World phylogeography and male-mediated gene flow in the sandbar shark, Carcharhinus plumbeus.

The sandbar shark, Carcharhinus plumbeus, is a large, cosmopolitan, coastal species. Females are thought to show philopatry to nursery grounds while males potentially migrate long distances, creating an opportunity for male-mediated gene flow that may lead to discordance in patterns revealed by mitochondrial DNA (mtDNA) and nuclear markers. While this dynamic has been investigated in elasmobranchs over small spatial scales, it has not been examined at a global level. We examined patterns of historical phylogeography and contemporary gene flow by genotyping 329 individuals from nine locations throughout the species' range at eight nuclear microsatellite markers and sequencing the complete mtDNA control region. Pairwise comparisons often resulted in fixation indices and divergence estimates of greater magnitude using mtDNA sequence data than microsatellite data. In addition, multiple methods of estimation suggested fewer populations based on microsatellite loci than on mtDNA sequence data. Coalescent analyses suggest divergence and restricted migration among Hawaii, Taiwan, eastern and western Australia using mtDNA sequence data and no divergence and high migration rates, between Taiwan and both Australian sites using microsatellite data. Evidence of secondary contact was detected between several localities and appears to be discreet in time rather than continuous. Collectively, these data suggest complex spatial/temporal relationships between shark populations that may feature pulses of female dispersal and more continuous male-mediated gene flow.

Genome-wide SNPs resolve spatiotemporal patterns of connectivity within striped marlin (Kajikia audax), a broadly distributed and highly migratory pelagic species.

Genomic methodologies offer unprecedented opportunities for statistically robust studies of species broadly distributed in environments conducive to high gene flow, providing valuable information for wildlife conservation and management. Here, we sequence restriction site-associated DNA to characterize genome-wide single nucleotide polymorphisms (SNPs) in a broadly distributed and highly migratory large pelagic fish, striped marlin (Kajikia audax). Assessment of over 4,000 SNPs resolved spatiotemporal patterns of genetic connectivity throughout the species range in the Pacific and, for the first time, Indian oceans. Individual-based cluster analyses identified six genetically distinct populations corresponding with the western Indian, eastern Indian, western South Pacific, and eastern central Pacific oceans, as well as two populations in the North Pacific Ocean (F ST = 0.0137-0.0819). F ST outlier analyses identified a subset of SNPs (n = 59) putatively under the influence of natural selection and capable of resolving populations separated by comparatively high degrees of genetic differentiation. Temporal collections available for some regions demonstrated the stability of allele frequencies over three to five generations of striped marlin. Relative migration rates reflected lower levels of genetic connectivity between Indian Ocean populations (m R ≤ 0.37) compared with most populations in the Pacific Ocean (m R ≥ 0.57) and highlight the importance of the western South Pacific in facilitating gene flow between ocean basins. Collectively, our results provide novel insights into rangewide population structure for striped marlin and highlight substantial inconsistencies between genetically distinct populations and stocks currently recognized for fisheries management. More broadly, we demonstrate that species capable of long-distance dispersal in environments lacking obvious physical barriers to movement can display substantial population subdivision that persists over multiple generations and that may be facilitated by both neutral and adaptive processes. Importantly, surveys of genome-wide markers enable inference of population-level relationships using sample sizes practical for large pelagic fishes of conservation concern.

Evaluation of biomarkers for monitoring thrombogenic potential of FXaI16L.

: A zymogen-like activated factor X variant (FXa) is being developed for treating acute bleeding conditions. Activated factor V is an essential cofactor to FXa for activating prothrombin to thrombin. Thrombi/emboli formation was observed microscopically in FXa toxicity studies in animals. The objective of this research was to evaluate candidate biomarkers for FXa-induced thrombi/emboli formation to inform safety monitoring and dose-escalation decisions in FXa clinical trials. Effects of intravenous FXa administration on platelets, fibrinogen, activated partial thromboplastin time (aPTT), prothrombin time (PT), D-dimer, tissue factor pathway inhibitor, thrombin : antithrombin complex, antithrombin, and factor V, and protein C (PC) activities were evaluated in mice, rats, and monkeys. Mice had endogenous factor V activity 10× that of monkeys and were overly sensitive to FXa-induced thrombi/emboli formation. In monkeys, decreases in fibrinogen and prolongation in aPTT and PT emerged as potential biomarkers for impending FXa-induced thrombi/emboli formation, based on association of changes with microscopically observable thrombi/emboli (0-97 thrombi/emboli per monkey). PC decreases, measured by a clot-based assay, were also observed. A similar reduction in PC activity, when measured by clot-based assay, was observed in a phase 1 clinical trial. However, an in-vitro experiment with human plasma spiked with increasing concentrations of FXa indicated dose-dependent FXa-induced interference with clot-based assays and no depletion of PC or S by FXa in non-clot-based assays. Nonclinical biomarker studies identified fibrinogen, aPTT and PT as potential biomarkers for monitoring the clinical safety of FXa. Results of clot-based assays with FXa treatment should be interpreted with caution.

