Blacktip shark Carcharhinus limbatus presence at fishing piers in South Carolina: association and environmental drivers.

We tagged 12 Carcharhinus limbatus with acoustic transmitters and monitored their presence at five piers along the north-east coast of South Carolina, USA in 2016 and four piers in 2017 using acoustic receivers. Data were analysed with pier association indices (PAI), mixed models and fast Fourier transformation analyses to identify potential factors related to residence time and presence at piers and any cyclical patterns in visits to piers. While the majority of monitored C. limbatus were infrequently detected at piers, three (25.0%) were highly associated with piers (PAI ≥ 0.50). Of the C. limbatus that were detected after initial capture, three (25.0%) recorded detection events only at the pier where they were tagged and two individuals (16.7%) recorded at least one detection event at all monitored piers. The best-fit model explaining C. limbatus residence time at piers included terms for pier location and diel cycle (wi = 0.88), whereas the best fit model explaining presence-absence of C. limbatus at piers included terms for tidal height, diel cycle, barometric pressure and angler count (wi = 0.98). Carcharhinus limbatus did not appear to display cyclical patterns in their visits to piers. Along the north-east coast of South Carolina, association of C. limbatus with piers is a phenomenon for a proportion of mature individuals, but continued research is necessary to understand if this behaviour is driven by attraction to and feeding on angler discards or increased foraging opportunities resulting from the attraction of potential prey to the physical structure provided by piers.

Blue crabs Callinectes sapidus as potential biological reservoirs for white spot syndrome virus (WSSV).

White spot syndrome virus (WSSV) is a virulent pathogen of cultured shrimp and was first detected in farms in South Carolina (USA) in 1997 and subsequently in wild shrimp in 1999. We screened groups of 1808 wild Atlantic white shrimp Litopenaeus setiferus and 300 blue crabs Callinectes sapidus collected from South Carolina, Georgia, and Florida for the presence of WSSV using the Shrimple® immunoassay-strip test, with all positives and random subsets of negatives tested by TaqMan real-time PCR and in infectivity bioassays. Of 87 shrimp and 11 crabs that tested positive using the Shrimple® test, only a single C. sapidus was confirmed to be infected with WSSV by PCR and the infectivity bioassay. The data indicate that the prevalence of WSSV in these species is low in these southeastern US regions, but that C. sapidus may serve as a biological reservoir.

Association behavior between sand tiger sharks and round scad is driven by mesopredators.

In marine systems, behaviorally-mediated indirect interactions between prey, mesopredators, and higher trophic-level, large predators are less commonly investigated than other ecologic interactions, likely because of inherent difficulties associated with making observations. Underwater videos (n = 216) from SharkCam, a camera installation sited beneath Frying Pan Tower, a decommissioned light house and platform, on a natural, hard bottom site approximately 50 km off Cape Fear, North Carolina, were used to investigate association behavior of round scad Decapterus punctatus around sand tiger sharks Carcharias taurus. Videos containing sand tiger sharks were analyzed for the simultaneous presence of round scad, and six species of scad mesopredators, with scad-shark interactions assigned to one of three categories of association: no visible interaction, loosely associated, or tightly associated. The likelihood of scad being loosely or tightly associated with sharks was significantly higher in the presence of scad mesopredators, suggesting that sharks provide a predation refuge for scad. This behaviorally-mediated indirect interaction has important implications for trophic energy transfer and mesopredator control on hard bottoms, as scad are one of the most abundant planktivorous fish on hard bottoms in the western Atlantic Ocean. Although we were not able to provide statistical evidence that sand tiger sharks also benefit from this association behavior, we have clear video evidence that round scad association conceals and attracts mesopredators, enhancing predation opportunities for sand tiger sharks. These interactions potentially yield additional trophic consequences to this unique association and highlight the value of exploring behaviorally-mediated interactions in marine communities.

Modelling interactions of acid-base balance and respiratory status in the toxicity of metal mixtures in the American oyster Crassostrea virginica.

