Vertebral cartilage of the clearnose skate, Raja eglanteria: Development, structure, ageing, and hormonal regulation of growth

Incremental marks formed in the vertebral cartilage of most sharks, skates and rays are widely used as indicators of age in elasmobranch growth studies. Such information is essential for fisheries management, thus vertebral ageing has become an invaluable tool for investigating elasmobranch life history. Unfortunately, lack of information on the processes that regulate vertebral growth and mineralization limit efforts to correlate episodic stimuli with increment production. to address this research need, this dissertation investigated these processes through a detailed study on the vertebral cartilage of the clearnose skate, Raja eglanteria. Histologic observations indicated that changes in appositional cartilage growth are the catalyst for increment formation. Observations by scanning electron microscopy supported this conclusion by demonstrating that growth increments differ in the proportion of cells to mineralized matrix. In contrast, elemental analysis of vertebrae using energy-dispersive spectrophotometry demonstrated no change in mineral concentration between seasonal growth layers. This observation was advantageous, because uptake of the radiotracer &\sp{lcub}45{rcub}&Ca was ineffective in estimating the rates of vertebral calcification in captive R. eglanteria. Failure of this standard method appeared to reflect the free exchange of calcium between skeletal and serologic reservoirs. An in vitro method for measuring vertebral growth was developed using incorporation of &\sp{lcub}35{rcub}&S-sulfate as a marker for cartilage matrix synthesis. Certain conserved among elasmobranchs and higher vertebrates. The growth hormone-dependent serum factor insulin-like growth factor-I (IGF-I) increased vertebral matrix synthesis, suggesting an important role for this hormone in regulating elasmobranch skeletal growth. In contrast, corticosterone reduced &\sp{lcub}35{rcub}&S-sulfate uptake in vertebral cartilage, suggesting an inhibitory role for glucocorticosteroids in elasmobranch chondrogenesis. Calcitonin also inhibited vertebral matrix synthesis and, in vivo, may play some role in skeletal development or mineral homeostasis. Finally, nutritional status also appeared to influence vertebral growth in vitro, perhaps indirectly through effects on hormone production. In sum, this dissertation identified changes in cartilage growth as the impetus for vertebral growth zone production in R. eglanteria. Physiological mechanisms that likely regulate vertebral growth are described in this pioneer study on elasmobranch skeletal growth

Why Are You So Slimy?

The epithelium of vertebrates is a complex tissue that houses a large variety of cells with different functions. One of its most important functions is protection and in many cases this function is achieved by exuding copious amounts of mucous that fight off diseases, harmful toxins and overall protect the species from external invaders. Mucous, made of mucin, is produced by specialized epithelial cells called goblet cells. Among vertebrates’, fishes are perhaps one of the major groups known to produce mucous, particularly stingrays. The Atlantic stingray, Dasyatis sabina is one of the few cartilaginous fishes known to inhabit both freshwater areas and saltwater areas. It has been suggested by that mucous production varies significantly among freshwater and saltwater populations. In this study we aim to characterize the epithelium of freshwater vs saltwater D. sabina by quantifying (if any) variations on goblet cells density among different populations (freshwater vs salt water). This study will contribute to a much-needed characterization of stingray epithelia that may be used as a baseline anatomical framework in future studies and will contribute to the limited epithelium related literature among lower vertebrates. In addition, this study will contribute to our understanding of the morphological bases for the differential mucous production among this populations to ultimately relate this to ecosystem differences and environmental issues including water pollution

Abundance and Distribution of Sharks in Northeast Florida Waters and Identification of Potential Nursery Habitat

Sharks are considered top predators in many marine ecosystems and can play an important role in structuring community ecology. As a result, it is necessary to understand the factors that influence their abundance and distribution. This is particularly important as fishery managers develop management plans for sharks that identify areas that serve as essential fish habitat, especially nursery habitat. However, our understanding of shark habitat use in northeast Florida waters is limited. The goal of this study was to characterize the abundance and distribution of sharks in northeast Florida estuaries and to examine the effect of abiotic factors on shark habitat use. A bottom longline survey conducted from 2009 to 2011 indicated that 11 shark species use the estuarine waters of northeast Florida during the summer months. Atlantic Sharpnose Sharks Rhizoprionodon terraenovae, Blacktip Sharks Carcharhinus limbatus, and Bonnetheads Sphyrna tiburo were the most abundant species and made up 81.4% of the total catch. Site, month, and bottom water temperature were the most important factors determining the presence and abundance of sharks and suggest both regional and seasonal variations in the use of northeast Florida waters. Depth, salinity, and dissolved oxygen were also important factors. Our data show that these waters serve as a nursery for Atlantic Sharpnose and Blacktip Sharks, with young-of-the-year and juveniles being present in the summer months. Limited tag-return data reveal that juvenile sharks remain in these waters throughout the summer and that some return in subsequent summers. This is the first study to characterize the abundance and distribution of sharks and identify potential nursery areas in northeast Florida estuaries. Received October 10, 2012; accepted March 7, 2013. © 2013 Copyright Taylor and Francis Group, LLC

