The presence of Apicomplexan parasites in king scallops (Pecten maximus) in Scottish waters.

The king scallop (Pecten maximus) is a commercially important species found around the United Kingdom coast. The association of an Apicomplexan-like parasite with mass mortality of Icelandic scallop (Chlamys islandica) in Iceland and the presence of identical parasites in king scallop (Pecten maximus) and queen scallop (Aequipecten opercularis) in Scotland raised serious concerns regarding the health of Scottish king scallops. Marine Scotland Science (MSS) conducted a survey in 2016 to assess the prevalence and the intensity of parasite infection in king scallops. King scallops were collected and sampled during the annual scallop dredge surveys in the Shetland Isles and the east and west coast of Scotland. The king scallop adductor muscle was macroscopically examined and tissue imprints taken to grade the intensity of infection. The parasite was present in the majority of the king scallops sampled in all surveyed areas: Shetland Isles 87.1%, east coast 76.0% and west coast of Scotland 64.1%. However, the parasitic infestations were light in intensity with the majority of the king scallops graded as 1 (≤20 zoites per microscopic field). No macroscopic changes in the adductor muscle were observed and histopathology examination revealed minor localized fiber degeneration of adjacent fibers to parasite clusters. The results suggested the parasite to be widespread around the Scottish coast and it appears to be able to live within the king scallop at low intensity of infection without causing significant downgrade of the adductor muscle (in terms of colour or texture) or mortality. The partial genome sequence of the parasite in king scallops from Scottish waters was identical to the one reported by Kristmundsson and Freeman (2018) in the Icelandic scallop in Icelandic waters.

Risk factors and prevalence of enteroparasitic diseases in shellfish pickers from a lake area in the Northeast of Brazil / Fatores de risco e prevalência de enteroparasitoses em marisqueiras de uma região lagunar no Nordeste do Brasil

INTRODUCTION: Intestinal parasitosis are a public health problem worldwide. There are several risk factors and a high association with some specific labor activities. OBJECTIVE: The present study assessed the risk factors and prevalence of enteroparasitic diseases in shellfish pickers from one district of Maceió, Alagoas state, Brazil. METHODS: Crosssectional study of 41 female shellfish pickers including parasitological tests in fecal samples and a questionnaire with objective and subjective questions. Sand samples from their working environment were also analyzed. RESULTS: At least one species of parasite was found in 19.51% of the fecal samples. Pathogenic species of Giardia lamblia, Trichuris trichiura, Schistosoma mansoni, Ascaris lumbricoides, Enterobius vermicularis, from the Ancylostomatidae family, and non-pathogenic species of Entamoeba coli were found. Polyparasitism was diagnosed in 37.5% of the positive samples. A total of 57.14% of sand samples contained hookworm larvae. Regarding the risk factor, low educational level was statistically associated to the presence of parasites (p<0.05). CONCLUSION: Greater investment in basic education is needed to increase the knowledge about preventive measures against parasitic diseases and the promotion food-handling courses in order to change existing inadequate habits in the community. Basic sanitation is also essential in preventing environmental contamination.

INTRODUÇÃO: As parasitoses intestinais representam um problema de saúde pública mundial. Estão associados inúmeros fatores de risco, bem como atividades laborais específicas. OBJETIVO: O presente estudo avaliou os fatores de risco e a prevalência de enteroparasitoses em marisqueiras de um bairro da cidade de Maceió, Alagoas, Brasil. MÉTODOS: Procedeu-se um estudo de corte transversal, sendo 41 marisqueiras avaliadas mediante exame parasitológico de fezes e questionário contendo questões discursivas e de múltipla escolha. Foram analisadas também amostras de areia do ambiente de trabalho das mesmas. RESULTADOS: A positividade para pelo menos uma espécie de parasito nas fezes foi de 19,51%. Foram encontradas as espécies patogênicas Giardia lamblia, Trichuris trichiura, Schistosoma mansoni, Ascaris lumbricoides, Enterobius vermicularis e da família Ancylostomatidae, e a espécie não patogênica, Entamoeba coli. Poliparasitismo foi diagnosticado em 37,5% dos exames positivos. Um total de 57,14% das amostras de areia continha larvas de ancilostomatídeos. Em relação aos fatores de risco, a baixa escolaridade foi estatisticamente associada à presença de parasitas (p<0,05). CONCLUSÃO: É necessário maior investimento na educação básica para aprimorar o conhecimento das formas de prevenção das parasitoses e a promoção de cursos de boas práticas de manipulação de alimentos, a fim de modificar hábitos errôneos já incorporados na comunidade. O saneamento básico também é fundamental para evitar contaminação ambiental.