Heavy metals, such as copper, zinc and cadmium, represent some of the most common and serious pollutants in coastal estuaries. In the present study, we used a combination of linear and artificial neural network (ANN) modelling to detect and explore interactions among low-dose mixtures of these heavy metals and their impacts on fundamental physiological processes in tissues of the Eastern oyster, Crassostrea virginica. Animals were exposed to Cd (0.001-0.400 microM), Zn (0.001-3.059 microM) or Cu (0.002-0.787 microM), either alone or in combination for 1 to 27 days. We measured indicators of acid-base balance (hemolymph pH and total CO(2)), gas exchange (Po(2)), immunocompetence (total hemocyte counts, numbers of invasive bacteria), antioxidant status (glutathione, GSH), oxidative damage (lipid peroxidation; LPx), and metal accumulation in the gill and the hepatopancreas. Linear analysis showed that oxidative membrane damage from tissue accumulation of environmental metals was correlated with impaired acid-base balance in oysters. ANN analysis revealed interactions of metals with hemolymph acid-base chemistry in predicting oxidative damage that were not evident from linear analyses. These results highlight the usefulness of machine learning approaches, such as ANNs, for improving our ability to recognize and understand the effects of sub-acute exposure to contaminant mixtures.

Time-course analysis of peroxinectin mRNA in the shrimp Litopenaeus vannamei after challenge with Vibrio campbellii.

Peroxinectin (Pox), which promotes cell adhesion and encapsulation of bacteria in crustaceans, is synthesized in granular and semigranular hemocytes. In this study, real-time PCR was used to quantify Pox transcripts in individual tissues of the Pacific white shrimp, Litopenaeus vannamei, over 48 h following injection of a sublethal dose of the shrimp pathogen Vibrio campbellii. The resulting data were used to infer the movements of hemocytes among the tissues in response to bacterial challenge. Over all times and treatments, Pox transcripts (ng total RNA)(-1) varied by orders of magnitude among individual tissues, such that circulating hemocytes >> gills >> heart lymphoid organ hepatopancreas approximately muscle. Relatively low constitutive expression of Pox in the lymphoid organ compared to circulating hemocytes, gills, and heart supports a primary role for this organ in bacteriostasis and degradation, rather than encapsulation of invasive bacteria. Numbers of Pox transcripts increased significantly at the injection site within 4 h and remained significantly elevated for 48 h, consistent with a rapid and sustained recruitment of hemocytes to the site of injection. Transcripts increased significantly in the gill but not in other tissues over the time-course of this experiment. These expression data reinforce the role of the gill in trapping and encapsulating invasive bacteria as a primary strategic focus during the early phase of the crustacean immune response and, by comparison with earlier studies of lysozyme expression in the same tissues, suggest differential roles for various tissues in a successful immune response.

Prevalence of the invasive Rhizocephalan parasite Loxothylacus panopaei in Eurypanopeus depressus in South Carolina and genetic relationships of the parasite in North and South Carolina.

The rhizocephalan barnacle Loxothylacus panopaei is a parasitic castrator of xanthid crabs that has invaded the U.S. Atlantic coast. It was transported to the Chesapeake Bay in the mid-1960s with mud crabs associated with Gulf coast oysters and has since spread north to Long Island Sound, New York, and south to Cape Canaveral, Florida. Here we report parasite prevalence at 3 South Carolina sites--2 from which the parasite had not been previously reported--and examine the genetic relationships of North and South Carolina L. panopaei populations relative to Gulf of Mexico and other Atlantic coast parasite populations. Total L. panopaei prevalence was 24.2% among all 3 sites, with monthly prevalence as high as 51.6% at Waties Island, South Carolina. Sequence analyses of North and South Carolina specimens revealed the presence of 4 cytochrome c oxidase subunit I haplotypes--3 commonly found in other invasive populations and 1 new haplotype found in a single specimen from the Rachel Carson Reserve in Carteret County, North Carolina--and indicate that the Carolina populations are a result of range expansion from the original Atlantic coast invasion.

Lysozyme gene expression by hemocytes of Pacific white shrimp, Litopenaeus vannamei, after injection with Vibrio.

