Novel preservation condition without ultra-low temperature is needed for the study of pathogen in marine fishes. Freeze-drying is such a method usually used for preservation of terrigenous bacteria. However, studies u...Novel preservation condition without ultra-low temperature is needed for the study of pathogen in marine fishes. Freeze-drying is such a method usually used for preservation of terrigenous bacteria. However, studies using freeze-drying method to preserving marine microorganisms remain very limited. In this study, we optimized the composition of protectants during the freeze-drying of Edwardsiella tarda, a fish pathogen that causes systemic infection in marine fishes. We found that the optimal composition of protectant mixture contained trehalose(8.0%), skim milk(12.0%), sodium citrate(2.0%), serum(12.0%) and PVP(2.0%). Orthogonal and interaction analyses demonstrated the interaction between serum and skim milk or sodium citrate. The highest survival rate of E. tarda was observed when the concentration of Na Cl was 10.0, 30.0 and between 5.0 and 10.0 g L^(-1) for preparing TSB medium, E. tarda suspension and protectant mixture, respectively. When E. tarda was frozen at-80℃ or-40℃ for 6 h, its survival rate was higher than that under other tested conditions. Under the optimized conditions, when the protectant mixture was used during freeze-drying process, the survival rate(79.63%–82.30%) of E. tarda was significantly higher than that obtained using single protectant. Scanning electron microscopy(SEM) image indicated that E. tarda was embedded in thick matrix with detectable aggregation. In sum, the protectant mixture may be used as a novel cryoprotective additive for E. tarda.展开更多
Due to the small volume of goat semen ejaculate, just a few doses of goat semen were produced when the sperm concentration is 100 × 10^/mL. The study was aimed to determine the viability of extended goat semen at...Due to the small volume of goat semen ejaculate, just a few doses of goat semen were produced when the sperm concentration is 100 × 10^/mL. The study was aimed to determine the viability of extended goat semen at refrigerated condition at 5 ℃ using varying sperm concentrations and evaluated if sperm concentration lower than 100 × 10^6/mL would affect the motility, viability and sperm morphology at refrigerated condition. Using an artificial vagina, ejaculated goat semen was collected from goat semen donor aged 1.5 year. Physical evaluation of the collected semen showed an average volume of 0.54 mL, mean pH of 6.8 and a milky white color with thick consistency indicative of high concentration. Fresh goat semen had an initial average of 76% with an average initial sperm concentration of 128 × 10^/mL. The semen was divided into four treatments： sperm concentration of 100× 10^/mL, 75 × 10^/mL, 50 × 10^/mL and 25 × 10^/mL, and were stored at 5 ℃ for a period of 10 d. The semen evaluation was performed for each of the four treatments every other day. Results showed that the sperm concentration of spermatozoa affected the duration of storage based on the sperm motility percentage, viability and morphology of spermatozoa. The extended goat semen with sperm concentration of 25 and 50 million sperm/mL is optimal for storage within 6 d that gave satisfactory percentage motile, viable and morphologically normal spermatozoa.展开更多
基金the National Natural Science Foundation of China (No. 31302206)Special Research Funds for Independent Innovation and Scientific & Technology Achievements Transformation of Shandong Province (No. 2014ZZCX06205)Agriculture Seed Improvement Project of Shandong Province
文摘Novel preservation condition without ultra-low temperature is needed for the study of pathogen in marine fishes. Freeze-drying is such a method usually used for preservation of terrigenous bacteria. However, studies using freeze-drying method to preserving marine microorganisms remain very limited. In this study, we optimized the composition of protectants during the freeze-drying of Edwardsiella tarda, a fish pathogen that causes systemic infection in marine fishes. We found that the optimal composition of protectant mixture contained trehalose(8.0%), skim milk(12.0%), sodium citrate(2.0%), serum(12.0%) and PVP(2.0%). Orthogonal and interaction analyses demonstrated the interaction between serum and skim milk or sodium citrate. The highest survival rate of E. tarda was observed when the concentration of Na Cl was 10.0, 30.0 and between 5.0 and 10.0 g L^(-1) for preparing TSB medium, E. tarda suspension and protectant mixture, respectively. When E. tarda was frozen at-80℃ or-40℃ for 6 h, its survival rate was higher than that under other tested conditions. Under the optimized conditions, when the protectant mixture was used during freeze-drying process, the survival rate(79.63%–82.30%) of E. tarda was significantly higher than that obtained using single protectant. Scanning electron microscopy(SEM) image indicated that E. tarda was embedded in thick matrix with detectable aggregation. In sum, the protectant mixture may be used as a novel cryoprotective additive for E. tarda.
文摘Due to the small volume of goat semen ejaculate, just a few doses of goat semen were produced when the sperm concentration is 100 × 10^/mL. The study was aimed to determine the viability of extended goat semen at refrigerated condition at 5 ℃ using varying sperm concentrations and evaluated if sperm concentration lower than 100 × 10^6/mL would affect the motility, viability and sperm morphology at refrigerated condition. Using an artificial vagina, ejaculated goat semen was collected from goat semen donor aged 1.5 year. Physical evaluation of the collected semen showed an average volume of 0.54 mL, mean pH of 6.8 and a milky white color with thick consistency indicative of high concentration. Fresh goat semen had an initial average of 76% with an average initial sperm concentration of 128 × 10^/mL. The semen was divided into four treatments： sperm concentration of 100× 10^/mL, 75 × 10^/mL, 50 × 10^/mL and 25 × 10^/mL, and were stored at 5 ℃ for a period of 10 d. The semen evaluation was performed for each of the four treatments every other day. Results showed that the sperm concentration of spermatozoa affected the duration of storage based on the sperm motility percentage, viability and morphology of spermatozoa. The extended goat semen with sperm concentration of 25 and 50 million sperm/mL is optimal for storage within 6 d that gave satisfactory percentage motile, viable and morphologically normal spermatozoa.
