The potential for cryopreserving larvae of the sea urchin, Evechinus chloroticus

Larvae of the sea urchin, Evechinus chloroticus, at varying stages of development, were assessed for their potential to survive cryopreservation. Ethylene glycol (EG) and dimethyl sulphoxide (Me2SO), at concentrations of 1-2 M, were evaluated as cryoprotectants (CPAs) in freezing regimes initially based on methods established for freezing larvae of other sea urchin species. Subsequent work varied cooling rate, holding temperature, holding time, and plunge temperature. Ethylene glycol was less toxic to larvae than Me2SO. However, no larvae survived freezing and thawing in EG. Larvae frozen in Me2SO at the gastrula stage and 4-armed pluteus stage regained motility post-thawing. The most successful freezing regime cooled straws containing larvae in 1.5 M Me2SO from 0 to -35 degrees C at 2.5 degrees C min(-1), held at -35 degrees C for 5 min, then plunged straws into liquid nitrogen. Motility was high 2-4 h post-thawing using this regime but decreased markedly within 24 h. Some 4-armed pluteus larvae that survived beyond this time developed through to metamorphosis and settled. Different Me2SO concentrations and supplementary trehalose did not improve long-term survival. Large variation in post-thaw survival was observed among batches of larvae produced from different females.     

Cryopreservation of seabream (Sparus aurata) spermatozoa

The aim of this research was to optimize protocols for freezing spermatozoa of seabream (Sparus aurata). All the phases of the cryopreservation procedure (sampling, choosing the cryoprotective extender, cooling, freezing, and thawing) were studied in relation to the species of spermatozoa under examination, so as to be able to restore on thawing the morphological and physiological characteristics of fresh semen. Seabream spermatozoa were collected by stripping and transported to the laboratory chilled (0-2 degrees C). Five cryoprotectants, dimethyl sulfoxide (Me(2)SO), ethylene glycol (EG), 1,2-propylene glycol (PG), glycerol, and methanol, were tested at concentrations between 5 and 15% by volume to evaluate their effect on the motility of semen exposed for up to 30 min at 26 degrees C. The less toxic cryoprotectants, 10% EG, 10% PG, and 5% Me(2)SO, respectively, were added to 1% NaCl to formulate the extenders for freezing. The semen was diluted 1:6 with the extender, inserted into 0.25-ml plastic straws by Pasteur pipette, and frozen using a cooling rate of either 10 or 15 degrees C/min to -150 degrees C followed by transfer and storage in liquid nitrogen (-196 degrees C). The straws were thawed at 15 degrees C/s. On thawing, the best motility was obtained with 5% Me(2)SO, although both 10% PG and EG showed good results; no differences were found between the two freezing gradients, although semen frozen with the 10 degrees C/min gradient showed a slightly higher and more prolonged motility.     

Effect of trehalose- and sucrose-based extenders on equine sperm quality after vitrification: Preliminary results

There has been a lack of research into equine sperm vitrification to date, but studies of other species suggest it may have significant potential. To evaluate the impact of various cryoprotectant agents (CPA) and vitrification on equine sperm quality, a controlled study was carried out. A total of 12 ejaculates were subjected to exposure to CPA and vitrification. Sperm was diluted in a range of CPA: fresh, control (BSA), sucrose (0.15M, 0.3M and 0.5M), trehalose (0.15M, 0.3M and 0.5M) and the combination of sucrose and trehalose (M1: 0.15M sucrose+0.5M trehalose; M2: 0.5M sucrose+0.15M trehalose). Sperm motility, viability, acrosome integrity and DNA fragmentation were assessed at the time of CPA exposure and after vitrification. The exposure of spermatozoa to various concentrations of sucrose and/or trehalose significantly reduced sperm motility, with lower concentrations resulting in higher sperm motility. Sperm viability and DNA fragmentation did not vary after exposure to CPA, but acrosome integrity fell significantly when spermatozoa were exposed to CPA with high osmolality. When spermatozoa were vitrified, motility values were significantly higher than those obtained during the exposure. Low concentrations of sucrose (0.15M and 0.3M) and trehalose (0.15M) showed the best progressive sperm motility. The vitrification-warmed procedure significantly reduced sperm viability and acrosome integrity, but DNA did not vary with any of CPA used. Equine sperm vitrification demonstrates a low capacity for preserving sperm motility, and extenders containing trehalose or sucrose at lower concentrations are associated with a better protective effect on sperm motility. After vitrification, acrosome and plasma membranes were severely impaired, while the DNA structure was maintained. Equine spermatozoa partially recover the motility after vitrification, but there is a need for further studies into the preservation of sperm membranes.     