Female sperm storage in the bonnethead Sphyrna tiburo oviducal gland: Immunolocalization of steroid hormone receptors in sperm storage tubules

Female sperm storage (FSS) has been demonstrated to occur in representatives from all major vertebrate groups and has been hypothesized to have several possible adaptive benefits that may maximize reproductive success of its practitioners. However, while the range of taxa that exhibit FSS and its possible evolutionary benefits have received significant attention in past years, the physiological mechanisms by which FSS occurs in vertebrates have only recently been explored. In this study, we examined the potential role of gonadal steroid hormones in regulating FSS in the bonnethead Sphyrna tiburo, a small hammerhead species in which females have been shown to be capable of storing male spermatozoa for up to 6 – 7 months following copulation. Like past studies on this species, we observed associations between plasma concentrations of the gonadal steroids 17β-estradiol, testosterone, and progesterone with FSS in female bonnetheads, suggesting roles for these hormones in regulating this process. Using immunohistochemistry, we also observed presence of androgen receptor, estrogen receptor alpha (ERα), and progesterone receptor in epithelial cells of sperm storage tubules in the bonnethead oviducal gland, as well as occurrence of ERα in stored spermatozoa, specifically during the sperm storage period. These results suggest that E2, T, and P4 may regulate certain aspects of FSS in bonnethead indirectly through actions on the female reproductive tract, whereas E2 may also have direct effects on sperm function. This is the first study on the regulation of FSS in sharks and has formed a basis for future work geared towards improving our understanding of this process in chondrichthyans

Resource-use dynamics of co-occurring chondrichthyans from the First Coast, North Florida, USA

Recent studies on shark assemblages on the northeast Florida and southeast Georgia coast (hereafter referred to collectively as the “First Coast”) have demonstrated differences in species and age-class composition of catch from previously characterized estuaries and newly surveyed area beaches, demonstrating that these regions may provide a critical habitat to different segments (i.e., life stages) of local shark populations. In this study, carbon and nitrogen stable isotopes (δ13C and δ15N) from muscle tissue and blood plasma were used to examine trophic dynamics (and temporal variability thereof) of the three dominant co-occurring species found along First Coast beaches (the Atlantic Sharpnose shark Rhizoprionodon terraenovae, Blacknose shark Carcharhinus acronotus and Blacktip shark Carcharhinus limbatus) to determine if they exhibit overlap in resource use along with spatial and temporal habitat use. Although considered spatially segregated from the beach species, a dominant, age-class species found in First Coast estuaries (juvenile Sandbar sharks Carcharhinus plumbeus) was also included in this analysis for comparison. Temporal variability of resource-use characteristics was detected at the species level. Resource-use overlap among species varied by tissue type and was generally higher for blood plasma, suggesting greater resource sharing over more recent time periods. Over longer time periods Atlantic Sharpnose and Blacktip sharks exhibited resource-use expansion, whereas Blacknose sharks exhibited a narrowing in resource use, suggesting a more specialized foraging strategy compared to the other species. The resource-use breadth of Sandbar sharks also expanded between blood plasma and muscle tissue. Significant size relationships were detected in Blacktip and Sandbar sharks, indicating ontogenetic resource shifts for both species. A diversity of highly productive resource pools likely support shark populations along the First Coast such that resource-use differentiation is not required to facilitate species co-occurrence. This work may shed light on understanding patterns of species co-occurrence as well as aid in future conservation efforts

Protein expression of hypoxia-inducible factor 1-alpha (HIF-1α) in spot (Leiostomus xanthurus) exposed to constant and diel-cycling hypoxia

Fish kills often occur overnight from low dissolved oxygen (DO) events, however many mortality events are of unknown cause, since little water quality monitoring occurs overnight. The hypoxia-inducible factor 1α (HIF-1α) protein is an excellent candidate as a biomarker for deciphering idiopathic fish kills. In this study, spot (Leiostomus xanthurus) were exposed to either constant or diel-cycling hypoxia, and HIF-1α expression was compared to normoxic control over three days. The results indicated that HIF-1α protein increased (p \u3c 0.005) in muscle tissue after three days exposure to both constant and a simulated diel-cycling hypoxic event in a laboratory setting when compared to normoxic control animals. In comparison, diel-cycling hypoxia produced higher HIF-1α protein concentrations than constant hypoxia in all treatments, suggesting that the fluctuations of DO induced the expression of protein. The results from this study pose implications for using HIF-1α as a biomarker in wild populations, as overnight hypoxic cycles may produce higher protein concentrations than a constant hypoxic event, which would enable the detection of otherwise unnoticed hypoxic stress. © 2012 Elsevier B.V

