Gold Nanoparticle Paper Immunoassays for Sensing the Presence of Vibrio parahaemolyticus in Oyster Hemolymph.

Seafood contamination with Vibrio bacteria is a problem for aquaculture, especially with oysters, which are often consumed raw. Current methods for diagnosing bacterial pathogens in seafood involve lab-based assays such as polymerase chain reaction or culturing, which are time consuming and must occur in a centralized location. Detection of Vibrio in a point-of-care assay would be a significant tool for food safety control measures. We report here a paper immunoassay that can detect the presence of Vibrio parahaemolyticus (Vp) in buffer and oyster hemolymph. The test uses gold nanoparticles conjugated to polyclonal anti-Vibrio antibodies in a paper-based sandwich immunoassay. A sample is added to the strip and wicked through by capillary action. If Vp is present, it results in a visible color at the test area that can be read out by eyes or a standard mobile phone camera. The assay has a limit of detection of 6.05 × 105 cfu/mL and a cost estimate of $5 per test. Receiver operating characteristic curves with validated environmental samples showed a test sensitivity of 0.96 and a specificity of 1.00. Because the assay is inexpensive and can be used on Vp directly without the requirement for culturing, or sophisticated equipment, it has the potential to be used in fieldable settings.

Foreign stingers: South American freshwater river stingrays Potamotrygon spp. established in Indonesia.

The pet trade is known to be one of the most important pathways of aquatic non-native species introduction and Indonesia is a significant trade partner. Popular ornamental South American river stingrays (Potamotrygon spp.) were introduced to Indonesia in the 1980s and the culture was established. Here we present a detailed Indonesian market and aquaculture survey, the volume of trade between January 2020 and June 2022, and the list of customer countries with the total amount of imported stingrays. Climate similarities between the native range of P. motoro and P. jabuti and Indonesia were analysed. A significant number of areas of Indonesian islands were identified as suitable for the establishment of this species. This was confirmed by the first record of probably established populations in the Brantas River (Java). In total 13 individuals including newborns were captured. The culture of potamotrygonid stingrays is unregulated in Indonesia, and the risk of the establishment of this predator and its potential spread is alarming for wildlife. Moreover, the first case of envenomation caused by Potamotrygon spp. in the wild outside of South America was recorded. The current condition is predicted as the 'tip of the iceberg' and continuous monitoring and mitigation of risks are strongly recommended.

The determination of thiocyanate in the blood plasma and holding water of Amphiprion clarkii after exposure to cyanide.

The illegal practice of cyanide fishing continues throughout the Indo-Pacific. To combat this destructive fishing method, a reliable test to detect whether a fish has been captured using cyanide (CN) is needed. We report on the toxicokinetics of acute, pulsed CN exposure and chronic thiocyanate (SCN) exposure, the major metabolite of CN, in the clownfish species, Amphiprion clarkii. Fish were pulse exposed to 50 ppm CN for 20 or 45 s or chronically exposed to 100 ppm SCN for 12 days and blood plasma levels of SCN were measured. SCN blood plasma levels reached a maximum concentration (301-468 ppb) 0.13-0.17 days after exposure to CN and had a 0.1 to 1.2 day half-life. The half-life of blood plasma SCN after chronic exposure to SCN was found to be 0.13 days. Interestingly, we observed that when a fish, with no previous CN or SCN exposure, was placed in holding water spiked to 20 ppb SCN, there was a steady decrease in the SCN concentration in the holding water until it could no longer be detected at 24 hrs. Under chronic exposure conditions (100 ppm, 12 days), trace levels of SCN (∼40 ppb) were detected in the holding water during depuration but decreased to below detection within the first 24 hrs. Our holding water experiments demonstrate that low levels of SCN in the holding water of A. clarkii will not persist, but rather will quickly and steadily decrease to below detection limits refuting several publications. After CN exposure, A. clarkii exhibits a classic two compartment model where SCN is eliminated from the blood plasma and is likely distributed throughout the body. Similar studies of other species must be examined to continue to develop our understanding of CN metabolism in marine fish before a reliable cyanide detection test can be developed.

Emerging COVID-19 impacts, responses, and lessons for building resilience in the seafood system.

