Marc Suquet

Oyster reproduction is affected by exposure to polystyrene microplastics

Significance Plastics are a contaminant of emerging concern accumulating in marine ecosystems. Plastics tend to break down into small particles, called microplastics, which also enter the marine environment directly as fragments from a variety of sources, including cosmetics, clothing, and industrial processes. Given their ubiquitous nature and small dimensions, the ingestion and impact of microplastics on marine life are a cause for concern, notably for filter feeders. Oysters were exposed to polystyrene microparticles, which were shown to interfere with energy uptake and allocation, reproduction, and offspring performance. A drop in energy allocation played a major role in this reproductive impairment. This study provides ground-breaking data on microplastic impacts in an invertebrate model, helping to predict ecological impact in marine ecosystems. Plastics are persistent synthetic polymers that accumulate as waste in the marine environment. Microplastic (MP) particles are derived from the breakdown of larger debris or can enter the environment as microscopic fragments. Because filter-feeder organisms ingest MP while feeding, they are likely to be impacted by MP pollution. To assess the impact of polystyrene microspheres (micro-PS) on the physiology of the Pacific oyster, adult oysters were experimentally exposed to virgin micro-PS (2 and 6 µm in diameter; 0.023 mg·L−1) for 2 mo during a reproductive cycle. Effects were investigated on ecophysiological parameters; cellular, transcriptomic, and proteomic responses; fecundity; and offspring development. Oysters preferentially ingested the 6-µm micro-PS over the 2-µm-diameter particles. Consumption of microalgae and absorption efficiency were significantly higher in exposed oysters, suggesting compensatory and physical effects on both digestive parameters. After 2 mo, exposed oysters had significant decreases in oocyte number (−38%), diameter (−5%), and sperm velocity (−23%). The D-larval yield and larval development of offspring derived from exposed parents decreased by 41% and 18%, respectively, compared with control offspring. Dynamic energy budget modeling, supported by transcriptomic profiles, suggested a significant shift of energy allocation from reproduction to structural growth, and elevated maintenance costs in exposed oysters, which is thought to be caused by interference with energy uptake. Molecular signatures of endocrine disruption were also revealed, but no endocrine disruptors were found in the biological samples. This study provides evidence that micro-PS cause feeding modifications and reproductive disruption in oysters, with significant impacts on offspring.

Nucleotide content, oxydative phosphorylation, morphology, and fertilizing capacity of turbot (Psetta maxima) spermatozoa during the motility period

The interdependence between motility, respiration, ATP production, and utilization was investigated in intact spermatozoa of turbot (Psetta maxima), a marine teleost. When spermatozoa were diluted in a hyperosmotic medium (>300 mOsmol/kg), they immediately became motile, and the intracellular concentration of ATP as well as the adenylate energy charge ratio dropped concomitant with the straight‐line velocity. The ADP and AMP levels increased from 1.4 to 8.0 nmole/108 cells and from 0.6 to 6.0 nmole/108 cells, respectively. Moreover, 31P‐NMR spectra recorded prior to the swimming phase revealed the presence of phosphomonoesters (PMEs) and phosphodiesters (PDEs), intracellular inorganic phosphate (Pi), and phosphocreatine (PCr). At the end of the motility period, PCr, PDE, and PME decreased, while the Pi level increased markedly. Following initiation of motility, O2 consumption of spermatozoa increased from 34.9 to 124.8 O2 nmole/109 spermatozoa/min. FCCP, an uncoupler of oxydative phosphorylation, did not significantly affect the respiratory rate of motile spermatozoa. Ouabain, a specific inhibitor of (Na+/K+)/ATPase, slightly decreased the respiration rate of motile spermatozoa, indicating that the major part of ATP catabolism was linked to dynein ATPase. Inhibitors of the respiratory chain (KCN, NaN3, NaHCO3–, oligomycin) reduced sperm respiration, percentage of motile cells, velocity, and adenylate contents. Following the reactivation of motility of demembranated spermatozoa, KCN, NaN3, NaHCO3– altered the flagellar beat frequency, demonstrating that these respiratory inhibitors possess action sites other than mitochondria. Mitochondrial oxydative phosphorylation is highly requested to produce energy required during motion. Nevertheless it is insufficient to maintain endogenous ATP stores. A second phase of motility was induced by a transfer of exhausted spermatozoa into an ionic medium of low osmolality (200 mOsmol/kg) for 30 min. Spermatozoa, once reactivated in AM, recovered 55% of initial motility and 31% of initial fertilization rate. In hypo‐osmotic medium, mitochondrial oxydative phosphorylation also induced ATP regeneration. Following activation of movement, several morphological changes were observed in the mitochondria and the midpiece. Mol. Reprod. Dev. 53:230–243, 1999.

