The expansion of molluscan aquaculture in the United States has induced a rising demand for molluscan seed to sustain it. Out-of-state transfers of seed from hatcheries and nurseries are considered a pathogen risk to both aquacultured and wild animals. Another obstacle to the maintenance of the industry is the small number of laboratories able to conduct molluscan health certification. We hypothesized that larger animals would have higher pathogen prevalences, as they proportionally filter more water. To investigate this, we analyzed health data of three pathogens: Perkinsus spp. (dermo disease) in eastern oysters (Crassostrea virginica), Mucochytrium quahogii (QPX disease) in (Mercenaria mercenaria), and Haplosporidium nelsoni (MSX disease) in both oysters and clams. Analyses included 1309 samples containing over 75,000 animals, which were categorized by life stage (gametes, larvae, seed, juveniles and adults), as well as the type of water treatment in which animals were reared. We found higher dermo prevalences (> 50 %) only in larger oysters (> 40 mm) that were reared in raw water. Hard clams did not present high prevalences of either dermo or QPX in any life stage, nor did MSX in oysters. There was a marked difference in prevalences between raw and treated water for all three pathogens, highlighting the importance of biosafety practices to ensure both product quality and reliable hatchery operations.

Disease freedom surveillance is challenging in settings where environmental conditions and population health can change abruptly. We propose a novel approach to freedom evaluation that is specifically suited to open environments. The approach merges information about pathogen introduction risk with information from pathogen surveillance to estimate assurance of freedom. While the logic is not new, the approach uses a generalizable model designed by an expert panel to estimate introduction probability, easing the need for costly, site-specific risk assessment. This introduction probability is then used to inform the time-adjusted value of historical surveillance data and adapt the consequent requirements (sampling volume and frequency) of future surveys. We apply the approach to an open water region on the Pacific Coast of the United States known for its mollusk production. Results support absence of the target pathogens-Marteilia refringens, Marteilioides chungmuensis, Ostreid herpesvirus 1, Perkinsus marinus, and Perkinsus olseni-for the host species, region, and time-period under investigation. Findings demonstrate (1) the ability to retain assurance in pathogen freedom through time in open systems, (2) the limitations of sample volume, and importance of sampling frequency, for pathogens with higher introduction risk, and (3) expert elicitation as a generalizable alternative to formal risk assessment.

Bonamia (Haplosporida) are oyster parasites capable of devastating oyster populations. The near-circumglobal distribution of the host generalist B. exitiosa has previously been associated with the natural and anthropogenic dispersal of broadly distributed non-commercial oysters in the Ostrea stentina species complex. Here, we took a global snapshot approach to explore the role of the widely introduced Pacific oyster Magallana gigas, a commercially important species that can be found on every continent except Antarctica, in transporting Bonamia. We screened 938 M. gigas individuals from 41 populations in this oyster’s native and non-native geographic range for presence of Bonamia DNA using PCR. B. exitiosa was the only species detected and only within 2 of 5 populations from southern California, USA (10 and 42% PCR prevalence). Therefore, M. gigas could have played a role in transporting B. exitiosa to California (if introduced) and/or maintaining B. exitiosa populations within California, but morphological confirmation of infection needs to be done to better understand the host-parasite dynamics within this system. We detected no Bonamia DNA within any other non-native M. gigas populations (n = 302) nor within native M. gigas populations in Japan and Korea (n = 582) and thus found no evidence to support the co-dispersal of M. gigas and other Bonamia species. Lower sample sizes within some populations and the non-systematic nature of our sampling design may have led to false negatives, especially in areas where Bonamia are known to occur. Nevertheless, this global snapshot provides preliminary guidance for managing both natural and farmed oyster populations.