Is cryopreservation a homogeneous process? Ultrastructure and motility of untreated, prefreezing, and postthawed spermatozoa of Diplodus puntazzo (Cetti).

This study subdivides the cryopreservation procedure for Diplodus puntazzo spermatozoa into three key phases, fresh, prefreezing (samples equilibrated in cryosolutions), and postthawed stages, and examines the ultrastructural anomalies and motility profiles of spermatozoa in each stage, with different cryodiluents. Two simple cryosolutions were evaluated: 0.17 M sodium chloride containing a final concentration of 15% dimethyl sulfoxide (Me(2)SO) (cryosolution A) and 0.1 M sodium citrate containing a final concentration of 10% Me(2)SO (cryosolution B). Ultrastructural anomalies of the plasmatic and nuclear membranes of the sperm head were common and the severity of the cryoinjury differed significantly between the pre- and the postfreezing phases and between the two cryosolutions. In spermatozoa diluted with cryosolution A, during the prefreezing phase, the plasmalemma of 61% of the cells was absent or damaged compared with 24% in the fresh sample (P < 0.001). In spermatozoa diluted with cryosolution B, there was a pronounced increase in the number of cells lacking the head plasmatic membrane from the prefreezing to the postthawed stages (from 32 to 52%, P < 0.01). In both cryosolutions, damages to nuclear membrane were significantly higher after freezing (cryosolution A: 8 to 23%, P < 0.01; cryosolution B: 5 to 38%, P < 0.001). With cryosolution A, the after-activation motility profile confirmed a consistent drop from fresh at the prefreezing stage, whereas freezing and thawing did not affect the motility much further and 50% of the cells were immotile by 60-90 s after activation. With cryosolution B, only the postthawing stage showed a sharp drop of motility profile. This study suggests that the different phases of the cryoprocess should be investigated to better understand the process of sperm damage.     

Osmotic sensitivity of canine spermatozoa

The objective of this study was to determine osmotic tolerance of canine spermatozoa. The study comprised three experiments: (1) spermatozoa suspended either in an egg yolk-citrate (EYC) extender or in Kenney skim milk extender were exposed to NaCl solutions ranging from 290 to 1500 mOsm; (2) spermatozoa suspended in EYC were exposed to 550 to 1500 mOsm solutions of glucose, galactose, or fructose; and (3) spermatozoa suspended in EYC or glucose-bovine serum albumin (G-BSA) were exposed to 0.6 M (approximately 900 mOsm) or 1.2 M (approximately 1600 mOsm) solutions of glycerol, ethylene glycol (EG), or dimethyl sulfoxide (Me(2)SO). In all experiments, motility and membrane integrity of spermatozoa were assessed after they were diluted into isotonic medium at 37 degrees C. Exposure of canine spermatozoa to solutions of either NaCl or monosaccharides at osmolalities >500 mOsm caused a significant reduction of motility (P<0.01). Motility of spermatozoa was more affected by osmotic stress than their membrane integrity. Osmotic sensitivity of canine spermatozoa was dependent on the type of extender; spermatozoa suspended in the Kenney extender were more resistant to osmotic stress than those suspended in the EYC extender. Despite their sensitivity to exposure to high concentrations of nonpermeating agents, canine spermatozoa were rather resistant to exposure to glycerol and EG. However, Me(2)SO was toxic to canine spermatozoa; motility was substantially reduced after spermatozoa were exposed to 0.6 M Me(2)SO. The type of extender also affected the sensitivity of canine spermatozoa to Me(2)SO; spermatozoa suspended in the EYC extender were more resistant than those suspended in G-BSA. It was concluded that canine spermatozoa are sensitive to osmotic stress, but are tolerant to shrinking and swelling caused by exposure to permeating cryoprotectants.     

