Hox and paraHox genes in bivalve molluscs

In this study, we sought the presence and analysed the sequences of the Hox and ParaHox genes in bivalve molluscs. The clustered Hox genes play a central role in anterior-posterior axial patterning in bilaterian metazoa, whereas the ParaHox gene cluster is a paralogue (evolutionary sister) of the Hox cluster.Using polymerase chain reaction (PCR)-based approaches, we isolated nine different sequences in five species belonging to three of the main bivalve subclasses: Ensis ensis and Tapes philippinarum (Heterodonta), Pecten maximus and Mytilus galloprovincialis (Pteriomorphia), and Yoldia eightsi (Protobranchia). Comparison with the Hox and ParaHox genes of other bilaterians, particularly lophotrochozoans, allowed us to attribute six of these sequences to the Hox gene cluster (one to paralog group [PG] 3 class, and five to the central class), two to the ParaHox cluster and one to the Gbx gene family.The results of our investigation seem to indicate that homeotic Hox and ParaHox gene clusters are homogeneous for both presence and characteristics in molluscs.     

The role of bivalve molluscs as tools in estuarine sediment toxicity testing: a review

Estuarine sediments frequently are repositories and therefore potential sources of anthropogenic contaminants. Many organic and metallic chemical compounds released into aquatic systems bind to particulates and so accumulate in the sediments, thus, sediments become repositories of contaminants in estuaries. These may also cause contamination through diffusion of porewater, resuspension of particulates and dispersal of benthic fauna. There is a need to assess the biological affects of these anthropogenic contaminants because they may be toxic to infauna and bottomfish. Sediment toxicity bioassays are a means for carrying out such an assessment and primarily provide data on toxicity by measuring the effects on the test organism. Existing sediment toxicity bioassays rely on a battery of aquatic toxicity tests, which are based on the extraction of pore water, and elutriate from sediments and then subjecting these sediment phases to toxicity testing regimes. Two estuarine bivalve molluscs, Scrobicularia plana and Tapes semidecussatus were used to assess the ecotoxicity of field-collected sediments from estuarine and coastal areas around the Irish and English Coast over a 3-year study period. A variety of endpoints were measured during the study including survival in air, behaviour, animal condition, biochemistry, soft tissue metal concentrations, lysosomal membrane integrity and histopathology. Of these endpoints, the most sensitive were survival, survival in air, lysosomal membrane integrity, behaviour and histopathology.     

Podocytes in bivalve molluscs: Morphological evidence for ultrafiltration

Summary In representatives from a survey of three major taxa of bivalves the pericardial glands were found in two distinct positions. In protobranches (Acila castrensis) and filibranch bivalves (Glycymeris subobsoleta, Chlamys hastata, Pecten caurinus, Placopecten magellanicus, Mytilus edulis andMytilus californianus) the pericardial glands are located on the auricular surface. In heterodonts (Mercenaria mercenaria, Clinocardium nuttallii andMya arenaria) the pericardial glands are found in an anterodorsal position to the pericardial cavity.The sites of ultrafiltration are described. They consist of podocytes with basally extending pedicels forming an interdigitating network apposed to a basal lamina. Other characteristics of this ultrafiltration barrier described are anionic sites on the basal lamina and presence of substructural components within the ultrafiltration slits between pedicels.The pathway for the ultrafiltrate in protobranchs and filibranchs is from the hemocoel through the basal lamina, through the ultrafiltration slits of the pedicel network, into the urinary spaces between the podocyte cell bodies and into the pericardial cavity. The pathway for the ultrafiltrate in heterodonts is from the hemocoel through the basal lamina, through the ultrafiltration slits of the pedicel network, into urinary spaces between the podocyte cell bodies, into the lumen of the pericardial gland tubules and into the pericardial cavity.All podocyte cells have electron dense granules, Golgi apparatus and vacuoles associated with their cytoplasm. Heterodont species have microvilli on the cell surfaces of the podocytes apposed to urinary spaces.In all cases the morphological sites of ultrafiltration were associated with the pericardial glands of the heart-pericardial complex.     

Environmental contaminants influencing immunefunction in marine bivalve molluscs

The increased observation of pollution induced disease conditions in marine organisms has led to a growing interest on the effects of environmental contaminants on the immune system. Most studies on modulation of the immune system in bivalves by pollutants have concentrated on the effects of heavy metals and polycyclic aromatic hydrocarbons (PAHs). The current literature on contaminant effects on specific components of the bivalve immune system is reviewed together with the effects on susceptibility to infection. Data are presented showing the effects on immune parameters of exposure to Vibrio tubiashi following pre-exposure to copper or cadmium. Mussels exposed to cadmium for 7 days followed by 7 days exposure to V. tubiashi demonstrated significantly higher numbers of circulating haemocytes compared with non-Vibrio-exposed groups. Similar experiments conducted with copper exposure for both 7 days and 7 weeks followed by V. tubiashi for 7 days demonstrated a significant decrease in the percentage of circulating eosinophils compared with basophilic cells for both short and long term exposures. The intracellular release of superoxide (NBT reduction) by haemocytes was stimulated in Vibrio-challenged mussels with no copper pre-exposure but was significantly reduced in mussels pre-exposed to 0·2 ppm of copper for 7 weeks. The mortalities for the copper experiments showed increased levels with increasing copper concentration and were consistently higher in the V. tubiashi challenged mussels which had also been exposed to copper.     

