Effects Of Sodium Diacetate And Kojic Acid On The Quality Of Boar Semen Of Guanzhong Black Pig During Normal Temperature Preservation

The quality of the diluted powder is very important in the porcine artificial insemination(AI),which determines the quality of diluted boar semen and the reproductive performance of the sow after AI.However,semen is very susceptible to bacterial contamination;and when semen is keeping stored or being transported,the oxidative stress(OS)which is induced by the imbalance of reactive oxygen species(ROS)and antioxidants can do harm to sperm,so it’s very essential to add some bacteriostats and antioxidants to the diluted powder.Currently,the dilution powder for the preservation of boar semen at room temperature are almost designed for the lean meat breeds,but for Guanzhong black pig and other cultivated varieties rarely.In this study,antimicrobial sodium diacetate(SDA)and antioxidant kojic acid were applied to the preservation of Guanzhong black pig semen at room temperature.The bacteriostatic test,sperm viability,plasma membrane integrity,mitochondrial integrity,Acrosome integrity and total antioxidant capacity(T-AOC)and some other detection indexes were experimented to explore the feasibility of adding SDA instead of gentamicin into diluted powder and the efficacy of kojic acid on the protection of semen,this study was aimed to provide a theoretical support for the research of dilution powder for guanzhong black pig semen.The following are the main results in this study:1.The results showed that more than 100 kinds of bacteria were detected in the process of preservation of guanzhong black pig’s diluted semen,most of which were gram-positive bacteria: Staphylococcus,Streptococcus Aerococcaceae and Bacillacea were accounted for 24.13%,13.39%,11.84% and 6.91%,respectively.However,with the prolong of storage time,the proportion of gram-negative bacteria especially Enterobacteriaceae was increased,and it came to 97.76% on the fifth day.2.Bacteriostatic test found that sodium diacetate had less inhibition effect on Escherichia coli than gentamicin(P < 0.05),although it had a certain inhibitory effect indeed.The inhibition zone diameter of sodium diacetate group was increased significantly in the surface of LB solid medium which coated with Acinetobacter lwoffii and Staphylococcus aureus with the increase of concentration(P < 0.05).In contrast,the OD value was significantly decreased with the increased concentration of sodium diacetate in LB liquid(P < 0.05).However,when the concentration increased to above 0.8 g/L,it had a similar inhibitory effect with gentamicin on Acinetobacter lwoffii and Staphylococcus aureus(P < 0.05).3.The addition of sodium diacetate at concentrations above 0.2 g/L had a significant effect on the osmotic pressure of the diluted semen(P < 0.05).The effect of semen preservation was firstly increased and then decreased when semen diluted in BTS solution added 0~0.8 g/L sodium diacetate.Adding 0.2 g/L sodium diacetate alone can effectively inhibit bacterial growth,improve sperm motility,plasma membrane,acrosome and mitochondrial integrity(P < 0.05).BTS solution with high concentrations(> 1 g/L)of sodium diacetate is not conducive to Guanzhong black pig sperm preservation.4.Adding 0.2 g/L sodium diacetate in BTS solution instead of 40% gentamicin can achieve the equivalent of 250 mg/L gentamicin on the preservation of semen.5.BTS solution with less than 0.1 g/L kojic acid can significantly improve the sperm plasma membrane,acrosome integrity,enhance the total antioxidant capacity of sperm(P < 0.05),especially 0.04 g/L group has the best performance for the sperm viability was 61.45% at the 5th day which can prolong the sperm storage time for another 2 days.In addition,the sperm preservation effect of 0.06 g / L group was not significantly different with 0.04 g/L group(P > 0.05).6.On the whole,the conbine of different concentrations of sodium diacetate with kojic acid is conducive to Guanzhong black pig sperm preservation compared to the blank control group,which can improve the survival rate,acrosome integrity rate.Nevertheless,the combination of the two has no significant effect than adding each of them alone.In summary,the Gram-negative bacteria appeared in the semen of Guanzhong black pig during the preservation at room temperature showed an increasing trend,which accounted for the vast majority of Enterobacteriaceae.Adding 0.2 g/L sodium diacetate in BTS solution could replace 40% gentamicin and promote the preservation of porcine semen.At the same time,adding 0.04 g/L kojic acid in BTS solution had the best antioxidation protection effect on sperm preservation at room temperature,but the combination of sodium diacetate and kojic acid had no difference.The results of this study are of great significance to solve the problem of antibiotic resistance and sperm oxidative damage in porcine semen preservation at room temperature,and provide experimental basis for independent diluted powder research.