Metallothionein gene identification and expression in the cockle ( Cerastoderma edule) under parasitism (trematodes) and cadmium contaminations

Parmi les organismes benthiques, les bivalves sont souvent utilisés comme bioindicateurs des pollutions environnementales, du fait de leur importante capacité de bioaccumulation des métaux lourds conduisant à l'induction des métallothionéines (MT). Les MT sont de petites protéines cytosoliques capables de fixer les métaux et qui sont impliquées dans l'homéostasie et la détoxication de ces métaux chez les organismes. Ces protéines peuvent être induites par un grand nombre de facteurs tels que les hormones, les stress physiques, le parasitisme. La quantification des MT en relation avec le parasitisme est rarement reportée dans la littérature, bien que le parasitisme soit omniprésent et très délétère chez les bivalves. De plus, seulement un petit nombre de gènes de MT ont été identifiés chez les mollusques. Cette étude décrit la caractérisation de la séquence partielle d'un gène de MT (Cemt1) chez la coque Cerastoderma edule, qui présente plus de 80 % d'homologie avec de nombreuses séquences de MT de bivalves. A partir de cette séquence, des amorces spécifiques, utilisables en PCR quantitative en temps réel, ont été déterminées. Le niveau d'expression du gène Cemt1, ainsi que la quantité de protéine MT ont été analysés chez des individus soumis à différents paramètres : sans parasites ou infestés par le trématode digène Himasthla elongata, et lors d'expositions au cadmium à 15 µg Cd L−1. Des résultats concordants ont été obtenus lors de la quantification des protéines MT et de l'analyse de l'expression du gène Cemt1. Ceux-ci mettent en évidence que les concentrations en MT augmentent significativement par les deux types de traitement (infestation parasitaire et exposition au cadmium).Among benthic organisms, bivalves are often used as bioindicators of environmental pollution because of their high bioaccumulation capacities for heavy metals leading to metallothioneins (MT) induction. MT are small cytosolic metal-binding proteins involved in metal homeostasis and detoxification in living organisms. These proteins can also be induced by a wide range of factors, such as hormones, physical stress, parasitism. MT quantification in relation to parasitism is rarely reported in literature, while parasites are omnipresent and have deleterious impacts on bivalves. Moreover, only a few number of MT genes have been characterized in molluscs. This study describes the partial sequence of the MT gene (Cemt1) in the edible cockle Cerastoderma edule. The cockle's MT cDNA was sequenced and showed over 80% homology to several other bivalve MT sequences. This sequence was then used to determine MT specific primers which can be used in quantitative real time PCR. MT protein and gene expression levels were quantified for individuals selected under different conditions: free from or infected by the digenean trematode Himasthla elongata, and under cadmium exposure at 15 mu g Cd L-1. Results evidenced that MT concentrations were significantly increased by both treatments; parasite infection and Cd exposure. Moreover, congruent results between MT protein and gene expression levels were obtained

Cockle Cerastoderma edule population mortality: Role of the digenean parasite Himasthla quissentensis

Infection patterns of the digenean trematode Himasthla quissetensis in its second intermediate host, the edible cockle Cerastoderma edule, and the impact of this parasite on cockle populations were quantified. A 2 yr transplant experiment and a 4 yr survey were conducted in Arcachon Bay, a coastal lagoon on the Atlantic SW coast of France. Cockles free of H. quissetensis were transplanted at La Canelette, a station where cockles were usually highly infected. Infection began in May, when the water temperature reached 17degreesC and stopped in November (12degreesC). There was no new infection between November and April. Monthly surveys at 2 stations, Arguin and La Canelette, confirmed that cockle infection by H. quissetensis occurred during the warmer period of the year. Only cockles with a shell length > 8 mm became infected; therefore, cockle growth rate, which differed at the 2 stations, was also a key factor explaining infection phenology. In winter, the mean parasite abundance of H quissetensis in cockles decreased at both stations. The mortality rate of parasites in cockles was very low, with no seasonal pattern, and did not explain this winter decrease. Consequently, the decrease of H. quissetensis was interpreted as mortality of heavily infected cockles. At Arguin, where the cockle population was characterised by moderate abundance, high growth rate and short lifespan, cockle mortality amounted to 93% during the first year of benthic life, with no significant contribution of parasites; in the following year (Year 2), cockle mortality was 85%, with 28% due to H. quissetensis infection. At La Canelette, the cockle population, characterised by low abundance and low growth rate, suffered almost 100% mortality in the first year, with no contribution by parasites, which started to infect cockles in the second year only; the remaining cockles had a long lifespan (up to 5 yr), and > 46% of their mortality was linked to H. quissetensis infection. It is concluded that both host growth rate and water temperature are important factors in the initiation of parasite infection and that the intensity of infection and its effect on host mortality closely depend on host growth and environmental factors

First Record and Description of Metacercariae of Curtuteria Arguinae N. Sp. (Digenea: Echinostomatidae), Parasite of Cockles Cerastoderma Edule (Mollusca: Bivalvia) in Arcachon Bay, France

