In Silico Analysis of Pacific Oyster (Crassostrea gigas) Transcriptome over Developmental Stages Reveals Candidate Genes for Larval Settlement.

Following their planktonic phase, the larvae of benthic marine organisms must locate a suitable habitat to settle and metamorphose. For oysters, larval adhesion occurs at the pediveliger stage with the secretion of a proteinaceous bioadhesive produced by the foot, a specialized and ephemeral organ. Oyster bioadhesive is highly resistant to proteomic extraction and is only produced in very low quantities, which explains why it has been very little examined in larvae to date. In silico analysis of nucleic acid databases could help to identify genes of interest implicated in settlement. In this work, the publicly available transcriptome of Pacific oyster Crassostrea gigas over its developmental stages was mined to select genes highly expressed at the pediveliger stage. Our analysis revealed 59 sequences potentially implicated in adhesion of C. gigas larvae. Some related proteins contain conserved domains already described in other bioadhesives. We propose a hypothetic composition of C. gigas bioadhesive in which the protein constituent is probably composed of collagen and the von Willebrand Factor domain could play a role in adhesive cohesion. Genes coding for enzymes implicated in DOPA chemistry were also detected, indicating that this modification is also potentially present in the adhesive of pediveliger larvae.

Colonization of Polystyrene Microparticles by Vibrio crassostreae: Light and Electron Microscopic Investigation.

Microplastics collected at sea harbor a high diversity of microorganisms, including some Vibrio genus members, raising questions about the role of microplastics as a novel ecological niche for potentially pathogenic microorganisms. In the present study, we investigated the adhesion dynamics of Vibrio crassostreae on polystyrene microparticles (micro-PS) using electronic and fluorescence microscopy techniques. Micro-PS were incubated with bacteria in different media (Zobell culture medium and artificial seawater) with or without natural marine aggregates. The highest percentage of colonized particles (38-100%) was observed in Zobell culture medium, which may be related to nutrient availability for production of pili and exopolysaccharide adhesion structures. A longer bacterial attachment (6 days) was observed on irregular micro-PS compared to smooth particles (<10 h), but complete decolonization of all particles eventually occurred. The presence of natural marine agreggates around micro-PS led to substantial and perennial colonization featuring monospecific biofilms at the surface of the aggregates. These exploratory results suggest that V. crassostreae may be a secondary colonizer of micro-PS, requiring a multispecies community to form a durable adhesion phenotype. Temporal assessment of microbial colonization on microplastics at sea using imaging and omics approaches are further indicated to better understand the microplastics colonization dynamics and species assemblages.

Impaired photoacclimation in a kleptoplastidic dinoflagellate reveals physiological limits of early stages of endosymbiosis.

Dinophysis dinoflagellates are predators of Mesodinium ciliates, from which they retain only the plastids of cryptophyte origin. The absence of nuclear photosynthetic cryptophyte genes in Dinophysis raises intriguing physiological and evolutionary questions regarding the functional dynamics of these temporary kleptoplastids within a foreign cellular environment. In an experimental setup including two light conditions, the comparative analysis with Mesodinium rubrum and the cryptophyte Teleaulax amphioxeia revealed that Dinophysis acuminata possessed a smaller and less dynamic functional photosynthetic antenna for green light, a function performed by phycoerythrin. We showed that the lack of the cryptophyte nucleus prevented the synthesis of the phycoerythrin α subunit, thereby hindering the formation of a complete phycoerythrin in Dinophysis. In particular, biochemical analyses showed that Dinophysis acuminata synthesized a poorly stable, incomplete phycoerythrin composed of chromophorylated ß subunits, with impaired performance. We show that, consequently, a continuous supply of new plastids is crucial for growth and effective photoacclimation in this organism. Transcriptome analyses revealed that all examined strains of Dinophysis spp. have acquired the cryptophyte pebA and pebB genes through horizontal gene transfer, suggesting a potential ability to synthesize the phycobilin pigments bound to the cryptophyte phycoerythrin. By emphasizing that a potential long-term acquisition of the cryptophyte plastid relies on establishing genetic independence for essential functions such as light harvesting, this study highlights the intricate molecular challenges inherent in the enslavement of organelles and the processes involved in the diversification of photosynthetic organisms through endosymbiosis.