New insights into the Manila clam - Perkinsus olseni interaction based on gene expression analysis of clam hemocytes and parasite trophozoites through in vitro challenges.

The Manila clam (Ruditapes philippinarum) is the bivalve species with the highest global production from both fisheries and aquaculture, but its production is seriously threatened by perkinsosis, a disease caused by the protozoan parasite Perkinsus olseni. To understand the molecular mechanisms underlying R. philippinarum-P. olseni interactions, we analysed the gene expression profiles of in vitro challenged clam hemocytes and P. olseni trophozoites, using two oligo-microarray platforms, one previously validated for R. philippinarum hemocytes and a new one developed and validated in this study for P. olseni. Manila clam hemocytes were in vitro challenged with trophozoites, zoospores, and extracellular products from P. olseni in vitro cultures, while P. olseni trophozoites were in vitro challenged with Manila clam plasma along the same time-series (1 h, 8 h, and 24 h). The hemocytes showed a fast activation of the innate immune response, particularly associated with hemocyte recruitment, in the three types of challenges. Nevertheless, different immune-related pathways were activated in response to the different parasite stages, suggesting specific recognition mechanisms. Furthermore, the analyses provided useful complementary data to previous in vivo challenges, and confirmed the potential of some proposed biomarkers. The combined analysis of gene expression in host and parasite identified several processes in both the clam and P. olseni, such as redox and glucose metabolism, protease activity, apoptosis and iron metabolism, whose modulation suggests cross-talk between parasite and host. This information might be critical to determine the outcome of the infection, thus highlighting potential therapeutic targets. Altogether, the results of this study aid understanding the response and interaction between R. philippinarum and P. olseni, and will contribute to developing effective control strategies for this threatening parasitosis.

Modeling Pathogen Dispersal in Marine Fish and Shellfish.

In marine ecosystems, oceanographic processes often govern host contacts with infectious agents. Consequently, many approaches developed to quantify pathogen dispersal in terrestrial ecosystems have limited use in the marine context. Recent applications in marine disease modeling demonstrate that physical oceanographic models coupled with biological models of infectious agents can characterize dispersal networks of pathogens in marine ecosystems. Biophysical modeling has been used over the past two decades to model larval dispersion but has only recently been utilized in marine epidemiology. In this review, we describe how biophysical models function and how they can be used to measure connectivity of infectious agents between sites, test hypotheses regarding pathogen dispersal, and quantify patterns of pathogen spread, focusing on fish and shellfish pathogens.

The aquatic animal pandemic crisis.

The growth of aquaculture over the past 50 years has been accompanied by the emergence of aquatic animal diseases, many of which have spread to become pandemic in countries or continents. An analysis of 400 emerging disease events in aquatic animals that were logged by the Centre for Environment, Fisheries and Aquaculture Science between 2002 and 2017 revealed that more than half were caused by viruses. However, in molluscs, most events were parasitic. Categorising these events indicated that the key processes underpinning emergence were the movement of live animals and host switching. Profiles of key pathogens further illustrate the importance of wild aquatic animals as the source of new infections in farmed animals. It is also clear that the spread of new diseases through the largescale movement of aquatic animals for farming, for food and for the ornamental trade has allowed many to achieve pandemic status. Many viral pathogens of fish (e.g. infectious salmon anaemia, viral haemorrhagic septicaemia) and shrimp (e.g. white spot syndrome virus) affect a large proportion of the global production of key susceptible species. Wild aquatic animal populations have also been severely affected by pandemic diseases, best exemplified by Batrachochytrium dendrobatidis, a fungal infection of amphibians, whose emergence and spread were driven by the movement of animals for the ornamental trade. Batrachochytrium dendrobatidis is now widespread in the tropics and subtropics and has caused local extinctions of susceptible amphibian hosts. Given the rising demand for seafood, aquacultural production will continue to grow and diseases will continue to emerge. Some will inevitably achieve pandemic status, having significant impacts on production and trade, unless there are considerable changes in global monitoring and the response to aquatic animal diseases.