Shellfish fisheries and aquaculture within the Chesapeake Bay (hereafter 'the Bay') and its tributaries have been historically impacted by disease and climate events. Climate-driven shifts in temperature and salinity can alter host-parasite dynamics, influencing outbreaks. Here, we explore the relationship between temperature, salinity and parasite distribution and abundance in the eastern oyster Crassostrea virginica-Perkinsus marinus system. We use long-term (30 yr) environmental data and P. marinus surveys in the Bay to identify (1) how climate affects P. marinus prevalence and intensity, (2) seasonal and climate-driven infection patterns, and (3) regional environmental influences on disease. We found significant relationships between P. marinus infection intensity, prevalence, increasing temperature and decreasing salinity. Our results indicated that there is an overall decreased abundance of P. marinus prevalence and intensity throughout the Bay driven by decreases in salinity over time, most prominently from 2003-2020. However, these temporal trends in prevalence and intensity vary largely by region, with some regions still experiencing high disease burden. Examining monthly environmental parameters reinforced the dominant role of salinity in driving disease patterns. Salinity had significant relationships with prevalence and intensity year-round, with the largest effects in late spring/early summer. Monthly temperatures had fewer significant relationships to prevalence and intensity, but the largest significant effects were seen in late winter/early spring. Notably, this study is the first to document that winter salinity influences fall parasite prevalence, sometimes exerting a greater effect than temperature. Continued and expanded monitoring of marine disease is crucial to understand how the changing climate is impacting disease.

The massive geographic expansion of terrestrial plant crops, livestock, and marine aquacultured species during the 19th and 20th centuries provided local economic benefits, stabilized food demands, and altered local ecosystems. The invasion history of these translocations remains uncertain for most species, limiting our understanding of their future adaptive potential and historical roles as vectors for coinvaded species. We provide a framework for filling this gap in invasion biology using the widely transplanted Pacific oyster as a case study. A two-dimensional summary of population-level variation in single nucleotide polymorphisms in native Japan reflected the geographical map of Japan and allowed identification of the source regions for the worldwide expansion. Pacific oysters proliferate in nonnative areas with environmental temperatures similar to those areas where native lineages evolved. Using Approximate Bayesian Computation, we ranked the likelihood of historical oyster or shipping vectors to explain current-day distribution of genotypes in 14 coinvaded algal and animal species. Oyster transplants were a more likely vector than shipping for six species, shipping activity was more likely for five species, and a vector was ambiguous for three species. Applying this approach to other translocated species should reveal similar legacy effects, especially for economically important foundation species that also served as vectors for nonnative species.

OBJECTIVE:The dinoflagellate Alexandrium monilatum forms blooms during summer in tributaries of the lower Chesapeake Bay. Questions persist about the potential for A. monilatum to negatively affect aquatic organisms. Its main toxin, goniodomin A (GDA), a polyketide macrolide, has been shown to have adverse effects on animals, for example through cytotoxicity and interaction with actin. METHODS:Eastern oysters Crassostrea virginica were exposed for 96 h to sublethal concentrations of A. monilatum (615 ± 47 cells/mL [average ± SD]; containing mainly intracellular GDA [215 ± 7.15 pg/cell] and to a lesser extent goniodomin B, goniodomin C, and GDA seco-acid as quantified by liquid chromatography coupled to tandem mass spectrometry) or to nontoxic phytoplankton or were unexposed. They were subsequently depurated for 96 h by exposure to nontoxic phytoplankton. Clearance rates were estimated, and oysters were sampled daily and tissue (gill, digestive gland, and remaining tissues) excised for analyses by histopathology, gene expression quantified by quantitative PCR, and goniodomin quantification. RESULT:A positive clearance rate, no mortality, and no tissue pathologies were observed in oysters exposed to A. monilatum. Goniodomin A was detected in gill 6 h after exposure (504 ± 329 μg/kg [average ± SE]) and to a lesser extent in the digestive gland and remaining soft tissues. In the digestive gland, a trend of transformation of GDA to GDA seco-acid was observed. The majority of toxins (≥83%) were depurated after 96 h. Expression of genes involved in oxidative response increased 14-fold after 6 h, and those involved in actin synthesis showed a 27-fold change after 24 h, while expression of apoptosis genes increased 6.9-fold after 96 h compared with the control (eastern oysters exposed to nontoxic phytoplankton). CONCLUSION:Exposure experiments (nonsublethal or chronic) should be carried out to better assess the threat of this species and toxins for eastern oysters and other marine organisms.

