caretta). Combined with satellite tracks of oceanic-stage loggerhead turtles, our work identifies the Sargasso Sea as an important nursery habitat for North Atlantic sea turtles, supporting a growing body of research that suggests oceanic-stage sea turtles are behaviourally more complex than previously assumed.

Characterizing stable isotope relationships between green turtle (Chelonia mydas) skin and unhatched eggs.

RATIONALE: Stable isotope analysis is used to understand the foraging habits and movements of a diverse set of organisms. Variability in stable isotope ratios among tissues derived from the same animal makes it difficult to compare data among study results in which different tissue types are evaluated. Isotopic relationships between two green turtle (Chelonia mydas) tissue types, skin and unhatched egg contents are unknown. Similarly, few data exist to evaluate the influence of time elapsed after oviposition (as a proxy for decomposition) on isotopic variability among unhatched eggs within the same nest. METHODS: Skin and unhatched egg contents were collected from 69 adult female green turtles and associated nests at the Archie Carr National Wildlife Refuge in Florida, USA. Values of δ13 C, δ15 N, and δ34 S were measured for both tissue types using a continuous flow isotope ratio mass spectrometer. Standardized major-axis (SMA) regression was used to generate conversion equations of carbon, nitrogen, and sulfur isotope ratios between the two tissue types. Model selection frameworks consisting of single-factor linear models were employed per isotope ratio to assess how egg time-in-nest affected intraclutch isotopic variability. RESULTS: Conversion equations for all three isotope ratios indicated significant relationships between skin and unhatched egg values, although model fits were lower than found in some studies examining similar patterns in other marine turtle species. The probability of increased intraclutch variability was significantly higher among eggs collected at longer intervals after deposition. CONCLUSIONS: This study reports the first-ever δ13 C and δ15 N conversion equations between skin and unhatched eggs for green turtles, and the first δ34 S conversion equation for any marine turtle species. SMA regression was used to directly convert tissue values bidirectionally, unlike equations generated using ordinary least-squares regression. Issues with increased intraclutch variability at later excavation dates highlight the importance of collecting unhatched eggs as soon as possible after hatchling emergence.

First satellite tracks of South Atlantic sea turtle 'lost years': seasonal variation in trans-equatorial movement.

In the South Atlantic Ocean, few data exist regarding the dispersal of young oceanic sea turtles. We characterized the movements of laboratory-reared yearling loggerhead turtles from Brazilian rookeries using novel telemetry techniques, testing for differences in dispersal during different periods of the sea turtle hatching season that correspond to seasonal changes in ocean currents. Oceanographic drifters deployed alongside satellite-tagged turtles allowed us to explore the mechanisms of dispersal (passive drift or active swimming). Early in the hatching season turtles transited south with strong southward currents. Late in the hatching season, when currents flowed in the opposite direction, turtles uniformly moved northwards across the Equator. However, the movement of individuals differed from what was predicted by surface currents alone. Swimming velocity inferred from track data and an ocean circulation model strongly suggest that turtles' swimming plays a role in maintaining their position within frontal zones seaward of the continental shelf. The long nesting season of adults and behaviour of post-hatchlings exposes young turtles to seasonally varying ocean conditions that lead some individuals further into the South Atlantic and others into the Northern Hemisphere. Such migratory route diversity may ultimately buffer the population against environmental changes or anthropologic threats, fostering population resiliency.

Passive drift or active swimming in marine organisms?

Predictions of organismal movements in a fluid require knowing the fluid's velocity and potential contributions of the organism's behaviour (e.g. swimming or flying). While theoretical aspects of this work are reasonably well-developed, field-based validation is challenging. A much-needed study recently published by Briscoe and colleagues in Proceedings of the Royal Society B compared movements and distribution of satellite-tracked juvenile sea turtles to virtual particles released in a data-assimilating hindcast ocean circulation model. Substantial differences observed between turtles and particles were considered evidence for an important role of active swimming by turtles. However, the experimental design implicitly assumed that transport predictions were insensitive to (i) start location, (ii) tracking duration, (iii) depth, and (iv) physical processes not depicted in the model. Here, we show that the magnitude of variation in physical parameters between turtles and virtual particles can profoundly alter transport predictions, potentially sufficient to explain the reported differences without evoking swimming behaviour. We present a more robust method to derive the environmental contributions to individual movements, but caution that resolving the ocean velocities experienced by individual organisms remains a problem for assessing the role of behaviour in organismal movements and population distributions.