The COVID-19 pandemic and subsequent lockdowns are creating health and economic crises that threaten food and nutrition security. The seafood sector provides important sources of nutrition and employment, especially in low-income countries, and is highly globalized allowing shocks to propagate. We studied COVID-19-related disruptions, impacts, and responses to the seafood sector from January through May 2020, using a food system resilience 'action cycle' framework as a guide. We find that some supply chains, market segments, companies, small-scale actors and civil society have shown initial signs of greater resilience than others. COVID-19 has also highlighted the vulnerability of certain groups working in- or dependent on the seafood sector. We discuss early coping and adaptive responses combined with lessons from past shocks that could be considered when building resilience in the sector. We end with strategic research needs to support learning from COVID-19 impacts and responses.

Food-Based Dietary Guidelines for Seafood Do Not Translate into Increased Long-Chain Omega-3 Levels in the Diet for U.S. Consumers.

Humans under-consume fish, especially species high in long-chain omega-3 fatty acids. Food-based dietary guidelines are one means for nations to encourage the consumption of healthy, nutritious food. Here, associations between dietary omega-3 consumption and food-based dietary guidelines, gross domestic product, the ranked price of fish, and the proportions of marine fish available at a national level were assessed. Minor associations were found between consumption and variables, except for food-based dietary guidelines, where calling out seafood in FBDGs did not associate with greater consumption. This relationship was explored for consumers in the United States, and it was observed that the predominant seafood they ate, shrimp, resulted in little benefit for dietary omega-3 consumption. Seafood is listed under the protein category in the U.S. Dietary Guidelines, and aggregating seafood under this category may limit a more complete understanding of its nutrient benefits beyond protein.

Nutrition and origin of US chain restaurant seafood.

BACKGROUND: Seafood has a nutritional profile that can be beneficial to human health, which gives it a role to play in healthy diets. In addition, because its production and harvesting can have fewer environmental impacts than some forms of animal protein, it can contribute to sustainable diets. However, the positive health and environmental outcomes are not guaranteed-they depend on how seafood is prepared and served and whether it is sourced from sustainable fisheries and aquaculture industries. OBJECTIVES: We examined the availability and nutritional attributes of seafood meals at chain restaurants in the United States. We assessed nutritional attributes by store type and geography. We also assessed menu labeling for species, production methods, and origin. METHODS: The study population was 159 chain restaurants with 100,948 branch locations in the United States. Data were harvested from online restaurant menus, and the nutritional profile of seafood meals was calculated. RESULTS: The average seafood menu item provides up to 49-61% of the total daily limit of saturated fat, 65% of the total daily limit of sodium, and 58-71% of total daily protein requirement for adult men and women. Restaurant chains located in the Deep South and Ohio River Valley, and casual dining chains nationally, carry seafood meals with more total calories and saturated fat per 100 g than other regions or chain types. Most menu items did not list origin or production methods, which is information that would help consumers make informed decisions. CONCLUSIONS: The added ingredients and cooking methods used at chain restaurants can attenuate the health benefits of seafood. We recommend reformulating menus to reduce portion sizes, total calories, added fat, and sodium content per meal and to improve consumer-facing information about origin and production methods.

Optimization of paper-based nanoparticle immunoassays for direct detection of the bacterial pathogen V. parahaemolyticus in oyster hemolymph.

The detection of foodborne pathogens is critical for disease control and infection prevention, especially in seafood consumed raw or undercooked. Paper-based diagnostic tools are promising for rapid fieldable detection and provide a readout by eye due to the use of gold nanoparticle immunoprobes. Here we study different strategies to overcome these challenges in a real biological matrix, oyster hemolymph, for the detection of the pathogenic bacteria Vibrio parahaemolyticus (Vp). Nanoparticle surface chemistry, nitrocellulose speed and blocking, running steps, and antibody concentrations on the NP and nitrocellulose were all studied. Their effect on paper immunoassay signal intensity was quantified to determine optimal conditions, which enabled the detection of Vp directly from hemolymph below pathogenic concentrations.

Did the movie Finding Dory increase demand for blue tang fish?

