Longzhen Zhang

The bioaccumulation of fluoride ion (F(-)) in Siberian sturgeon (Acipenser baerii) under laboratory conditions.

A 90-d growth trial was conducted to determine the accumulation of fluoride ions (F(-)) in juvenile Siberian sturgeon (Acipenser baerii). Fish were exposed to nominal F(-) concentrations of 4, 10, 25, 62.5 mg L(-1) (added as NaF), along with a control group. Fish growth was determined and F(-) concentrations in bone, cartilage, skin, gill, muscle, liver, gut and pylorus were analyzed by fluoride specific electrode method. Fish growth was inhibited for groups exposed to 10, 25, and 62.5 mg F(-) L(-1) compared to the control group. Significant increases in F(-) concentrations were observed in bone, cartilage, skin and gill of Siberian sturgeon exposed to 4 mg F(-) L(-1). The highest F(-) concentration was in bone (3204.4 mg kg(-1), dry weight), followed by cartilage (1401.2 mg kg(-1)), gill (389.4 mg kg(-1)), skin (100.1 mg kg(-1)). Muscle, liver, gut and pylorus did not show increasing accumulation with increasing F(-) levels in the water, but liver, gut, pylorus accumulated higher F(-) concentration compared to muscle. It is summarized that F(-) accumulates mainly in bone and cartilage in Siberian sturgeon, with 10 mg F(-) L(-1) in water harmful to growth. Even 4 mg F(-) L(-1) may be harmful to Siberian sturgeon growth during a long-term waterborne exposure.

Osmotic and ionic regulation and Na+/K+-ATPase, carbonic anhydrase activities in mature Chinese mitten crab, Eriocheir sinensis H. Milne Edwards, 1853 (Decapoda, Brachyura) exposed to different salinities

Mature Chinese mitten crabs, Eriocheir sinensis H. Milne Edwards, 1853, are exposed to brackish water or seawater as an obligatory part of their reproductive migration. We investigated the changes in haemolymph osmolality, haemolymph Na+, Cl−, Ca2+ and K+ concentrations, as well as gill Na+/K+-ATPase and carbonic anhydrase activity in both mature females and males, when in freshwater (0.3‰ salinity, 8 mOsm (kg H2O)−1) and after step-wise acclimation to salinities ranging from 7 to 35‰ (175-1044 mOsm (kg H2O)−1). Both females and males strongly hyper-regulated their haemolymph osmolality over a salinity range of 0.3-21‰, the isosmotic point approaching 28‰ salinity, and slightly hypo-regulated at 35‰ salinity. Haemolymph Na+ and Cl− levels correlated positively with haemolymph osmolality. Ca2+ and K+ levels, however, were maintained at a relatively narrow range and showed a contrary trend in their response to salinity. This indicated that Na+ and Cl− are the main contributors to increasing haemolymph osmolality. Reduced Na+/K+-ATPase and carbonic anhydrase activity in the posterior gills at elevated salinity suggests a direct physiological response to a decrease of Na+ intake. No sexual differences existed for all parameters except for significantly higher haemolymph Ca2+ and K+ levels and CA activity in females than in males at some salinity levels. It is suggested that mature Eriocheir sinensis is a strong hyper-osmoregulator but a weak hypo-osmoregulator. No sexual differences appeared to exist in osmotic and ionic regulation of mature Eriocheir sinensis exposed to different salinities.