Biomarkers reveal sea turtles remained in oiled areas following the Deepwater Horizon oil spill.

Assessments of large-scale disasters, such as the Deepwater Horizon oil spill, are problematic because while measurements of post-disturbance conditions are common, measurements of pre-disturbance baselines are only rarely available. Without adequate observations of pre-disaster organismal and environmental conditions, it is impossible to assess the impact of such catastrophes on animal populations and ecological communities. Here, we use long-term biological tissue records to provide pre-disaster data for a vulnerable marine organism. Keratin samples from the carapace of loggerhead sea turtles record the foraging history for up to 18 years, allowing us to evaluate the effect of the oil spill on sea turtle foraging patterns. Samples were collected from 76 satellite-tracked adult loggerheads in 2011 and 2012, approximately one to two years after the spill. Of the 10 individuals that foraged in areas exposed to surface oil, none demonstrated significant changes in foraging patterns post spill. The observed long-term fidelity to foraging sites indicates that loggerheads in the northern Gulf of Mexico likely remained in established foraging sites, regardless of the introduction of oil and chemical dispersants. More research is needed to address potential long-term health consequences to turtles in this region. Mobile marine organisms present challenges for researchers to monitor effects of environmental disasters, both spatially and temporally. We demonstrate that biological tissues can reveal long-term histories of animal behavior and provide critical pre-disaster baselines following an anthropogenic disturbance or natural disaster.

Determining origin in a migratory marine vertebrate: a novel method to integrate stable isotopes and satellite tracking.

Stable isotope analysis is a useful tool to track animal movements in both terrestrial and marine environments. These intrinsic markers are assimilated through the diet and may exhibit spatial gradients as a result of biogeochemical processes at the base of the food web. In the marine environment, maps to predict the spatial distribution of stable isotopes are limited, and thus determining geographic origin has been reliant upon integrating satellite telemetry and stable isotope data. Migratory sea turtles regularly move between foraging and reproductive areas. Whereas most nesting populations can be easily accessed and regularly monitored, little is known about the demographic trends in foraging populations. The purpose of the present study was to examine migration patterns of loggerhead nesting aggregations in the Gulf of Mexico (GoM), where sea turtles have been historically understudied. Two methods of geographic assignment using stable isotope values in known-origin samples from satellite telemetry were compared: (1) a nominal approach through discriminant analysis and (2) a novel continuous-surface approach using bivariate carbon and nitrogen isoscapes (isotopic landscapes) developed for this study. Tissue samples for stable isotope analysis were obtained from 60 satellite-tracked individuals at five nesting beaches within the GoM. Both methodological approaches for assignment resulted in high accuracy of foraging area determination, though each has advantages and disadvantages. The nominal approach is more appropriate when defined boundaries are necessary, but up to 42% of the individuals could not be considered in this approach. All individuals can be included in the continuous-surface approach, and individual results can be aggregated to identify geographic hotspots of foraging area use, though the accuracy rate was lower than nominal assignment. The methodological validation provides a foundation for future sea turtle studies in the region to inexpensively determine geographic origin for large numbers of untracked individuals. Regular monitoring of sea turtle nesting aggregations with stable isotope sampling can be used to fill critical data gaps regarding habitat use and migration patterns. Probabilistic assignment to origin with isoscapes has not been previously used in the marine environment, but the methods presented here could also be applied to other migratory marine species.

First satellite tracks of neonate sea turtles redefine the 'lost years' oceanic niche.

