Vibrio vulnificus and Vibrio parahaemolyticus in Oysters under Low Tidal Range Conditions: Is Seawater Analysis Useful for Risk Assessment?

Human-pathogenic Vibrio bacteria are acquired by oysters through filtering seawater, however, the relationships between levels of these bacteria in measured in oysters and overlying waters are inconsistent across regions. The reasons for these discrepancies are unclear hindering our ability to assess if -or when- seawater samples can be used as a proxy for oysters to assess risk. We investigated whether concentrations of total and human pathogenic Vibrio vulnificus (vvhA and pilF genes) and Vibrio parahaemolyticus (tlh, tdh and trh genes) measured in seawater reflect concentrations of these bacteria in oysters (Crassostrea virginica) cultured within the US lower Chesapeake Bay region. We measured Vibrio spp. concentrations using an MPN-qPCR approach and analyzed the data using structural equation modeling (SEM). We found seawater concentrations of these bacteria to predictably respond to temperature and salinity over chlorophyll a, pheophytin or turbidity. We also inferred from the SEM results that Vibrio concentrations in seawater strongly predict their respective concentrations in oysters. We hypothesize that such seawater-oyster coupling can be observed in regions of low tidal range. Due to the ease of sampling and processing of seawater samples compared to oyster samples, we suggest that under low tidal range conditions, seawater samples can foster increased spatial and temporal coverage and complement data associated with oyster samples

Influence of oyster genetic background on levels of human-pathogenic Vibrio spp.

Human-pathogenic Vibrio bacteria are common inhabitants of oyster tissues, but our understanding of factors driving the wide range of concentrations found in individual oysters is extremely limited. We examined the influence of oyster sex and parasitism in light of their profound effects on oyster tissues against a backdrop of eastern oysters, Crassostrea virginica, from two diploid and two triploid aquacultured lines. This allowed us to examine not only the effect of oyster ploidy but also of oyster genetics, a factor never investigated with regard to human-pathogenic Vibrio species. We measured levels of total Vibrio vulnificus (vvhA), and of total (tlh) and pathogenic (tdh+, trh+) V. parahaemolyticus, in each oyster, and analyzed the data through generalized linear mixed-effects models. A key outcome of these analyses was the consistent inclusion of oyster line as a predictor variable across Vibrio targets. A potential effect of Perkinsus marinus infections and/or oyster sex was also suggested, although the combination of variables varied with Vibrio target. This study suggests that the influence of oyster genetic background should be further investigated, and that the dynamics of human-pathogenic Vibrio spp. in oysters is likely driven by multiple, interacting factors, some of which may be under oyster host genetic control

High Salinity Relay as a Postharvest Processing Strategy To Reduce Vibrio vulnificus Levels in Chesapeake Bay Oysters (Crassostrea virginica)

In 2009 the U.S. Food and Drug Administration (FDA) announced its intention to implement postharvest processing (PHP) methods to eliminate Vibrio vulnificus from oysters intended for the raw, half-shell market that are harvested from the Gulf of Mexico during warmer months. FDA-approved PHP methods can be expensive and may be associated with unfavorable responses from some consumers. A relatively unexplored PHP method that uses relaying to high salinity waters could be an alternative strategy, considering that high salinities appear to negatively affect the survival of V. vulnificus. During relay, however, oysters may be exposed to rapid and large salinity increases that could cause increased mortality. In this study, the effectiveness of high salinity relay to reduce V. vulnificus to (MPN) per g and the impact on oyster mortality were assessed in the lower Chesapeake Bay. Two relay experiments were performed during the summer and fall of 2010. Oysters collected from three grow-out sites, a low salinity site (14 to 15 practical salinity units [psu]) and two moderate salinity sites (22 to 25 psu), were relayed directly to a high salinity site (≥30 psu) on Virginia\u27s Eastern Shore. Oysters were assayed for V. vulnificus and Vibrio parahaemolyticus (another Vibrio species of concern) densities at time 0 prior to relay and after 7 and 14 days of relay, using the FDA MPN enrichment method combined with detection by real-time PCR. After 14 days, both V. vulnificus and V. parahaemolyticus densities were ≤0.8 MPN/g, and decreases of 2 to 3 log in V. vulnificus densities were observed. Oyster mortalities were low (

Landscape-Level Variation in Disease Susceptibility Related to Shallow-Water Hypoxia

