245 OOCYTE RECOVERY BY OVUM PICK UP AND EMBRYO PRODUCTION IN MURRAH AND NILI-RAVI BUFFALOES (BUBALUS BUBALIS) IMPORTED IN CHINA

Abstract

In this preliminary study, in vitro embryo production and cryopreservation in two river type buffaloes (Murrah and Nili-Ravi) imported into China have been carried out. The objective of the study was enhancement of the genetic merit and productive performances of imported river buffaloes in conjunction with the utilization of local swamp buffaloes. In order to improve milk and meat production in China local swamp buffaloes (2n = 48), which are the predominant subspecies, have been crossbred with imported river buffaloes (Murrah and Nili-Ravi: 2n = 50). At present, several hundred thousand crossbred heads have been produced, and although both males and females can reproduce with 2n = 49 crossbred buffaloes, their reproductive performances are significantly reduced when compared to 2n = 50 buffaloes. As an alternative approach, a program of embryo production in river buffaloes and transfer into both river and swamp buffaloes has been implemented at the Guangxi Buffalo Research Institute, in Nanning, P.R. China. Some preliminary results are presented: from a start-up experiment, a total of 46 river buffaloes were subjected to 2 to 3 ovum pickup sessions at 4-day intervals. A total of 750 antral follicles were punctured and 495 (66%) cumulus-oocyte complexes (COCs) were retrieved. Only COCs characterized by at least one layer of granulosa cells (n = 451; 91.1%) were considered for in vitro maturation (IVM). COCs were matured in TCM 199 + 10% FCS, 0.5 μg/mL FSH, 5 μg/mL LH, and 1 μg/mL estradiol in the presence of cysteamine (50 μM) at 39°C under 5% CO₂ in humidified air for 24 h. Of the initial 451 COCs matured, only 277 could be considered for in vitro fertilization (IVF). IVF was performed at 39°C under CO₂ in humidified air in TALP medium supplemented with 0.2 mM penicillamine, 0.1 mM hypotaurine, and 0.01 mM heparin. Frozen/thawed sperm from a tested bull was treated by swim-up procedure and used at a final concentration of 20 million/mL. Following 20 to 22 h of co-incubation, presumptive zygotes were cultured in SOF medium, supplemented with essential and non-essential amino acids and 8 mg/mL BSA, in a gas atmosphere of 5% CO₂, 7% O₂, and 88% N₂. A total of 41 (14.8%) blastocysts were produced, of which 33 were vitrified and 8 transferred immediately into available swamp and river buffalo recipients. Two calves were born (25%) from the transfer of fresh embryos into one river and one swamp buffalo. In vitro embryo production in the buffalo species is still characterized by a high degree of variable results. However, these preliminary results reinforce the need to implement newly developed reproductive technologies not only for speeding up genetic gain of already productive species, but also for the utilization of local breeds characterized by reduced productive performance.