Few at-sea behavioural data exist for oceanic-stage neonate sea turtles, a life-stage commonly referred to as the sea turtle 'lost years'. Historically, the long-term tracking of small, fast-growing organisms in the open ocean was logistically or technologically impossible. Here, we provide the first long-term satellite tracks of neonate sea turtles. Loggerheads (Caretta caretta) were remotely tracked in the Atlantic Ocean using small solar-powered satellite transmitters. We show that oceanic-stage turtles (i) rarely travel in Continental Shelf waters, (ii) frequently depart the currents associated with the North Atlantic Subtropical Gyre, (iii) travel quickly when in Gyre currents, and (iv) select sea surface habitats that are likely to provide a thermal benefit or refuge to young sea turtles, supporting growth, foraging and survival. Our satellite tracks help define Atlantic loggerhead nursery grounds and early loggerhead habitat use, allowing us to re-examine sea turtle 'lost years' paradigms.

Predicting the distribution of oceanic-stage Kemp's ridley sea turtles.

The inaccessibility of open ocean habitat and the cryptic nature of small animals are fundamental problems when assessing the distribution of oceanic-stage sea turtles and other marine animals sharing similar life-history traits. Most methods that estimate patterns of abundance cannot be applied in situations that are extremely data limited. Here, we use a movement ecology framework to generate the first predicted distributions for the oceanic stage of the Kemp's ridley sea turtle (Lepidochelys kempii). Our simulations of particle dispersal within ocean circulation models reveal substantial annual variation in distribution and survival among simulated cohorts. Such techniques can help prioritize areas for conservation, and supply inputs for more realistic demographic models attempting to characterize population trends.

Modeling juvenile sea turtle bycatch risk in commercial and recreational fisheries.

Understanding the drivers of fisheries bycatch is essential for limiting its impacts on vulnerable species. Here we present a model to estimate the relative magnitude of sea turtle bycatch in major coastal fisheries across the southeastern US based on spatiotemporal variation in fishing effort and the simulated distributions of juvenile Kemp's ridley (Lepidochelys kempii) and green (Chelonia mydas) sea turtles recruiting from oceanic to nearshore habitats. Over the period modeled (1996-2017), bycatch in recreational fisheries was estimated to be greater than the sum of bycatch that occurred in commercial fisheries that have historically been considered high risks to turtles (e.g., those using trawls, gillnets, and bottom longlines). Prioritizing engagement with recreational anglers to reduce bycatch could be especially beneficial to sea turtle populations. Applying lessons learned from efforts to protect turtles in commercial fisheries may help meet the challenges that arise from the large, diffuse recreational fishing sector.

Adaptive evolution of major histocompatibility complex class I immune genes and disease associations in coastal juvenile sea turtles.

Characterizing polymorphism at the major histocompatibility complex (MHC) genes is key to understanding the vertebrate immune response to disease. Despite being globally afflicted by the infectious tumour disease fibropapillomatosis (FP), immunogenetic variation in sea turtles is minimally explored. We sequenced the α 1 peptide-binding region of MHC class I genes (162 bp) from 268 juvenile green (Chelonia mydas) and 88 loggerhead (Caretta caretta) sea turtles in Florida, USA. We recovered extensive variation (116 alleles) and trans-species polymorphism. Supertyping analysis uncovered three functional MHC supertypes corresponding to the three well-supported clades in the phylogeny. We found significant evidence of positive selection at seven amino acid sites in the class I exon. Random forest modelling and risk ratio analysis of Ch. mydas alleles uncovered one allele weakly associated with smooth FP tumour texture, which may be associated with disease outcome. Our study represents the first characterization of MHC class I diversity in Ch. mydas and the largest sample of sea turtles used to date in any study of adaptive genetic variation, revealing tremendous genetic variation and high adaptive potential to viral pathogen threats. The novel associations we identified between MHC diversity and FP outcomes in sea turtles further highlight the importance of evaluating genetic predictors of disease, including MHC and other functional markers.

Genetic variation among sea turtle life stages and species suggests connectivity among ocean basins.