Au cours des 50 dernières années, la forte croissance qu'a connue l'aquaculture est allée de pair avec l'émergence de nombreuses maladies affectant les animaux aquatiques, dont certaines se sont propagées jusqu'à devenir pandémiques à l'échelle nationale ou continentale. L'analyse de 400 événements sanitaires survenus chez des animaux aquatiques et consignés entre 2002 et 2017 par le Centre for Environment, Fisheries and Aquaculture Science a déterminé l'origine virale de plus de la moitié d'entre eux. Toutefois, chez les mollusques la plupart des événements analysés étaient d'ordre parasitaire. Le classement des événements par catégories a montré que les principaux processus sous-jacents à cette émergence étaient liés aux transferts d'animaux vivants et à la colonisation de nouveaux hôtes par les agents pathogènes. Les profils des agents pathogènes majeurs illustrent le rôle des espèces aquatiques sauvages en tant que sources d'infections nouvelles chez les animaux aquatiques d'élevage. Il apparaît clairement que la propagation de nouvelles maladies à la faveur des transferts massifs d'animaux aquatiques à des fins d'élevage, de production alimentaire ou de commerce d'espèces d'ornement a conféré un statut pandémique à nombre de ces maladies. De nombreux virus affectant les poissons (par ex., le virus de l'anémie infectieuse du saumon, le virus de la septicémie hémorragique virale) et les crevettes (par ex., le virus du syndrome des points blancs) ont une incidence majeure sur de vastes segments de la production mondiale d'espèces sensibles cruciales. Les populations sauvages d'animaux aquatiques sont également touchées par des maladies pandémiques, dont l'exemple type est l'infection à Batrachochytrium dendrobatidis, une affection fongique des amphibiens dont l'émergence et la propagation sont le fruit des transferts d'animaux aquatiques destinés au commerce aquariophile. Batrachochytrium dendrobatidis est désormais largement présent dans les eaux tropicales et subtropicales où il est responsable d'extinctions locales parmi les espèces d'amphibiens sensibles. La croissance de la production aquacole se poursuivra afin de répondre à une demande toujours plus forte en poissons et fruits de mer, entraînant l'émergence continue de nouvelles maladies. Si des changements déterminants ne sont pas introduits dans la surveillance exercée au niveau mondial sur les maladies des animaux aquatiques et dans la réponse qui leur est apportée, certaines de ces maladies vont inéluctablement acquérir une dimension pandémique avec des conséquences importantes sur la production et le commerce.

El crecimiento de la acuicultura en los últimos 50 años se ha acompañado de la aparición de enfermedades de los animales acuáticos, que en muchos casos se han propagado hasta llegar a ser pandémicas en ciertos países o continentes. Tras analizar 400 episodios de enfermedades emergentes de animales acuáticos registrados entre 2002 y 2017 por el Centre for the Environment, Fisheries and Aquaculture Science, los autores constataron que más de la mitad de esos episodios fueron causados por virus, si bien en el caso de los moluscos la mayoría de ellos eran parasitarios. De la clasificación de esos episodios se desprende que los procesos básicos que subyacen a su aparición son los desplazamientos de animales vivos y los cambios de anfitrión. El perfil de los principales patógenos revela además la importancia que revisten los animales acuáticos silvestres como fuente de nuevas infecciones de los animales de acuicultura. También está claro que la propagación de nuevas enfermedades por el movimiento a gran escala de animales acuáticos con fines de producción acuícola, consumo alimentario o comercio de animales ornamentales ha propiciado que muchas de ellas adquieran carácter pandémico. Muchos patógenos víricos de los peces (como el virus de la anemia infecciosa del salmón o el de la septicemia hemorrágica viral) y camarones (como el virus del síndrome de las manchas blancas) afectan a una gran parte de la producción mundial de las principales especies sensibles. Las poblaciones silvestres de animales acuáticos también se han visto afectadas de gravedad por enfermedades pandémicas, como ejemplifica perfectamente la infección por Batrachochytrium dendrobatidis, micosis de los anfibios cuya aparición y propagación fue alimentada por el comercio y el consiguiente movimiento de animales con fines ornamentales. Este hongo, muy extendido ahora en las regiones tropicales y subtropicales, ha causado la extinción en ciertas áreas de especies anfibias sensibles. Habida cuenta de la creciente demanda de alimentos de origen marino, la producción acuícola seguirá creciendo y también seguirán surgiendo enfermedades. Inevitablemente, algunas de ellas se harán pandémicas y resultarán muy dañinas para la producción y el comercio, a menos que haya cambios de calado en los sistemas mundiales de vigilancia y respuesta ante las enfermedades de los animales acuáticos.

The parasitic dinoflagellate Hematodinium perezi infecting mudflat crabs, Helice tientsinensis, in polyculture system in China.

