Lipid extraction is often necessary in stable isotope analysis, and general patterns for when it is necessary need to be more fully explored. We studied the effects of lipid extraction on delta 13C and delta 15N ratios in liver, muscle, and skin of West-Indian manatees (Trichechus manatus) and bottlenose dolphins (Tursiops truncates), two ecologically important species in the Gulf of Mexico that occupy different trophic groups. We also performed a meta-analysis to more broadly determine how the effects lipid extraction vary across taxa, tissues, and trophic groups. Lipid extraction did not affect delta 13C ratios of the three tissues in manatees but significantly affected all tissues in dolphins. The effects of lipid extraction on delta 13C ratios increased with lipid content of tissues and was greatest in liver, followed by skin and muscle. Lipid extraction had no effect on delta 15N ratios of any tissue in either manatees or dolphins. Across 103 species, lipid extraction significantly affected delta 13C ratios in all three tissues we reviewed, and the effects were stronger in lipid-rich tissues. Across species, lipid extraction significantly affected delta 15N ratios in muscles, marginally in liver, but not at all in skin. Finally, lipid extraction had a much smaller effect on herbivores than on carnivores or omnivores, which may explain why lipid extraction did not affect manatee tissues. Researchers have assumed that tissues with C:N ratios > 3.5 require lipid extraction, but we found this was not always true. Our results strengthen the growing body of evidence that the necessity of lipid extraction is tissue- and species-specific. Furthermore, trophic group may forecast the necessity for lipid extraction, possibly due to differences in diet composition or the physiology of lipid synthesis, both of which may lead to variation in lipid composition and subsequently affect the need for extraction.

Manatees are herbivorous mammals found throughout the southeastern United States and are currently expanding their range to higher latitudes. Their diets have been extensively studied throughout the Florida peninsula, but little is known about what they eat at the edges of their range, in areas such as the northcentral Gulf of Mexico (nGOM). To better understand manatee diets in the nGOM, we took vegetation surveys and behavioral observations of tagged manatees approximately weekly in the nGOM during the warm months and at least bi-monthly during the cold months (outside of the nGOM). In addition, opportunistic observations of untagged manatees were conducted. We analyzed stable isotope ratios (δ13C and δ15N) in vegetation and epidermis, muscle, and liver of deceased manatees to better understand the types of vegetation that contributed to manatee diet. Throughout the study manatees were most often observed in freshwater/brackish habitats, eating freshwater vegetation. Studies of manatees along the west coast of Florida indicate that seagrasses make up 48% of manatee diet, however our results indicate that freshwater submerged aquatic vegetation (SAV) was the primary diet source of manatees in the nGOM (from MS to Apalachicola, FL). Preliminary results indicate that tape grass (Vallisneria sp.) was the most common plant observed in areas manatees were spending time, with Southern naiad and Eurasian watermilfoil as the second most common. Initial stable isotope results showed that δ13C values of manatee tissues and vegetation ranged from -24 to -11, indicating that manatees consume a broad diet. δ15N values were ranged from 3-11, suggesting manatees may exhibit some degree of carnivory, either directly or indirectly via consumption of epibionts found on vegetation. Understanding manatee diet in the nGOM will provide information about resources that may be important to manatees as their population grows and they continue to expand their range.

The northern coast of the Gulf of Mexico contains many large estuaries that convey freshwater to northern Gulf ecosystems, including Mobile Bay. Mobile Bay is the sixth largest drainage basin among watersheds within the United States and has the third largest freshwater discharge. Periods of intense rainfall within the watershed can rapidly change the salinity and forage communities in the bay. Currently, it is unknown how the common bottlenose dolphin (Tursiops truncatus) that resides within Mobile Bay responds to these pulses of freshwater. Dolphins may alter their diet or foraging patterns in response to freshwater exposure, which could affect body condition and lead to death. To study the effects of freshwater influx on dolphins in a freshwater-dominated estuary, I will sample skin, muscle, liver, and teeth and quantify body condition of dolphins that stranded dead around Mobile Bay from 2011 to 2018.  This 8-year period includes multiple wet and dry seasons to enable comparisons of diet and body condition relative to freshwater influence. I will use stable isotope ratios in dolphins and prey species and mixing models to compare the short- and long-term shifts in the diets of the dolphins between wet and dry periods. I will use trace element signatures along with sclerochronological techniques in teeth to relate changes in diet to freshwater sources through time within individual dolphins. Photographic analyses and histology (when possible) will further determine the prevalence of pathologies related to freshwater exposure. This research will aid understanding of how highly mobile and high trophic-level, long-lived marine species manage potentially stressful low-salinity conditions associated with residency in freshwater-dominated estuarine environments. These data also provide a baseline to assess effects of other stressors (e.g., oil or other contaminant spills, cold stress, hurricanes, freshwater diversion) that are common to coastal waters, particularly in the Gulf of Mexico.