Regional genetic differentiation of mitochondrial lineages occurs in migratory species with natal philopatry such as sea turtles. However, early juvenile dispersal represents a key opportunity for gene flow and colonization of new regions through founder events, making it an important yet under-studied life stage. To assess connectivity among sea turtle life stages and ocean basins, we sequenced mitochondrial DNA (mtDNA) fragments from 35 juveniles sampled in the Gulf of Mexico from the rarely observed dispersal stage across three species: green turtles (Chelonia mydas; n = 30), hawksbills (Eretmochelys imbricata; n = 3), and loggerheads (Caretta caretta; n = 2). We estimated green turtle rookery contributions using a many-to-many Bayesian mixed stock analysis that incorporated dispersal probabilities based on rookery size and transport via ocean currents. We assembled a gene tree including 709 distinct mtDNA control region haplotypes from the literature for all seven extant sea turtle species to assess gaps in life-stage data across ocean basins, as well as contextualize the lineages we sampled from dispersing juveniles. Our results indicate a high likelihood that green turtles sampled in the Gulf of Mexico originated from rookeries along the coast of Mexico, with smaller contributions from Costa Rica and Suriname. The gene tree analysis yielded species-level relationships consistent with those presented previously, while intra-species relationships between lineages and ocean basins differed, particularly within loggerhead and green turtle clades. Our results highlight the lack of genetic data from juvenile sea turtles, especially the early dispersal stage, and the potential for these data to answer broader questions of connectivity and diversification across species and lineages.

Gene expression changes with tumor disease and leech parasitism in the juvenile green sea turtle skin transcriptome.

Emerging infectious diseases are a major threat to biodiversity in the 21st century. Fibropapillomatosis (FP) is an epithelial tumor disease that affects immature and adult marine turtles worldwide, particularly green turtles (Chelonia mydas). We know little about the host factors contributing to FP susceptibility, in part because transcriptomic studies that compare transcript expression in turtles with and without FP are lacking. Here, we performed RNA-Seq on healthy skin tissue from immature C. mydas in the Indian River Lagoon, Florida, USA, comparing turtles (1) with and without FP and (2) with and without leech parasites, a putative vector of FP. We assembled a de novo C. mydas skin transcriptome to identify transcripts with significant differential expression (DE) across FP and leech categories. Significant DE transcripts were found across FP and leech comparisons, including 10 of the same transcripts with DE across both comparisons. Leech-positive individuals significantly upregulated different immune and viral interaction transcripts than did leech-negative individuals, including viral interaction transcripts associated with herpesvirus interactions. This finding strengthens the role of marine leeches as mechanical vectors of Chelonid herpesvirus 5 (ChHV5) which has been implicated as a causative agent of FP. FP-positive turtles upregulated several tumor progression and suppression transcripts relative to FP-negative turtles, which had no significant DE tumor progression transcripts. FP-positive turtles also upregulated significantly more protein interaction transcripts than FP-negative turtles. DE transcripts across leech comparisons showed no functional enrichment, whereas DE transcripts across FP comparisons showed some GO terms were enriched in FP-positive and FP negative turtles. Notably, only FP-negative turtles were enriched for GO terms involved in acquired and inflammatory immune gene regulation. Overall, our DE transcripts included several candidate genes that may play important roles in C. mydas resistance to or recovery from FP, highlighting that transcriptomics provides a promising venue to understand this impactful disease. Continued investigation of C. mydas responses to FP and leech affliction is imperative for species persistence and the conservation of marine ecosystems worldwide due to the essential role of sea turtles in ecosystem function and stability.

Molecular evolution of fibropapilloma-associated herpesviruses infecting juvenile green and loggerhead sea turtles.

