This study was conducted at the Department of Surgery and Obstetrics, BAU during the period from January to December 2001 under the BAURES project, Evaluation of Multiple Ovulation and Embryo-Transfer Technique in Black Bengal goat in Bangladesh. The eleven Black Bengal goat of 1 and 2 years old, weighing 10-15 kg were flushed as a donor in three times (Flush I, 5 Flush II 2, and Flush III 4) in one years time within the MOET treatment. The number of recipient does was also 11 (Flush I 4, Flush II, 4 and Flush III, 3). The goats were housed in well-ventilated, concrete floor; house with adequate light. The goats were fed sufficient green grass and a concentrate mixture containing equal amounts of wheat bran, crushed wheat, soybean bran, maize grain at the added with 2% iodinated salt and 1% vitamin and mineral premix at the rate of 0.5 kg/head. The goats were acclimatized for 3 months and checked for normal cycle before giving treatment.
Synchronization of oestrus: Synchronization of oestrus was done with two intramuscular injection of 1 ml equivalent to 2 mg Gabbrostim® (Alfaprostol, VETEM SPA, Porto Empedocle, Italy) at 11 days apart in both donor and recipient goats. The donor and recipient goats were observed for signs oestrus at 4h interval from 700h to 2000h. Onset of oestrus was detected on the basis of the behavioral and clinical signs.
Superovulatory response: Three doses of PMSG (Folligon®, intervet International B.V., Boxmer, Netherlands) 900 IU (n = 5), 800 IU (n = 2) and 700 IU (n = 4) were used for superovulation. The superovulation treatment was initiated at Day 10 or Day 11 of the oestrous cycle (Day 0 = day of oestrus). A luteolytic dose of synthetic prostaglandin F2α (alfaprostol, Gabbrostim®, 2 mg, VETEM S.P.A, Porto Empedocle, Italy) was injected intramuscularly 48 hours after injection of PMSG to induce oestrus.
Natural service and artificial insemination: Both natural service and artificial insemination were performed to fertilize the superovulated ovum. The donor goats were inseminated at 6 hour after the onset of oestrus. Then the goats were allowed to serve by the buck at about 6 hour after insemination.
Embryo collection: Embryos were collected from donor goats by surgical embryo collection method on Day 6 or 7 after natural service and AI. A sedative, chlorpromazine hydrochloride (Largactil®, Rhone-Poulenc, Bangladesh), was injected IM at the dose rate of 0.1 mg/kg body weight approximately 15-30 minutes before surgery. The rear half of the abdominal wall, anterior to the udder, was clipped, shaved, scrubbed with antiseptic solution (Hexisol®; 0.5% chlorhexidine gluconate, ACI, pharmaceuticals, Ltd, Bangladesh) and dried. Paravertebral nerves were blocked by injecting 12 ml of 2% lignocaine hydrochloride, 4 ml in last thoracic, 3 ml in both first and second lumbar and 2 ml in third lumbar vertebral spinal nerve (Jasocaine®, Jayson pharmaceuticals, Ltd, Bangladesh). The skin and underline tissue was desensitized by infiltrating 5 ml of 2% lignocaine hydrochloride.Post collection care and management: The skin wounds of all donors goats were covered with benzoin seal. The donor goats were immediately injected with 2mg alfaprostol and 0.5 gm streptomycin with 5 lac penicillin (Streptopen®, 0.5gm, Renata Animal Health, Dhaka, Bangladesh) to induce oestrus and protect from microbial infection.
Selection and grading of collected embryos: The flushed out concentrated medium was divided in to two portion and placed in a searching petridish (100-mm diameter, CORNING® New York, USA) and examined under a stereomicroscope (20X, Binocular Stereomicroscope®, XTS-110, China) to visualize the embryos. The identified embryos were individually picked up with a micropipette controller (Assistent Micro Classic® No. 558, Kart Hecht GmbH & Co. KG, Germany) and placed in a small sized petri dish (35 mm diameter, FALCON®, Betcon Dickinson Company, Japan) containing fresh, filtere-sterilized (0.22μm pore size, Minisart®, Minitub GmbH, Tiefenbach b. Landshut, Germany), medium. The embryos were washed 4 times in the same way and evaluated subsequently. The embryos were classified in four groups according to the method described by Flores- Foxworth (1997) and Bari et al.(1999). Grade 1- the embryo was nearly perfect with more than 98% of the cell mass active and healthy, Grade 2- about 70 to 98% of the cell mass was active and healthy and some extruded blastomeres were found, Grade 3- a poor quality embryo with less than 70% of the cell mass active and healthy and several extruded blastomeres were present, Grade 4- degenerated embryos with no active cell mass and distinct plasma membrane.
The data were analyzed statistically by the paired t-test. The donor goat was suspended in head down position in the wooden made cradle. The abdomen was covered with a sterile towel cloth attached with towel clips. A 1-2 inch para-midline incision was made immediately cranial to the udder using regional paravertebral and local anaesthesia.