Regeneration of lost siphon tissues in the tellinacean bivalve Nuttallia olivacea

The inhalant siphon of the tellinacean bivalve Nuttallia olivacea is an important prey item for juvenile stone flounder Platichthys bicoloratus in estuaries in Japan. We examined quantitative siphon regeneration of N. olivacea in rearing experiments of siphon-removed bivalves (> 30 mm shell length) both in the laboratory and in their natural habitat. Under laboratory conditions, siphon-removed bivalves regenerated lost tissues quantitatively at 15 and 25 °C 1 mo after siphon removal, although regeneration was incomplete. A 3-mo caging experiment in the field showed that great regeneration occurred in siphon-removed bivalves. However, the siphon weight of removed bivalves was significantly smaller than that of non-amputated bivalves, suggesting the incomplete regeneration. In a 1-mo caging experiment, bivalves that had approximately 15% of their siphons amputated were selected at some intervals to illustrate the quantitative regeneration process. Estimated daily siphon production was remarkably high only a few days after amputation. It decreased greatly thereafter, but regeneration was not completed within 30 d. These results indicate that bivalves regenerate siphons rapidly just after losing siphon tissues and then regeneration is slowed down before it is completed.     

A clockwork mollusc: Ultradian rhythms in bivalve activity revealed by digital photography

Time-lapse digital images can be acquired and archived using web-cameras, allowing non-invasive analysis of behavior patterns of bivalve molluscs at ultradian (sub-daily) time-scales over long intervals. These records can be analyzed directly by a human operator or through properly calibrated image analysis software. Preliminary results using species of marine and freshwater bivalves identify several ultradian biological rhythms of similar duration. Wavelet analysis indicates strong periodicity in mantle and siphon activity in the 3 to 7 min range, with longer duration shell contraction periods at 60-90 min. The recurrence of these rhythms among marine and freshwater bivalve species maintained under constant (but differing) conditions suggests the influence of common intrinsic drivers (chemico-physical mechanisms or biological clocks). Sub-daily growth increments preserved in the shells of rapidly growing bivalve species are potentially related to these biological rhythms, with implications for shell growth, biomineralization, and the temporal resolution of paleoclimate proxy data.     

Brown shrimp (Crangon crangon, L.) functional response to density of different sized juvenile bivalves Macoma balthica (L.)

Variability in infaunal bivalve abundance in the Wadden Sea is largely determined by recruitment variability. Post-settlement, but pre-recruitment bivalve mortality is high and related to the occurrence of their most abundant predator, the brown shrimp Crangon crangon. To investigate if the mortality patterns of newly settled bivalves can be explained by the foraging behavior of brown shrimp, we carried out experiments on shrimp functional response to three size classes of juveniles of the Baltic Tellin Macoma balthica. The functional response curves for all three prey sizes (0.62 mm, 0.73 mm, and 0.85 mm) were the hyperbolic Holling's type II. The attack rate was highest for the smallest prey size (a = 0.31, medium and large prey a = 0.22); the handling time was longest for the largest prey size (T_h = 29 s, small and medium prey T_h = 15 s). Thus, a large body size is advantageous for the bivalves over the whole density range. Knowledge of individual foraging behavior is needed to model predation mortality of bivalves. The consumption rates in the experiment were theoretically high enough to account for M. balthica mortality in the field.     

Growth rate and age effects on Mya arenaria shell chemistry: Implications for biogeochemical studies

