Fertilization success of Siberian sturgeon semen cryopreserved using glucose-methanol extender
Introduction
Several sturgeon species are close to extinction due to cutting migration routes, overfishing and pollution. Application of sperm cryopreservation to sturgeon aquaculture enables gene banking of endangered populations, facilitate artificial fertilization and reduce the stock of maintained males. Recently, a simple and effective cryopreservation procedure with the use of glucose-methanol extender was established for rainbow trout semen (Ciereszko et al., 2014). The aim of this study was to test the possibility of application of a simple glucose-methanol extender to cryopreservation of Siberian sturgeon semen.
Material and methods
Semen of Siberian sturgeon was cryopreserved in 0.25-ml straws in glucose-methanol extender at a ratio of 1:1 (semen: extender). In the first experiment the effect of glucose at concentrations 0, 0.10, 0.15, 0.20 and 0.30 M on sperm motility of fresh and cryopreserved semen was tested. The extender consisting of 0.1 M glucose and 15% methanol was used in the second experiment aimed to test sperm motility and fertilizing ability after cryopreservation. The sperm motility was examined for the fresh-diluted, equilibrated for 30 min and frozen-thawed sperm (n=6). The eggs were fertilized with fresh and thawed semen with spermatozoa to egg ratio 100 000:1. The fertilization success was established by calculating the percentage of hatched larvae. Results are expressed as mean ± SD. Data were subjected to repeated measures one-way ANOVA followed by Tukey's post hoc test. The hatching success was compared using t-test.
Results
Glucose concentration in the extender did not have any effect on the fresh-diluted semen but significantly affected post-thaw sperm motility parameters of Siberian sturgeon (Fig. 1).
The extender consisting of 0.10 and 0.15 M glucose and 15% methanol provided high and consistent results of post-thaw sperm motility (57 and 58%) and the first extender was arbitrally selected the subsequent experiments. The application of the cryopreservation procedure resulted in high post-thaw sperm motility (56 and 58% for 0 and 30 min equilibration, respectively; Fig. 2). Sperm motility was not affected by equilibration period both for fresh-diluted and cryopreserved semen. The fertilization success of cryopreserved semen (31%) was similar to that of fresh semen (36%; Fig. 3).
Discussion and conclusion
Our data demonstrated that fertilizing ability of frozen-thawed Siberian sturgeon semen, cryopreserved using the simple glucose-methanol extender, can be comparable to the values obtained for fresh semen. As such, this new protocol can be alternative method to previously described by Glogowski et al. (2002). Semen could be equilibrated at least 30 min before cryopreservation without loss of quality which allows quite long handling time before freezing. Further studies should be focused on scaling up this efficient cryopreservation technique for facilitation of artificial reproduction of Siberian sturgeon.
Acknowledgements
This work was supported by funds of the National Science Centre granted on research project 2011/01/D/NZ9/03738 and funds appropriated to the Institute of Animal Reproduction and Food Research, Polish Academy of Sciences.
References
Ciereszko, A., Dietrich, G.J., Nynca, J., Dobosz, S., Zalewski, T., 2014. Cryopreservation of rainbow trout semen using a glucose-methanol extender. Aquaculture, 420-421: 275-281.
Glogowski, J., Kolman, R., Szczepkowski, M., Horvath, A., Urbanyi, B., Sieczyński, P., Rzemieniecki, A., Domagała, J., Demianowicz, W., Kowalski, R., Ciereszko, A., 2002. Fertilization rate of Siberian sturgeon (Acipenser baerii, Brandt) milt cryopreserved with methanol. Aquaculture, 211: 367-373.