Enhanced scanning electron microscopy images using muscovite mica, an example with Rhizaria.

Muscovite mica sheets were used as a support to capture scanning electron microscopy pictures of marine biological samples. The physical properties of the cleaved muscovite mica provide a clean background, which greatly reduces the postprocessing of images, thereby enhancing them and resulting in impressive images. We chose siliceous Rhizaria for this investigation due to their morphological diversity and elaborate skeletons. RESEARCH HIGHLIGHTS: Muscovite mica sheet was used as a SEM support for biological sample Physical properties of muscovite mica sheet can provide native dark, gray, and white background of SEM images Muscovite mica sheet should be considered for image production in the context of art and science intersection.

First subcellular localization of the amnesic shellfish toxin, domoic acid, in bivalve tissues: Deciphering the physiological mechanisms involved in its long-retention in the king scallop Pecten maximus.

Domoic acid (DA), the phycotoxin responsible for amnesic shellfish poisoning (ASP), is an excitatory amino acid naturally produced by at least twenty-eight species of the bloom-forming marine diatoms Pseudo-nitzschia spp. Suspension feeders, such as bivalve mollusks, can accumulate and lengthy retain high amounts of DA in their tissues, threatening human health and leading to extensive-prolonged fishery closures, and severe economic losses. This is particularly problematic for the king scallop Pecten maximus, which retains high burdens of DA from months to years compared to other fast-depurator bivalves. Nonetheless, the physiological and cellular processes responsible for this retention are still unknown. In this work, for the first time, a novel immunohistochemical techniques based on the use of an anti-DA antibody was successfully developed and applied for DA-detection in bivalve tissues at a subcellular level. Our results show that in naturally contaminated P. maximus following a Pseudo-nitzschia australis outbreak, DA is visualized mainly within small membrane-bounded vesicles (1 - 2.5 µm) within the digestive gland cells, identified as autophagosomic structures by means of immune-electron microscopy, as well as in the mucus-producing cells, particularly those from gonad ducts and digestive tract. Trapping of DA in autophagososomes may be a key mechanism in the long retention of DA in scallops. These results and the development of DA-immunodetection are essential to provide a better understanding of the fate of DA, and further characterize DA contamination-decontamination kinetics in marine bivalves, as well as the main mechanisms involved in the long retention of this toxin in P. maximus.

Towards eco-friendly marine antifouling biocides - Nature inspired tetrasubstituted 2,5-diketopiperazines.

Marine biofouling plagues all maritime industries at vast economic and environmental cost. Previous and most current methods to control biofouling have employed highly persistent toxins and heavy metals, including tin, copper, and zinc. These toxic methods are resulting in unacceptable environmental harm and are coming under immense regulatory pressure. Eco-friendly alternatives are urgently required to effectively mitigate the negative consequence of biofouling without causing collateral harm. Amphiphilic micropeptides have recently been shown to exhibit excellent broad-spectrum antifouling activity, with a non-toxic mode of action and innate biodegradability. The present work focused on incorporating the pharmacophore derived from amphiphilic micropeptides into a 2,5-diketopiperazine (DKP) scaffold. This privileged structure is present in a vast number of natural products, including marine natural product antifoulants, and provides advantages of synthetic accessibility and adaptability. A novel route to symmetrical tetrasubstituted DKPs was developed and a library of amphiphilic 2,5-DKPs were subsequently synthesised. These biodegradable compounds were demonstrated to be potent marine antifoulants displaying broad-spectrum activity in the low micromolar range against a range of common marine fouling organisms. The outcome of planned coating and field trials will dictate the future development of the lead compounds.