Since 2012, frequent outbreaks of Hematodinium diseases have significantly impacted sustainable culture of marine crabs Portunus trituberculatus in the coastal areas of Shandong Peninsula. The mechanisms of the Hematodinium parasite epizootics in polyculture pond systems remain to be explored and alternate crustacean hosts are speculated to play important roles in transmission and epizootiology of the disease. To investigate their possible role in transmission, the common wild mudflat crabs Helice tientsinensis were sampled from the waterway connecting to polyculture ponds in Huangdao, Qingdao, China and diagnosed for Hematodinium infection. Hematodinium infection was found in H. tientsinensis collected in the waterway from April-November 2018, with a prevalence of 5.8-31.7%. In addition, 23.1% of H. tientsinensis sampled from the adjacent polyculture pond were infected during the peak of the Hematodinium epizootic in July. Amoeboid trophonts or prespores were observed in the hemolymph of the Hematodinium-infected crabs and histopathological changes were observed in major organs (e.g. hepatopancreas, heart, gill and muscle). The ITS1 rRNA of Hematodinium sp. infecting H. tientsinensis shared 99-100% similarity to isolates infecting P. trituberculatus and Penaeus monodon in local polyculture ponds, and are included in a monophylogenetic clade, Hematodinium perezi genotype II, in the phylogenetic tree. The results further showed that this generalist parasite was infecting various types of marine crustaceans in the coastal waters of China, and that mudflat crabs H. tientsinensis may serve as an important alternate host during epizootics of Hematodinium parasites in the polyculture system.

Novel Insights on the Toxicity of Phycotoxins on the Gut through the Targeting of Enteric Glial Cells.

In vitro and in vivo studies have shown that phycotoxins can impact intestinal epithelial cells and can cross the intestinal barrier to some extent. Therefore, phycotoxins can reach cells underlying the epithelium, such as enteric glial cells (EGCs), which are involved in gut homeostasis, motility, and barrier integrity. This study compared the toxicological effects of pectenotoxin-2 (PTX2), yessotoxin (YTX), okadaic acid (OA), azaspiracid-1 (AZA1), 13-desmethyl-spirolide C (SPX), and palytoxin (PlTX) on the rat EGC cell line CRL2690. Cell viability, morphology, oxidative stress, inflammation, cell cycle, and specific glial markers were evaluated using RT-qPCR and high content analysis (HCA) approaches. PTX2, YTX, OA, AZA1, and PlTX induced neurite alterations, oxidative stress, cell cycle disturbance, and increase of specific EGC markers. An inflammatory response for YTX, OA, and AZA1 was suggested by the nuclear translocation of NF-κB. Caspase-3-dependent apoptosis and induction of DNA double strand breaks (γH2AX) were also observed with PTX2, YTX, OA, and AZA1. These findings suggest that PTX2, YTX, OA, AZA1, and PlTX may affect intestinal barrier integrity through alterations of the human enteric glial system. Our results provide novel insight into the toxicological effects of phycotoxins on the gut.

Comparison of PCR assays to detect Toxoplasma gondii oocysts in green-lipped mussels (Perna canaliculus).

Toxoplasma gondii is recognised as an important pathogen in the marine environment, with oocysts carried to coastal waters in overland runoff. Currently, there are no standardised methods to detect T. gondii directly in seawater to assess the extent of marine ecosystem contamination, but filter-feeding shellfish may serve as biosentinels. A variety of PCR-based methods have been used to confirm presence of T. gondii DNA in marine shellfish; however, systematic investigations comparing molecular methods are scarce. The primary objective of this study was to evaluate analytical sensitivity and specificity of two nested-PCR (nPCR) assays targeting dhps and B1 genes and two real-time (qPCR) assays targeting the B1 gene and a 529-bp repetitive element (rep529), for detection of T. gondii. These assays were subsequently validated for T. gondii detection in green-lipped mussel (Perna canaliculus) haemolymph using oocyst spiking experiments. All assays could reliably detect 50 oocysts spiked into mussel haemolymph. The lowest limit of detection was 5 oocysts using qPCR assays, with the rep529 primers performing best, with good correlation between oocyst concentrations and Cq values, and acceptable efficiency. Assay specificity was evaluated by testing DNA from closely related protozoans, Hammondia hammondi, Neospora caninum, and Sarcocystis spp. Both nPCR assays were specific to T. gondii. Both qPCR assays cross-reacted with Sarcocystis spp. DNA, and the rep529 primers also cross-reacted with N. caninum DNA. These studies suggest that the rep529 qPCR assay may be preferable for future mussel studies, but direct sequencing is required for definitive confirmation of T. gondii DNA detection.

