Effect of Bull and Sperm Preparation Method on in Vitro Fertilization of Buffalo Oocytes

The present study was designed to compare fertilization rates following oocyte exposure to spermatozoa from different buffalo bulls, using three sperm preparation methods i.e. percoll density gradient, swim-up (modified Ca free Tyrode’s medium and TALP medium) and sodium citrate washing prior to co-incubation with oocytes. Buffalo oocytes were aspirated from ovarian follicles within 1.5 to 2 hours after slaughter. They were matured in TCM-199 supplemented with 20% oestrus buffalo serum drops under mineral oil in CO2 incubator at 39°C for 24 hours. Matured oocytes were transferred to the fertilization droplets and inseminated with 1x10 capacitated sperms prepared by different experimental methods. Data collected on recovered sperm motility immediately after treatment and 24 hours after incubation (at 37°C) and cleavage rate of co-incubated oocytes were subjected to statistical analysis. The percentage of motile spermatozoa was significantly higher (P<0.05) in semen samples prepared by swimup method in Ca free Tyrode’s medium compared to other experimental techniques. Bull wise comparison showed significantly higher (P<0.05) motility in bull B1 (50.50 ± 5.92%), followed by bull B2 (46.46 ± 5.99%) and B3 (45.96 ± 5.79%). Fertilization rate was also significantly (P<0.05) higher for spermatozoa prepared by Swim-up method in Ca free Tyrode’s medium (63.75 ± 2.81%), followed by sodium citrate (26.70 ± 5.08%), swim-up TALP (29.14 ± 3.74%) and Percoll gradient density (23.89 ± 3.88%). Fertilization rate was significantly higher (P<0.05) in oocytes inseminated with semen from bull B1 (43.43 ± 8.59%), followed by B2 (33.38 ± 9.95%) and B3 (30.80 ± 9.56%). The results of present study indicate that bulls and sperm preparation methods differ in their contribution to in vitro fertilization rate. Further studies are suggested to ascertain the factors responsible for such specific effects.