Cryopreservation of turbot (Scophthalmus maximus) spermatozoa.

The aim of this study was to develop a method for cryopreserving turbot semen and to compare sperm motility characteristics, metabolic status and fertilization capacity of frozenthawed and fresh semen. The best results were obtained when spermatozoa were diluted at a 1:2 ratio with a modified Mounib extender, supplemented with 10% BSA and 10% DMSO. For freezing sperm samples, straws were placed at 6.5 cm above the surface of liquid nitrogen (LN) and plunged in LN. The straws were thawed in water bath at 30 degrees C for 5 sec. Use of this simple method resulted in a 60 to 80% reactivation rate of the thawed spermatozoa. Although the percentage of motile spermatozoa in the frozen-thawed semen samples was significantly lower than in fresh semen, spermatozoa velocity and respiratory rate remained unchanged. The process of cryopreservation significantly decreased intracellular ATP content. The fertilization rate of frozen-thawed spermatozoa was significantly lower than that of fresh spermatozoa, but it increased with sperm concentration.

Cryopreservation of sea bass (Dicentrarchus labrax) spermatozoa in experimental and production simulating conditions

Abstract A sperm cryopreservation protocol adapted from turbot, was tested on sea bass using either 250-μL straws or 1.5-mL cryovials. A dilution to 1/3 in Mounib s extender and a cooling rate of −65 °C·min −1 allowed frozen sperm to recover an initial motility similar to that of fresh sperm at thawing; however, significant differences in motility ( P n = 10 fish semen) were observed at further post-activation times, the motility decrease being faster in thawed sperm. At the experimental scale, triplicate inseminations of 2-mL aliquots (approximately 2 000 eggs) showed a significant fertility decay of thawed sperm compared to that of fresh sperm ( P n = 12 fish semen) when a discriminating 35·10 3 spermatozoa to egg ratio was applied. When 70·10 3 and 200·10 3 spermatozoa per egg were provided in the same experimental conditions, no significant difference appeared between the fertilisation rates of fresh and thawed sperm. In order to validate the procedure for production or cryobank purpose, a scaled-up protocol was established. Two and 50 mL batches of eggs (approximately 2·10 3 and 50·10 3 eggs, respectively) were inseminated in triplicate using either fresh or thawed individual sperms of 5 males with 200·10 3 spermatozoa per egg. The mean fertility decreased by 23.5 % due to cryopreservation. This decline was explained by the loss of fertility of only one sperm, and only in large-volume conditions, probably due to the delay of use after thawing.

Artificial insemination in turbot (Scophthalmus maximus): determination of the optimal sperm to egg ratio and time of gamete contact

Abstract In high-quality batches of eggs (mean egg viability rate 89.6%), fertilization success (FS = number of 4-cell-stage eggs/number of viable eggs) was maximal when the ratio of spermatozoa to egg was above 6 × 10 3 . For lower ratios, FS decreased and became highly variable. In lower-quality batches of eggs (mean egg viability rates 72.0 and 75.9%), FS was variable even for the highest sperm to egg ratios. For the ratio of 1.5 × 10 3 spermatozoa per egg, the occurrence of maximum FS in relation to increasing contact time between gametes was scattered and mainly recorded after 2 and 3 min. For the ratio of 6 × 10 3 spermatozoa per egg, maximum FS was mainly observed after a contact of 1 min between gametes. In commercial production, a minimum ratio of 6 × 10 3 spermatozoa per egg and a contact time between gametes of 3 min is recommended for the artificial insemination of turbot eggs.