The chemical composition of bivalve shells can reflect that of their environment, making them useful indicators of climate, pollution, and ecosystem changes. However, biological factors can also influence chemical properties of biogenic carbonate. Understanding how these factors affect chemical incorporation is essential for studies that use elemental chemistry of carbonates as indicators of environmental parameters. This study examined the effects of bivalve shell growth rate and age on the incorporation of elements into juvenile softshell clams, Mya arenaria. Although previous studies have explored the effects of these two biological factors, reports have differed depending on species and environmental conditions. In addition, none of the previous studies have examined growth rate and age in the same species and within the same study. We reared clams in controlled laboratory conditions and used solution-based inductively coupled plasma mass spectrometry (ICP-MS) analysis to explore whether growth rate affects elemental incorporation into shell. Growth rate was negatively correlated with Mg, Mn, and Ba shell concentration, possibly due to increased discrimination ability with size. The relationship between growth rate and Pb and Sr was unresolved. To determine age effects on incorporation, we used laser ablation ICP-MS to measure changes in chemical composition across shells of individual clams. Age affected incorporation of Mn, Sr, and Ba within the juvenile shell, primarily due to significantly different elemental composition of early shell material compared to shell accreted later in life. Variability in shell composition increased closer to the umbo (hinge), which may be the result of methodology or may indicate an increased ability with age to discriminate against ions that are not calcium or carbonate. The effects of age and growth rate on elemental incorporation have the potential to bias data interpretation and should be considered in any biogeochemical study that uses bivalves as environmental indicators.     

Host specificity and population dynamics of a sponge-endosymbiotic bivalve

We assessed the host-use pattern of the sponge-endosymbiotic bivalve Vulsella vulsella and its demographic consequences in an inland sea in Okinawa Island, Japan. Vulsella vulsella utilized only one massive globular sponge species Spongia sp. as a host, and no Spongia sp. without V. vulsella were found. Individual sponges contained 9-248 live bivalves and 0-222 dead bivalves. The densities of live and dead bivalves in individual sponges were approximately constant irrespective of sponge size, indicating that available space is very scarce inside each sponge. The size distribution of bivalves was skewed to small, young individuals less than 30 mm in shell height, although the estimated largest possible size was 106 mm. The bivalve population at each sampling date was composed of three yearly cohorts, and recruitment of juveniles occurred in the summer. The bivalves became sexually mature as males within one year after recruitment and changed sex from male to female as they grew. The size and sex distributions of the bivalve were largely similar among sponges regardless of sponge size, suggesting that the recruitment, growth, longevity, and sex change of the bivalve were strictly regulated, probably by the high water temperature and strong waves generated by typhoons in summer months.     

Predator-prey interactions from in situ time-lapse observations of a sublittoral mussel bed in the Gulf of Trieste (Northern Adriatic)

Hexaplex trunculus (Linnaeus, 1758) is one of the most abundant and widespread muricid gastropods in the Northern Adriatic Sea, but relatively little is known about the feeding ecology of this predator. We examined the activity of H. trunculus on a sublittoral mussel bed at 24 m depth through in situ time-lapse observations and bulk samples. The camera photographed a 0.25 m² section of the mussel bed at 6-min intervals for ~ 23 h. Photos were examined frame-by-frame for gastropod movement and activities, especially interactions between H. trunculus and Mytilus galloprovincialis (Lamarck, 1819). Our survey indicates high activity-levels of H. trunculus on the sea floor: all gastropods made minor movements, most made major movements, and most left the field of view during the study-interval. On average, individuals remained stationary for only 7.3 h. Two predation attempts on Mytilus involving conspecific competition were documented, and one Hexaplex was consuming a mussel at the onset of the deployment. Additionally, 487 M. galloprovincialis from four diver-taken 0.25 m² quadrates were measured and examined for traces of marginal chipping and drilling predation. Mytilus from surface samples ranged from 11.1 mm to 95.5 mm in length, and one of the four samples had a significantly different average shell length from the others. 114 H. trunculus were collected and measured. Hexaplex ranged from 22.1 mm to 86.1 mm and the mean shell length did not differ among samples, though they were overwhelmingly medium and large. Predation frequency (the ratio of successfully preyed upon bivalves to the total number of bivalves sampled) is high at the studied site (> 55%), and large gastropods preferred a chipping mode of predation to drilling, supporting earlier laboratory studies showing a preference for M. galloprovincialis and this predation strategy. Prey effectiveness (the ratio of failed predatory attacks to total predatory attacks) is also high (63.8%), and no evidence of a size refuge was found. Feeding in H. trunculus is highly facultative, calling for caution when using drill holes to estimate predation intensities; whenever possible, traces of multiple predation modes should be considered.     

