Short-term resilience to climate-induced temperature increases for equatorial sea turtle populations.

Projection models are being increasingly used to manage threatened taxa by estimating their responses to climate change. Sea turtles are particularly susceptible to climate change as they have temperature-dependent sex determination and increased sand temperatures on nesting beaches could result in the 'feminisation' of hatchling sex ratios for some populations. This study modelled likely long-term trends in sand temperatures and hatchling sex ratios at an equatorial nesting site for endangered green turtles (Chelonia mydas) and critically endangered hawksbill turtles (Eretmochelys imbricata). A total of 1078 days of sand temperature data were collected from 28 logger deployments at nest depth between 2018 and 2022 in Papua New Guinea (PNG). Long-term trends in sand temperature were generated from a model using air temperature as an environmental proxy. The influence of rainfall and seasonal variation on sand temperature was also investigated. Between 1960 and 2019, we estimated that sand temperature increased by ~0.6°C and the average hatchling sex ratio was relatively balanced (46.2% female, SD = 10.7). No trends were observed in historical rainfall anomalies and projections indicated no further changes to rainfall until 2100. Therefore, the sex ratio models were unlikely to be influenced by changing rainfall patterns. A relatively balanced sex ratio such as this is starkly different to the extremely female-skewed hatchling sex ratio (>99% female) reported for another Coral Sea nesting site, Raine Island (~850 km West). This PNG nesting site is likely rare in the global context, as it is less threatened by climate-induced feminisation. Although there is no current need for 'cooling' interventions, the mean projected sex ratios for 2020-2100 were estimated 76%-87% female, so future interventions may be required to increase male production. Our use of long-term sand temperature and rainfall trends has advanced our understanding of climate change impacts on sea turtles.

Development and application of species-specific cell-based bioassays to assess toxicity in green sea turtles.

Despite the detection of a wide range of contaminants in the blood of green turtle populations foraging in three locations of northern Queensland - Upstart Bay, Cleveland Bay and the Howick Group of Reefs, little is known about the effects of these contaminants on turtle health. Newly developed cell-based bioassays using green turtle primary cell cultures provide an ethical, reproducible, and high-throughput method for assessing the risk of chemical exposure sea turtles. In this project, the toxicity of six priority metals (Mn, Co, Mo, As, Sb, Cu) and blood extracts from foraging turtles were tested in two bioassays adapted to green turtle primary skin and liver cells. Cytotoxicity of metals and blood extracts was measured in primary skin fibroblast cells using a resazurin assay. Glutathione-S-transferase (GST) activity was measured in primary skin fibroblasts and primary liver epithelial cells following exposure to metals and blood extracts. Arsenic, molybdenum, cobalt and copper were found to be cytotoxic to green turtle skin cells. Only manganese, cobalt and copper were found to alter GST activity, predominantly in skin cells, indicating a higher sensitivity of green turtle skin cells compared to liver cells. Effect concentrations of metals in both bioassays were above concentrations found in turtle blood. Turtle blood extracts from the three foraging grounds showed differences in cytotoxicity and GST activity. In both assays, blood extracts of turtles from Upstart Bay were the most toxic, followed by those from Cleveland Bay, then the Howick Reefs, suggesting turtles from Upstart Bay and Cleveland Bay may be at risk from current concentrations of organic contaminants. This study demonstrates that species-specific cell-based bioassays can be used effectively to assess chemical risk in sea turtles and their foraging grounds, and could be applied to assess chemical risk in other marine wildlife.

Coastal bays and coral cays: Multi-element study of Chelonia mydas forage in the Great Barrier Reef (2015-2017).

There is increasing interest in understanding potential impacts of complex pollutant profiles to long-lived species such as the green sea turtle (Chelonia mydas), a threatened megaherbivore resident in north Australia. Dietary ingestion may be a key exposure route for metals in these animals and marine plants can accumulate metals at higher concentrations than the surrounding environment. We investigated concentrations of 19 metals and metalloids in C. mydas forage samples collected from a group of offshore coral cays and two coastal bays over a period of 2-3 years. Although no samples exceeded sediment quality guidelines, coastal forage Co, Fe, and V concentrations were up to 2-fold higher, and offshore forage Sr concentrations were ~3-fold higher, than global seagrass means. Principal Component Analysis differentiated coastal bay from coral cay forage according to patterns consistent with underlying terrigenous-type or marine carbonate-type sediment geochemistry, such that coastal bay forage was higher in Fe, Co, Mn, Cu, and Mo (and others) but forage from coral cays was higher in Sr and U. Forage from the two coastal bays was differentiated according to temporal variation in metal profiles, which may be associated with a more episodic sediment disturbance regime in one of the bays. For all study locations, some forage metal concentrations were higher than previously reported in the global literature. Our results suggest that forage metal profiles may be influenced by the presence of some metals in insoluble forms or bound to ultra-fine sediment particles adhered to forage surfaces. Metal concentrations in Great Barrier Reef forage may be present at levels higher than expected from the global seagrass literature and appear strongly influenced by underlying sediment geochemistry.

Green turtle (Chelonia mydas) population demographics at three chemically distinct foraging areas in the northern Great Barrier Reef.