Effects of different vitrificant solutions on the embryos of the Chinese mitten crab Eriocheir sinensis (Decapoda, Brachyura)

Vitrification could provide a promising tool for the cryopreservation of embryos. However, in order to achieve a vitrifiable medium, a high concentration of permeable cryoprotectants must be used, and the incorporation of high-molecular-weight compounds should also be considered. In the present study, the toxicity of six vitrificant solutions was determined in Eriocheir sinensis H. Milne Edwards, 1853 embryos at five developmental stages (cleavage, blastula, gastrula, eyed stage and heart beating stage). Six vitrifying solutions were prepared with seawater of salinity 15 and the cryoprotectants propylene glycol (PG), methanol (MeOH), dimethyl sulfoxide (DMSO) and dimethylformamide (DMF) in different proportions (A: 30% PG + 20% DMF; B: 30% MeOH + 20% DMF; C: 30% PG + 20% MeOH; D: 30% PG + 10% MeOH + 10% DMF; E: 20% DMSO + 30% PG; F: 20% DMSO + 30% MeOH). The impact of each of these solutions on E. sinensis embryos was then assessed and compared. Vitrifying solution “A” produced the highest survival rate, albeit lower than that of the control. A five-step equilibration of embryos in all six vitrifying solutions resulted in higher survival rates than equilibration in 3 or 2 steps. The tolerance of E. sinensis embryos to the vitrifying solutions increased with embryonic maturity, cleavage and blastula stage embryos being very sensitive to vitrifying solutions, and a two-step equilibration of embryos in the six vitrifying solutions resulted in no survival at all. Embryos in the eyed stage and heart beating stage showed a higher tolerance to vitrifying solutions as compared to the other stages, with the survival rates achieved over 50% and 60%, respectively. Cleavage stage and gastrula stage embryos did not survive after cryopreservation, although we acquired a percentage of transparent embryos (generally an indication of successful cryopreservation). Fifteen viable embryos were eventually recovered from 361 freeze-thawed embryos, all from eyed stage and heart beating stage embryos. One larva with apparently normal morphology hatched successfully from those 15 embryos.

Substrate color preference and feeding by juvenile Chinese sturgeon Acipenser sinensis: exploration of a behavioral adaptation

Experiments were conducted in artificial stream tanks with juvenile Chinese sturgeon, Acipenser sinensis, captured in the Yangtze Estuary to test two null hypotheses (1) juveniles have no preference for white or black substrates, and (2) white or black substrate color has no effect on juvenile feeding efficiency on the barcheek goby,Rhinogobius giurinus, a dark-colored prey of wild juveniles. Juveniles strongly preferred white substrate. However, in feeding tests, feeding efficiency by juveniles was similar on white or black substrates. Thus, substrate color had no effect on feeding efficiency. Results suggest the innate preference for white substrate is not a visual adaptation for capturing prey on a contrasting background, but is an adaptation predisposing juveniles to seek light-colored sandy substrate where forage abundance is greatest.

Effects of extracellular environment factors on the motility in Japanese eel spermatozoa.

We have examined the effects in vitro of the ionic composition, pH and temperature on the motility by the spermatozoa of the Japanese eel (Anguilla japonica). Milt was obtained from 10 males that had been artificially matured by repeated injections of hCG. Sperm motility was monitored with a VHS video recorder and a video camera connected to a microscope. The results showed that most of the sperm were highly motile in 250–700 mM NaCl, 250–650 mM KCl and 350–550 mM CaCl2 solution. The longest duration of sperm motility recorded in 500 mM NaCl, 250 mM KCl and 350 mM CaCl2 solution. Sperm was not motile when suspended at pH 2, sperm motility was observed at pH 3, there was a relatively higher percentage of motile sperm in solutions at pH 4–12 (above 80%). The motility and duration increased within 18–24°C and decreased at the range of 24–30°C. Appropriate K+ ion concentration in the active medium could enhance the percent motility and duration of eel spermatozoa.