Comparison of Cryptosporidium oocyst recovery methods for their applicability for monitoring of consumer-ready fresh shellfish.

Growing demand for fresh, unprocessed food favours the emergence of Cryptosporidium infections in humans. Mainly it is food of plant origin or unpasteurized milk which have been involved in food-borne outbreaks of cryptosporidiosis. So far consumption of shellfish contaminated with Cryptosporidium were not associated with human infections although such as possibility exists. In this study an attempt was undertaken to evaluate the analytical performance of three commonly used methods for recovery of Cryptosporidium oocysts from shellfish: i) pepsin digestion of shellfish in conjunction with immunomagnetic separation (IMS) of oocysts (method A), ii) pepsin-HCl treatment of shellfish homogenate without IMS (method B), and iii) a strainer method with direct oocyst extraction and separation from shellfish tissue using IMS (method C). Each method's performance was assessed according to the ISO standard requirements by testing shellfish homogenates seeded with different numbers of C. parvum oocysts. Two groups of parameters were compared, encompassing precision (coefficient of variation (CV)) and accuracy of measurements. These were described by linear regression models allowing calculation of the methods' limits of detection (LOD) and quantification (LOQ). In addition, oocyst recovery efficiencies from shellfish were calculated for each method. All three compared methods allowed for at least 66% recovery of Cryptosporidium oocysts from the tested samples. The best recovery (83.3-100%) in the whole range of tested suspensions was obtained for method C. The accuracy of method B was better (linearity of r2 = 0.9996 in the full measurement range) than that of method A (r2 = 0.968). Method C showed the best accuracy (r2 = 1) and precision (CV 0.2-14.1). Compared to other methods it was also characterised by the best LOD and LOQ, attaining ≅4 and ≅12 oocysts per 3 g of tested shellfish sample respectively. Despite a lack of the ability of method A to give the proportional results in oocysts recovery (non-linearity of the method) compared to the reference values, it achieved the highest LOD and LOQ values among the tested methods. As demonstrated here, the most efficient method for extraction of Cryptosporidium oocysts from shellfish tissues was method C employing sample homogenisation and separation of oocysts from tissue debris using IMS. Used alone this method does not in fact allow for identification of Cryptosporidium species but delivers quantitative results concerning the level of food contamination by parasites.

Toxoplasma gondii and Other Zoonotic Protozoans in Mediterranean Mussel ( Mytilus galloprovincialis) and Blue Mussel ( Mytilus edulis): A Food Safety Concern?

Mediterranean mussels ( Mytilus galloprovincialis) and blue mussels ( Mytilus edulis) are among the most consumed fishery products, but they are frequent vehicles of foodborne infection worldwide. In this study, we investigated the occurrence and seasonality of zoonotic protozoans in mussels farmed or sold at retail outlets in Italy. We collected and tested 1,440 M. galloprovincialis and 180 M. edulis. Pooled samples were molecularly tested for Giardia duodenalis, Cryptosporidium spp., and Toxoplasma gondii and then sequenced. Sixty-two (45.9%; 95% confidence interval, 37.5 to 54.3%) mussel pools tested positive for one or more of the investigated pathogens. Both Mytilus species and samples from all the investigated areas harbored pathogens. Mussels were statistically more contaminated by Cryptosporidium spp., followed by T. gondii and G. duodenalis assemblage A, and M. galloprovincialis was more contaminated than M. edulis ( P < 0.01). Contamination was more likely in mussels at retail outlets ( P < 0.05) than in those from farms and in mussels collected in spring ( P < 0.01) than in other seasons. This is the first report of T. gondii found in M. galloprovincialis in Italy and in M. edulis in Europe. The detection of zoonotic protozoans in a widely consumed food source indicates the need for a more detailed microbiological risk analysis, especially considering that bivalve mollusks are often consumed raw worldwide.

Survey of Gymnophalloides seoi Metacercariae in Natural and Cultured Oysters from Several Western Coastal Areas, Korea.