Multilocus genetic analysis of Cryptosporidium in naturally contaminated bivalve molluscs

AIMS: To evaluate the application of discriminatory multilocus PCR procedures for the characterization of Cryptosporidium in samples of naturally contaminated bivalve molluscan shellfish. METHODS AND RESULTS: Nucleic acid was extracted from 22 shellfish previously identified as contaminated with Cryptosporidium spp. and subjected to PCR-based analysis for two independent fragments of the Cryptosporidium oocyst wall protein (COWP) gene, three microsatellite markers (ML 1, GP 15 and MS 5) and an extra-chromosomal small double-stranded RNA (dsRNA). Overall, at least one COWP gene fragment was amplified from all 22 samples, 21 amplified the dsRNA and 14 at least one of the three microsatellite loci. More than one dsRNA or microsatellite allele was detected in 50% of samples. The majority of samples were contaminated with Cryptosporidium parvum types circulating in both humans and livestock. A novel dsRNA element was identified in one sample, which did not amplify any of the three microsatellite loci investigated. CONCLUSIONS: Multilocus analysis of Cryptosporidium can be applied to DNA extracted from naturally contaminated shellfish. SIGNIFICANCE AND IMPACT OF STUDY: This multilocus genetic analysis highlights that filter feeder molluscs are a potential source of cryptosporidial oocysts, which may be infectious to humans.     

Protein gradients in byssal threads of some marine bivalve molluscs

Many marine bivalve molluscs produce byssal threads for attachment to solid substrata. Small (less than 10 mm) consecutive sections of the byssal threads of Mytilus edulis, M. californianus, Geukensia demissa, Atrina vexillum, and A. rigida were analyzed by amino acid analysis to determine if chemical composition remains constant as a function of location in thread segments. Nonlinear longitudinal protein gradients, probably involving collagen and an elastic protein, were found in the Mytilus species. In these, collagen peaks in the distal third of the thread. In Geukensia and the Atrina species, although the two differed greatly in composition, there is a clear nonvariability in composition of the thread within each species as a function of location in the thread. The adhesive plaque at the tip of the thread of all species examined differs substantially in composition from the remainder of the thread. Protein gradients in the threads of some bivalves may reflect specific adaptations evolved to respond to exposed habitats in high-energy environments.     

Anaerobic metabolism of bivalve molluscs during exposure to air

The anaerobic metabolism of Mytilus edulis, Cardium edule, Scrobicularia plana and Macoma balthica was investigated. On exposure to the atmosphere, all of these species were found to be able to utilise ¹⁴CO₂. This suggests that these species gape during exposure to the atmosphere. A comparative study on the pattern of ¹⁴C incorporation suggests that there is a similarity between M. edulis and S. plana in the extent of the utilization of the succinate pathway during exposure to air. However C. edule and M. balthica were also similar in the extent of utilizing the succinate pathway, even though there were significant species differences between the similar animals. It is suggested that exposure in S. plana represents a stressful situation and that this species might react to this stress by utilizing the succinate pathway. A higher incorporation of radiocarbon into alanine by M. balthica could be due to high activity of the enzymes that control the reactions leading to production of radioactive pyruvate.     

Analysis of different methods to estimate fecundity in bivalve molluscs

Abstract

We analyzed the advantages and disadvantages of three methods to estimate potential fecundity in the bivalve molluscs Pinctada mazatlanica and Atrina maura, both species having commercial value in northwestern Mexico. Data of gonad samples collected during the breeding season in March 1999 (P. mazatlanica) and March 2003 (A. maura) were processed with histological techniques combined with Cavalieri’s Principle; stereological analysis of the gonad with the caliper method; and estimation of the theoretical radius of oocytes. These estimates were compared to a reference value obtained from direct counts of the number of oocytes stripped from the gonad. Compared to other methods, potential fecundity determined with the caliper method was more accurate in both species: A. maura (9.8–15 × 10⁶ cells ind^?1 and P. mazatlanica (10.8–17 × 10⁶ cells ind^?1). The potential of the caliper method, combined with image analysis software may offer accurate estimates of aspects of reproduction in different bivalve species, which has direct applications in hatcheries and conservation programs.     

The adenosine triphosphate content of some marine bivalve molluscs

The total ATP-content of representatives of 23 bivalve families was found to vary from 1.26–0.26 % of the dry tissue weight, with the dry tissue containing about 40 % carbon. Representatives from certain families consistently had more than average and others less than average ATP values, and there were greater differences between species from different families than between closely related species. Variations in the oxygen uptake of whole animals accounted for little of the observed variation in ATP-content. The ATP-content of individual tissues from the same individual showed wide variation, as did that of the same tissue from different species. The highest values of ATP were found in muscle tissues. High maintained levels of ATP were associated with the ability of the species to use energy rapidly for short periods, for example, in escape responses.     