Co-infecting parasites modify infection outcomes in the wild. However, it is unclear how multiple environmental factors influence co-infection. The Chesapeake Bay metapopulation of the eastern oyster, Crassostrea virginica, provides an opportunity to test the importance of co-infection across heterogeneous environments because multiple parasites infect oysters across a broad salinity gradient. This study leverages Maryland and Virginia oyster monitoring for a large-scale survey of four co-infecting organisms, including two tissue parasites and two shell bio-eroding parasites. We diagnosed infection in 440 oysters across 16 paired harvested and unharvested reefs and tested the importance of co-infecting organisms for each parasite relative to environmental conditions, host traits, and marine spatial management. Microscopic visual methods were used to diagnose prevalence and intensity of tissue infections with Perkinsus marinus (the causative agent of dermo disease) and Haplosporidium nelsoni (the causative agent of MSX disease). Macroscopic visual methods were used to diagnose prevalence and intensity of shell infections with Cliona boring sponges and blister-inducing Polydora worms. For the three oyster parasites that were detected [H. nelsoni infections were absent in all oysters], salinity was the overall strongest predictor, corresponding to bay-wide patterns of parasite prevalence and/or intensity. Despite high environmental and spatial variation, co-infections corresponded to altered prevalence and/or intensity for all three oyster parasites. The correlational patterns suggest that P. marinus acts as a lynchpin in co-infection, as its intensity increased with Cliona sponge prevalence and P. marinus co-infection predicted higher Polydora blister intensity. Oyster shell height, reef habitat, and harvest status also predicted parasite prevalence and intensity, further reflecting the multivariate drivers of infections in this system. Unharvested reefs had greater vertical habitat structure and higher intensities of Cliona sponge infections, but no differences in the prevalence of any of the three parasites. Spatial patterns unexpectedly show that reef-level predictors of parasite patterns were more important than differences between tributaries. This correlational survey provides novel insights through the statistical relationships between the three oyster parasites, environmental conditions, host traits, and human resource management. New and more detailed scenarios are needed to expand disease ecological theory to encompass co-infection in anthropogenically impacted wildlife populations.

With the increasing affordability of next-generation sequencing technologies, genotype-by-sequencing has become a cost-effective tool for ecologists and conservation biologists to describe a species' evolutionary history. For host–parasite interactions, genotype-by-sequencing can allow the simultaneous examination of host and parasite genomes and can yield insight into co-evolutionary processes. The eastern oyster, Crassostrea virginica, is among the most important aquacultured species in the United States. Natural and farmed oyster populations can be heavily impacted by ‘dermo’ disease caused by an alveolate protist, Perkinsus marinus. Here, we used restricted site-associated DNA sequencing (RADseq) to simultaneously examine spatial population genetic structure of host and parasite. We analysed 393 single-nucleotide polymorphisms (SNPs) for P. marinus and 52,100 SNPs for C. virginica from 36 individual oysters from the Gulf of Mexico (GOM) and mid-Atlantic coastline. All analyses revealed statistically significant genetic differentiation between the GOM and mid-Atlantic coast populations for both C. virginica and P. marinus, and genetic divergence between Chesapeake Bay and the outer coast of Virginia for C. virginica, but not for P. marinus. A co-phylogenetic analysis confirmed significant coupled evolutionary change between host and parasite across large spatial scales. The strong genetic divergence between marine basins raises the possibility that oysters from either basin would not be well adapted to parasite genotypes and phenotypes from the other, which would argue for caution with regard to both oyster and parasite transfers between the Atlantic and GOM regions. More broadly, our results demonstrate the potential of RADseq to describe spatial patterns of genetic divergence consistent with coupled evolution.

As the start of the supply chain for the aquaculture industry, hatcheries are a crucial component in the success of oyster and northern quahog (hard clam) aquaculture on the East Coast of the US. Intermittent failures in hatchery production slow industry growth and reduce profits. To begin investigations into the possible role of algal toxins in hatchery production failure, post-treatment hatchery water from one research and four commercial hatcheries in lower Chesapeake Bay, USA, was sampled for (1) toxin presence and (2) harmful algal bloom (HAB) cell enumeration. Overall, seven toxin classes, likely produced by six different HAB species, were detected in posttreatment hatchery water, despite a lack of visually identifiable HAB cells within the facility. Toxins detected include pectenotoxin-2, goniodomin A, karlotoxin-1 and karlotoxin-3, okadaic acid and dinophysistoxin-1, azaspiracid-1 and azaspiracid-2, brevetoxin-2, and microcystin-LR. In a second, more targeted study, two batches of source water were followed and sampled at each step of a water-treatment process in the VIMS Aquaculture Genetics and Breeding Technology Center research hatchery in Gloucester Point, Virginia, USA. Two treatment steps showed particular promise for decreasing the concentrations of the three toxins detected in the source water, 24-h circulation through sand filters and activated charcoal filtration. Toxin concentrations of pectenotoxin-2, 3.53 ?? 0.56 pg mL-1, okadaic acid, 6.14 ?? 0.69 pg mL-1, and dinophysistoxin-1, 1.88 ?? 0.0 pg mL-1, were low in the source water. The sand filtration step decreased these concentrations by 49???62%. Activated charcoal filtration subsequently brought the concentrations down to <0.5 pg mL-1, successfully removing another 87???99% of toxins from incoming water. With toxin breakthrough now documented in commercial hatchery facilities during non-bloom conditions, future studies are needed to investigate breakthrough and water-treatment options during more-intense bloom conditions, as well as the potential interactions of algal toxins with other stressors in a potentially multifactorial etiology underlying hatchery production failures.

