Effects of demineralization on the stable isotope analysis of bone samples.

RATIONALE: The sampling of sequential, annually formed bone growth layers for stable carbon (δ(13)C values) and nitrogen (δ(15)N values) isotope analysis (SIA) can provide a time series of foraging ecology data. To date, no standard protocol exists for the pre-SIA treatment of cortical samples taken from fresh, modern, bones. METHODS: Based on the SIA of historical bone, it is assumed that fresh bone samples must be pre-treated with acid prior to SIA. Using an elemental analyzer coupled to an isotope ratio mass spectrometer to measure stable carbon and nitrogen ratios, we tested the need to acidify cortical bone powder with 0.25 M HCl prior to SIA to isolate bone collagen for the determination of δ(13)C and δ(15)N values. We also examined the need for lipid extraction to remove potential biases related to δ(13)C analysis, based on a C:N ratio threshold of 3.5. RESULTS: It was found that acidification of micromilled cortical bone samples from marine turtles does not affect their δ(15)N values, and the small effect acidification has on δ(13)C values can be mathematically corrected for, thus eliminating the need for pre-SIA acidification of cortical bone. The lipid content of the cortical bone samples was low, as measured by their C:N ratios, indicating that lipid extracting cortical bone samples from modern marine turtles is unnecessary. CONCLUSIONS: We present a standard protocol for testing fresh, modern cortical bone samples prior to SIA, facilitating direct comparison of future studies. Based on the results obtained from marine turtle bones, pre-acidification and lipid removal of cortical bone are not recommended. This is especially useful as there is frequently not enough bone material removed via micromilling of sequential growth layers to accommodate both acid treatment and SIA.

Age and residency duration of loggerhead turtles at a North Pacific bycatch hotspot using skeletochronology.

For migratory marine animals, like sea turtles, effective conservation can be challenging because key demographic information such as duration of life stages and exposure to spatially explicit threats in different habitats are often unknown. In the eastern Pacific near the Baja California Peninsula (BCP), Mexico, tens of thousands of endangered North Pacific loggerhead sea turtles (Caretta caretta) concentrate at a foraging area known to have high rates of fishery bycatch. Because stage survivorship of loggerheads in the BCP will vary significantly depending on the number of years spent in this region, we applied skeletochronology to empirically estimate residency duration in this loggerhead hotspot. The observed age distribution obtained from skeletochronology analysis of 146 dead-stranded loggerheads ranged from three to 24 years old, suggesting a BCP residency of >20 years. Given the maximum estimated age and a one-year migration to western Pacific nesting beaches, we infer age-at-maturation for BCP loggerheads at ~25 years old. We also examine survivorship at varying BCP residency durations by applying our findings to current annual mortality estimates. Predicted survivorship of loggerheads spending over 20 years in this BCP foraging habitat is less than 10%, and given that ~43,000 loggerhead turtles forage here, a significant number of turtles are at extreme risk in this region. This is the first empirical evidence supporting estimated age-at-maturation for BCP North Pacific loggerheads, and the first estimates of BCP stage survivorship. Our findings emphasize the urgent need for continued and effective international conservation efforts to minimize bycatch of this endangered species.