Gymnophalloides seoi (Digenea: Gymnophallidae) is a human intestinal trematode contracted by eating raw oysters (Crassostrea gigas) in the Republic of Korea (=Korea). It has been known to be highly endemic in Aphae Island, Shinan-gun, Jeollanam-do (Province). However, recent epidemiological status of G. seoi has not been reported since the 1990s. In this study, we investigated the prevalence of G. seoi metacercariae in natural and cultured oysters collected from 3 islands and 2 coastal areas in western parts of Korea. The oysters were examined using the artificial digestion method followed by stereomicroscopy. The overall positive rate of G. seoi metacercariae in natural oysters was 66.0% (99/150), and the oysters collected from Yubu Island showed the highest infection rate (74.0%). However, the metacercarial density per oyster was relatively low (1.5-2.4 per oyster). By contrast, no metacercaria was found in cultured oysters purchased from 2 coastal areas in Chungcheongnam-do. Thus, we could confirm that natural oysters produced from 3 western coastal islands are infected with G. seoi metacercariae, whereas cultured oysters purchased from 2 coastal areas were free from infection.

Tapeworm larvae in Artemia franciscana (Crustacea: Anostraca) in the Godolphin lakes of Dubai (United Arab Emirates) throughout an annual cycle.

A total of 1840 brine shrimps (Artemia franciscana) were examined for cestode larvae at monthly intervals between November 2015 and June 2016. Of these, 663 (36.03%) specimens were infected with cysticercoids of seven cestode species in numbers between one and sixteen. During the first four months of examination, the percentage of infected shrimps was low but rose significantly with increasing temperatures in March, reaching maximum values in May. Flamingolepis liguloides and Flamingolepis flamingo showed the highest prevalence overall, at 25.3 and 10.7%, respectively. The intensity of infection was 1-10 and 1-4 cysticercoids, respectively. Eurycestus avoceti, Wardium stellorae, Gynandrotaenia stammeri, Anomotaenia tringae and Confluaria podicipina occurred at lower prevalence of 4.5, 3.2, 1.7, 0.3 and 0.05%, respectively. Up to four species were detected in one host.

Perkinsus olseni in the short neck yellow clam, Paphia malabarica (Chemnitz, 1782) from the southwest coast of India.

Parasites of the genus Perkinsus predominantly infect bivalves, and two species among them, P. olseni and P. marinus, are notifiable to OIE. P. olseni infections are known to cause extensive damage to wild as well as farmed bivalves globally with enormous implications to its fishery. Consequent to the initiation of a surveillance programme for aquatic animal diseases in India, Perkinsus infections were observed in many species of bivalves. The present paper describes P. olseni infections in the short neck yellow clam, Paphia malabarica from the southwest coast of India. Diagnosis of the parasite was carried out using Ray's Fluid Thioglycollate Medium culture, histology, in-situ hybridisation and molecular taxonomy. Pathology of infection and development of zoospores is also described. This forms the first report of a P. olseni infection in P. malabarica. High prevalence and intensity of infection of Perkinsus in clams raises concerns, as clam reserves in this geographical area sustain fisheries and the livelihoods of local fishing communities.

The facultatively parasitic ciliated protozoan, Tetrahymena glochidiophila (Lynn, 2018), causes a reduction in viability of freshwater mussel glochidia.

This study investigated the effect of a previously uncharacterized species of ciliated protozoan, Tetrahymena glochidiophila, on the viability of glochidia from three species of freshwater mussel (Lampsilis siliquoidea, Lampsilis fasciola, and Lampsilis cardium). Over the course of 72 h, the viability of glochidia exposed to T. glochidiophila declined by >60% while the decline in the viability of uninfected glochidia was <10%. The density of T. glochidiophila increased >1000-fold during the experiment in treatments with infected glochidia. Lampsilis cardium glochidia were also either exposed to gill rinsate or gill contents from infected gravid female L. siliquoidea for the purpose of elucidating the location of the greatest density of ciliates within infected mussels. Glochidia exposed to gill contents declined significantly (p < 0.05) more than glochidia exposed to gill rinsate. Finally, a clone of ciliates was isolated from infected glochidia and cultured on bacterized medium. The clonal culture was then used to expose uninfected glochidia for the purpose of further confirming a parasitic relationship between glochidia and T. glochidiophila. The viability of glochidia exposed to T. glochidiophila from the clonal culture declined significantly relative to uninfected glochidia but not to the extent of glochidia exposed to ciliates from the gills of infected L. siliquoidea.

Parasites in two coexisting bivalves of the Patagonia coast, southwestern Atlantic Ocean: The Puelche oyster (Ostrea puelchana) and false oyster (Pododesmus rudis).