Sperm motility in turbot, Scophthalmus marimus: initiation of movement and changes with time of swimming characteristics

SynopsisTurbot sperm motility is observed using dark field microscopy and stroboscopic illumination combined with video recording. Sperm motility is triggered by dilution of spermatozoa in sea water or in non ionic media (glucose or saccharose), presenting osmotic pressure ranging from 300 to 2100 mOsmol. The percentage of motile spermatozoa reaches 100% under conditions of osmotic pressure of 300 to 1100 mOsmol and pH close to 8.0. In full sea water, glucose or saccharose solutions an agglutination of spermatozoa is observed; this is prevented by addition of bovine serum albumin (5 mg ml−1). Immediately after transfer in activation solutions, 100% spermatozoa are motile in most samples freshly stripped. This percentage drops suddenly between 15 and 30% after 70 to 100 sec. The beat frequency remains at a constant value of 50 Hz during 40 s post activation and then drops suddenly between 15 and 30 Hz. The spermatozoa velocity is about 200 micrometers s−1 during 30 to 40 s and then declines to a stable value of 100 micrometers s−1 at 50 s post activation. After 1.20 mn, more and more spermatozoa become motionless. The minimum calculated and averaged distance covered during 1.20 min, is about 12 mm. The high performances of turbot spermatozoa motility are interpreted as a compensatory mechanism for the low sperm production.

Effect of urine on semen quality in turbot (Psetta maxima)

Abstract The deleterious effects of urine contamination on the quality of spermatozoa were observed in turbot ( Psetta maxima ). In order to overcome this problem, two methods of sperm collection were compared to evaluate on urine contamination. When collected by stripping, the mean contamination rate of sperm by urine was 15.3% (urine volume: sperm volume). The catherization of ureter prior to sperm collection significantly decreased the urine contamination to 9.3%. The composition of urine was measured in starved and fed juvenile turbots during a 24-h period. Both urea concentrations and pH varied in relation with the diet and showed significant daily variations ( P μ m s −1 to 160 μ m s −1 when measured at 10-s post-activation ( P

Selection method of new candidates for finfish aquaculture: the case of the French Atlantic, the Channel and the North Sea coasts

At present, European marine fish farming is based on sea bream and sea bass. A trend for diversification is sustained by the diversity of environmental conditions, by the availability of new production techniques such as recirculating systems, by an increase in rearing yields, by new market trends and by the possibilities to reduce risks of disease outbreak. Already reared fish species were chosen considering a limited number of criteria such as a high selling price and the availability of juveniles or breeders in the wild. This paper proposes a new selection method of fish species as candidates for aquaculture development on the French Atlantic, the Channel and the North Sea coasts. Using a three-phase procedure, candidates were selected among 20 000 fish species. Final classification was carried out using a panel of 22 criteria, taking into account inquiries conducted in France with aquaculturists but also with the main actors of distribution and transformation channels and with consumers. Cod (Gadus morhua) was selected by the present work as the first candidate for aquaculture development on the western coasts of France. This work also highlights the high interest of Gadoids for aquaculture development in this area.

Reliable assessment of overripening in turbot (Scophthalmus maximus) by a simple pH measurement

Abstract The pH of ovarian fluid was around 8.1 in turbot spawn collected by stripping shortly after ovulation. It decreased down to 7.1 when overripening developed. A highly significant correlation ( r =0.92, P

Long-term effects of the cryopreservation of turbot (Psetta maxima) spermatozoa

The survival of turbot eggs and the rearing capacities of larvae stemmed from artificial fertilization practices using frozen-thawed spermatozoa were evaluated. Furthermore, the viability of sperm samples stored during a 9 month period in liquid nitrogen was assessed. No significant difference in the fertilization rate, hatching rate, survival and wet weight of 10-day old larvae were observed using fresh or frozen-thawed spermatozoa. The motility recorded at 10 s and 60 s post-activation and the fertilization capacity of frozen-thawed spermatozoa were not significantly decreased during a 9 month storage period in liquid nitrogen. These results confirm the high quality of the turbot spermatozoa stemmed from the cryopreservation process, allowing their use for routine aquaculture practices.