Sora L. Kim

Using Stable Isotope Analysis to Understand the Migration and Trophic Ecology of Northeastern Pacific White Sharks (Carcharodon carcharias)

The white shark (Carcharodon carcharias) is a wide-ranging apex predator in the northeastern Pacific (NEP). Electronic tagging has demonstrated that white sharks exhibit a regular migratory pattern, occurring at coastal sites during the late summer, autumn and early winter and moving offshore to oceanic habitats during the remainder of the year, although the purpose of these migrations remains unclear. The purpose of this study was to use stable isotope analysis (SIA) to provide insight into the trophic ecology and migratory behaviors of white sharks in the NEP. Between 2006 and 2009, 53 white sharks were biopsied in central California to obtain dermal and muscle tissues, which were analyzed for stable isotope values of carbon (δ13C) and nitrogen (δ15N). We developed a mixing model that directly incorporates movement data and tissue incorporation (turnover) rates to better estimate the relative importance of different focal areas to white shark diet and elucidate their migratory behavior. Mixing model results for muscle showed a relatively equal dietary contribution from coastal and offshore regions, indicating that white sharks forage in both areas. However, model results indicated that sharks foraged at a higher relative rate in coastal habitats. There was a negative relationship between shark length and muscle δ13C and δ15N values, which may indicate ontogenetic changes in habitat use related to onset of maturity. The isotopic composition of dermal tissue was consistent with a more rapid incorporation rate than muscle and may represent more recent foraging. Low offshore consumption rates suggest that it is unlikely that foraging is the primary purpose of the offshore migrations. These results demonstrate how SIA can provide insight into the trophic ecology and migratory behavior of marine predators, especially when coupled with electronic tagging data.

Methods to collect, preserve, and prepare elasmobranch tissues for stable isotope analysis

Stable isotope analysis has the potential to expand our understanding of elasmobranch ecology. However, elasmobranchs share unique traits (i.e., retention of urea, lack of adipose tissue, cartilaginous skeletons) that require modified preparation techniques. Alternative tissue collection and preservation methods would allow sampling from ichthyology collections and at remote locations. We compared different collection, preservation, and preparation techniques to identify treatments that yielded robust isotopic data. Blood components collected in tubes coated with lithium heparin (an anti-coagulant) were not isotopically distinct from blood collected in no-additive tubes. Compared to frozen muscle, ethanol-treated muscle had altered δ13C values, but similar δ15N values. Finally, we removed lipids and urea with petroleum ether and deionized water, respectively. Although untreated and treated muscle had similar amino acid compositions, treated muscle preferentially lost 14N and had greater C:N ratios. These results indicate that urea affects isotope ratios and that water treatment removes urea without altering muscle protein composition. Although not exhaustive, our study begins to address the need for elasmobranch-specific methods.

Isotopic incorporation rates for shark tissues from a long-term captive feeding study

SUMMARY Stable isotope analysis has provided insight into the dietary and habitat patterns of many birds, mammals and teleost fish. A crucial biological parameter to interpret field stable isotope data is tissue incorporation rate, which has not been well studied in large ectotherms. We report the incorporation of carbon and nitrogen into the tissues of leopard sharks (Triakis semifasciata). Because sharks have relatively slow metabolic rates and are difficult to maintain in captivity, no long-term feeding study has been conducted until the point of isotopic steady state with a diet. We kept six leopard sharks in captivity for 1250 days, measured their growth, and serially sampled plasma, red blood cells and muscle for stable carbon and nitrogen isotope analysis. A single-compartment model with first-order kinetics adequately described the incorporation patterns of carbon and nitrogen isotopes for these three tissues. Both carbon and nitrogen were incorporated faster in plasma than in muscle and red blood cells. The rate of incorporation of carbon into muscle was similar to that predicted by an allometric equation relating isotopic incorporation rate to body mass that was developed previously for teleosts. In spite of their large size and unusual physiology, the rates of isotopic incorporation in sharks seem to follow the same patterns found in other aquatic ectotherms.