Chelonid Alphaherpesvirus 5 (ChHV5) has long been associated with fibropapillomatosis (FP) tumor disease in marine turtles. Presenting primarily in juvenile animals, FP results in fibromas of the skin, connective tissue, and internal organs, which may indirectly affect fitness by obstructing normal turtle processes. ChHV5 is near-universally present in tumorous tissues taken from affected animals, often at very high concentrations. However, there is also considerable asymptomatic carriage amongst healthy marine turtles, suggesting that asymptomatic hosts play an important role in disease ecology. Currently, there is a paucity of studies investigating variation in viral genetics between diseased and asymptomatic hosts, which could potentially explain why only some ChHV5 infections lead to tumor formation. Here, we generated a database containing DNA from over 400 tissue samples taken from green and loggerhead marine turtles, including multiple tissue types, a twenty year time span, and both diseased and asymptomatic animals. We used two molecular detection techniques, quantitative (q)PCR and nested PCR, to characterize the presence and genetic lineage of ChHV5 in each sample. We found that nested PCR across multiple loci out-performed qPCR and is a more powerful technique for determining infection status. Phylogenetic reconstruction of three viral loci from all ChHV5-positive samples indicated widespread panmixia of viral lineages, with samples taken across decades, species, disease states, and tissues all falling within the same evolutionary lineages. Haplotype networks produced similar results in that viral haplotypes were shared across species, tissue types and disease states with no evidence that viral lineages associated significantly with disease dynamics. Additionally, tests of selection on viral gene trees indicated signals of selection dividing major clades, though this selection did not divide sample categories. Based on these data, neither the presence of ChHV5 infection nor neutral genetic divergence between viral lineages infecting a juvenile marine turtle is sufficient to explain the development of FP within an individual.

Foraging and recruitment hotspot dynamics for the largest Atlantic loggerhead turtle rookery.

Determining patterns of migratory connectivity for highly-mobile, wide-ranging species, such as sea turtles, is challenging. Here, we combined satellite telemetry and stable isotope analysis to estimate foraging locations for 749 individual loggerheads nesting along the east central Florida (USA) coast, the largest rookery for the Northwest Atlantic population. We aggregated individual results by year, identified seven foraging hotspots and tracked these summaries to describe the dynamics of inter-annual contributions of these geographic areas to this rookery over a nine-year period. Using reproductive information for a subset of turtles (n = 513), we estimated hatchling yields associated with each hotspots. We found considerable inter-annual variability in the relative contribution of foraging areas to the nesting adults. Also reproductive success differed among foraging hotspots; females using southern foraging areas laid nests that produced more offspring in all but one year of the study. These analyses identified two high priority areas for future research and conservation efforts: the continental shelf adjacent to east central Florida and the Great Bahama Bank, which support higher numbers of foraging females that provide higher rates of hatchling production. The implementation of the continuous-surface approach to determine geographic origins of unknown migrants is applicable to other migratory species.

Direct evidence of swimming demonstrates active dispersal in the sea turtle "lost years".

Although oceanic dispersal in larval and juvenile marine animals is widely studied, the relative contributions of swimming behavior and ocean currents to movements and distribution are poorly understood [1-4]. The sea turtle "lost years" [5] (often referred to as the surface-pelagic [6] or oceanic [7] stage) are a classic example. Upon hatching, young turtles migrate offshore and are rarely observed until they return to coastal waters as larger juveniles [5]. Sightings of small turtles downcurrent of nesting beaches and in association with drifting organisms (e.g., Sargassum algae) led to this stage being described as a "passive migration" during which turtles' movements are dictated by ocean currents [5-10]. However, laboratory and modeling studies suggest that dispersal trajectories might also be shaped by oriented swimming [11-15]. Here, we use an experimental approach designed to directly test the passive-migration hypothesis by deploying pairs of surface drifters alongside small green (Chelonia mydas) and Kemp's ridley (Lepidochelys kempii) wild-caught turtles, tracking their movements via satellite telemetry. We conclusively demonstrate that these turtles do not behave as passive drifters. In nearly all cases, drifter trajectories were uncharacteristic of turtle trajectories. Species-specific and location-dependent oriented swimming behavior, inferred by subtracting track velocity from modeled ocean velocity, contributed substantially to individual movement and distribution. These findings highlight the importance of in situ observations for depicting the dispersal of weakly swimming animals. Such observations, paired with information on the mechanisms of orientation, will likely allow for more accurate predictions of the ecological and evolutionary processes shaped by animal movement.