Diel-cycling hypoxia is widespread in shallow portions of estuaries and lagoons, especially in systems with high nutrient loads resulting from human activities. Far less is known about the effects of this form of hypoxia than deeper-water seasonal or persistent low dissolved oxygen. We examined field patterns of diel-cycling hypoxia and used field and laboratory experiments to test its effects on acquisition and progression of Perkinsus marinus infections in the eastern oyster, Crassostrea virginica, as well as on oyster growth and filtration. P. marinus infections cause the disease known as Dermo, have been responsible for declines in oyster populations, and have limited success of oyster restoration efforts. The severity of diel-cycling hypoxia varied among shallow monitored sites in Chesapeake Bay, and average daily minimum dissolved oxygen was positively correlated with average daily minimum pH. In both field and laboratory experiments, diel-cycling hypoxia increased acquisition and progression of infections, with stronger results found for younger (1-year-old) than older (2-3-year-old) oysters, and more pronounced effects on both infections and growth found in the field than in the laboratory. Filtration by oysters was reduced during brief periods of exposure to severe hypoxia. This should have reduced exposure to waterborne P. marinus, and contributed to the negative relationship found between hypoxia frequency and oyster growth. Negative effects of hypoxia on the host immune response is, therefore, the likely mechanism leading to elevated infections in oysters exposed to hypoxia relative to control treatments. Because there is considerable spatial variation in the frequency and severity of hypoxia, diel-cycling hypoxia may contribute to landscape-level spatial variation in disease dynamics within and among estuarine systems

Genetic Population Structure of US Atlantic Coastal Striped Bass (Morone saxatilis)

Genetic population structure of anadromous striped bass along the US Atlantic coast was analyzed using 14 neutral nuclear DNA microsatellites. Young-of-the-year and adult striped bass (n = 1114) were sampled from Hudson River, Delaware River, Chesapeake Bay, North Carolina, and South Carolina. Analyses indicated clear population structure with significant genetic differentiation between all regions. Global multilocus F-ST was estimated at 0.028 (P \u3c 0.001). Population structure followed an isolation-by-distance model and temporal sampling indicated a stable population structure more than 2 years at all locations. Significant structure was absent within Hudson River, whereas weak but significant genetic differences were observed between northern and southern samples in Chesapeake Bay. The largest and smallest effective striped bass population sizes were found in Chesapeake Bay and South Carolina, respectively. Coalescence analysis indicated that the highest historical gene flow has been between Chesapeake Bay and Hudson River populations, and that exchange has not been unidirectional. Bayesian analysis of contemporary migration indicated that Chesapeake Bay serves as a major source of migrants for Atlantic coastal regions from Albemarle Sound northward. In addition to examining population genetic structure, the data acquired during this project were capable of serving as a baseline for assigning fish with unknown origin to source region

Implication d'hôtes bentho-pélagiques dans le cycle de Marteilia refringens parasite de l'huître plate Ostrea edulis: mise en place d'outils de détection et de validation de sa présence chez des hôtes potentiels

L'huître plate, Ostrea edulis a subit depuis les années 1970 deux graves épizooties provoquées par les parasites protozoaires, Bonamia ostreae et Marteilia refringens. Si, pour le premier de ces parasites, des huîtres plates tolérantes ont pu être sélectionnées, la marteiliose ne peut être contrôlée que par une gestion du risque de contamination. Dans ce cadre, la connaissance du cycle parasitaire de Marteilia refringens est primordiale. Les travaux réalisés à ce sujet ont permis de poser l'hypothèse d'un cycle hétéroxène. Au cours de notre étude la recherche du parasite a été effectuée sur des espèces du zooplancton, de la méiofaune et de la macrofaune prélevées au sein d'un biotope sélectif, les claires ostréicoles. Le parasite a été recherché par la technique de polyméristion en chaîne (PCR) au sein des différentes espèces de ces groupes. La détection en PCR du parasite au sein d'une espèce de la macrofaune Cereus pedunculatus (Cnidaires, Actiniaires) a été confirmée par différentes techniques (Southern blot, séquençage, hybridation in situ et hi stologie). Cette espèce pourrait constituer un hôte potentiel de Marteilia refringens. Cette étude ouvre de nouvelles perspectives pour l'étude du cycle parasitaire de Marteilia refringens

Infection dynamics of Marteilia refringens in flat oyster Ostrea edulis and copepod Paracartia grani in a claire pond of Marennes-Oleron Bay