Subclavian versus femoral arterial cannulations during extracorporeal membrane oxygenation: A propensity-matched comparison.

BACKGROUND: During peripheral extracorporeal veno-arterial membrane oxygenation (VA-ECMO) support, subclavian arterial cannulation provides, in comparison to femoral arterial cannulation, an anterograde flow which may prevent from left ventricular (LV) distention and improve outcomes. We aimed to compare the effectiveness of subclavian cannulation to femoral cannulation in reducing LV overdistension consequences, hemostatic complications and mortality. METHODS: This retrospective study conducted in two intensive care units of the Lille academic hospitals from January 2013 to December 2019 included 372 non-moribund adult patients supported by VA-ECMO. The primary endpoint was a new onset of pulmonary edema (PO) or LV unloading. Secondary endpoints were myocardial recovery, serious bleeding (according to Extracorporeal Life Support Organization definition), thrombotic complications (a composite of stroke, cannulated limb or mesenteric ischemia, intracardiac or aortic-root thrombosis) and 28 day mortality. Differences in outcomes were analyzed using propensity score matching (PSM) and inverse probability of treatment weighting adjustment (IPTW). RESULTS: As compared to femoral cannulation (n = 320 patients), subclavian cannulation (n = 52 patients) did not reduce the occurrence of new onset of PO or LV unloading after PSM [HR 0.99 (95% CI 0.51-1.91)]. There was no other difference in outcomes in PSM cohort. In IPTW adjustment cohort, subclavian cannulation was associated with reduced recovery and increased serious bleeding with four accidental decannulations observed. CONCLUSION: Subclavian artery cannulation was not associated with reduced LV distension related complications, thrombotic complications and 28 day mortality. Rather, it may increase serious bleeding and accidental decannulations, and reduce recovery. Therefore, subclavian cannulation should be limited to vascular accessibility issues.

Antifouling Activity of Meroterpenes Isolated from the Ascidian Aplidium aff. densum.

The settlement and growth of fouling organisms on man-made surfaces can be prevented by the application of antifouling paints containing active compounds (biocides, heavy metals), most of which are toxic to non-target organisms. As part of our research program in chemical ecology and blue biotechnology, we are conducting studies to investigate the natural defence mechanisms of marine organisms that are free from epibionts, with the aim of isolating molecules involved in surface defence that could be good candidates as antifouling agents. Ascidians were selected for our investigation because previous studies have shown that they contain abundant and diverse secondary metabolites, which play a defensive role and have been applied to drug discovery. It is therefore relevant to study the role of such secondary metabolites in surface protection. In this study, 5 meroterpenoids (cordiachromene A, didehydroconicol, epiconicol, methoxyconidiol, conidione) from Aplidium aff. densum (ascidian) were investigated as potential antifoulants towards the inhibition of bacterial growth and settlement inhibition of barnacles. Cardiochromene A (IC50 barnacle settlement = 6.04 µg/mL; MIC Gram positive = 125 µg/mL; MIC Gram negative = 32 µg/mL) and epiconicol (IC50 barnacle settlement = 8.05 µg/mL; MIC Bacillus = 63 µg/mL; MIC other strains = 32 µg/mL) were the most promising compounds among those tested in this study.

Evaluation of Anti-Activated Factor X Activity and Activated Partial Thromboplastin Time Relations and Their Association with Bleeding and Thrombosis during Veno-Arterial ECMO Support: A Retrospective Study.