Preliminary studies on molecular mechanism of salinity regulation of Na+, K+-ATPase α-subunit in gills of juvenile Chinese sturgeon(Acipenser sinensis)

In order to explore the molecular regulation mechanism of Na+,K+-ATPase(NKA)α-subunit in gills of juvenile Chinese sturgeon at salinity 10,partial NKA α-subunit gene sequence was cloned,and the abundance of α-subunit mRNA,NKA activity,ion concentration and osmolality were examined at 3,6,12,24,48,72,96 hours post-transfer.Results showed that:in experimental group(fresh water-salinity water,FW-SW),the expression of NKA α-subunit mRNA and NKA activity increased significantly(P0.05)during the adaptation phase(0-12 h),and the concentration of Na+,Cl-and osmolality rose to a maximum level at 12 h post-transfer.In regulatory phase(12-24 h),the amounts of NKA α-subunit mRNA and NKA activity fell to a relatively lower level which was still higher than that of fish in control group(P0.05).The change both in ion concentration and osmolality were in accordance with the change of NKA and mRNA expression.In conclusion,gill Na+,K+-ATPase in Chinese sturgeon plays an important role in osmolality regulation by changing α-subunit mRNA expression.The different α-subunit mRNA expression can lead to the variation of enzyme activity which can further regulate the ion concentration and osmolality.

Copper and Zinc Toxicity Estimates for Juvenile Chinese Sturgeon Acipenser sinensis in the Yangtze River Estuary

Acute toxicity estimates of copper (Cu) and zinc (Zn) in juvenile Chinese sturgeon were studied in the Yangtze River estuary and using the static test method to assess Cu and Zn toxicity. The results showed that Cu was more severely toxic to Chinese sturgeon than Zn. The Cu LC50 after 24-, 48-, 72and 96-h exposures to Chinese sturgeon were 0.0606, 0.0414, 0.0289 and 0.0217 mg·L, respectively, whereas these values were 3.590, 2.514, 1.759 and 1.200 mg·L for Zn, respectively. Estimated safe concentrations of Cu and Zn were 0.00217 and 0.1200 mg·L, respectively. However, the Cu and Zn standards for fisheries water quality in China are 0.01 and 0.10 mg·L, respectively. The liver was the major accumulation site for Cu. Cu content in the liver of juveniles in the control group (no Cu) was the highest, followed by the gills, whereas the lowest content was detected in cartilage. The fastest accumulator of Cu was the liver, followed by the stomach, and cartilage and skin had the lowest contents. Superoxide dismutase (SOD) and catalase (CAT) activities in the liver tissues of juveniles were relatively low for the first 24 h, but the activities recovered gradually from 24 to 48 h and decreased after 48 h to levels lower than the contro1 group. These changes in SOD and CAT activities indicate that juvenile Chinese sturgeon were severely affected by Cu. These data show that Chinese sturgeon juveniles face the threat of Cu contamination and this alerts officials to the dangerous situation.

Analysis of fatty acid composition in different tissues of adult female Tridentiger trigonocephalus collected from the Yangtze Estuary

Research was made in this paper on the moisture,lipid content and fatty acid composition of muscle,liver and ovary of adult female Tridentiger trigonocephalus collected from the Yangtze Estuary at Ⅴ stage of ovary development.The results indicated that:(1) The moisture content in muscle,liver and ovary was reduced successively,and the value was significantly different among these three tissues(P0.05).The total lipid content in these three tissues was(1.32%±0.08%),(7.75%±0.38%),and(26.73%±0.59%),respectively.There was significant difference in total lipid content among these three tissues,and the highest was in ovary,followed by liver,and the lowest was in muscle.(2) The content of SFA in muscle was significant higher than that in liver and ovary.There was significant difference in SFA content among three tissues(P0.05).About the MUFA,C16:1 content of liver is the highest.The C16:1 content in muscle,liver and ovary were(11.90%±0.71%),(15.28%±2.42%) and(9.49%±0.37%) respectively,and there was significant difference among these three tissues(P0.05).The content of PUFA,3PUFA,6PUFA in ovary was significantly higher than that in muscle and liver(P0.05),but showed no significant difference between muscle and liver(P0.05).The total content of EPA and DHA in muscle,liver and ovary were increased successively.The total content of EPA and DHA in these three tissues was(30.17%±1.02%),(30.53%±0.50%) and(34.33%±0.71%),respectively.The content of EPA and DHA in ovary was significantly higher than that in muscle and liver(P0.05),but showed no significant difference between muscle and liver(P0.05).The ratio of EPA to DHA(EPA/DHA) in these three tissues was 1.50,1.75 and 1.59 respectively,and the ratio of total 3 polyunsaturated fatty acids to total 6 polyunsaturated fatty acids(∑ 3PUFA/∑ 6PUFA) in these three tissues was 6.44,8.05 and 7.00,respectively.The ratios of EPA/DHA and ∑ 3PUFA/∑ 6PUFA were highest in liver,followed by ovary,and the lowest was in muscle.

