Silvia Battistella

Mediterranean Mussel Gene Expression Profile Induced by Okadaic Acid Exposure

Seasonal seawater temperature increases define optimal growth conditions for Dinoflagellate species which can reach high concentrations in water column and also in filter-feeding organisms like Mytilus galloprovincialis. Commonly produced by Dinophysis and Prorocentrum spp., okadaic acid (OA) and its analogues are responsible for the Diarrheic Shellfish Poisoning (DSP) syndrome in humans. Closure of shellfishing grounds is therefore recommended by the EU when DSP toxin levels in shellfish exceed 16 μg OA 100 g(-1) flesh. Despite not being responsible for casualties either in humans or mussels, DSP outbreaks are considered natural events causing health and economic issues due to the frequency of their occurrence. Since gene expression studies offer a wide range of different solutions, we used a mussel cDNA microarray to evaluate gene expression changes in the digestive gland of mussels fed for five weeks with OA-contaminated nutrient. Among the differentially expressed genes we observed a general up-regulation of transcripts coding for stress proteins, proteins involved in cellular synthesis, and a few not annotated proteins. Overall, at the first time point analyzed we identified 58 candidate transcripts for OA-induced stress in mussels, half of which have unknown function. In this paper we present the first gene expression analysis performed on Mediterranean mussels exposed to okadaic acid. The characterization of these transcripts could be useful for the identification of an early physiological response to OA exposure.

Hemocytes and immunological reactions in crustaceans

Abstract The role of lectins in the cellular immune response in crustaceans is comparatively reviewed and, in particular, the role of proPO‐system in immunological mechanisms is discussed.

Apis mellifera ligustica, Spinola 1806 as bioindicator for detecting environmental contamination: a preliminary study of heavy metal pollution in Trieste, Italy

Honeybees have become important tools for the ecotoxicological assessment of soil, water and air metal contamination due to their extraordinary capacity to bioaccumulate toxic metals from the environment. The level of heavy metal pollution in the Trieste city was monitored using foraging bees of Apis mellifera ligustica from hives owned by beekeepers in two sites strategically located in the suburban industrial area and urban ones chosen as control. The metal concentration in foraging bees was determined by inductively coupled plasma-mass spectrometry. The chemical analysis has identified and quantified 11 trace elements accumulated in two different rank orders: Zn> Cu > Sr > Bi > Ni > Cr > Pb = Co > V > Cd > As in foraging bees from the suburban site and Zn > Cu > Sr > Cr > Ni > Bi > Co = V > Pb > As > Cd in bees from urban site. Data revealed concentrations of Cr and Cu significantly higher and concentration of Cd significantly lower in bees from urban sites. The spatial difference and magnitude order in heavy metal accumulation along the urban-suburban gradient are mainly related to the different anthropogenic activity within sampled sites and represent a risk for the human health of people living in the city. We discussed and compared results with the range of values reported in literature.

Immune challenges trigger cellular and humoral responses in adults of Pterostichus melas italicus (Coleoptera, Carabidae).

The present study focuses on the ability of Pterostichus melas italicus Dejean to mount cellular and humoral immune responses against invading pathogens. Ultrastructural analyses revealed the presence of five morphologically distinct types of hemocytes: prohemocytes, plasmatocytes, granulocytes, oenocytoids and macrophage-like cells. Differential hemocyte counts showed that plasmatocytes and granulocytes were the most abundant circulating cell types and plasmatocytes exhibited phagocytic activity following the latex bead immune challenge. Macrophage-like cells were recruited after the immune challenge to remove exhausted phagocytizing cells, apoptotic cells and melanotic capsules formed to immobilize the latex beads. Total hemocyte counts showed a significant reduction of hemocytes after latex bead treatment. Phenoloxidase (PO) assays revealed an increase of total PO in hemolymph after immune system activation with lipopolysaccharide (LPS). Moreover, the LPS-stimulated hemocytes showed increased protein expression of inducible nitric oxide synthase, indicating that the cytotoxic action of nitric oxide was engaged in this antimicrobial collaborative response. These results provide a knowledge base for further studies on the sensitivity of the P. melas italicus immune system to the environmental perturbation in order to evaluate the effect of chemicals on non-target species in agroecosystems.

Identification of an induced lectin involved in capsule formation in Squilla mantis (Crastacea, Stomatopoda)

Abstract The process of capsule formation in Squilla mantis involves the direct action of granulocytes in the recognition of foreign bodies. Evidence was presented that a relevant role in this recognition is played by a lectin (lect‐I), which is synthesized and secreted by granulocytes after the activation of the immunologic reaction.

Recognition of a 31,000 molecular weight protein synthesized by the mussel Mytilus galloprovincialis Lmk. during Dinophysis spp. algal bloom in the Gulf of Trieste (upper Adriatic Sea)

During Dinophysis spp. bloom, a histological alteration in the digestive gland of Mytilus galloprovincialis Lmk. occurs. Electrophoretic analysis of the water-soluble proteins of the hepatopancreas showed the presence of a low mol. wt protein, which appeared after the ingestion of algae by mussels. The probable relationship between the toxicity of Dinophysis and the synthesis of the new protein by mussels prompted us to produce a monoclonal antibody against this protein because it should be useful for the identification of mussel toxicity.

Effects of X-ray irradiation on haemocytes of Procambarus clarkii (Arthropoda: Decapoda) males

Abstract The red swamp crayfish Procambarus clarkii is an invasive alien species spreading worldwide. The sterile male release technique (SMRT) is among the methods used to contrast the growth of P. clarkii populations within invaded areas. In this study males underwent X-ray sterilisation with a dose of 40 Gy and their immunocompetence was analysed in comparison to untreated animals to ascertain whether radiation can affect welfare parameters other than reproductive organs. The present research investigated the immune function in P. clarkii males in term of (1) morphological haemocyte characterisation by transmission electron microscopy to identify the main phagocyting haemocyte after in vivo artificial non-self challenge with latex beads; (2) total and differential haemocyte counts; and (3) basal and total phenoloxidase activities as components of the humoral defence. Three types of circulating haemocytes were characterised via transmission electron microscopy: hyaline, semigranular and granular haemocytes. The ultrastructural features of haemocyte granules allowed the characterisation of a fourth type of haemocyte, the medium granule haemocyte. In vivo artificial non-self-challenge with latex beads identified the semigranular haemocytes as primarily involved in phagocyting activity. Circulating haemocytes of males irradiated with a dose of 40 Gy, after 20 days, showed a significantly lower diameter in the granules of hyaline and semigranular haemocytes, but no other evident ultrastructural alterations in comparison with un-irradiated animals were found. Irradiated males showed a significant decrease of about 80% of circulating haemocytes and an increase in frequency of semigranular and granular haemocytes. No significant differences in basal and total phenoloxidase activity were recorded and this could, in part, explain the good survival level of irradiated males despite the drastic decline of the haemocyte number. This study represents the basis to appraise whether SMRT affects important functions, such as those of the immune system, in addition to altering the gonad tissue.

Crustacean Immunity: The Modulation of Stress Responses

Abstract This chapter provides an overview of the main stress responses modulated by neuropeptides of the crustacean hyperglycemic hormone (CHH) family. The current scenario of upstream regulation of CHHs is presented here. Numerous sequence variants, splicing-isoforms, and posttranslational modifications of CHHs point to a wide functional diversification of these peptides. Other than the classical main CHH-mediated cascade resulting in energy mobilization, new aspects focused on CHH chirality are demanding deeper efforts on the characterization of their target and mode of action. Moreover, new CHH variants with immune-related functions have been recently described, broadening further the role of such a neuropeptides family.