The purpose of this study was to quantify the gene expression of lysozyme, an important antibacterial protein produced by shrimp hemocytes, within tissues of Litopenaeus vannamei Boone in response to a pathogen challenge. We quantified lysozyme transcripts with a real-time PCR method and used these data, along with total hemocyte counts, to infer patterns of hemocyte trafficking during the immune response. Transcript expression was detected by in situ hybridization of mRNA in circulating hemocytes, and within tissues with high hemocyte concentrations. Lysozyme gene expression was monitored in 5 tissues and in circulating hemocytes for 48 h following challenge with the shrimp pathogen Vibrio campbellii Baumann. The results suggest that lysozyme is expressed in most if not all hemocytes in circulation and in peripheral tissues. Injection with V. campbellii produced a significant decrease in transcriptional signal in circulating hemocytes and peripheral tissues 4 h after injection. Over the same early time period lysozyme signal increased significantly in the muscle at the site of injection and remained high for the duration of the time-course, suggesting that hemocytes are recruited to the site of injection early during the course of the immune response. After 4 h, lysozyme signal increased in circulating hemocytes and tissues, with a return to control levels noted for all tissues except the muscle at the site of injection.

In vitro response of the striped bass natural resistance-associated macrophage protein, Nramp, to LPS and Mycobacterium marinum exposure.

Mycobacteriosis in Chesapeake Bay (USA) striped bass Morone saxatilis is an ongoing disease problem with important economic implications for a large commercial and recreational fishery. Additionally, striped bass serve as a reservoir of potential mycobacterial zoonoses. Recently, we described a striped bass gene homolog of the natural resistance-associated macrophage protein family (MsNramp), which is responsible for resistance to mycobacterial infections in mice. Striped bass MsNramp is strongly induced in peritoneal exudate cells (PE) in vivo after intraperitoneal injection with Mycobacterium spp. The purpose of the present study was to investigate short-term in vitro MsNramp expression and reactive oxygen intermediate (ROI) production in primary cultures of adherent PE after exposure to bacterial lipopolysaccharide (LPS), or live- or heat-killed (HK) Mycobacterium marinum. PE expressed significantly higher levels of MsNramp at 4 and 24 h post-treatment with live and HK M. marinum. MsNramp response to LPS was dose-dependent in these cells, with maximum expression at 4 h and 20 microg/ml LPS. Treatment of PE with LPS resulted in increased intracellular superoxide anion levels, whereas treatment with live M. marinum caused a significant depression. This study is the first report of induction of a teleost Nramp in vitro by mycobacteria, and supports findings of teleost Nramp induction by LPS.

Mycobacterium-inducible Nramp in striped bass (Morone saxatilis).

In mammals, the natural resistance-associated macrophage protein 1 gene, Nramp1, plays a major role in resistance to mycobacterial infections. Chesapeake Bay striped bass (Morone saxatilis) is currently experiencing an epizootic of mycobacteriosis that threatens the health of this ecologically and economically important species. In the present study, we characterized an Nramp gene in this species and obtained evidence that there is induction following Mycobacterium exposure. The striped bass Nramp gene (MsNramp) and a 554-amino-acid sequence contain all the signal features of the Nramp family, including a topology of 12 transmembrane domains (TM), the transport protein-specific binding-protein-dependent transport system inner membrane component signature, three N-linked glycosylation sites between TM 7 and TM 8, sites of casein kinase and protein kinase C phosphorylation in the amino and carboxy termini, and a tyrosine kinase phosphorylation site between TM 6 and TM 7. Phylogenetic analysis most closely grouped MsNramp with other teleost Nramp genes and revealed high sequence similarity with mammalian Nramp2. MsNramp expression was present in all tissues assayed by reverse transcription-PCR. Within 1 day of injection of Mycobacterium marinum, MsNramp expression was highly induced (17-fold higher) in peritoneal exudate (PE) cells compared to the expression in controls. The levels of MsNramp were three- and sixfold higher on days 3 and 15, respectively. Injection of Mycobacterium shottsii resulted in two-, five-, and threefold increases in gene expression in PE cells over the time course. This report is the first report of induction of an Nramp gene by mycobacteria in a poikilothermic vertebrate.