BACKGROUND: We aimed to investigate the relationship between anti-activated Factor X (anti-FXa) and activated Partial Thromboplastin Time (aPTT), and its modulation by other haemostasis co-variables during veno-arterial extracorporeal membrane oxygenation (VA-ECMO) support. We further investigated their association with serious bleeding and thrombotic complications. METHODS: This retrospective single-center study included 265 adults supported by VA-ECMO for refractory cardiogenic shock from January 2015 to June 2019. The concordance of anti-FXa and aPTT and their correlations were assessed in 1699 paired samples. Their independent associations with serious bleeding or thrombotic complications were also analysed in multivariate analysis. RESULTS: The concordance rate of aPTT with anti-FXa values was 50.7%, with 39.3% subtherapeutic aPTT values. However, anti-FXa and aPTT remained associated (ß = 0.43 (95% CI 0.4-0.45) 10-2 IU/mL, p < 0.001), with a significant modulation by several biological co-variables. There was no association between anti-FXa nor aPTT values with serious bleeding or with thrombotic complications. CONCLUSION: During VA-ECMO, although anti-FXa and aPTT were significantly associated, their values were highly discordant with marked sub-therapeutic aPTT values. These results should favour the use of anti-FXa. The effect of biological co-variables and the failure of anti-FXa and aPTT to predict bleeding and thrombotic complications underline the complexity of VA-ECMO-related coagulopathy.

Author Correction: Proteinaceous secretion of bioadhesive produced during crawling and settlement of Crassostrea gigas larvae.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The dinoflagellate Alexandrium minutum affects development of the oyster Crassostrea gigas, through parental or direct exposure.

Harmful algal blooms are a threat to aquatic organisms and coastal ecosystems. Among harmful species, the widespread distributed genus Alexandrium is of global importance. This genus is well-known for the synthesis of paralytic shellfish toxins which are toxic for humans through the consumption of contaminated shellfish. While the effects of Alexandrium species upon the physiology of bivalves are now well documented, consequences on reproduction remain poorly studied. In France, Alexandrium minutum blooms have been recurrent for the last decades, generally appearing during the reproduction season of most bivalves including the oyster Crassostrea gigas. These blooms could not only affect gametogenesis but also spawning, larval development or juvenile recruitment. This study assesses the effect of toxic A. minutum blooms on C. gigas reproduction. Adult oysters were experimentally exposed to A. minutum, at environmentally realistic concentrations (102 to 103 cells mL-1) for two months during their gametogenesis and a control group, not exposed to A. minutum was fed with a non-toxic dinoflagellate. To determine both consequences to next generation and direct effects of A. minutum exposure on larvae, the embryo-larval development of subsequent offspring was conducted with and without A. minutum exposure at 102 cells mL-1. Effects at each stage of the reproduction were investigated on ecophysiological parameters, cellular responses, and offspring development. Broodstock exposed to A. minutum produced spermatozoa with decreased motility and larvae of smaller size which showed higher mortalities during settlement. Embryo-larval exposure to A. minutum significantly reduced growth and settlement of larvae compared to non-exposed offspring. This detrimental consequence on larval growth was stronger in larvae derived from control parents compared to offspring from exposed parents. This study provides evidence that A. minutum blooms, whether they occur during gametogenesis, spawning or larval development, can either affect gamete quality and/or larval development of C. gigas, thus potentially impacting oyster recruitment.

Proteinaceous secretion of bioadhesive produced during crawling and settlement of Crassostrea gigas larvae.

Bioadhesion of marine organisms has been intensively studied over the last decade because of their ability to attach in various wet environmental conditions and the potential this offers for biotechnology applications. Many marine mollusc species are characterized by a two-phase life history: pelagic larvae settle prior to metamorphosis to a benthic stage. The oyster Crassostrea gigas has been extensively studied for its economic and ecological importance. However, the bioadhesive produced by ready to settle larvae of this species has been little studied. The pediveliger stage of oysters is characterized by the genesis of a specific organ essential for adhesion, the foot. Our scanning electron microscopy and histology analysis revealed that in C. gigas the adhesive is produced by several foot glands. This adhesive is composed of numerous fibres of differing structure, suggesting differences in chemical composition and function. Fourier transformed infrared spectroscopy indicated a mainly proteinaceous composition. Proteomic analysis of footprints was able to identify 42 proteins, among which, one uncharacterized protein was selected on the basis of its pediveliger transcriptome specificity and then located by mRNA in situ hybridization, revealing its potential role during substrate exploration before oyster larva settlement.