Burrowing behaviour of the softshell clam (Mya arenaria) following erosion and transport

Erosion and transport of juvenile individuals may alter the distribution pattern of intertidal bivalves. The burrowing success of recently transported juvenile softshell clams (Mya arenaria) was studied in a laboratory flume under a wide range of hydrosedimentary environments. Juvenile individuals (5-20 mm) were observed under a simulated 30 min slack tide before initiating the flow for a period of 60 min. Five different free-stream velocities (0, 3, 5, 10 and 24 cm s^− 1) and four sediment types (mud, sandy-mud, sand and gravel) were used. The mean proportion of juvenile clams that initiated (MPI) or completed (MPC) a burial decreased with increasing shell length. Erosion from the sediment was more important in large juveniles suggesting that large juveniles may have more difficulty successfully relocating once transported. The MPI increased with increasing flow speed in experimental runs held at speed < 24 cm s^− 1. This was observed in all sediment types. Most individuals were unable to burrow at 24 cm s^− 1 because they got eroded. The MPC also increased with increasing flow speed in mud, sandy-mud and sand. The MPC's response to flow was more complex in gravel because of a shell length × flow speed interaction effect. Our observations suggest that water movement may induce the burrowing behaviour of recently eroded juvenile clams. Results are discussed in an ecological and aquacultural context.     

Predatory impact of the green crab (Carcinus maenas Linnaeus) on post-settlement winter flounder (Pseudopleuronectes americanus Walbaum) as revealed by immunological dietary analysis

Predation on flatfish during the early juvenile stage is an important factor regulating year-class strength and recruitment. In this study, immunological dietary analysis was performed on green crabs (Carcinus maenas) collected from the Niantic River, Connecticut, in an effort to evaluate the predatory impact of this species on post-settlement winter flounder (Pseudopleuronectes americanus). Through the use of species-specific antiserum, winter flounder proteins were identified in 4.8% of the green crab stomachs analyzed (n = 313, size range = 14-74 mm carapace width, CW), revealing that crabs ≥ 29 mm CW are predators of post-settlement winter flounder in natural populations. The most significant factor underlying the predator-prey interaction was the relative size relationship between species, such that the incidence of winter flounder remains in the stomach contents of green crabs was positively correlated with predator-to-prey size ratio. Results from dietary analysis were incorporated into a deterministic model to estimate the average daily instantaneous mortality and cumulative mortality of winter flounder owing to green crab predation. Accordingly, green crabs may account for 0.4% to 7.7% (mean = 2.2%) of the daily mortality of winter flounder and consume 1.1% to 32.3% (mean = 10.2%) of the flounder year-class. Model simulations further indicate that variations in green crab abundance and size-structure account for the greatest variability in winter flounder mortality. Relative to other macro-crustacean predators, however, predation by green crabs has a minimal effect on winter flounder survival, due in large part to the low densities of these crabs in temperate estuaries.     

Diel feeding rhythms,daily ration,and seasonal changes thereof in marbled flounder Pseudopleuronectes yokohamae

This paper aims to assess the diel feeding pattern and seasonal variation in the daily ration of immature and mature marbled flounder (Pseudopleuronectes yokohamae). A day‐night collection by bottom trawls was conducted in Sendai Bay in July 2014. Marbled flounder (131–493 mm total length; Number of collected individuals = 1830, Number of analyzed individuals = 497) fed mainly on polychaetes during the day. At night, stomach content weight decreased with time, but the weight and proportion of bivalve siphons were consistently higher at night than during the day, suggesting nocturnal feeding by the flounder on bivalve siphons. Daily ration was greater in females (<300 mm: 2.6%–3.2% body weight; ≥300 mm: 1.5%–2.5%) than in males (<300 mm: 1.7%–2.6%; ≥300 mm: 1.3%–1.9%). Seasonal surveys were also carried out, and the greater ration in females than males were consistent throughout the year, suggesting that greater growth in females than males attributes to the greater food intake of females. The ration was highest in June, especially for large individuals, although water temperature in June was lower than that in September. These results indicate that the amount of food intake is related to the annual life cycle of the marbled flounder.     