Physiological stress and post-release mortality of white marlin (Kajikia albida) caught in the United States recreational fishery.

White marlin, a highly migratory pelagic marine fish, support important commercial and recreational fisheries throughout their range in the tropical and subtropical Atlantic Ocean. More than 10 000 individuals can be caught annually in the United States recreational fishery, of which the vast majority are captured on circle hooks and released alive. The probability of post-release mortality of white marlin released from circle hooks has been documented to be <0.02, but the associated physiological stress resulting from capture and handling techniques has not been characterized despite its importance for understanding the health of released fish. We examined the physiological response of 68 white marlin caught on circle hooks in the recreational fishery and followed the fate of 22 of these fish with pop-up satellite archival tags programmed to release after 30 days. Measures of plasma sodium, chloride, glucose and lactate concentrations taken from fish that were briefly and consistently (mean = 120 s, standard deviation = 40 s) removed from the water increased with angling time, but post-release mortality was inversely related to angling time. The probability of post-release mortality was predicted by elevated plasma potassium concentrations and was more than 10 times greater than has been previously reported for white marlin caught on circle hooks that were not removed from the water. This disparity in estimates of post-release mortality suggests that removal of fish from the water for physiological sampling greatly heightens stress, disrupts homeostasis and thus increases the probability of post-release mortality. Our results demonstrate that elevated concentrations of plasma potassium predict mortality in white marlin and that the probability of post-release mortality is highly dependent on post-capture handling procedures.

Fisheries conservation on the high seas: linking conservation physiology and fisheries ecology for the management of large pelagic fishes.

Populations of tunas, billfishes and pelagic sharks are fished at or over capacity in many regions of the world. They are captured by directed commercial and recreational fisheries (the latter of which often promote catch and release) or as incidental catch or bycatch in commercial fisheries. Population assessments of pelagic fishes typically incorporate catch-per-unit-effort time-series data from commercial and recreational fisheries; however, there have been notable changes in target species, areas fished and depth-specific gear deployments over the years that may have affected catchability. Some regional fisheries management organizations take into account the effects of time- and area-specific changes in the behaviours of fish and fishers, as well as fishing gear, to standardize catch-per-unit-effort indices and refine population estimates. However, estimates of changes in stock size over time may be very sensitive to underlying assumptions of the effects of oceanographic conditions and prey distribution on the horizontal and vertical movement patterns and distribution of pelagic fishes. Effective management and successful conservation of pelagic fishes requires a mechanistic understanding of their physiological and behavioural responses to environmental variability, potential for interaction with commercial and recreational fishing gear, and the capture process. The interdisciplinary field of conservation physiology can provide insights into pelagic fish demography and ecology (including environmental relationships and interspecific interactions) by uniting the complementary expertise and skills of fish physiologists and fisheries scientists. The iterative testing by one discipline of hypotheses generated by the other can span the fundamental-applied science continuum, leading to the development of robust insights supporting informed management. The resulting species-specific understanding of physiological abilities and tolerances can help to improve stock assessments, develop effective bycatch-reduction strategies, predict rates of post-release mortality, and forecast the population effects of environmental change. In this synthesis, we review several examples of these interdisciplinary collaborations that currently benefit pelagic fisheries management.

Ultrasound assisted dispersal of a copper nanopowder for electroless copper activation.

