No abstract

The success of buffalo invitro embryo production (IVEP) depends, in part, on the number and competence of oocytes. Superstimulation with FSH has been linked to oocyte quality and embryo yield, and anti-Müllerian hormone (AMH) has been proposed as an indicator of ovarian reserve. The objective was to evaluate FSH stimulation in one ovum pick-up (OPU) cycle conducted 7 or 14 days after follicular ablation; we additionally analysed residual FSH treatment effect carried over to a subsequent aspiration cycle. Finally, we evaluated the relationship between plasma AMH and FSH response. Nine multiparous, cycling Murrah donors (603±7.0kg, body condition score=3.5) were subjected to complete follicular ablation followed by OPU 1 week later (T0 Control), and plasma AMH was determined at this point. They were then randomly allocated to FSH stimulation (3 days b.i.d. in descending doses, total 300mg of pFSH, Vetpharm, with the last injection 48h before OPU) with aspiration conducted 7 (T7 FSH, n=5) or 14 days (T14 FSH, n=4) post-follicular ablation. The residual effect (T Residual, n=5) was evaluated in one subsequent, nonstimulated OPU cycle conducted 1 week later (9 days after last FSH administration). For plasma AMH, concentration was measured using ELISA kit (cat. no. AL114, AnshLabs; sensitivity 1.0 pgmL−1, interassay variation <5%, additional validation conducted with serial dilutions of buffalo plasma, r=0.98 against standard curve). Continuous data were analysed with ANOVA and Tukey's post hoc comparisons; categorical data were analysed using Chi-squared test. Pearson coefficient was used to evaluate correlation between AMH, follicular reserve, and invitro performance. Superovulatory treatment resulted in a higher number of follicles and good quality oocytes (grades 1 and 2) per donor, irrespective of OPU interval (T7 FSH: 3.0±2.55 and T14 FSH: 1.5±0.5 vs. T0 Control: 0.45±0.38 and T Residual: 0.4±0.4; P<0.05). There were more cleaved embryos (T7 FSH: 26%, T14 FSH; 30% vs. T0 Control: 7% and T Residual: 20%) and blastocysts (T7 FSH: 21%, T14 FSH; 30% vs. T0 Control: 5% and T Residual: 0; P<0.05) from FSH-treated donors. There was a strong correlation (r=0.9439) between follicle count and plasma AMH, and weaker relationships for oocytes recovered (r=0.3975), cleaved embryos (r=0.2317), or blastocysts (r=0.0106). Results indicate that FSH treatment with a 48-h coasting period results in significant improvements in follicular population, oocyte quality, and embryonic development; these effects are observed independently of the interval between initial follicular ablation and OPU. No residual effects of FSH treatment were observed, indicating that the beneficial effects of superstimulation are not carried over to the following OPU cycle. In conclusion, a strong correlation coefficient was established between initial plasma AMH level and follicular response to FSH treatment, indicating that it may be used as a selection criterion for animals likely to exhibit a good response to superstimulatory treatment.

It is well known that buffaloes have seasonal anoestrus and that the use of melatonin reduces the effects of season in reproduction, articularly among sheep and goats. This study evaluates the use of melatonin on water buffaloes to increase pregnancy rates in a fixed-time insemination programme. The study was conducted on an Argentinian buffalo herd, located in the Corrientes Province, during an unfavourable reproductive season. Artificial insemination cycles were performed in September and December 2018. Sixty animals were selected, and melatonin was administered to 29 females at a dose of 18 mg/50 kg. Ten days after the melatonin implant, ovulation synchronization was started. Animals were inseminated with frozen semen of a single bull. Pregnancy was evaluated by ultrasound 35 days after insemination. The pregnancy rate of the two repetitions was 34.62% for September and 6.4% for December (P=0.007). No significant differences were found in the parameters associated with follicular development and formation of the corpus luteum. Analysis of the effects of melatonin on the two replicates showed that there was no statistically significant effect of ovarian response regarding the analysed follicular development parameters. There was also no effect on pregnancy rates (17.85% vs. 20.68%; P=0.15), or on follicular development for treatment and control. In conclusion, this study was not able to demonstrate an effect of melatonin on the ovarian response of buffaloes as reported by other authors. Other factors, such as environmental conditions of subtropical areas and species specificities, may have a more significant effect on buffalo endocrinology. These factors should be evaluated to improve the results of fixed-timed artificial insemination programmes.