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.

Bivalve mollusks including oysters have low metabolic potential and are therefore susceptible to accumulating high levels of lipophilic organic contaminants such as polycyclic aromatic hydrocarbons (PAHs). Human exposure to PAHs via consumption of this important commercial shellfish can be a serious public health concern in areas where high PAH contamination exists. Previous PAH immunohistochemical studies have been limited to laboratory‐based exposures focusing on one or a few individual PAH compounds. To date, such studies have yet to explore PAH accumulation in oysters, known to have some of the highest levels of PAHs across different food products. Using a monoclonal antibody selective for a range of three‐ to five‐ring PAHs, we present a method to detect and localize complex mixtures of PAHs in oyster tissues via fluorescent immunohistochemistry. Observed immunofluorescence intensity followed a similar trend as measured levels of PAHs in oyster interstitial fluid from PAH‐contaminated sites and oysters exposed to the water accommodated fraction of crude oil. This method will be valuable in understanding internal partitioning mechanisms of PAH‐exposed oysters and will have important applications in studies on PAH distribution in the tissues of additional organisms for environmental, medical, or veterinary purposes. Environ Toxicol Chem 2023;42:475–480. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Zoothamnium intermedium is an obligate epibiont ciliate and has been found in a diverse array of hosts and environments. Different studies have reported conflicting distribution patterns and host preferences, even though studies in Chesapeake Bay have suggested that the ciliate has a strong host specificity for two calanoid copepod species. We examined the life cycle, host preferences, and ecological conditions conducive to Z. intermedium presence on copepods in Chesapeake Bay, the largest estuary in North America. The York River tributary was sampled biweekly from fall 2014 through summer 2015 for plankton, peritrichs and bacteria in the water column. Bacterial abundance in the water column peaked in fall and late spring, coinciding with increased abundance and species richness of non-epibiont peritrichs. Among the plankton, only the calanoid copepods Acartia tonsa and Centropages hamatus were colonized by Z. intermedium. The peritrich epibiont displayed higher colonization rates on C. hamatus even when A. tonsa was far more abundant. Multivariate correlation analysis of infestation prevalence on A. tonsa showed a strong correlation with dissolved oxygen, salinity and water temperature. Such correlations, along with differences in host species biology, might be driving the seasonality of this epibiotic relationship.

AIMS:To investigate the relationships between individual health status of oysters, particularly with regard to parasitic infection, and variability in abundance of human-pathogenic Vibrio species.METHODS AND RESULTS:Aquacultured eastern oysters, Crassostrea virginica, were analysed individually for infection by the protozoan parasite Perkinsus marinus through quantitative PCR, and total Vibrio vulnificus and total and pathogenic Vibrio parahaemolyticus abundance was assessed using a most probable number (MPN)-qPCR approach. Additionally, perspective on general oyster health and other parasitic infections was obtained through histopathology. Perkinsus marinus infection and human-pathogenic Vibrio species levels were not correlated, but through histology, analyses revealed that oysters infected by Haplosporidium nelsoni harboured more V. vulnificus.CONCLUSIONS:The highly prevalent parasite P. marinus had little influence on human-pathogenic Vibrio species levels in eastern oysters, but the less prevalent parasite, H. nelsoni, may influence V. vulnificus levels, highlighting the potential nuances of within-oyster dynamics of Vibrio species.SIGNIFICANCE AND IMPACT OF THE STUDY:Human-pathogenic bacteria continue to be a concern to the oyster industry and causes for individual oyster variation in bacterial levels remain unknown. The major oyster pathogen P. marinus does not appear to affect levels of these bacteria within oysters, suggesting that other factors may influence Vibrio spp. levels in oysters.