Cryopreservation of European catfish Silurus glanis sperm: sperm motility, viability, and hatching success of embryos

The aim of this study was to elaborate cryopreservation methods for ex situ conservation of European catfish. The success of sperm cryopreservation was evaluated by post-thaw sperm motility and velocity, percentage of live spermatozoa and fertility (hatching rates) using frozen/thawed sperm. The best hatching rates of 82-86% were obtained with sperm stored for 5 h before freezing in immobilizing solution and frozen with Me2SO in concentrations of 8, 10, and 12%, or with a mixture of 5% Me2SO and 5% propandiole. These results did not significantly differ from the fresh sperm control sample. The percentage of live spermatozoa in frozen/thawed sperm did not correlate with hatching rate or motility of spermatozoa, but was negatively correlated with velocity of spermatozoa (r=-0.47, P=0.05). The percentage motility in frozen/thawed sperm ranged from 8 to 62%, when sperm was stored in immobilizing solution 5h before freezing. The average value in the fresh sperm (control) was 96%. The frozen/thawed sperm motility rate significantly correlated with the hatching rate (r=0.76, P=0.0002), but not with the percentage of live spermatozoa (r=0.16, P=0.52) or the sperm velocity (r=0.07, P=0.79). The velocity of frozen/thawed spermatozoa ranged from 37 to 85 microm/s, whereby methanol concentrations of 7.5 and 10% resulted in highest velocities. Freezing sperm volumes of 1-4 ml did not affect the quality of frozen/thawed sperm.     

Gamete cryobanks for laboratory research: Developing a rapid and easy-to-perform protocol for the cryopreservation of the sea urchin Paracentrotus lividus (Lmk, 1816) spermatozoa

Gamete cryopreservation is a biotechnology that can guarantee a continuous supply of gametes, regardless of the seasonal reproductive cycle. In this study we developed a protocol for the cryopreservation of the sea urchin Paracentrotuslividus spermatozoa, with a view to the creation of cryobanks of semen to be used as a model system in laboratory research and ecotoxicological tests. All the key phases of the procedure were separately considered and the effect on sperm motility was evaluated by means of computer assisted analysis. The best results were obtained using 7% dimethylsulfoxide in 1% NaCl plus 0.04 M trehalose as the extender, at a freezing rate of −20 °C/min. On thawing, in semen samples cryopreserved in accordance with this protocol the velocity parameters of the sub-population of rapid sperm (best performing spermatozoa) did not significantly differ from semen on collection; in addition also the fertilization ability was restored, and about 50% of normal developed plutei larvae were obtained by thawed semen. The developed protocol is rapid and easy-to-perform; moreover, the use of gametes from reared urchins makes it unnecessary to continuously collect specimens from natural populations, making this procedure a promising starting point for the creation of alternative and more sustainable methodologies in laboratory research on sea urchin gametes and embryos.     

Improvement of post-thaw sperm survivals using liquid nitrogen vapor in a spermcasting oyster Ostrea angasi

