Evidence of separate influence of moon and sun on light synchronization of mussel's daily rhythm during the polar night.

Marine organisms living at high latitudes are faced with a light climate that undergoes drastic annual changes, especially during the polar night (PN) when the sun remains below the horizon for months. This raises the question of a possible synchronization and entrainment of biological rhythms under the governance of light at very low intensities. We analyzed the rhythms of the mussel Mytilus sp. during PN. We show that (1) mussels expressed a rhythmic behavior during PN; (2) a monthly moonlight rhythm was expressed; (3) a daily rhythm was expressed and influenced by both sunlight and moonlight; and (4) depending on the different times of PN and moon cycle characteristics, we were able to discriminate whether the moon or the sun synchronize the daily rhythm. Our findings fuel the idea that the capability of moonlight to synchronize daily rhythms when sunlight is not sufficient would be a crucial advantage during PN.

Growth and behaviour of blue mussels, a re-emerging polar resident, follow a strong annual rhythm shaped by the extreme high Arctic light regime.

Polar regions are currently warming at a rate above the global average. One issue of concern is the consequences on biodiversity in relation to the Northward latitudinal shift in distribution of temperate species. In the present study, lasting almost two years, we examined two phenological traits, i.e. the shell growth and behavioural rhythm of a recently re-established species in the high Arctic, the blue mussel Mytilus sp. We compared this with a native species, the Islandic scallop Chlamys islandica. We show marked differences in the examined traits between the two species. In Mytilus sp., a clear annual pattern of shell growth strongly correlated to the valve behaviour rhythmicity, whereas C. islandica exhibited a shell growth pattern with a total absence of annual rhythmicity of behaviour. The shell growth was highly correlated to the photoperiod for the mussels but weaker for the scallops. The water temperature cycle was a very weak parameter to anticipate the phenology traits of both species. This study shows that the new resident in the high Arctic, Mytilus sp., is a highly adaptive species, and therefore a promising bioindicator to study the consequences of biodiversity changes due to global warming.

Bivalve mollusc circadian clock genes can run at tidal frequency.

Marine coastal habitats are complex cyclic environments as a result of sun and moon interactions. In contrast with the well-known circadian orchestration of the terrestrial animal rhythmicity (approx. 24 h), the mechanism responsible for the circatidal rhythm (approx. 12.4 h) remains largely elusive in marine organisms. We revealed in subtidal field conditions that the oyster Crassostrea gigas exhibits tidal rhythmicity of circadian clock genes and clock-associated genes. A free-running (FR) experiment showed an endogenous circatidal rhythm. In parallel, we showed in the field that oysters' valve behaviour exhibited a strong tidal rhythm combined with a daily rhythm. In the FR experiment, all behavioural rhythms were circatidal, and half of them were also circadian. Our results fuel the debate on endogenous circatidal mechanisms. In contrast with the current hypothesis on the existence of an independent tidal clock, we suggest that a single 'circadian/circatidal' clock in bivalves is sufficient to entrain behavioural patterns at tidal and daily frequencies.

Noise pollution limits metal bioaccumulation and growth rate in a filter feeder, the Pacific oyster Magallana gigas.

Shipping has increased dramatically in recent decades and oysters can hear them. We studied the interaction between noise pollution and trace metal contamination in the oyster Magallana gigas. Four oyster-groups were studied during a 14-day exposure period. Two were exposed to cadmium in the presence of cargo ship-noise ([Cd++]w ≈ 0.5 µg∙L-1; maximum sound pressure level 150 dBrms re 1 µPa), and 2 were exposed only to cadmium. The Cd concentration in the gills ([Cd]g) and the digestive gland ([Cd]dg), the valve closure duration, number of valve closures and circadian distribution of opening and closure, the daily shell growth-rate and the expression of 19 genes in the gills were studied. Oysters exposed to Cd in the presence of cargo ship-noise accumulated 2.5 times less Cd in their gills than did the controls without ship noise and their growth rate was 2.6 times slower. In the presence of ship noise, oysters were closed more during the daytime, and their daily valve activity was reduced. Changes in gene activity in the gills were observed in 7 genes when the Cd was associated with the ship noise. In the absence of ship noise, a change in expression was measured in 4 genes. We conclude that chronic exposure to cargo ship noise has a depressant effect on the activity in oysters, including on the volume of the water flowing over their gills (Vw). In turn, a decrease in the Vw and valve-opening duration limited metal exposure and uptake by the gills but also limited food uptake. This latter conclusion would explain the slowing observed in the fat metabolism and growth rate. Thus, we propose that cargo ship noise exposure could protect against metal bioaccumulation and affect the growth rate. This latter conclusion points towards a potential risk in terms of ecosystem productivity.

How annual course of photoperiod shapes seasonal behavior of diploid and triploid oysters, Crassostrea gigas.