The purpose of this study was to compare the parasites of two coexisting bivalves, the edible Puelche oyster (Ostrea puelchana) and the false oyster (Pododesmus rudis) that lives attached to O. puelchana shells, and to investigate their host specificity. Samples from wild populations, 465 O. puelchana and 131 P. rudis, were collected seasonally during two years in the San José Gulf (northern Patagonia, Argentina) and were processed using standard histological techniques. To increase the natural low prevalences of Bonamia spp. and Perkinsus spp. that are present in wild populations, an in situ experiment was performed by maintaining captive sentinel bivalves at high densities inside a plastic mesh bag to enhance parasite transmission. Polymerase chain reaction (PCR) assays were used to test for apparent Bonamia sp. infections among captive sentinel O. puelchana specimens (n = 80), and Ray's fluid thioglycollate medium (RFTM) assays and histological immunoassays tested for apparent Perkinsus sp. infections among captive sentinel P. rudis specimens (n = 100). Despite histological observations that revealed the presence of microcells resembling Bonamia sp. infecting hemocytes of some Puelche oysters, PCR assays did not confirm that parasite identification. Among captive sentinel P. rudis that showed histological evidence of Perkinsus sp. infections, neither RFTM nor immunoassays confirmed such parasites. Ostrea puelchana from wild populations were occasional hosts for both Rickettsia-like organism (RLOs) and Urastoma-like turbellarians. In contrast, six parasite taxa infected P. rudis from coexisting populations, including RLOs, Urastoma-like turbellarians, an intracellular gregarine species, Nematopsis-like oocysts, an unidentified coccidian and a Perkinsus qugwadi-like protozoan. These results demonstrated specific infection patterns of the identified parasites in relation to their hosts.

Haplosporidium pinnae sp. nov., a haplosporidan parasite associated with mass mortalities of the fan mussel, Pinna nobilis, in the Western Mediterranean Sea.

This study provides morphological and molecular characterization of a new species, Haplosporidium pinnae), very likely responsible for mass mortality of fan mussels, Pinna nobilis, in the Western Mediterranean Sea. The parasite was found in dead or moribund P. nobilis but did not occur in healthy fan mussels from locations that were not affected by abnormal mortality. Histological examination of infected fan mussels showed uninucleate cells of a haplosporidan parasite throughout the connective tissue and hemolymph sinuses of the visceral mass and binucleate cells and, rarely, multinucleate plasmodia were also detected in the connective tissue. Additionally, stages of sporulation occurred in the epithelium of the host digestive gland tubules. Spores were slightly ellipsoidal with a hinged operculum in one pole. Typical haplosporosomes were not found with TEM but vesicles with two concentric membranes resembling haplosporosomes were abundant in the cytoplasm of the multinucleate plasmodia occurring in host digestive gland tubules. SEM analysis showed multiple structures on the spore surface; some spores had two or four long tape-like filaments attached to the spore wall. Phylogenetic analysis based on the SSU rDNA sequence placed this parasite within a large clade including species of the order Haplosporida, not in the Bonamia/Minchinia subclade or the subclade containing most Haplosporidium species, but within a subclade of Haplosporidium sp. from Penaeus vannamei. Our results suggested that H. pinnae and the parasite of P. vannamei may represent a distinct new genus within the order Haplosporida.

Mytilus trossulus and hybrid (M. edulis-M. trossulus) - New hosts organisms for pathogenic microalgae Coccomyxa sp. from the Estuary and northwestern Gulf of St. Lawrence, Canada.

During summer 2014-2017, wild mytilid mussels, highly infested with the pathogenic Coccomyxa-like microalgae, were collected along the Estuary and northwestern part of Gulf of St. Lawrence (Québec, Canada). Molecular identification showed that algae can be assigned to a single taxon, Coccomyxa sp. (KJ372210), whereas hosts are represented by Mytilus edulis, M. trossulus and hybrid between these two species. This is the first record of M. trossulus and hybrid among hosts of this pathogenic alga. Our results are indicative of a possible widespread distribution of Coccomyxa sp. in the Lower St. Lawrence Estuary and along coastal waters of Canadian Maritime provinces.

First report of Toxoplasma gondii sporulated oocysts and Giardia duodenalis in commercial green-lipped mussels (Perna canaliculus) in New Zealand.