Low survival of cryopreserved sperm impedes the application of cryopreservation technique in spermcasting oyster species. This study developed a simple method of liquid nitrogen vapor freezing to improve post-thaw sperm survival in the spermcasting oyster Ostrea angasi. The results indicate that the permeable cryoprotectants, dimethyl sulfoxide (DMSO), ethylene glycol (EG) and propylene glycol (PG) were non-toxic to sperm up to 20% concentration and 90 min exposure whereas methanol at 10% or higher was toxic to sperm for any exposure over 30 min. Among the treatments with permeable cryoprotectants, 15% EG produced the highest post-thaw sperm motility. Sperm motility was further improved by the addition of non-permeable cryoprotectants (trehalose and glucose), with 15% EG + 0.2 M trehalose resulting in the highest post-thaw sperm motility among all the combinations evaluated. The durations of 20, 30 and 60 min equilibrations produced a higher post-thaw sperm motility and plasma membrane integrity (PMI) than 10 min. Higher post-thaw motility and PMI were achieved by freezing sperm at the 8 cm height from the liquid nitrogen surface than at the 2, 4, 6, 10 or 12 cm height. Holding sperm for 10 min in liquid nitrogen vapor produced higher post-thaw motility and PMI than for 2, 5 or 20 min. The cryopreservation protocol developed in this study improved both post-thaw motility and PMI of O. angasi sperm at least 15% higher than those cryopreserved using programmable freezing method. Liquid nitrogen vapor freezing might have greater applicability in improving post-thaw sperm quality of spermcasting oyster species.     

Impact of supplementation of semen extender with antioxidants on the quality of chilled or cryopreserved Arabian stallion spermatozoa

The aim of the present study was to evaluate the effects of supplementation of semen extender with various non-enzymatic antioxidants on the quality of cooled or cryopreserved Arabian stallion spermatozoa. Semen collected from four pure Arabian stallions was centrifuged at 600g for 15 min. Spermatozoa were then diluted in INRA-82 extender supplemented with bovine serum albumin (BSA; 0, 10, 15 and 20 mg/mL) or trehalose (0, 75, 100 and 150 mM) or zinc sulphate (0, 100, 150 and 200 μM). The diluted semen was then either cooled at 5 °C or cryopreserved in 0.5–ml plastic straws. After cooling or thawing, sperm motility, viability, sperm abnormalities, viability index, and plasma membrane integrity were evaluated. The results showed that supplementation of semen extender with 150 mM trehalose or with 200 μM zinc sulphate significantly (P < 0.05) improved motility, viability, sperm membrane integrity and acrosome status in Arabian stallion spermatozoa after cooling or after freezing and thawing compared with controls (non-supplemented media) or with those supplemented with other concentrations of trehalose or zinc sulphate. Supplementation of semen extender with BSA did not improve sperm motility or cryosurvival of Arabian stallion spermatozoa after cooling or after freezing and thawing. In conclusion, supplementation of semen extender with non-enzymatic antioxidants (trehalose or zinc sulphate) improved the quality of chilled and frozen/thawed Arabian stallion spermatozoa. The most beneficial effects occur when semen diluent was supplemented with 150 mM trehalose or 200 μM zinc sulphate.     

Spatio-temporal variation in the genetic composition of wild populations of pearl oyster (Pinctada margaritifera cumingii) in French Polynesia following 10 years of juvenile translocation

Abstract The genetic impact of the cultural practice of spat collection and translocation between genetically distinct stocks of black-lipped pearl oyster, Pinctada margaritifera cumingii, was studied by comparing samples collected in the 1980s and 2000s from seven atolls in French Polynesia. An amova revealed homogenization of the previously genetically distinct wild stocks of Tuamotu-Gambier and Society archipelagos (the indices of genetic differentiation among archipelagos and among populations within archipelagos, respectively, Phi(CT) and Phi(ST), decreased from 0.032* and 0.025*, respectively, to 0.006(NS) and 0.007(NS)). These results suggest high success of spontaneous reproduction in farms, probably due to the very high density of cultivated pearl oysters, and underline the importance of genetic monitoring of future hatchery produced stocks.     