Representations of wildlife in television and films have long been hypothesized to shape human-wildlife interactions. A recent example is Pixar's film Finding Dory, which featured a blue tang fish (Paracanthurus hepatus) as the main character and was widely reported in the popular press to have increased the number of such fish in the pet trade. We use Bayesian posterior predictive counterfactual models to evaluate the movie's effect on three metrics of societal behaviour. Although there was an increase in global online searches for the blue tang 2-3 weeks after the movie, we find no substantial evidence for an increase in imports of blue tang fish into the US, or in number of visitors to US aquaria compared to counterfactual expectations. It is vital that an evidence-based discourse is used when communicating potential impacts of popular culture on human-wildlife relationships to avoid loss of credibility and misdirection of conservation resources.

An abundance of seafood consumption studies presents new opportunities to evaluate effects on neurocognitive development.

The relationship between seafood eaten during pregnancy and neurocognition in offspring has been the subject of considerable scientific study for over 25 years. Evaluation of this question led two scientific advisory committees to the Dietary Guidelines for Americans (DGAC), the Food and Agriculture Organization of the United Nations with the World Health Organization (FAO/WHO), Health Canada, the European Food Safety Authority (EFSA), and the U.S. Food and Drug Administration (FDA) to conclude through 2014 that seafood consumed by pregnant women is likely to benefit the neurocognitive development of their children. The evidence they reviewed included between four and ten studies of seafood consumption during pregnancy that reported beneficial associations. In contrast there are now 29 seafood consumption studies available describing over 100,000 mothers-child pairs and 15 studies describing over 25,000 children who ate seafood. A systematic review of these studies using Nutrition Evaluation Systematic Review methodology is warranted to determine whether recent research corroborates, builds on, or significantly alters the previous conclusions. Studies that evaluate the integrated effects of seafood as a complete food more directly and completely evaluate impacts on neurocognition as compared to studies that evaluate individual nutritients or toxicological constituents in isolation. Here we address how the findings could add to our understanding of whether seafood consumed during pregnancy and early childhood affects neurocognition, including whether such effects are clinically meaningful, lasting, related to amounts consumed, and affected by any neurotoxicants that may be present, particularly mercury, which is present at varying levels in essentially all seafood. We provide the history, context and rationale for reexamining these questions in light of currently available data.

Ocean acidification alters morphology of all otolith types in Clark's anemonefish (Amphiprion clarkii).

Ocean acidification, the ongoing decline of surface ocean pH and [CO 3 2 - ] due to absorption of surplus atmospheric CO2, has far-reaching consequences for marine biota, especially calcifiers. Among these are teleost fishes, which internally calcify otoliths, critical elements of the inner ear and vestibular system. There is evidence in the literature that ocean acidification increases otolith size and alters shape, perhaps impacting otic mechanics and thus sensory perception. Here, larval Clark's anemonefish, Amphiprion clarkii (Bennett, 1830), were reared in various seawater pCO2/pH treatments analogous to future ocean scenarios. At the onset of metamorphosis, all otoliths were removed from each individual fish and analyzed for treatment effects on morphometrics including area, perimeter, and circularity; scanning electron microscopy was used to screen for evidence of treatment effects on lateral development, surface roughness, and vaterite replacement. The results corroborate those of other experiments with other taxa that observed otolith growth with elevated pCO2, and provide evidence that lateral development and surface roughness increased as well. Both sagittae exhibited increasing area, perimeter, lateral development, and roughness; left lapilli exhibited increasing area and perimeter while right lapilli exhibited increasing lateral development and roughness; and left asterisci exhibited increasing perimeter, roughness, and ellipticity with increasing pCO2. Right lapilli and left asterisci were only impacted by the most extreme pCO2 treatment, suggesting they are resilient to any conditions short of aragonite undersaturation, while all other impacted otoliths responded to lower concentrations. Finally, fish settlement competency at 10 dph was dramatically reduced, and fish standard length marginally reduced with increasing pCO2. Increasing abnormality and asymmetry of otoliths may impact inner ear function by altering otolith-maculae interactions.

Prevalence of alopecia in gray seals Halichoerus grypus atlantica in Massachusetts, USA, 2004-2013.