The catchments of the Great Barrier Reef (GBR) have experienced significant modifications in recent decades, leading to increases in sources of pollutants and declines in coastal water quality. As coastal waters of the GBR support some of the highest density green turtle (Chelonia mydas) foraging populations in the western Pacific Ocean, understanding the effects of contaminants on GBR green turtle populations is a priority. In 2012, elevated strandings of green turtles in the Upstart Bay region instigated the WWF's collaborative Rivers to Reef to Turtles (RRT) project to investigate if coastal pollutants are compromising green turtle health. Important to interpreting these investigations into toxicology and health is understanding the demographics of the green turtle populations being investigated. In three green turtle foraging grounds, Cleveland Bay (CLV), Upstart Bay (UPB) and the Howick Group of Reefs (HWK), this study explored population size, age class structure, sex ratio, growth rates, body condition and diet, as well as indices of turtle health, such as plastron barnacle loads and eye lesions. The three foraging populations had similar age class structure and adult sex ratios to other green turtle foraging populations in the GBR. Somatic growth rate was nonlinear, peaking in immature turtles, and was much slower in turtles foraging at HWK compared to the other two sites. This may have been due to differences in food source, which was supported by the observed dietary shifts between seagrass and algae in HWK turtles, compared to a consistently seagrass diet in CLV and UPB turtles. There were also small differences in body condition between sites, as well as differences in barnacle loads, eye lesions and occurrence of fibropapilloma tumors. This study provides important information on green turtle foraging ground population dynamics in the northern GBR, and context for the other papers in this special issue.

Monitoring the health of green turtles in northern Queensland post catastrophic events.

Between 2014 and 2017, the Rivers to Reef to Turtles (RRT) project examined the health of green turtles at two coastal sites impacted by urban and agricultural human activities (Cleveland and Upstart Bays) and one proposed pristine site (Howick Group of Reefs) in northern Queensland, Australia, through blood biochemistry and haematology, plasma protein electrophoresis, and clinical assessments including body condition and barnacle counts. Furthermore, cases of mortality were subjected to comprehensive postmortem examination. In an attempt to advance diagnostics, associations between specific contaminants and health of turtles in this region were tested. No comprehensive health assessments had been conducted at these sites prior to this study. The coastal Cleveland and Upstart Bays both demonstrated effects likely to be in response to stressors suspected to be anthropogenic in origin (elevated total white cell counts and creatinine kinase levels across the populations, respectively). This was associated with a suite of trace elements, in particular cobalt. While these indicators of stress resolved by the final year of the study, a chronic stressor was suspected to be persisting with ongoing low albumin: globulin. Necropsies did not elucidate any specific diseases. Although body condition index did not closely correlate with site health, barnacle counts in juvenile turtles may prove a reliable indicator of site health. Based on previously established indicators of poor health, barnacle counts showed that 10% of the population was in poor health at Upstart Bay and nearly 20% of the population at Cleveland Bay. This is above what would be expected for a normal population. Overall, the health component of this study suggested that the pristine turtle population was healthy and the coastal turtle populations were under active stressors, possibly caused by anthropogenic effectors such as chemical pollutants, when initially examined in 2014. These stressors resolved by the conclusion of the study in 2017; but chronic stressors remained absent in the pristine site and present within each of the studied coastal populations.

Effect-based approach for screening of chemical mixtures in whole blood of green turtles from the Great Barrier Reef.

Organisms are exposed to mixtures of both known and unknown chemicals which are diverse and variable, and thus difficult and costly to characterise and monitor using traditional target analyses. The objective of this study was to validate and apply in vitro effect-based methods by which whole blood can be used to screen internal exposure to such complex chemical mixtures. For this study, we used whole blood of green sea turtles (Chelonia mydas). To ensure the chemical mixture in blood is transferred with minimal losses or bias, we tested a modified QuEChERS extraction method specifically developed for multi- and non-target instrument analysis. The extracts were dosed to a battery of in vitro bioassays (AhR-CAFLUX, AREc32, NFκB-bla, VM7Luc4E2, Microtox), each with a different mode of action (e.g., AhR receptor mediated xenobiotics, NrF2-mediated oxidative stress, NFκB mediated response to inflammation, estrogen activity and baseline toxicity oxidative stress, respectively) in order to cover a wide spectrum of chemicals. Results confirmed the absence of interferences of the blood extract with the responses of the different assays, thus indicating the methods' compatibility with effect-based screening approaches. To apply this approach, whole blood samples were collected from green turtles foraging in agricultural, urban and remote areas of the Australian Great Barrier Reef. The effect-based screening revealed significant differences in exposure, with higher induction of AhR-CAFLUX, AREc32 and Microtox assays in turtles from the agricultural foraging ground. Overall, these results corroborated with concurrent health, target and non-target analyses in the same animals performed as part of a larger program. This study provides evidence that the proposed effect-based approach is suitable for screening and evaluating internal exposure of organisms to chemical mixtures. The approach could be valuable for advancing understanding on multiple levels ranging from identification of priority chemicals in effect-directed investigations to exploring relationships between exposure and disease, not only in sea turtles, but in any organism.