Trehalose improves rabbit sperm quality during cryopreservation

High levels of reactive oxygen species are associated with spermatozoa cryopreservation, which bring damage to functional spermatozoa. The aim of the present study was to investigate whether and how the freezing extenders supplemented with trehalose was beneficial for the survival of rabbit spermatozoa. semen was diluted with Tris-citrate-glucose extender addition of different concentrations of trehalose. Addition of 100 mM trehaose significantly improved post-thaw rabbit sperm parameters, such as motility, acrosome integriy, membrane integrity and mitochondrial membrane potential. Moreover, when freezing extenders supplemented with trehalose, activities of catalase (CAT), superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) of post-thaw spermatozoa were enhanced, meanwhile, reactive oxygen species (ROS) level and Malondialdehyde (MDA) content were decreased. The results suggest that freezing extenders supplemented with 100 mM trehalose resulted in less ROS level and MDA content, higher motility and mitochondrial membrane potential as well as the integrity of acrosome and plasma membrane. Supplementation of trehalose with freezing extenders is beneficial to the rabbit breeding industry.     

Cryopreservation of fetal skin is improved by extracellular trehalose

In this study, we tested a non-permeating cryoprotectant, trehalose, in combination with dimethyl sulfoxide (Me(2)SO) in the cryopreservation of human fetal skin and compared it to Me(2)SO and glycerol, protocols that are routinely used by skin banks. The viability of fetal skin from four groups (fresh, and cryopreserved with glycerol, Me(2)SO, or trehalose/Me(2)SO) were evaluated using an in vitro membrane integrity assay and by transplantation to immunodeficient mice. The membrane integrity assay showed a 90% integrity in fresh, unfrozen fetal skin. The number of intact cells dropped to 23 and 44% in fetal skin cryopreserved with glycerol and Me(2)SO, respectively. When trehalose was added to the cryopreservation medium containing Me(2)SO, the membrane integrity rose to 65%. When transplanted to immunodeficient mice, fetal skin cryopreserved with trehalose/Me(2)SO showed a graft performance indistinguishable from fresh unfrozen fetal skin and strikingly better graft take than that of fetal skin cryopreserved with Me(2)SO or glycerol only. These results suggest that cryopreservation protocols routinely used the skin banks can be improved by combining sugars such as trehalose with a permeating cryoprotectant.     

Effects of glycine and alanine on short-term storage and cryopreservation of striped bass (Morone saxatilis) spermatozoa

Three experiments were designed to examine the effects of the amino acids glycine and alanine on short-term storage and cryopreservation of striped bass spermatozoa. In the first experiment, the effect of glycine on post-equilibration motility was evaluated. In the presence of 2.5 or 5.0% Me(2)SO, glycine treatments (25, 50, and 75 mM) yielded higher (P<0.05) post-equilibration motility at all equilibration times examined compared to the control. There was no difference (P>0.05) among these three glycine treatments. In the second experiment, glycine and alanine at concentrations of 25, 50, 75, or 100mM were evaluated for post-thaw motility in the presence of 2.5 or 5% Me(2)SO. When compared to the control, both the glycine and alanine treatments showed positive effects on post-thaw motility at all concentrations tested. The highest (P<0.05) post-thaw motility was achieved with 50mM glycine or 75 mM alanine using 5% Me(2)SO. No interaction (P>0.05) between Me(2)SO and glycine or alanine was observed, indicating that the effect of glycine or alanine was independent of the concentrations of Me(2)SO. In the third experiment, glycine was evaluated for sperm motility, after short-term refrigerated storage and after cryopreservation of the same refrigerated semen. Sperm motility decreased after 24h of refrigerated storage in 50mM glycine treatment and the control, when compared to fresh sperm motility. However, 50mM glycine treatment yielded higher (P<0.01) sperm motility after both 24 and 48 h of storage as well as higher (P<0.01) post-thaw motility when compared to the control. An average of 30+/-2.9% and 16+/-2.4% post-thaw motility was achieved with the 50mM glycine treatment after 24 and 48 h of refrigerated semen, respectively.     

Sensitivity of newly transformed juveniles of the freshwater pearl mussel: Margaritifera margaritifera to acute toxicity of a wide range of contaminants

Hydrobiologia - The freshwater pearl mussel (FWPM) Margaritifera margaritifera is a bivalve mollusk critically endangered in Europe. There is a special concern about the conservation of this...     