There has been an increase in the presence of alopecia among gray seals Halichoerus grypus atlantica in Massachusetts, USA. To understand the prevalence and describe the presentation of this condition, data records and photographs of 10070 gray seals from 2004-2013 were reviewed; there were sufficient data to confidently assess the presence or absence of alopecia in 2134 seals. Mild hair loss presented in multifocal patches with minimal to no skin lesions or erythema. In severe cases, alopecia was concentrated over the dorsal head, neck, and shoulders and extended down the ventrum, affecting >50% of the body. Associated skin lesions and erythema were often present. Alopecia was documented in 7.1% of the surveyed seals, and was centered in Nantucket. Alopecia was more prevalent in stranded/sighted animals in spring and summer, with 81% of cases documented from April to July. There was no sex bias, and weanlings were the most affected age class (38%). The etiology for alopecia in Massachusetts gray seals is unknown. Possible causes of alopecia in wildlife are infectious disease, nutritional deficiencies, endocrinopathies, or chronic physiologic stress. High population density around Nantucket may escalate intraspecific competition for resources, which may indirectly lead to stress-induced immunosuppression or nutritional deficiencies. Crowded haul-out sites increase the opportunity for disease transmission. The weanling age class may be prone to alopecia due to naïve immune systems and inexperienced foraging capabilities. Diagnostic sample collection from gray seals will be required to characterize the etiology, pathogenesis, and significance of alopecia in this population.

The skin microbiome of cow-nose rays (Rhinoptera bonasus) in an aquarium touch-tank exhibit.

Public aquaria offer numerous educational opportunities for visitors while touch-tank exhibits offer guests the ability to directly interact with marine life via physical contact. Despite the popularity of touch-tanks, there is a paucity of research about animal health in these exhibits and, in particular, there is little research on the microbial communities in these highly interactive exhibits. Microbial community structure can have implications for both host health and habitat function. To better understand the microbiome of a touch-tank we used high-throughput sequencing of the 16S rRNA gene to analyze the microbial community on the dorsal and ventral surfaces of cow-nose rays (Rhinoptera bonasus) as well as their environment in a frequently visited touch-tank exhibit at the New England Aquarium. Our analyses revealed a distinct microbial community associated with the skin of the ray that had lower diversity than the surrounding habitat. The ray skin was dominated by three orders: Burkholderiales (∼55%), Flavobacteriales (∼19%), and Pseudomonadales (∼12%), taxonomic groups commonly associated with other fish species. Our results provide a survey of ray-associated bacterial communities in a touch-tank environment, thereby laying the foundation for future studies examining the role of potential challenges to ray microbiota and their associated health.

ASSESSMENT OF SERUM 25-HYDROXYVITAMIN D CONCENTRATIONS IN TWO COLLECTIONS OF CAPTIVE GORILLAS (GORILLA GORILLA GORILLA).

Serum 25-hydroxyvitamin D concentrations were assessed in subadult to adult captive lowland gorillas ( Gorilla gorilla gorilla) (n = 26) at two institutions with different husbandry and management practices. Serum 25-hydroxyvitamin D (25[OH]D) concentrations for gorillas managed predominantly indoors was low (14.2 ± 5.9 ng/ml), despite consuming commercial biscuits fortified with vitamin D3. Concentrations of 25(OH)D in gorillas with near daily outdoor access were significantly higher than gorillas managed indoors, although many individuals still had serum values below concentrations recommended for adult humans. Consideration should be given to assessing 25(OH)D concentrations in all captive gorillas and providing specific supplementation, particularly to juveniles without access to direct sunlight.

A transdisciplinary approach to the initial validation of a single cell protein as an alternative protein source for use in aquafeeds.

The human population is growing and, globally, we must meet the challenge of increased protein needs required to feed this population. Single cell proteins (SCP), when coupled to aquaculture production, offer a means to ensure future protein needs can be met without direct competition with food for people. To demonstrate a given type of SCP has potential as a protein source for use in aquaculture feed, a number of steps need to be validated including demonstrating that the SCP is accepted by the species in question, leads to equivalent survival and growth, does not result in illness or other maladies, is palatable to the consumer, is cost effective to produce and can easily be incorporated into diets using existing technology. Here we examine white shrimp (Litopenaeus vannamei) growth and consumer taste preference, smallmouth grunt (Haemulon chrysargyreum) growth, survival, health and gut microbiota, and Atlantic salmon (Salmo salar) digestibility when fed diets that substitute the bacterium Methylobacterium extorquens at a level of 30% (grunts), 100% (shrimp), or 55% (salmon) of the fishmeal in a compound feed. In each of these tests, animals performed equivalently when fed diets containing M. extorquens as when fed a standard aquaculture diet. This transdisciplinary approach is a first validation of this bacterium as a potential SCP protein substitute in aquafeeds. Given the ease to produce this SCP through an aerobic fermentation process, the broad applicability for use in aquaculture indicates the promise of M. extorquens in leading toward greater food security in the future.