The protozoan parasite Marteilia refringens has been partly responsible for the severe decrease in the production of the European flat oyster Ostrea edulis Linnaeus in France since the 1970s. The calanoid copepod Paracartia grani Sars was recently found to be a host for M refringens in French shallow-water oyster ponds (\u27claires\u27). This study reconsidered M refringens transmission dynamics in the light of this finding, taking into account not only oyster infection dynamics and environmental factors but also data concerning the copepod host. P. grani population dynamics in the claire under study revealed that this species is the dominant planktonic copepod in this confined ecosystem. During winter, M refringens overwintered in O. edulis, with P. grani existing only as resting eggs in the sediment. The increase in temperature in spring controlled and synchronized both the release of M refringens sporangia in the oyster feces, and the hatching of the benthic resting eggs of the copepod. Infection of oysters by M refringens was limited to June, July and August, coinciding with (1) the highest temperature recorded in the claire, and (2) the highest abundance of P. grani. PCR detection of M refringens in P. grani during the summer period was linked to the release of parasite sporangia by the oyster. Our results are supported by previous results on the effective transmission of this parasite from the oyster to the copepod

Influence of Gonadal Stage and Ploidy on Human-Pathogenic Vibrio Levels in the Oyster Crassostrea virginica - Final Report

To determine whether oyster reproductive status and particularly ploidy influences levels of human-pathogenic Vibrio spp. in oyster tissues was the overall aim of this project Our project objectives were three-fold: 1) To determine the relationship between oyster gonadal stage and levels of humanpathogenic Vibrio spp. in two separate pairs of genetically related diploid and triploid oysters through analyses of individual oysters conducted during peak Vibrio season in the York River, Virginia; 2) To determine the relationship between oyster ploidy and levels of humanpathogenic Vibrio spp. in those same oysters; and 3) To determine the degree to which farm-level marine aquaculture practices change based on results of this project.

Does the slipper limpet (Crepidula fornicata, L.) impair oyster growth and zoobenthos biodiversity? A revisited hypothesis

International audienceThe Prosobranch Gastropod Crepidula fornicata was introduced into Great Britain at the end of the 19th century from North America, upon imported oysters Crassostrea virginica. Since then, it has invaded sheltered coasts of the North West Atlantic and Mediterranean Sea. C. fornicata proliferation has often generated social conflicts due to three main causes: (1) trophic competition with other suspension feeders, e.g. the cultivated oyster Crassostrea gigas; (2) spatial competition with macrozoobenthos; and (3) enhancement of silt and clay sedimentation. The effects of C. fornicata on C. gigas growth and on macrozoobenthic density and diversity have been documented through field experiments in an oyster park of Arcachon Bay (France). Densities of C. gigas and biomass of C. fornicata were manipulated over a period of 247 days within field enclosures at low water level to test: (1) oyster growth, condition index and mortality; (2) zoobenthic community alterations (abundance, biomass, species richness). From this small-scale experiment, it was shown that none of these investigated parameters was significantly affected by the presence of C. fornicata. However, faunal assemblages were modified in enclosures compared to external bare sands, due to adding a hard substrata over a soft sediment

Real-Time PCR for Detection and Quantification of the Protistan Parasite Perkinsus marinus in Environmental Waters

The protistan parasite Perkinsus marinus is a severe pathogen of the oyster Crassostrea virginica along the east coast of the United States. Very few data have been collected, however, on the abundance of the parasite in environmental waters, limiting our understanding of P. marinus transmission dynamics. Real-time PCR assays with SybrGreen I as a label for detection were developed in this study for quantification of P. marinus in environmental waters with P. marinus species-specific primers and of Perkinsus spp. with Perkinsus genus-specific primers. Detection of DNA concentrations as low as the equivalent of 3.3 × 10(−2) cell per 10-μl reaction mixture was obtained by targeting the multicopy internal transcribed spacer region of the genome. To obtain reliable target quantification from environmental water samples, removal of PCR inhibitors and efficient DNA recovery were two major concerns. A DNA extraction kit designed for tissues and another designed for stool samples were tested on environmental and artificial seawater (ASW) samples spiked with P. marinus cultured cells. The stool kit was significantly more efficient than the tissue kit at removing inhibitors from environmental water samples. With the stool kit, no significant difference in the quantified target concentrations was observed between the environmental and ASW samples. However, with the spiked ASW samples, the tissue kit demonstrated more efficient DNA recovery. Finally, by performing three elutions of DNA from the spin columns, which were combined prior to target quantification, variability of DNA recovery from different samples was minimized and more reliable real-time PCR quantification was accomplished