Comprehensive Screening Links Halogenated Organic Compounds with Testosterone Levels in Male Delphinus delphis from the Southern California Bight.

While environmental pollutants have been associated with changes in endocrine health in cetaceans, efforts to link contaminant exposure with hormones have largely been limited to a list of known, targeted contaminants, overlooking minimally characterized or unknown compounds of emerging concern. To address this gap, we analyzed a suite of potential endocrine disrupting halogenated organic compounds (HOCs) in blubber from 16 male short-beaked common dolphins ( Delphinus delphis) with known maturity status collected from fishery bycatch in the Southern California Bight. We employed a suspect screening mass spectrometry-based method to investigate a wide range of HOCs that were previously observed in cetaceans from the same region. Potential endocrine effects were assessed through the measurement of blubber testosterone. We detected 167 HOCs, including 81 with known anthropogenic sources, 49 of unknown origin, and 37 with known natural sources. The sum of 11 anthropogenic and 4 unknown HOC classes were negatively correlated with blubber testosterone. Evidence suggests that elevated anthropogenic HOC load contributes to impaired testosterone production in mature male D. delphis. The application of this integrative analytical approach to cetacean contaminant analysis allows for inference of the biological consequences of accumulation of HOCs and prioritization of compounds for future environmental toxicology research.

Blubber cortisol: a potential tool for assessing stress response in free-ranging dolphins without effects due to sampling.

When paired with dart biopsying, quantifying cortisol in blubber tissue may provide an index of relative stress levels (i.e., activation of the hypothalamus-pituitary-adrenal axis) in free-ranging cetacean populations while minimizing the effects of the act of sampling. To validate this approach, cortisol was extracted from blubber samples collected from beach-stranded and bycaught short-beaked common dolphins using a modified blubber steroid isolation technique and measured via commercially available enzyme immunoassays. The measurements exhibited appropriate quality characteristics when analyzed via a bootstraped stepwise parallelism analysis (observed/expected = 1.03, 95%CI: 99.6 - 1.08) and showed no evidence of matrix interference with increasing sample size across typical biopsy tissue masses (75-150 mg; r(2) = 0.012, p = 0.78, slope = 0.022 ng(cortisol deviation)/ul(tissue extract added)). The relationships between blubber cortisol and eight potential cofactors namely, 1) fatality type (e.g., stranded or bycaught), 2) specimen condition (state of decomposition), 3) total body length, 4) sex, 5) sexual maturity state, 6) pregnancy status, 7) lactation state, and 8) adrenal mass, were assessed using a Bayesian generalized linear model averaging technique. Fatality type was the only factor correlated with blubber cortisol, and the magnitude of the effect size was substantial: beach-stranded individuals had on average 6.1-fold higher cortisol levels than those of bycaught individuals. Because of the difference in conditions surrounding these two fatality types, we interpret this relationship as evidence that blubber cortisol is indicative of stress response. We found no evidence of seasonal variation or a relationship between cortisol and the remaining cofactors.

First Assessment of the Sex Ratio for an East Pacific Green Sea Turtle Foraging Aggregation: Validation and Application of a Testosterone ELISA.

Determining sex ratios of endangered populations is important for wildlife management, particularly species subject to sex-specific threats or that exhibit temperature-dependent sex determination. Sea turtle sex is determined by incubation temperature and individuals lack external sex-based traits until sexual maturity. Previous research utilized serum/plasma testosterone radioimmunoassays (RIA) to determine sex in immature/juvenile sea turtles. However, there has been a growing application of enzyme-linked immunosorbent assay (ELISA) for wildlife endocrinology studies, but no study on sea turtles has compared the results of ELISA and RIA. This study provides the first sex ratio for a threatened East Pacific green sea turtle (Chelonia mydas) foraging aggregation, a critical step for future management of this species. Here, we validate a testosterone ELISA and compare results between RIA and ELISA of duplicate samples. The ELISA demonstrated excellent correspondence with the RIA for providing testosterone concentrations for sex determination. Neither assay proved reliable for predicting the sex of reproductively active females with increased testosterone production. We then applied ELISA to examine the sex ratio of 69 green turtles foraging in San Diego Bay, California. Of 45 immature turtles sampled, sex could not be determined for three turtles because testosterone concentrations fell between the ranges for either sex (females: 4.1-113.1 pg/mL, males: 198.4-2,613.0 pg/mL) and these turtles were not subsequently recaptured to enable sex determination; using a Bayesian model to predict probabilities of turtle sex we predicted all three 'unknowns' were female (> 0.86). Additionally, the model assigned all turtles with their correct sex (if determined at recapture) with 100% accuracy. Results indicated a female bias (2.83F:1M) among all turtles in the aggregation; when focusing only on putative immature turtles the sex ratio was 3.5F:1M. With appropriate validation, ELISA sexing could be applied to other sea turtle species, and serve as a crucial conservation tool.