International audienceA new Himasthlinae species, Curtuteria arguinae, is described as metacercariae from the cockle Cerastoderma edule (L.), collected at Banc d'Arguin (southwestern France). These metacercariae encysted preferentially in the mantle and also in the foot of cockles. Encysted and chemically excysted metacercariae were studied by light microscopy and scanning electron microscopy, respectively. Excysted metacercariae were elongated and curved ventrally. They bore a 33-spine circumoral collar. Sensory papillae were arranged around the oral sucker and also symmetrically along the ventral surface body, from the collar to the acetabulum. The dorsal and ventral tegument surfaces were densely packed with similar pointed spines. The posterior end of the body was without any spines. Among the Curtuteria species described previously, only Curtuteria haematopodis Smogorjewskaja and Iskova, 1966 had the same number of circumoral collar spines. A 6-yr field survey showed that the cockle population at Banc d'Arguin was subjected to a summer infection of C. arguinae. Curtuteria arguinae phenology of infection is characterized by interannual variability and seasonality (beginning in July–August and maximum in autumn). The first intermediate and final hosts remain unknown

Cockle infection by Himasthla quissetensis – II. The theoretical effects of climate change

Numerous marine populations experience parasite pressure. This is the case of the cockles Cerastoderma edule which are often highly infected by trematode macroparasites. These parasites display a complex life cycle, with a succession of free-living and parasitic stages. Climate, and in particular temperature, is an important modulator of the transmission dynamics of parasites. Consequently, global change is thought to have implications for the epidemiology of infectious diseases. Using Himasthla quissetensis, a dominant parasite of cockles as 2nd intermediate host in Arcachon Bay (France), we used mathematical models of parasite emergence (cercariae) and parasite infection (metacercariae) in cockles as a function of water temperature, in order to study different scenarios of temperature increases. Globally, with a + 0.5 °C to + 6.0 °C simulation, cumulated emergence of cercariae and accumulation of metacercariae tended to decrease or stagnate, respectively. This is the consequence of a trade-off between sooner (spring) and later (autumn) cercariae emergence/infestation on one hand, and a longer inhibition period of cercariae emergence/infestation during the hottest days in summer. Using sea water temperature in Oualidia (Morocco) where mean annual sea temperature is 3 °C higher than in Arcachon Bay, our model predicted infestation all year long (no seasonality). The model gave a correct estimation of the total number of parasites that was expected in cockles. Conversely, observed infestation in Oualidia followed a seasonal pattern like in Arcachon Bay. These results suggest that, if temperature is a strong driver of parasite transmission, extrapolation in the framework of climate change should be performed with caution

Effect of intermediate host size (Cerastoderma edule) on infectivity of cercariae of Himasthla quissetensis (Echinostomatidae: Trematoda)

The edible cockle (Cerastoderma edule), a common marine bivalve in semi-sheltered sandflats, is a natural host for many parasite species of the genus Himasthla (Echinostomatidae: Trematoda). In a previous paper, Wegeberg et al. (1999) showed segregation of cockle infection by three Himasthla species (H. interrupta, H. continua, H. elongata) in relation to host tissue (foot, mantle, siphon) and host size (1·5 to 14 mm shell length). Following the same experimental procedure, a fourth species, H. quissetensis, an introduced dominant parasite of cockles in Arcachon Bay (south-west France) was investigated. The infection pattern was very similar to the patterns shown by H. elongata and H. continua. Cercariae (the free-living stage shed from prosobranch snails and encysting as metacercariae in bivalves) were most successful in the shell-length range of 6–14 mm, where 74% of the added cercariae were recovered as metacercariae. The comparison with the other Himasthla species supports the previous conclusion that the efficiency of cercariae to infect cockles depends on host size

Gene transfer strategy to promote axonal regeneration in the injured spinal cord : towards clinical application

International audienceIn spinal cord injury (SCI), reactive astrocytes build-up a dense cellular structure, the glial scar, that forms both physical and chemical barriers considered to be detrimental for axonal regeneration. Reactive astrocytes present hypertrophic processes resulting mainly from increased expression of glial fibrillary acidic protein (GFAP) and vimentin, two astrocyte cytoskeletal intermediate filament proteins. It had been shown, in mice knockout for both GFAP and vimentin, that due to the absence of glial reactivity, reinnervation and partial motor recovery occurred after lateral spinal cord hemisection. In order to translate this proof of principle into a therapeutic strategy, we have developed an experimental approach based on the use of siRNA carried by lentiviral vectors. We have developed lentiviral vectors Lv-siGFAP and Lv-siVIM which inhibit the in vitro formation of a glial scar in a Scratch model of lesion. Here we report that in a mouse model of SCI, intraspinal application of Ientiviral vectors Lv-shGFAP and Lv-shVIM efficiently reduces glial reactivity by silencing GFAP alone or associated with vimentin, permits axonal regrowth and improves functional motor recovery. With the objective of translation to clinics, we are adapting the work performed on mice to a large animal model (domestic swine), in order to mimic the clinical settings. We thus first designed and generated lentiviral vectors Lv-siGFAPsw able to inhibit GFAP expression in swine astrocytes. In parallel, we started to define the experimental surgical procedures of lentiviral transduction in the swine injured spinal cord by hemisection