In this work, we study if ploidy (i.e. number of copies of chromosomes) in the oyster Crassostrea gigas may introduce differences in behavior and in its synchronization by the annual photoperiod. To answer to the question about the effect of the seasonal course of the photoperiod on the behavior of C. gigas according to its ploidy, we quantified valve activity by HFNI valvometry in situ for 1 year in both diploid and triploid oysters. Chronobiological analyses of daily, tidal and lunar rhythms were performed according the annual change of the photoperiod. In parallel, growth and gametogenesis status were measured and spawning events were detected by valvometry. The results showed that triploids had reduced gametogenesis, without spawning events, and approximately three times more growth than diploids. These differences in physiological efforts could explain the result that photoperiod (daylength and/or direction of daylength) differentially drives and modulates seasonal behavior of diploid and triploid oysters. Most differences were observed during long days (spring and summer), where triploids showed longer valve opening duration but lower opening amplitude, stronger daily rhythm and weaker tidal rhythm. During this period, diploids did major gametogenesis and spawning whereas triploids did maximal growth. Differences were also observed in terms of moonlight rhythmicity and neap-spring tidal cycle rhythmicity. We suggest that the seasonal change of photoperiod differentially synchronizes oyster behavior and biological rhythms according to physiological needs based on ploidy.

The sense of hearing in the Pacific oyster, Magallana gigas.

There is an increasing concern that anthropogenic noise could have a significant impact on the marine environment, but there is still insufficient data for most invertebrates. What do they perceive? We investigated this question in oysters Magallana gigas (Crassostrea gigas) using pure tone exposures, accelerometer fixed on the oyster shell and hydrophone in the water column. Groups of 16 oysters were exposed to quantifiable waterborne sinusoidal sounds in the range of 10 Hz to 20 kHz at various acoustic energies. The experiment was conducted in running seawater using an experimental flume equipped with suspended loudspeakers. The sensitivity of the oysters was measured by recording their valve movements by high-frequency noninvasive valvometry. The tests were 3 min tone exposures including a 70 sec fade-in period. Three endpoints were analysed: the ratio of responding individuals in the group, the resulting changes of valve opening amplitude and the response latency. At high enough acoustic energy, oysters transiently closed their valves in response to frequencies in the range of 10 to <1000 Hz, with maximum sensitivity from 10 to 200 Hz. The minimum acoustic energy required to elicit a response was 0.02 m∙s-2 at 122 dBrms re 1 µPa for frequencies ranging from 10 to 80 Hz. As a partial valve closure cannot be differentiated from a nociceptive response, it is very likely that oysters detect sounds at lower acoustic energy. The mechanism involved in sound detection and the ecological consequences are discussed.

In the darkness of the polar night, scallops keep on a steady rhythm.

Although the prevailing paradigm has held that the polar night is a period of biological quiescence, recent studies have detected noticeable activity levels in marine organisms. In this study, we investigated the circadian rhythm of the scallop Chlamys islandica by continuously recording the animal's behaviour over 3 years in the Arctic (Svalbard). Our results showed that a circadian rhythm persists throughout the polar night and lasts for at least 4 months. Based on observations across three polar nights, we showed that the robustness and synchronicity of the rhythm depends on the angle of the sun below the horizon. The weakest rhythm occurred at the onset of the polar night during the nautical twilight. Surprisingly, the circadian behaviour began to recover during the darkest part of the polar night. Because active rhythms optimize the fitness of an organism, our study brings out that the scallops C. islandica remain active even during the polar night.

Evidence for a plastic dual circadian rhythm in the oyster Crassostrea gigas.

Although a significant body of literature has been devoted to the chronobiology of aquatic animals, how biological rhythms function in molluscan bivalves has been poorly studied. The first objective of this study was to determine whether an endogenous circadian rhythm does exist in the oyster, Crassostrea gigas. The second objective was to characterize it in terms of robustness. To answer these questions, the valve activity of 15 oysters was continuously recorded for 2 mo in the laboratory under different entrainment and free-running regimes using a high-frequency noninvasive valvometer. The present work demonstrates the presence of a circadian rhythm in the oyster Crassostrea gigas. First, oysters were entrained by 12 L:12 D conditions. Then, free-running conditions (D:D and L:L) indicated that the most frequently observed period ranged from 20 to 28 h, the circadian range. That endogenous circadian rhythm was characterized as weak. Indeed, the period (τ) of the individual animals exhibited high plasticity in D:D and L:L, and the animals immediately followed a 4-h phase advance or delay. Additionally, C. gigas appeared as a dual organism: all oysters were nocturnal at the beginning of the laboratory experiment (January), whereas they were diurnal at the end (March). That shift was progressive. Comparison with a full-year in situ record showed the same behavioral duality as observed in the laboratory: the animals were nocturnal in autumn-winter and diurnal in spring-summer. The significant advantage of a plastic and dual circadian rhythm in terms of adaptability in a highly changing environment is discussed.