Ontogenetic and among-individual variation in foraging strategies of northeast Pacific white sharks based on stable isotope analysis

There is growing evidence for individuality in dietary preferences and foraging behaviors within populations of various species. This is especially important for apex predators, since they can potentially have wide dietary niches and a large impact on trophic dynamics within ecosystems. We evaluate the diet of an apex predator, the white shark (Carcharodon carcharias), by measuring the stable carbon and nitrogen isotope composition of vertebral growth bands to create lifetime records for 15 individuals from California. Isotopic variations in white shark diets can reflect within-region differences among prey (most importantly related to trophic level), as well as differences in baseline values among the regions in which sharks forage, and both prey and habitat preferences may shift with age. The magnitude of isotopic variation among sharks in our study (>5‰ for both elements) is too great to be explained solely by geographic differences, and so must reflect differences in prey choice that may vary with sex, size, age and location. Ontogenetic patterns in δ15N values vary considerably among individuals, and one third of the population fit each of these descriptions: 1) δ15N values increased throughout life, 2) δ15N values increased to a plateau at ∼5 years of age, and 3) δ15N values remained roughly constant values throughout life. Isotopic data for the population span more than one trophic level, and we offer a qualitative evaluation of diet using shark-specific collagen discrimination factors estimated from a 3+ year captive feeding experiment (Δ13Cshark-diet and Δ15Nshark-diet equal 4.2‰ and 2.5‰, respectively). We assess the degree of individuality with a proportional similarity index that distinguishes specialists and generalists. The isotopic variance is partitioned among differences between-individual (48%), within-individuals (40%), and by calendar year of sub-adulthood (12%). Our data reveal substantial ontogenetic and individual dietary variation within a white shark population.

A Simple and Non-Invasive Method for Nuclear Transformation of Intact-walled Chlamydomonas reinhardtii

Genetic engineering in microalgae is gaining attraction but nuclear transformation methods available so far are either inefficient or require special equipment. In this study, we employ positively charged nanoparticles, 3-aminopropyl-functionalized magnesium phyllosilicate (aminoclay, approximate unit cell composition of [H2N(CH2)3]8Si8Mg6O12(OH)4), for nuclear transformation into eukaryotic microalgae. TEM and EDX analysis of the process of transformation reveals that aminoclay coats negatively-charged DNA biomolecules and forms a self-assembled hybrid nanostructure. Subsequently, when this nanostructure is mixed with microalgal cells and plated onto selective agar plates with high friction force, cell wall is disrupted facilitating delivery of plasmid DNA into the cell and ultimately to the nucleus. This method is not only simple, inexpensive, and non-toxic to cells but also provides efficient transformation (5.03×102 transformants/µg DNA), second only to electroporation which needs advanced instrumentation. We present optimized parameters for efficient transformation including pre-treatment, friction force, concentration of foreign DNA/aminoclay, and plasticity of agar plates. It is also confirmed the successful integration and stable expression of foreign gene in Chlamydomonas reinhardtii through molecular methods.

Unifying latitudinal gradients in range size and richness across marine and terrestrial systems

Many marine and terrestrial clades show similar latitudinal gradients in species richness, but opposite gradients in range size—on land, ranges are the smallest in the tropics, whereas in the sea, ranges are the largest in the tropics. Therefore, richness gradients in marine and terrestrial systems do not arise from a shared latitudinal arrangement of species range sizes. Comparing terrestrial birds and marine bivalves, we find that gradients in range size are concordant at the level of genera. Here, both groups show a nested pattern in which narrow-ranging genera are confined to the tropics and broad-ranging genera extend across much of the gradient. We find that (i) genus range size and its variation with latitude is closely associated with per-genus species richness and (ii) broad-ranging genera contain more species both within and outside of the tropics when compared with tropical- or temperate-only genera. Within-genus species diversification thus promotes genus expansion to novel latitudes. Despite underlying differences in the species range-size gradients, species-rich genera are more likely to produce a descendant that extends its range relative to the ancestors range. These results unify species richness gradients with those of genera, implying that birds and bivalves share similar latitudinal dynamics in net species diversification.