Fine-scale transition to lower bacterial diversity and altered community composition precedes shell disease in laboratory-reared juvenile American lobster.

The American lobster Homarus americanus supports a valuable commercial fishery in the Northeastern USA and Maritime Canada; however, stocks in the southern portion of the lobster's range have shown declines, in part due to the emergence of shell disease. Epizootic shell disease is a bacterially induced cuticular erosion that renders even mildly affected lobsters unmarketable because of their appearance, and in more severe cases can cause mortality. Despite the importance of this disease, the associated bacterial communities have not yet been fully characterized. We sampled 2 yr old, laboratory-reared lobsters that displayed signs of shell disease at the site of disease as well as at 0.5, 1, and 1.5 cm away from the site of disease to determine how the bacterial community changed over this fine spatial scale. Illumina sequencing of the 16S rRNA gene revealed a distinct bacterial community at the site of disease, with significant reductions in bacterial diversity and richness compared to more distant sampling locations. The bacterial community composition 0.5 cm from the site of disease was also altered, and there was an observable decrease in bacterial diversity and richness, even though there were no signs of disease at that location. Given the distinctiveness of the bacterial community at the site of disease and 0.5 cm from the site of disease, we refer to these communities as affected and transitionary, and suggest that these bacteria, including the previously proposed causative agent, Aquimarina 'homaria', are important for the initiation and progression of this laboratory model of shell disease.

Expanding our understanding of the trade in marine aquarium animals.

The trade of live marine animals for home and public aquaria has grown into a major global industry. Millions of marine fishes and invertebrates are removed from coral reefs and associated habitats each year. The majority are imported into the United States, with the remainder sent to Europe, Japan, and a handful of other countries. Despite the recent growth and diversification of the aquarium trade, to date, data collection is not mandatory, and hence comprehensive information on species volume and diversity is lacking. This lack of information makes it impossible to study trade pathways. Without species-specific volume and diversity data, it is unclear how importing and exporting governments can oversee this industry effectively or how sustainability should be encouraged. To expand our knowledge and understanding of the trade, and to effectively communicate this new understanding, we introduce the publically-available Marine Aquarium Biodiversity and Trade Flow online database (https://www.aquariumtradedata.org/). This tool was created to communicate the volume and diversity of marine fishes and/or invertebrates imported into the US over three complete years (2008, 2009, and 2011) and three partial years (2000, 2004, 2005). To create this tool, invoices pertaining to shipments of live marine fishes and invertebrates were scanned and analyzed for species name, species quantities, country of origin, port of entry, and city of import destination. Here we focus on the analysis of the later three years of data and also produce an estimate for the entirety of 2000, 2004, and 2005. The three-year aggregate totals (2008, 2009, 2011) indicate that just under 2,300 fish and 725 invertebrate species were imported into the US cumulatively, although just under 1,800 fish and 550 invertebrate species were traded annually. Overall, the total number of live marine animals decreased between 2008 and 2011. In 2008, 2009, and 2011, the total number of individual fish (8.2, 7.3, and 6.9 million individuals) and invertebrates (4.2, 3.7, and 3.6 million individuals) assessed by analyzing the invoice data are roughly 60% of the total volumes recorded through the Law Enforcement Management Information System (LEMIS) dataset. Using these complete years, we back-calculated the number of individuals of both fishes and invertebrates imported in 2000, 2004, and 2005. These estimates (9.3, 10.8, and 11.2 million individual fish per year) were consistent with the three years of complete data. We also use these data to understand the global trade in two species (Banggai cardinalfish, Pterapogon kauderni, and orange clownfish, Amphiprion ocellaris / percula) recently considered for Endangered Species Act listing. Aquariumtradedata.org can help create more effective management plans for the traded species, and ideally could be implemented at key trade ports to better assess the global trade of aquatic wildlife.

Fight Fungi with Fungi: Antifungal Properties of the Amphibian Mycobiome.