Molecular Identification and Functional Characteristics of Peptide Transporters In the Bonnethead Shark (\u3ci\u3eSphyrna tiburo\u3c/i\u3e)

Elasmobranchs are considered to be top marine predators, and in general play important roles in the transfer of energy within marine ecosystems. Despite this, little is known regarding the physiological processes of digestion and nutrient absorption in these fishes. One topic that is particularly understudied is the process of nutrient uptake across the elasmobranch gastrointestinal tract. Given their carnivorous diet, the present study sought to expand knowledge on dietary nutrient uptake in elasmobranchs by focusing on the uptake of products of protein digestion. To accomplish this, a full-length cDNA encoding peptide transporter 1 (PepT1), a protein previously identified within the brush border membrane of vertebrates that is responsible for the translocation of peptides released during digestion by luminal and membrane-bound proteases, was isolated from the bonnethead shark (Sphyrna tiburo). A cDNA encoding the related peptide transporter PepT2 was also isolated from S. tiburo using the same methodology. The presence of PepT1 was then localized in multiple components of the bonnethead digestive tract (esophagus, stomach, duodenum, intestine, rectum, and pancreas) using immunohistochemistry. Vesicle studies were used to identify the apparent affinity of PepT1 and to quantify the rate of dipeptide uptake by its H+-dependent cotransporter properties. The results of this study provide insight into the properties of peptide uptake within the bonnethead gut, and can facilitate future work on physiological regulation of protein metabolism and absorption including how these processes may vary in elasmobranchs that exhibit different feeding strategies

Preliminary Assessment of Total Mercury in Tissues of Marine Associated Avifauna in the South Florida Region

Background/Question/Methods Marine associated avifauna (seabirds) are exposed to wide ranges of environmental contaminants due to spending a large portion of their time in the aquatic environment. Vectors of exposure to metal pollution include external contact, by inhalation, and most generally by ingestion of food and incidental seawater. Seabirds are often considered marine ecosystem bioindicators due to their high trophic level, long lives, and wide geographic ranges. All seabirds are not equally vulnerable to contaminants due to their generalist diets accompanied with varying abilities to excrete, metabolize, or sequester xenobiotics (foreign chemical substances). We examined the concentration of metals in the kidney, liver, breast muscle, and feathers of seven species of adult seabirds commonly found in South Florida: brown pelican Pelecanus occidentalis (n=15), northern gannet Morus bassanus (n=15), double-crested cormorant Phalacrocorax auritus (n=15), royal tern Thalasseus maximus (n=18), osprey Pandion haliaetus (n=15), herring gull Larus argentatus (n=8), and laughing gull Leucophaeus atricilla (n=16). Specimens were collected from four wildlife rehabilitation centers after the bird has died due to trauma or illness; specimens were frozen at death at the respective centers and dissected in the lab. Total mercury analysis used a Direct Mercury analyzer (DMA-80). Results/Conclusions Mercury is a highly toxic nonessential heavy metal that is emitted into the atmosphere by both natural and anthropogenic sources. Total mercury concentrations are thought to be equivalent to methylated mercury in tissues of organisms. Preliminary results were highly variable between species, individuals, and tissue types. Total mercury in the breast muscle, kidney, liver, and feathers ranged from 0.559 to 9.57 µg/g (dry wt), 2.06 to 12.16 µg/g (dry wt), 2.18 to 42.67 µg/g (dry wt), and 1.57 to 11.93 µg/g (dry wt), respectively. In the liver and kidney, values of methylmercury between 20 and 30 µg/g (wet wt) are associated with morbidity and mortality effects in terrestrial bird species. In feathers, total mercury concentrations of 5 µg/g (dry wt) are similarly associated with sublethal systemic and reproductive problems. Royal terns had the highest total mercury in three tissues whereas both gull species had the lowest values in all tissues. Additionally, cormorants and royal tern feathers had total mercury levels above the 5 µg/g threshold. Further assessment will help confirm whether seabird species in South Florida face a significant threat from metal pollution. Variability in metal concentrations between species and tissue types will be discussed in conjunction with concurrent stable isotope analyses in the context of trophic relationships