A novel role for dpp in the shaping of bivalve shells revealed in a conserved molluscan developmental program

During the molluscan evolution leading to the bivalves, the single dorsal shell was doubled. To elucidate the molecular developmental basis underlying this prominent morphological transition, we described the cell cleavage and expression patterns of three genes, brachyury, engrailed, and dpp in the Japanese spiny oyster Saccostrea kegaki, and examined the function of dpp in this species. The cleavage pattern of the S. kegaki embryo was nearly the same as the previously described pattern of other bivalve species, suggesting that the pattern itself is highly important for the establishment or the maintenance of the bivalve body plan. The expression pattern of a brachyury homolog in S. kegaki (SkBra) was similar to the pattern in gastopods even at the single cell level despite the deep divergence of gastropods and bivalves. Engrailed and dpp were previously found to be expressed around the shell anlagen in gastropods. Like that of gastropods, an engrailed homolog in S. kegaki (SkEn) was found to be expressed around the shell anlagen. However, the dpp homolog in S. kegaki (SkDpp) was expressed only in the cells along the dorsal midline. ZfBMP4 treatment experiments revealed the importance of dpp in establishing the characteristic shape of the bivalve shell anlagen.     

PREDATOR-PREY INTERACTIONS BETWEEN LEPSIELLA (BEDEVA) PAIVAE (GASTROPODA: MURICIDAE) AND KATELYSIA SCALARINA (BIVALVIA: VENERIDAE) IN PRINCESS ROYAL HARBOUR, WESTERN AUSTRALIA

On a sandy beach at Shoal Bay in Princess Royal Harbour, Albany,southwestern Western Australia, lives a small muricid gastropodthat feeds virtually monotonically on the overwhelmingly dominantresident bivalve Katelysia scalarina. Lepsiella paivae livesburied in the sand and attacks its prey within it. Because ofits small size (<13 mm shell height), bivalve prey isalso small and this study demonstrates a preference for K. scalarinaof 5 mm shell length, i.e. juveniles. Laboratory experimentsalso suggested a possible preference for attack of the rightvalve. Lepsiella paivae can and does, however, attack largerprey (up to 15 mm shell length), but cannot consume themcompletely. A second visit to Princess Royal Harbour in theAustral winter, when there was no juvenile K. scalarina present,showed L. paivae to be attacking at the sand surface, also bydrilling, the small (<4 mm) gastropod Hydrococcus brazieri(Hydrococcidae). SEM studies of experimentally determined drillholes of L. paivae show them to be of variable form, some straightsided, others bevelled (like a naticid) and <500 µmin diameter. On this sheltered Southern Ocean beach, therefore,L. paivae has specialized to attack juvenile bivalves by burrowingafter them. It can, however, attack other species opportunisticallyon the sand surface when seasonally favoured juvenile bivalveprey are not present. (Received 8 January 2005; accepted 16 March 2005)     

A noninvasive method for extracting bivalve DNA from the water-filled mantle cavity

Genetic studies play a great role for determining the biology of bivalves, particularly those covering population genetics, phylogeny, breeding, stock management, and conservation. However, DNA sampling methods that require removal of bivalves from the water and/or opening of their shells often cause stress and damage to bivalves, which can be lethal. The invasiveness of DNA sampling has made it difficult to conduct genetic studies in threatened species, rare species, and/or breeding lineages. In the present study, we developed a non-invasive method for bivalve DNA sampling using the water-filled mantle cavity (WMC). Our method can extract DNA from a small WMC sample (about 100 µl), collected using a fine needle and syringe without opening the shell. We demonstrated that the WMC sample contains intact mitochondrial and nuclear DNA. DNA contamination from other organisms, such as adjacent bivalve individuals, did not affect the resulting PCR and DNA sequencing analyses. Finally, the individuals from whom WMC was collected remained alive for more than 2 months after the experiments. This non-invasive method will be of great assistance in investigating the genetics of bivalves.