Evidence from shark teeth for a brackish Arctic Ocean in the Eocene greenhouse

Temperatures are rising at an unprecedented rate in the Arctic, and affecting the sensitive hydrological cycle with increased freshwater inputs to the marine environment. A deep-time analogue often used to understand and predict the environmental and ecological impacts of global warming is the early-middle Eocene greenhouse (ca. 53–38 Ma). Although the terrestrial Arctic during this interval is well documented, marine records are rare and largely based on one central Arctic Ocean site at Lomonosov Ridge. Here we estimate a mean paleosalinity of 12.7 PSU for coastal waters of the western Arctic Ocean by incorporating δ 18 O CO 3 values of shark teeth with estimates of Eocene Arctic paleotemperature and freshwater δ 18 O values into a salinity model. This paleosalinity is substantially lower than estimates by others for the Eocene central Arctic Ocean (21–25 PSU) and today9s Arctic Ocean (32–35 PSU). Our results provide the first Eocene salinity estimate for the western Arctic Ocean and, when considered with prior results from Lomonosov Ridge, suggest a reduced surface salinity, but larger salinity gradient (∼10 PSU) across the Eocene Arctic Ocean during the early–middle Eocene greenhouse. From an ecologic perspective, while today9s lamniform sharks are largely intolerant of low salinity, Eocene species were abundant in the brackish Arctic Ocean, suggesting that past environmental tolerances were much greater than today.

Unique Biochemical and Mineral Composition of Whale Ear Bones

Cetaceans are obligate aquatic mammals derived from terrestrial artiodactyls. The defining characteristic of cetaceans is a thick and dense lip (pachyosteosclerotic involucrum) of an ear bone (the tympanic). This unique feature is absent in modern terrestrial artiodactyls and is suggested to be important in underwater hearing. Here, we investigate the mineralogical and biochemical properties of the involucrum, as these may hold clues to the aquatic adaptations of cetaceans. We compared bioapatites (enamel, dentine, cementum, and skeletal bone) of cetaceans with those of terrestrial artiodactyls and pachyosteosclerotic ribs of manatees (Sirenia). We investigated organic, carbonate, and mineral composition as well as crystal size and crystallinity index. In all studied variables, bioapatites of the cetacean involucrum were intermediate in composition and structure between those of tooth enamel on the one hand and those of dentine, cementum, and skeletal bone on the other. We also studied the amino acid composition of the cetacean involucrum relative to that of other skeletal bone. The central involucrum had low glycine and hydroxyproline concentrations but high concentrations of nonessential amino acids, unlike most bone samples but similar to the tympanic of hippos and the (pachyosteosclerotic) ribs of manatees. These amino acid results are evidence of rapid bone development. We hypothesize that the mineralogical and amino acid composition of cetacean bullae differs from that of other bone because of (1) functional modifications for underwater sound reception and (2) structural adaptations related to rapid ossification.

Discrimination Factors and Incorporation Rates for Organic Matrix in Shark Teeth Based on a Captive Feeding Study

Sharks migrate annually over large distances and occupy a wide variety of habitats, complicating analysis of lifestyle and diet. A biogeochemical technique often used to reconstruct shark diet and environment preferences is stable isotope analysis, which is minimally invasive and integrates through time and space. There are previous studies that focus on isotopic analysis of shark soft tissues, but there are limited applications to shark teeth. However, shark teeth offer an advantage of multiple ecological snapshots and minimum invasiveness during removal because of their distinct conveyor belt tooth replacement system. In this study, we analyze δ13C and δ15N values of the organic matrix in leopard shark teeth (Triakis semifasciata) from a captive experiment and report discrimination factors as well as incorporation rates. We found differences in tooth discrimination factors for individuals fed different prey sources (mean ± SD; Δ13Csquid = 4.7‰ ± 0.5‰, Δ13Ctilapia = 3.1‰ ± 1.0‰, Δ15Nsquid = 2.0‰ ± 0.7‰, Δ15Ntilapia = 2.8‰ ± 0.6‰). In addition, these values differed from previously published discrimination factors for plasma, red blood cells, and muscle of the same leopard sharks. Incorporation rates of shark teeth were similar for carbon and nitrogen (mean ± SE; λC = 0.021 ± 0.009, λN = 0.024 ± 0.007) and comparable to those of plasma. We emphasize the difference in biological parameters on the basis of tissue substrate and diet items to interpret stable isotope data and apply our results to stable isotope values from blue shark (Prionace glauca) teeth to illustrate the importance of biological parameters to interpret the complex ecology of a migratory shark.