Changes in non-specific immune parameters of mature Chinese mitten crab(Eriocheir sinensis) in response to elevated salinities

Mature Chinese mitten crabs,Eriocheir sinensis,are exposed to brackish water or coastal waters as an obligatory part during adult reproductive migration.In this study,we investigated the changes in serum total protein and oxyhemocyanin contents,alkaline phosphatase(AKP),superoxide dismutase(SOD) and phenoloxidase(PO) activities in both mature male and female E.when in freshwater and after step-wise acclimation to the following salinities: 7,14,21,28,35.Increasing salinities caused a gradual decrease in serum total protein and oxyhemocyanin contents.Serum AKP activity was elevated slightly at 7,thereafter it dropped slowly by the end of experiment.Serum SOD activity showed a slight decrease after increase of salinity until 21,it was activated above 21.Serum PO activity decreased gradually with the increase of salinity in males;however,it maintained a stable level above 21 in females.All tested parameters in serum except for SOD were significantly higher in females than those in males.It is suggested that elevated salinity induced a decrease in serum total protein and oxyhemocyanin contents.Serum immune enzyme activities in mature E.sinensis will be affected when the salinity was near to or above 28,which could exert a negative effect on reproduction.Further studies are needed to understand the biochemical adaptation of mature E.sinensis,during their reproductive migration acclimation to elevated salinities.Sex differences should not be ignored.

Effects of cryopreservation on exterior shape and interior structure of Chinese mitten crab (Eriocheir sinensis) embryos

In order to deeply understand the effects of cryopreservation on the configuration of embryos,we chose the Eriocheir sinensis as the object,and observed the exterior shapes and interior structure of different developmental stages embryos after treatment with cryoprotectants and cryopreservation by means of microscope,slice and electron microscopy.Results showed that:(1) Cleavage embryos swelled clearly after treatment with vitrifying solutions by two-step methods,exterior shape of the embryos had no significant difference compared to the control group by three-step equilibration methods,some yolk overflowed from the inner cells and cells were damaged seriously after cryopreservation.(2) Exterior shape of gastrula stage embryos was similar to the control group after treatment with two-step equilibration methods,surface of gastrula stage embryo treated with cryoprotectants crinkled and shaped reticulation structure by scanning electron microscopy(SEM),white gobbet appeared in cells of embryos and fringe of cell was coarse and protuberant by transmission electron microscopy(TEM).After cryopreservation,color of embryos was pink,cell became opaque,the brim of cell membrane was blurry and shaped floss,segmental yolk was disintegrated into granule,the surface of embryos was sunken and wrinkled.(3) Cell membrane of embryos broke off,ice holes of different size appeared inside embryos,visible vacuoles appeared inside cells by slice observation.Ice holes in cells were very clear and some vacuoles appeared,about 80% mitochondria dissolved,breach of cells was very obvious by TEM.Exterior shape of the original zoea stage embryos had no difference compared to that of the control group after treatment with cryoprotectants by microscope and SEM.After cryopreservation,above 95% tissues shaped dispersion,partial yolk was disintegrated into granule,cell membrane of embryos broke off,ice holes appeared inside embryos.Surface of 90% embryos was sunken and wrinkled,but the cuticle of 10% embryos retained slick and intact by SEM.The results implied that the original zoea stage of embryos was the appropriate stage for cryopreservation.