     

Effects of juvenile non-indigenous Carcinus maenas on the growth and condition of juvenile Cancer irroratus

The Atlantic rock crab, Cancer irroratus, is a commercially fished species and a critical prey item for the American lobster, Homarus americanus, in Atlantic Canada. The recent invasion of European green crab, Carcinus maenas, may have significant effects on the growth and condition of native C. irroratus, because both species overlap spatially and temporally and have similar habitat and dietary requirements. To examine such potential effects, we measured the growth of juvenile C. irroratus in the presence of juvenile C. maenas over a period of 4 months (growing season), under the following species combinations: (1) one C. irroratus (10-25 mm CW); (2) two C. irroratus (10-25 mm CW); (3) one C. irroratus (10-25 mm CW) and one C. maenas (10-15 mm CW). Morphological measurements included pre- and post-molt carapace width, chela height, abdomen width (mm), weight (g), and estimates of molt increment (%) and intermolt duration (days). Analysis of the hepatopancreas for % lipid content at the end of the experiment provided an estimate of physiological condition. The effect of the presence of C. maenas on the growth of C. irroratus shifted from negative to positive, when C. irroratus reached CW of 19-22 mm and gained a presumably significant size advantage over C. maenas. The positive effect resulted from increased energy intake through crab consumption. In the absence of crab consumption, the presence of a second crab (conspecific or C. maenas) had no effect on growth. C. irroratus consumed crabs more frequently when the second individual was a green crab than a conspecific. Consumption of C. maenas had a pronounced effect on the growth rate of C. irroratus, resulting in shorter intermolt periods and larger percent molt increments than in the presence of a conspecific. Therefore, the presence of juvenile C. maenas does not appear to have a prolonged negative effect on the growth of C. irroratus; rather, it may provide an additional food item as rock crabs grow, as long as encounters between the two species occur at high enough rates.     

Feeding habits of juvenile slime flounder <Emphasis Type="Italic">Microstomus achne</Emphasis> in the coastal area of southern Hokkaido

A total of 45 juvenile [30.0–57.4 mm total length (TL)] slime flounder Microstomus achne were collected in the coastal area of southern Hokkaido from April to July in 2001 and April to June in 2002. Their diets were analyzed. Slime flounder juveniles of 30.0–39.9 mm TL fed predominantly on small crustaceans (gammarid amphipods, harpacticoids and cumaceans) and those of 40.0–57.4 mm TL on gammarid amphipods, cumaceans and polychaetes. The major prey items changed with growth from small crustaceans (e.g., harpacticoids) to polychaetes, although gammarid amphipods were the major prey items throughout the juvenile period (30.0–57.4 mm TL).     

Studies on wound healing,and an estimation of the rate of regeneration,of the siphon of Scrobicularia plana (da Costa)

Wound closure of the siphon of Scrobicularia plana (da Costa), after amputation of the tip, is complete within 48 h. Once the lesion has been plugged by connective tissue new epithelial tissue begins to grow and is fully formed 96 h after wounding. As the epithelial cells are being formed, regeneration of the siphon commences at a rate of ≈ 6 mg per wk. When expressed in terms of percentage weight increase of the total siphon weight (20%) this rate is comparable with that calculated for other bivalves. The siphon continues to grow until the original size is attained. When small amounts of siphon are removed (15% of total siphon), regeneration is of little energetic cost, even when animals are starved. When greater amounts are, however, removed from unfed animals (e.g. 50% of the total siphon) regeneration takes place at the expense of the body condition, the animal using 0.6 kJ of body energy to replace 0.2 kJ of siphon.     

Ontogenic changes in tolerance to hypoxia and energy metabolism of larval and juvenile Japanese flounder Paralichthys olivaceus