Pollution of marine ecosystems with the protozoan parasites Toxoplasma gondii, Cryptosporidium spp. and Giardia duodenalis can be studied using bivalve shellfish as biosentinels. Although evidence suggests that these parasites are present in New Zealand coastal waters, the extent of protozoal pollution has not been investigated. This study used optimised molecular methods to detect the presence of Cryptosporidium spp., G. duodenalis and T. gondii in commercially sourced green-lipped mussel (Perna canaliculus), an endemic species found throughout coastal New Zealand. A nested polymerase chain reaction was validated for detection of T. gondii DNA and applied to 104 commercially sourced mussels. Thirteen mussels were positive for T. gondii DNA with an estimated true prevalence of 16.4% using Bayesian statistics, and the presence of T. gondii in mussels was significantly associated with collection during the summer compared with that in the winter (P = 0.003). Consumption of contaminated shellfish may also pose a health risk for humans and marine wildlife. As only sporulated T. gondii oocysts can be infectious, a reverse transcriptase-polymerase chain reaction was used to confirm presence of a sporozoite-specific marker (SporoSAG), detected in four mussels. G. duodenalis assemblage B, known to be pathogenic in humans, was also discovered in 1% mussels, tested by polymerase chain reaction (n = 90). Cryptosporidium spp. was not detected in the sampled mussel haemolymph. Results suggest that New Zealand may have high levels of coastal contamination with T. gondii, particularly in summer months, and that naturally exposed mussels can ingest and retain sporulated oocysts, further establishing shellfish consumption as a health concern.

Evaluation of Rapid, Early Warning Approaches to Track Shellfish Toxins Associated with Dinophysis and Alexandrium Blooms.

Marine biotoxin-contaminated seafood has caused thousands of poisonings worldwide this century. Given these threats, there is an increasing need for improved technologies that can be easily integrated into coastal monitoring programs. This study evaluates approaches for monitoring toxins associated with recurrent toxin-producing Alexandrium and Dinophysis blooms on Long Island, NY, USA, which cause paralytic and diarrhetic shellfish poisoning (PSP and DSP), respectively. Within contrasting locations, the dynamics of pelagic Alexandrium and Dinophysis cell densities, toxins in plankton, and toxins in deployed blue mussels (Mytilus edulis) were compared with passive solid-phase adsorption toxin tracking (SPATT) samplers filled with two types of resin, HP20 and XAD-2. Multiple species of wild shellfish were also collected during Dinophysis blooms and used to compare toxin content using two different extraction techniques (single dispersive and double exhaustive) and two different toxin analysis assays (liquid chromatography/mass spectrometry and the protein phosphatase inhibition assay (PP2A)) for the measurement of DSP toxins. DSP toxins measured in the HP20 resin were significantly correlated (R² = 0.7-0.9, p < 0.001) with total DSP toxins in shellfish, but were detected more than three weeks prior to detection in deployed mussels. Both resins adsorbed measurable levels of PSP toxins, but neither quantitatively tracked Alexandrium cell densities, toxicity in plankton or toxins in shellfish. DSP extraction and toxin analysis methods did not differ significantly (p > 0.05), were highly correlated (R² = 0.98-0.99; p < 0.001) and provided complete recovery of DSP toxins from standard reference materials. Blue mussels (Mytilus edulis) and ribbed mussels (Geukensia demissa) were found to accumulate DSP toxins above federal and international standards (160 ng g-1) during Dinophysis blooms while Eastern oysters (Crassostrea virginica) and soft shell clams (Mya arenaria) did not. This study demonstrated that SPATT samplers using HP20 resin coupled with PP2A technology could be used to provide early warning of DSP, but not PSP, events for shellfish management.

Alteration of host-pathogen interactions in the wake of climate change - Increasing risk for shellfish associated infections?

The potential for climate-related spread of infectious diseases through marine systems has been highlighted in several reports. With this review we want to draw attention to less recognized mechanisms behind vector-borne transmission pathways to humans. We have focused on how the immune systems of edible marine shellfish, the blue mussels and Norway lobsters, are affected by climate related environmental stressors. Future ocean acidification (OA) and warming due to climate change constitute a gradually increasing persistent stress with negative trade-off for many organisms. In addition, the stress of recurrent hypoxia, inducing high levels of bioavailable manganese (Mn) is likely to increase in line with climate change. We summarized that OA, hypoxia and elevated levels of Mn did have an overall negative effect on immunity, in some cases also with synergistic effects. On the other hand, moderate increase in temperature seems to have a stimulating effect on antimicrobial activity and may in a future warming scenario counteract the negative effects. However, rising sea surface temperature and climate events causing high land run-off promote the abundance of naturally occurring pathogenic Vibrio and will in addition, bring enteric pathogens which are circulating in society into coastal waters. Moreover, the observed impairments of the immune defense enhance the persistence and occurrence of pathogens in shellfish. This may increase the risk for direct transmission of pathogens to consumers. It is thus essential that in the wake of climate change, sanitary control of coastal waters and seafood must recognize and adapt to the expected alteration of host-pathogen interactions.