Trehalose-enhanced fluidity of the goat sperm membrane and its protection during freezing

In an attempt to find a suitable freezing method for goat semen, two experiments were conducted to study the influence of trehalose on the cryopreservation of goat spermatozoa. In experiment 1, goat spermatozoa were frozen in trehalose extender (0.375 M) alone (100%) or at different combinations of trehalose with Tris-citric acid-glucose (TCG) extender (0%, 25%, 50%, 75%). Final concentrations of 20% (v:v) egg yolk and 4% (v:v) glycerol were employed in the extenders (osmolality = 370, pH = 7). Sperm motility was assessed using a computer-assisted sperm analysis system and acrosome integrity was assessed using fluorescein isothiocyanate-labeled peanut agglutinin (FITC-PNA). The sperm-motility parameters improved significantly by increasing the concentration of trehalose (P < 0.05) and significantly high recovery rates for the motility parameters were also achieved by a high concentration of trehalose (P < 0.05). Motility of the frozen-thawed spermatozoa after a 3-h incubation improved significantly with increasing concentrations of trehalose in the extender (P < 0.05). The 75% and 100% trehalose extenders yielded a significant increase in the percentage of spermatozoa with intact acrosome (P < 0.05). In experiment 2, merocyanine 540/Yo-Pro staining was used to study the influence of trehalose on membrane fluidity compared with that of sucrose and TCG. Percentage of cells with high merocyanine fluorescence was significantly higher in spermatozoa treated with trehalose than sucrose or TCG (P < 0.05), indicating a significantly highest membrane fluidity of sperm samples extended with trehalose solution. We thus conclude that the substitution of a Tris-citric acid diluent composition with trehalose significantly improves the freezability of goat spermatozoa. Furthermore, the cryoprotective effects of trehalose observed in this study may be due to enhanced sperm membrane fluidity before freezing.     

Suitability of cryoprotectants and impregnation protocols for embryos of Japanese whiting Sillago japonica

The purpose of this study was to examine the suitability of cryoprotectant agent (CPA) impregnation protocols for the embryos of Japanese whiting (Sillago japonica), a small-sized, easy-to-rear, and prolific marine fish which may constitute a suitable experimental material for the development of cryopreservation methods for fish embryos. Our immediate goals were to assess the toxicity and permeability of various CPAs to whiting embryos of different developmental stages. Exposure of gastrula, somites, tail elongation, and pre-hatching embryos to 10%, 15%, and 20% solutions of propylene glycol (PG), methanol (MeOH), dimethyl sulfoxide (Me2SO), dimethylformamide (DFA), ethylene glycol (EG), and glycerol (Gly) in artificial sea water (ASW; 33 psu) for 20 min revealed that CPA toxicity for whiting embryos increased in the order of PG相似文献   

Development of a simple sperm cryopreservation model using a chemically defined medium and goat cauda epididymal spermatozoa

This investigation was carried out to develop a simple sperm cryopreservation model using a chemically defined synthetic medium (modified Ringer's solution) and mature goat cauda epididymal sperm as the model system. Rates of cooling, freezing, and maximum freezing temperature were manipulated with the help of a computer-controlled programmable biofreezer. Highly motile goat cauda sperm dispersed in a modified Ringer's solution was subjected to the freezing protocol: cooling 0.25 degrees C min(-1) to 5 degrees C, 5 degrees C min (-1) to -20 degrees C, 20 degrees C min(-1) to -100 degrees C, prior to plunging into liquid nitrogen. In the absence of any cryoprotective agent, all of the spermatozoa lost their motility. Addition of glycerol (0.22 to 0.87 M) caused a dose-dependent increase of sperm motility recovery. The highest recovery of forward and total motility was (32 and 35%, respectively) at 0.87 M. Further increase of the glycerol concentration caused a marked decrease in motility. Changes in the cooling rate particularly before and during freezing had a notable effect on the sperm motility recovery. There was no or low recovery (0-18%) of sperm motility when the cells were transferred directly to liquid nitrogen from the initial two cooling stages. The data demonstrate the importance of all of the cooling stages in the cryopreservation of the cells. Like glycerol, dimethyl sulfoxide (Me(2)SO) and ethylene glycol also showed a dose-dependent increase in motility recovery as well as a biphasic curve of cryoprotection. At optimal concentrations, dimethyl sulfoxide (1.00 M) and ethylene glycol (1.29 M) were effective in recovering sperm motility to the extent of 20 and 13%, respectively. Thus these reagents have markedly lower cryoprotection potential than glycerol.     