Emerging infectious diseases caused by fungal taxa are increasing and are placing a substantial burden on economies and ecosystems worldwide. Of the emerging fungal diseases, chytridomycosis caused by the fungus Batrachochytrium dendrobatidis (hereafter Bd) is linked to global amphibian declines. Amphibians have innate immunity, as well as additional resistance through cutaneous microbial communities. Despite the targeting of bacteria as potential probiotics, the role of fungi in the protection against Bd infection in unknown. We used a four-part approach, including high-throughput sequencing of bacterial and fungal communities, cultivation of fungi, Bd challenge assays, and experimental additions of probiotic to Midwife Toads (Altyes obstetricans), to examine the overlapping roles of bacterial and fungal microbiota in pathogen defense in captive bred poison arrow frogs (Dendrobates sp.). Our results revealed that cutaneous fungal taxa differed from environmental microbiota across three species and a subspecies of Dendrobates spp. frogs. Cultivation of host-associated and environmental fungi realved numerous taxa with the ability to inhibit or facilitate the growth of Bd. The abundance of cutaneous fungi contributed more to Bd defense (~45% of the fungal community), than did bacteria (~10%) and frog species harbored distinct inhibitory communities that were distinct from the environment. Further, we demonstrated that a fungal probiotic therapy did not induce an endocrine-immune reaction, in contrast to bacterial probiotics that stressed amphibian hosts and suppressed antimicrobial peptide responses, limiting their long-term colonization potential. Our results suggest that probiotic strategies against amphibian fungal pathogens should, in addition to bacterial probiotics, focus on host-associated and environmental fungi such as Penicillium and members of the families Chaetomiaceae and Lasiosphaeriaceae.

Cuticles of European and American lobsters harbor diverse bacterial species and differ in disease susceptibility.

Diseases of lobster shells have a significant impact on fishing industries but the risk of disease transmission between different lobster species has yet to be properly investigated. This study compared bacterial biofilm communities from American (Homarus americanus) and European lobsters (H. gammarus), to assess both healthy cuticle and diseased cuticle during lesion formation. Culture-independent molecular techniques revealed diversity in the bacterial communities of cuticle biofilms both within and between the two lobster species, and identified three bacterial genera associated with shell lesions plus two putative beneficial bacterial species (detected exclusively in healthy cuticle or healing damaged cuticle). In an experimental aquarium shared between American and European lobsters, heterospecific transmission of potentially pathogenic bacteria appeared to be very limited; however, the claws of European lobsters were more likely to develop lesions when reared in the presence of American lobsters. Aquarium biofilms were also examined but revealed no candidate pathogens for environmental transmission. Aquimarina sp. 'homaria' (a potential pathogen associated with a severe epizootic form of shell disease) was detected at a much higher prevalence among American than European lobsters, but its presence correlated more with exacerbation of existing lesions rather than with lesion initiation.

A comparison of the structure of American (Homarus americanus) and European (Homarus gammarus) lobster cuticle with particular reference to shell disease susceptibility.

The integument of arthropods is an important first-line defence against the invasion of parasites and pathogens. Once damaged, this can be subject to colonisation by microbial agents from the surrounding environment, which in crustaceans can lead to a condition termed shell disease syndrome. This condition has been reported in several crustacean species, including crabs and lobsters. The syndrome is a progressive condition where the outer cuticle becomes pitted and eroded, and in extreme cases is compromised, leaving animals susceptible to septicaemia. This study examined the susceptibility of juvenile American (Homarus americanus) and European (Homarus gammarus) lobsters to shell disease, as a result of mechanical damage. Scanning electron microscopy was used as a method to identify differences in the cuticle structure and consequences of mechanical damage. Claw regions were aseptically punctured, whilst carapaces were abraded using sterile sandpaper, to mimic natural damage. After a period of between 10 and 12 weeks, lobsters were sacrificed, fixed and stored for later examination. The carapace and claws of juvenile American lobsters were shown to be thinner and more vulnerable to abrasion damage than their European counterparts. In addition, the number and distribution of setal pits and pore canal openings also differed between the two species of lobster. Mechanical damage resulted in the formation of shell disease lesions on the claw and carapace of both lobster species. However, American lobsters, unlike their European counterparts, had extensive bacterial colonisation on the margins of these lesions. Overall, it is concluded that the cuticle of the American lobster is more susceptible to damage and resulting microbial colonisation. This may have implications for susceptibility of both species of lobster to shell disease syndrome.