Changes in tolerances to hypoxia and sodium azide, an indicator of cellular respiration, and activities of various energy metabolism-related chemical components were studied in Japanese flounder Paralichthys olivaceus during its early life stages from 3.5 to 20.5 mm in total length (TL). They showed flexion stage around 10.4 mm TL. Lethal levels of hypoxia increased with growth from 3.5 to 8 mm total TL, and the levels remained high in larvae, until 10.4 mm TL, decreased significantly thereafter. The 50% lethal concentration of sodium azide temporarily increased at 4.5 mm TL, diminished drastically between 4.5 and 10.4 mm TL, and then increased again in post-flexion larvae. Cytochrome c oxidase activity was highest in larvae around flexion, at 10.4 mm TL, and subsequently decreased. In post-flexion larvae at 13.0 mm TL, lactate dehydrogenase (LDH) and creatine kinase activities increased; LDH activity decreased at the juvenile stage. The adenosine triphosphate content and energy charge in fish were consistently higher in the larval stage than in the juvenile stage. These results indicated that, from just before flexion to the post-flexion stage, the energy metabolism of larvae is higher due to activated aerobic and subsequent anaerobic metabolism for metamorphosis; as a consequence, hypoxia tolerance in fish is the lowest during the increase of aerobic metabolism just before and around flexion.     

Autotomy reduces feeding, energy storage and growth of the sea star Stichaster striatus

We evaluated the effect of autotomy on feeding, energy storage and growth of juvenile Stichaster striatus kept in the laboratory for five months with a limited supply of the mussel Semimytilus algosus. Autotomy strongly decreased feeding, energy storage and growth. Intact juveniles showed a ∼ 3 fold higher feeding rate than autotomized individuals throughout the experiment. Intact juveniles also had a higher (∼ 5 fold) energy content per pyloric caeca in each arm. This was mainly due to higher lipid content, the main proximate constituent of pyloric caeca. Intact juveniles showed a greater growth rate and reached a greater size than autotomized individuals, more evident for underwater mass than radius length. The reduced capacity to feed reduced energy intake in autotomized individuals. However, low energy reserves along with low growth in autotomized sea stars, support the hypothesis that juveniles of this species allocate energy to regeneration to the detriment of growth. This was also supported by the ∼ 25% of arm length regeneration after 5 mo. Remaining small could increase risk of lethal predation, however, S. striatus may reduce predation risk by using crevices and kelp holdfasts as refuges from predators. Given the strong impact of autotomy on feeding, regeneration of arms to recover full capacity to forage and grow seems a better strategy for juvenile S. striatus, than merely growing.     

Regeneration of oral siphon pigment organs in the ascidian Ciona intestinalis

Ascidians have powerful capacities for regeneration but the underlying mechanisms are poorly understood. Here we examine oral siphon regeneration in the solitary ascidian Ciona intestinalis. Following amputation, the oral siphon rapidly reforms oral pigment organs (OPO) at its distal margin prior to slower regeneration of proximal siphon parts. The early stages of oral siphon reformation include cell proliferation and re-growth of the siphon nerves, although the neural complex (adult brain and associated organs) is not required for regeneration. Young animals reform OPO more rapidly after amputation than old animals indicating that regeneration is age dependent. UV irradiation, microcautery, and cultured siphon explant experiments indicate that OPOs are replaced as independent units based on local differentiation of progenitor cells within the siphon, rather than by cell migration from a distant source in the body. The typical pattern of eight OPOs and siphon lobes is restored with fidelity after distal amputation of the oral siphon, but as many as 16 OPOs and lobes can be reformed following proximal amputation near the siphon base. Thus, the pattern of OPO regeneration is determined by cues positioned along the proximal distal axis of the oral siphon. A model is presented in which columns of siphon tissue along the proximal-distal axis below pre-existing OPO are responsible for reproducing the normal OPO pattern during regeneration. This study reveals previously unknown principles of oral siphon and OPO regeneration that will be important for developing Ciona as a regeneration model in urochordates, which may be the closest living relatives of vertebrates.     

Seasonal and ontogenetic diet shift in juvenile stone flounder Platichthys bicoloratus

The diet of juvenile (0 year old) stone flounder Platichthys bicoloratus after settlement was investigated in an estuarine habitat. The principal prey items included harpacticoids and palps of polychaetes in February and March and thereafter shifted to siphons of the bivalve Nuttallia olivacea from April to June. This seasonal change in diet occurred irrespective of body size, suggesting that other factors, such as morphological development of the predator or vulnerability of prey, induced the diet shift.