Field chronobiology of a molluscan bivalve: how the moon and sun cycles interact to drive oyster activity rhythms.

The present study reports new insights into the complexity of environmental drivers in aquatic animals. The focus of this study was to determine the main forces that drive mollusc bivalve behavior in situ. To answer this question, the authors continuously studied the valve movements of permanently immersed oysters, Crassostrea gigas, during a 1-year-long in situ study. Valve behavior was monitored with a specially build valvometer, which allows continuously recording of up to 16 bivalves at high frequency (10 Hz). The results highlight a strong relationship between the rhythms of valve behavior and the complex association of the sun-earth-moon orbital positions. Permanently immersed C. gigas follows a robust and strong behavior primarily driven by the tidal cycle. The intensity of this tidal driving force is modulated by the neap-spring tides (i.e., synodic moon cycle), which themselves depend of the earth-moon distance (i.e., anomalistic moon cycle). Light is a significant driver of the oysters' biological rhythm, although its power is limited by the tides, which remain the predominant driver. More globally, depending where in the world the bivalves reside, the results suggest their biological rhythms should vary according to the relative importance of the solar cycle and different lunar cycles associated with tide generation. These results highlight the high plasticity of these oysters to adapt to their changing environment.

Relationship between valve activity, microalgae concentration in the water and toxin accumulation in the digestive gland of the Pacific oyster Crassostrea gigas exposed to Alexandrium minutum.

The complexity of the relationships between Alexandrium minutum (A.m.) concentration in the water ([A.m.](w)), Paralytic Shellfish Poisoning contamination in the digestive gland ([PSP](dg)) and valve behavior was explored in oysters Crassostrea gigas. Two experiments were conducted, during which oysters' valve behaviour were analyzed. Oysters, first acclimated for 10-days with the non harmful microalgae Heterocapsa triquetra (H.t.), were exposed to four microalgae mixtures at constant total concentrations of 10×10(3)cells ml(-1) (experiment-1) and 5×10(3)cells ml(-1) (experiment-2): 100% A.m.; 50% A.m.-50% H.t.; 25% A.m.-75% H.t.; 100% H.t. At the end of experiment-2, [PSP](dg) were measured. At 10×10(3)cells ml(-1), the microalgal ingestion decreased (p<0.05) with increasing [A.m.](w) but not at 5×10(3)cells ml(-1) (p>0.05). The frequency of microclosures specifically increased with [A.m.](w) (p<0.05) and the opening duration with [PSP](dg) (p<0.0001). Oysters exhibiting the maximum increase in opening duration also exhibited the highest [PSP](dg). The results are discussed in terms of oyster physiology and origin of the behavioral response.

Water quality assessment by means of HFNI valvometry and high-frequency data modeling.

The high-frequency measurements of valve activity in bivalves (e.g., valvometry) over a long period of time and in various environmental conditions allow a very accurate study of their behaviors as well as a global analysis of possible perturbations due to the environment. Valvometry uses the bivalve's ability to close its shell when exposed to a contaminant or other abnormal environmental conditions as an alarm to indicate possible perturbations in the environment. The modeling of such high-frequency serial valvometry data is statistically challenging, and here, a nonparametric approach based on kernel estimation is proposed. This method has the advantage of summarizing complex data into a simple density profile obtained from each animal at every 24-h period to ultimately make inference about time effect and external conditions on this profile. The statistical properties of the estimator are presented. Through an application to a sample of 16 oysters living in the Bay of Arcachon (France), we demonstrate that this method can be used to first estimate the normal biological rhythms of permanently immersed oysters and second to detect perturbations of these rhythms due to changes in their environment. We anticipate that this approach could have an important contribution to the survey of aquatic systems.

Estimation of potential and limits of bivalve closure response to detect contaminants: application to cadmium.

Bivalve closure responses to detect contaminants have often been studied in ecotoxicology as an aquatic pollution biosensor. We present a new laboratory procedure to estimate its potential and limits for various contaminants and animal susceptible to stress. The study was performed in the Asiatic clam Corbicula fluminea and applied to cadmium. To take into account the rate of spontaneous closures, we integrated stress problems associated with fixation by a valve in common apparatus and the spontaneous rhythm associated with circadian activity to focus on conditions with the lowest probability of spontaneous closing. Moreover, we developed an original system by impedance valvometry, using light-weight impedance electrodes, to study free-ranging animals in low-stress conditions and a new analytical approach to describe valve closure behavior as a function of response time and concentration of contaminant. In C. fluminea, we show that cadmium concentrations above 50 microg/L can be detected within less than 1 h, concentrations down to 16 microg/L require 5 h of integration time, and values lower than 16 microg/L cannot be distinguished from background noise. Our procedure improved by a factor of six the cadmium sensitivity threshold reported in the literature. Problems of field applications are discussed.