Detection of Cryptosporidium oocysts in bivalve molluscs destined for human consumption

Clams (Dosinia exoleta, Ruditapes philippinarum, Venerupis pullastra, Venerupis rhomboideus, Venus verrucosa), mussels (Mytilus galloprovincialis), and oysters (Ostrea edulis) were tested for the presence of Cryptosporidium sp. oocysts using various stain techniques and a commercially available kit containing fluorescein isothiocyanate-conjugated monoclonal antibodies. All molluscs were harvested in northwest Spain (Galicia) except for R. philippinarum, which was from Italy, and 1 of the 6 oyster samples, which was from England. The results showed the presence of Cryptosporidium sp. oocysts in all of the molluscan species destined for human consumption.     

Mixed function oxygenases and xenobiotic detoxication/toxication systems in bivalve molluscs

Components of a xenobiotic detoxication/toxication system involving mixed function oxygenases are present inMytilus edulis. Our paper critically reviews the recent literature on this topic which reported the apparent absence of such a system in bivalve molluscs and attempts to reconcile this viewpoint with our own findings on NADPH neotetrazolium reductase, glucose-6-phosphate dehydrogenase, aldrin epoxidation and other reports of the presence of mixed function oxygenases. New experimental data are presented which indicate that some elements of the detoxication/toxication system inM. edulis can be induced by aromatic hydrocarbons derived from crude oil. This includes a brief review of the results of long-term experiments in which mussels were exposed to low concentrations of the water accommodated fraction of North Sea crude oil (7.7–68 μg 1⁻¹) in which general stress responses such as reduced physiological scope for growth, cytotoxic damage to lysosomal integrity and cellular damage are considered as characteristics of the general stress syndrome induced by the toxic action of the xenobiotics. In addition, induction in the blood cells of microsomal NADPH neotetrazolium reductase (associated with mixed function oxygenases) and the NADPH generating enzyme glucose-6-phosphate dehydrogenase are considered to be specific biological responses to the presence of aromatic hydrocarbons. The consequences of this detoxication/toxication system forMytilus edulis are discussed in terms of the formation of toxic electrophilic intermediate metabolites which are highly reactive and can combine with DNA, RNA and proteins with subsequent damage to these cellular constituents. Implications for neoplasms associated with the blood cells are also discussed. Finally, in view of the increased use of mussel species in pollutant monitoring programmes, the induction phenomenon which is associated with microsomal enzymes in the blood cells is considered as a possible tool for the detection of the biological effects of environmental contamination by low concentrations of certain groups of organic xenobiotics.     

Recent bivalve molluscs of the genus Calyptogena (Vesicomyidae)

The genus Calyptogena (Bivalvia: Vesicomyidae) comprises highlyspecialized bivalves living in symbiosis with sulphur-oxidizingbacteria in reducing habitats. In this study, the genus is revisedusing shell and anatomical features. The work is based on typematerial, as well as on the extensive collection of vesicomyidsobtained during twelve expeditions to the Pacific and IndianOceans. Nine Recent species are ascribed to the genus Calyptogena,four of which are new: C. pacifica Dall, 1891, C. fausta Okutani,Fujikura & Hashimoto, 1993, C. rectimargo Scarlato, 1981,C. valdiviae (Thiele & Jaeckel, 1931), C. gallardoi Sellanes& Krylova, 2005, C. goffrediae n. sp., C. starobogatovin. sp., C. makranensis n. sp. and C. costaricana n. sp. Thecharacteristic features of Calyptogena are: shell up to 90 mmin length, elongate-elliptical or elongate; presence of escutcheon;presence of broad posterior ramus (3b) of right subumbonal cardinaltooth as well as right posterior nymphal ridge; absence of pallialsinus as a result of attachment of intersiphonal septal retractorimmediately adjacent to ventral surface of posterior adductor;absence of processes on inner vulva of inhalant siphon; presenceof inner demibranch only, with descending and ascending lamellaewith interlamellar septa not divided into separate tubes. Themost closely related taxa to Calyptogena are probably the genusIsorropodon Sturany, 1896, and the group of species representedby ‘Calyptogena’ phaseoliformis Métivier,Okutani & Ohta, 1986. These groups have several charactersin common, namely absence of pallial sinus, presence of singleinner pair of demibranchs and absence of processes on innervulva of inhalant siphon. The worldwide distribution of thegenus Calyptogena suggests that methane seeps at continentalmargins are the major dispersal routes and that speciation waspromoted by geographical isolation. Recent species diversityand fossil records indicate that the genus originated in thePacific Ocean. Sufficient data to discuss the distribution atspecies level exist only for C. pacifica, which has a remarkablynarrow bathymetric range. Published studies on the physiologyof C. pacifica suggest that adaptation to a specific geochemicalenvironment has led to coexisting vesicomyid genera. The bacteria-containinggill of C. pacifica and other Calyptogena species is one ofthe most specialized in the family Vesicomyidae and may reflectthese ecological adaptations. (Received 23 December 2005; accepted 3 April 2006)