Cryopreservation of sperm in farmed Australian greenlip abalone Haliotis laevigata

This study investigated factors important to the development of the liquid nitrogen (LN) vapor sperm cryopreservation technique in farmed greenlip abalone Haliotis laevigata, including (1) cryoprotectant agent (CPA) toxicity; (2) cooling temperature (height above LN surface); (3) thawing temperature; (4) sperm to egg ratio; and (5) sugar supplementation, using sperm motility, fertilization rate or integrity/potential of sperm components and organelles as quality assessment indicators. Results suggested that among the single CPAs evaluated 6% dimethyl sulfoxide (Me2SO) would be the most suitable for sperm cryopreservation in this species. The highest post-thaw sperm motility was achieved with the sperm that had been exposed to LN vapor for 10 min at 5.2 cm above the LN surface, thawed and recovered in 60 and 18 °C seawater bathes, respectively after at least 2 h storage in LN. The highest fertilization rates were achieved at a sperm to egg ratio of 10,000:1 or 15,000:1. Addition of 1% glucose or 2% sucrose produced significantly higher post-thaw sperm motility than 6% Me2SO alone. Among the three cryoprotectant solutions further trialled, 6% Me2SO + 1% glucose produced the highest fertilization rate of 83.6 ± 3.7%. Evaluation of sperm has shown that the addition of glucose could significantly improve the sperm plasma membrane integrity and mitochondrial membrane potential. These results demonstrated a positive role of glucose in the improvement of sperm cryopreservation in farmed greenlip abalone.     

Cryoprotectants lead to phenotypic adaptation to freeze-thaw stress in Lactobacillus delbrueckii ssp. bulgaricus CIP 101027T

This study was conducted to investigate the ability of cryoprotective chemicals to induce phenotypic cryoadaptation in Lactobacillus delbrueckii ssp. bulgaricus CIP 101027T. Tolerance to negative temperature stress (freezing at -20 degrees C and thawing at 37 degrees C) was induced by pretreatment with Me(2)SO, glycerol, lactose, sucrose, and trehalose. Interestingly, Me(2)SO has a significantly greater cryoprotective effect than glycerol. Furthermore, lactose, sucrose, and trehalose, often referred to as osmotica, were shown to have greater cryoadaptive than cryoprotective properties. These results suggest that bacteria such as L. delbrueckii ssp. bulgaricus could be phenotypically adapted to freezing and thawing by an osmotic stress applied prior to freeze-thaw stress.     

Control of biofouling on pearl oysters Pinctada imbricata using wax and Chinese herbs

Abstract

Biofouling poses severe challenges to pearl oyster Pinctada imbricata culture in China, and controlling it is both labor- and capital-intensive. The antifouling properties of wax, and wax mixed with Chinese herbs, sprayed onto pearl oyster shell surfaces during peak biofouling seasons were evaluated. Pearl oysters coated with three wax treatments (plain wax, Chinaberry seed extract, Chinese honeylocust fruit extract) and a control (no treatment), were cultured in nets for up to 60?days. Mortality rate, fouling organism and pearl-oyster weights, and shell height are reported for individual oysters on each of six sampling dates. With the exception of oysters submerged for 12?days, all oysters were significantly affected by treatment type and submersion duration. Fouling weight increased more rapidly over time in the control-treatment oysters. Wax-based coatings deterred fouling-organism settlement on oysters for at least 2?months during the intensive fouling season, reducing mortality and not adversely effecting growth.