This paper describes the ultrasound assisted dispersal of a low wt./vol.% copper nanopowder mixture and determines the optimum conditions for de-agglomeration. A commercially available powder was added to propan-2-ol and dispersed using a magnetic stirrer, a high frequency 850 kHz ultrasonic cell, a standard 40 kHz bath and a 20 kHz ultrasonic probe. The particle size of the powder was characterized using dynamic light scattering (DLS). Z-Average diameters (mean cluster size based on the intensity of scattered light) and intensity, volume and number size distributions were monitored as a function of time and energy input. Low frequency ultrasound was found to be more effective than high frequency ultrasound at de-agglomerating the powder and dispersion with a 20 kHz ultrasonic probe was found to be very effective at breaking apart large agglomerates containing weakly bound clusters of nanoparticles. In general, the breakage of nanoclusters was found to be a factor of ultrasonic intensity, the higher the intensity the greater the de-agglomeration and typically micron sized clusters were reduced to sub 100 nm particles in less than 30 min using optimum conditions. However, there came a point at which the forces generated by ultrasonic cavitation were either insufficient to overcome the cohesive bonds between smaller aggregates or at very high intensities decoupling between the tip and solution occurred. Absorption spectroscopy indicated a copper core structure with a thin oxide shell and the catalytic performance of this dispersion was demonstrated by drop coating onto substrates and subsequent electroless copper metallization. This relatively inexpensive catalytic suspension has the potential to replace precious metal based colloids used in electronics manufacturing.

Distribution and habitat associations of billfish and swordfish larvae across mesoscale features in the Gulf of Mexico.

Ichthyoplankton surveys were conducted in surface waters of the northern Gulf of Mexico (NGoM) over a three-year period (2006-2008) to determine the relative value of this region as early life habitat of sailfish (Istiophorus platypterus), blue marlin (Makaira nigricans), white marlin (Kajikia albida), and swordfish (Xiphias gladius). Sailfish were the dominant billfish collected in summer surveys, and larvae were present at 37.5% of the stations sampled. Blue marlin and white marlin larvae were present at 25.0% and 4.6% of the stations sampled, respectively, while swordfish occurred at 17.2% of the stations. Areas of peak production were detected and maximum density estimates for sailfish (22.09 larvae 1000 m(-2)) were significantly higher than the three other species: blue marlin (9.62 larvae 1000 m(-2)), white marlin (5.44 larvae 1000 m(-2)), and swordfish (4.67 larvae 1000 m(-2)). The distribution and abundance of billfish and swordfish larvae varied spatially and temporally, and several environmental variables (sea surface temperature, salinity, sea surface height, distance to the Loop Current, current velocity, water depth, and Sargassum biomass) were deemed to be influential variables in generalized additive models (GAMs). Mesoscale features in the NGoM affected the distribution and abundance of billfish and swordfish larvae, with densities typically higher in frontal zones or areas proximal to the Loop Current. Habitat suitability of all four species was strongly linked to physicochemical attributes of the water masses they inhabited, and observed abundance was higher in slope waters with lower sea surface temperature and higher salinity. Our results highlight the value of the NGoM as early life habitat of billfishes and swordfish, and represent valuable baseline data for evaluating anthropogenic effects (i.e., Deepwater Horizon oil spill) on the Atlantic billfish and swordfish populations.

Atlantic bluefin tuna (Thunnus thynnus) population dynamics delineated by organochlorine tracers.

Atlantic bluefin tuna (ABFT) are highly valued and heavily exploited, and critical uncertainties regarding their population structure hinder effective management. Evidence supports the existence of two breeding populations of ABFT; a western population in the Gulf of Mexico and an eastern population in the Mediterranean Sea; both of which migrate and mix in the North Atlantic. Conventional tagging studies suggest low rates of trans-Atlantic migrations; however, electronic tagging and stable isotopes in otoliths indicate stock mixing up to 57% between management zones delineated by 45 degrees W longitude. Here we show that organochlorine pesticides and polychlorinated biphenyls (PCBs) can be used as tracers of bluefin tuna foraging grounds in the North Atlantic and confirm that stock mixing of juvenile tuna within the U.S. Mid Atlantic Bight is indeed high (33-83% eastern origin), and is likely spatially and temporally variable. We further demonstrate that >10% of the Mediterranean population is migratory, that young bluefin tuna migrate from the Mediterranean to western Atlantic foraging grounds as early as age 1, and then return to the Mediterranean Sea as young as age 5, presumably to breed. The tracer method described here provides a novel means for distinguishing bluefin tuna populations and ontogenetic shifts in migration in the North Atlantic.

Genetic identity of YOY bluefin tuna from the eastern and western Atlantic spawning areas.

We used 320 young-of-the-year (YOY) specimens of the highly migratory and overfished Atlantic bluefin tuna, Thunnus thynnus, Linnaeus 1758, to evaluate the hypothesis that Atlantic bluefin tuna comprises 2 stocks with spawning grounds in the Gulf of Mexico and in the Mediterranean Sea. Significant genetic differentiation at 8 nuclear microsatellite loci (F(ST) = 0.0059, P = 0.0005) and at the mitochondrial control region (Phi(ST) = 0.0129, P = 0.0139) was detected among YOY Atlantic bluefin tuna captured on spawning grounds in the Gulf of Mexico (n = 40) versus the western (n = 255) and eastern (n = 25) basins of the Mediterranean Sea. The genetic divergence among spawning populations, combined with the extensive trans-Atlantic movements reported for juvenile and adult Atlantic bluefin tuna, indicates a high degree of spawning site fidelity. Recognition of genetically distinct populations necessitates independent management of Atlantic bluefin tuna on spawning grounds and warrants evaluation of the level of mixing of populations on feeding grounds. The genetic pattern might not have been detected unless juvenile specimens or actively spawning adults had been sampled.

