Induction of apoptosis by UV in the flat oyster, Ostrea edulis

Apoptosis is a fundamental feature in the development of many organisms and tissue systems. It is also a mechanism of host defense against environmental stress factors or pathogens by contributing to the elimination of infected cells. Hemocytes play a key role in defense mechanisms in invertebrates and previous studies have shown that physical or chemical stress can increase apoptosis in hemocytes in mollusks. However this phenomenon has rarely been investigated in bivalves especially in the flat oyster Ostrea edulis. The apoptotic response of hemocytes from flat oysters, O. edulis, was investigated after exposure to UV and dexamethasone, two agents known to induce apoptosis in vertebrates. Flow cytometry and microscopy were combined to demonstrate that apoptosis occurs in flat oyster hemocytes. Investigated parameters like intracytoplasmic calcium activity, mitochondrial membrane potential and phosphatidyl-serine externalization were significantly modulated in cells exposed to UV whereas dexamethasone only induced an increase of DNA fragmentation. Morphological changes were also observed on UV-treated cells using fluorescence microscopy and transmission electron microscopy. Our results confirm the apoptotic effect of UV on hemocytes of O. edulis and suggest that apoptosis is an important mechanism developed by the flat oyster against stress factors.

Genome Wide Association Studies of Phagocytosis and the Cellular Immune Response in Drosophila melanogaster.

Phagocytosis of bacteria by specialized blood cells, known as hemocytes, is a vital component of Drosophila cellular immunity. To identify novel genes that mediate the cellular response to bacteria, we conducted three separate genetic screens using the Drosophila Genetic Reference Panel (DGRP). Adult DGRP lines were tested for the ability of their hemocytes to phagocytose the Gram-positive bacteria Staphylococcus aureus or the Gram-negative bacteria Escherichia coli. The DGRP lines were also screened for the ability of their hemocytes to clear S. aureus infection through the process of phagosome maturation. Genome-wide association analyses were performed to identify potentially relevant single nucleotide polymorphisms (SNPs) associated with the cellular immune phenotypes. The S. aureus phagosome maturation screen identified SNPs near or in 528 candidate genes, many of which have no known role in immunity. Three genes, dpr10, fred, and CG42673, were identified whose loss-of-function in blood cells significantly impaired the innate immune response to S. aureus. The DGRP S. aureus screens identified variants in the gene, Ataxin 2 Binding Protein-1 (A2bp1) as important for the cellular immune response to S. aureus. A2bp1 belongs to the highly conserved Fox-1 family of RNA-binding proteins. Genetic studies revealed that A2bp1 transcript levels must be tightly controlled for hemocytes to successfully phagocytose S. aureus. The transcriptome of infected and uninfected hemocytes from wild type and A2bp1 mutant flies was analyzed and it was found that A2bp1 negatively regulates the expression of the Immunoglobulin-superfamily member Down syndrome adhesion molecule 4 (Dscam4). Silencing of A2bp1 and Dscam4 in hemocytes rescues the fly’s immune response to S. aureus indicating that Dscam4 negatively regulates S. aureus phagocytosis. Overall, we present an examination of the cellular immune response to bacteria with the aim of identifying and characterizing roles for novel mediators of innate immunity in Drosophila. By screening panel of lines in which all genetic variants are known, we successfully identified a large set of candidate genes that could provide a basis for future studies of Drosophila cellular immunity. Finally, we describe a novel, immune-specific role for the highly conserved Fox-1 family member, A2bp1.

Exposure to toxic Alexandrium minutum activates the detoxifying and antioxidant systems in gills of the oyster Crassostrea gigas

Harmful algal blooms of Alexandrium spp. dinoflagellates regularly occur in French coastal waters contaminating shellfish. Studies have demonstrated that toxic Alexandrium spp. disrupt behavioural and physiological processes in marine filter-feeders, but molecular modifications triggered by phycotoxins are less well understood. This study analyzed the mRNA levels of 7 genes encoding antioxidant/detoxifying enzymes in gills of Pacific oysters (Crassostrea gigas) exposed to a cultured, toxic strain of A. minutum, a producer of paralytic shellfish toxins (PST) or fed Tisochrysis lutea (T. lutea, formerly Isochrysis sp., clone Tahitian (T. iso)), a non-toxic control diet, in four repeated experiments. Transcript levels of sigma-class glutathione S-transferase (GST), glutathione reductase (GR) and ferritin (Fer) were significantly higher in oysters exposed to A. minutum compared to oysters fed T. lutea. The detoxification pathway based upon glutathione (GSH)-conjugation of toxic compounds (phase II) is likely activated, and catalyzed by GST. This system appeared to be activated in gills probably for the detoxification of PST and/or extra-cellular compounds, produced by A. minutum. GST, GR and Fer can also contribute to antioxidant functions to prevent cellular damage from increased reactive oxygen species (ROS) originating either from A. minutum cells directly, from oyster hemocytes during immune response, or from other gill cells as by-products of detoxification.

Plejotropowe działanie neuropeptydów allatostatynowych w regulacji procesów fizjologicznych u chrząszczy

Wydział Biologii

Multibiomarker assessment of cadmium-based quantum dots effects in the marine mussel Mytilus galloprovincialis

Tese de Doutoramento, Ciências do Mar, da Terra e do Ambiente, Ramo: Ciências e Tecnologias do Ambiente, Especialização em Ecotoxicologia, Faculdade de Ciências e Tecnologia, Universidade do Algarve, 2016

Efeito da salinidade, densidade de estocagem e da infecção hipodermal e necrose hematopoiética (IHHN) na imunidade do camarão Litopenaeus vannamei cultivados em fazendas do Rio Grande do Norte

The main problem faced by the shrimp industry are the infectious diseases. The hypodermal and hematopoietic necrosis infection (IHHN) is one of the major cause of disease in the cultured shrimp, Litopenaeus vannamei. Environmental changes involving water quality, oxygen concentration, salinity, temperature, stocking density, presence of pathogens, among others, triggering a stressing condition for the cultured shrimp, weakening them and allowing the outbreak of diseases. The stress on the animal leads to a change in the molecules immune response components, which can be used as indicators of shrimp health. Thus, the objective of the present study was to evaluate the effect of salinity, stocking density and IHHNV infection on the L. vannamei shrimp. The immune parameters used to check the shrimp health were the total hemocytes counts (THC), the agglutinating activity (AA) and the clotting time (CT) of the serum of shrimp. These parameters were analyzed in healthy and IHHNV-infected shrimp, grown in low (0-0.5 ), medium (19-24 ) and high (> 38 ) salinity, and extensive (7-12 cam.m-2), semi-intensive (15-25 cam.m-2) and intensive (33-45 cam.m -2) stocking density. The IHHNV infection rate was significantly higher in low salinity (P<0.005) and intensive density (P<0.005), both stressful conditions for L. vannamei. Low salinity significantly increased THC (P<0.05) and decreased and CT (P<0.05) in healthy and infected shrimp, but AA (P<0.05) significantly decreased in healthy shrimp at medium salinity. Culture intensification did not affect the THC, AA and CT of healthy and infected shrimp (P>0.05). The IHHNV infection did not affect any immune parameters of shrimp cultured at different salinities and stocking densities. It is necessary to emphasize that this study was conducted in shrimp grown in ponds, where several environmental factors are acting simultaneously. Thus, further studies are needed about the influence of other environmental factors on the immune parameters of shrimp cultured in pond