Genetic polyandry and sexual conflict in the sandbar shark, Carcharhinus plumbeus, in the western North Atlantic and Gulf of Mexico.

To investigate patterns of polyandry in the sandbar shark (Carcharhinus plumbeus), 20 pregnant females were sampled from the western North Atlantic and Gulf of Mexico. Five species-specific microsatellite markers were used to genotype each shark and its litter. Of 20 litters, 17 (85%) were shown to have multiple sires. In multiply sired litters, the estimated minimum number of sires ranged from two to five with an average of 2.3 males per litter. Regression analysis did not demonstrate a significant relationship between female reproductive success and female body size or sire number and female body size. There was a high incidence of reproductive skew noted in litters, and two groups of males with significantly different mean reproductive success were observed. Analyses using Bateman's principles suggest that there is less direct benefit for females that acquire multiple mates than for males who bias paternity within litters. In light of past morphological and behavioural studies, these data suggest that patterns of polyandry in elasmobranchs may be determined by coercive mating, and that breeding behaviour has likely evolved in the context of sexual conflict.

GEOGRAPHIC DISTRIBUTION OF MOLECULAR VARIANCE WITHIN THE BLUE MARLIN (MAKAIRA NIGRICANS): A HIERARCHICAL ANALYSIS OF ALLOZYME, SINGLE-COPY NUCLEAR DNA, AND MITOCHONDRIAL DNA MARKERS.

This study presents a comparative hierarchical analysis of variance applied to three classes of molecular markers within the blue marlin (Makaira nigricans). Results are reported from analyses of four polymorphic allozyme loci, four polymorphic anonymously chosen single-copy nuclear DNA (scnDNA) loci, and previously reported restriction fragment length polymorphisms (RFLPs) of mitochondrial DNA (mtDNA). Samples were collected within and among the Atlantic and Pacific Oceans over a period of several years. Although moderate levels of genetic variation were detected at both polymorphic allozyme (H = 0.30) and scnDNA loci (H = 0.37), mtDNA markers were much more diverse (h = 0.85). Allele frequencies were significantly different between Atlantic and Pacific Ocean samples at three of four allozyme loci and three of four scnDNA loci. Estimates of allozyme genetic differentiation (θO ) ranged from 0.00 to 0.15, with a mean of 0.08. The θO values for scnDNA loci were similar to those of allozymes, ranging from 0.00 to 0.12 with a mean of 0.09. MtDNA RFLP divergence between oceans (θO = 0.39) was significantly greater than divergence detected at nuclear loci (95% nuclear confidence interval = 0.04-0.11). The fourfold smaller effective population size of mtDNA and male-mediated gene flow may account for the difference observed between nuclear and mitochondrial divergence estimates.

Microsatellite and mitochondrial DNA analyses of Atlantic bluefin tuna (Thunnus thynnus thynnus) population structure in the Mediterranean Sea.

Genetic variation was surveyed at nine microsatellite loci and the mitochondrial control region (868 bp) to test for the presence of genetic stock structure in young-of-the-year Atlantic bluefin tuna (Thunnus thynnus thynnus) from the Mediterranean Sea. Bluefin tuna were sampled over a period of 5 years from the Balearic and Tyrrhenian seas in the western basin of the Mediterranean Sea, and from the southern Ionian Sea in the eastern basin of the Mediterranean Sea. Analyses of multilocus microsatellite genotypes and mitochondrial control region sequences revealed no significant heterogeneity among collections taken from the same location in different years; however, significant spatial genetic heterogeneity was observed across all samples for both microsatellite markers and mitochondrial control region sequences (FST=0.0023, P=0.038 and PhiST=0.0233, P=0.000, respectively). Significant genetic differentiation between the Tyrrhenian and Ionian collections was found for both microsatellite and mitochondrial markers (FST=0.0087, P=0.015 and PhiST=0.0367, P=0.030, respectively). These results suggest the possibility of a genetically discrete population in the eastern basin of the Mediterranean Sea.