Martin Nikolovski, Monika Dovenska, Ksenija Ilievska, Nikola Adamov, Branko Atanasov, Miroslav Radeski, Daniela Kirovski, Vladimir Petkov and Toni Dovenski
Reduced glutathione (GSH) and homologous ram seminal plasma (HSP), used as additives in cryopreserving (CP) media prior to freezing, showed conflicting results in retaining structural integrity and progressive motility in post-thawed ram spermatozoa. The aims of this research were: (1) to assess the effect of GSH and/or HSP supplementation via soybean-lecithin CP extender on cryopreserved ram spermatozoa viability, morphology and motility pattern; and (2) to assess the effect of incubation in the context of the previous aim. Quantitatively and qualitatively, homogenized and pooled ram ejaculates (N=10) were extended with one of the following extenders: control (C) – tris-based, GSH and HSP-free, experimental-1 (E1) – C + GSH 5 mM, experimental-2 (E2) – C + HSP 20 % and experimental-3 (E3) - GSH 5 mM + HSP 20 %. Following thawing, samples were taken at 0- and 3-hours from each group (n=10) and were assessed for spermatozoa viability, morphology, and motility pattern. C-0h samples yielded a spermatozoa population with low viability, altered head morphology and highly deviated motility pattern. E3-3h samples yielded spermatozoa with unaffected viability, head morphology and high progressive motility. In conclusion, E3 extender added to cryopreserved-thawed ram spermatozoa is most efficient in obtaining high viability, unaltered head morphology, and progressive motility.
Branko Atanasov, Ljupco Mickov, Igor Esmerov, Ksenija Ilievska, Martin Nikolovski and Toni Dovenski
Postpartum anestrus is a physiological phenomenon in high-producing dairy cows. Static ovaries have been related as major contributors for its occurrence causing a significant reproductive problem to the dairy industry. Different treatment methods have been employed with inconsistent rate of success in initiation of cyclicity, requiring further investigations in order to achieve satisfactory results. The aim of the present study was to compare the ovarian response in cows diagnosed with static ovaries, more than 60 days postpartum using two different hormonal treatment (GnRH and eCG) methods. A total of 58 acyclic cows (no CL, follicles<8mm, P4<0.5ng/mL) were randomly divided into three groups: GnRH (Group 1, n=23), eCG (Group 2, n=23) and Controls (n=12), and allocated thereafter, into subgroups according to the applied doses of GnRH (100μg or 250μg); eCG (750 IU or 1000 IU) whilst control group cows were left untreated. Daily follicular growth rate and treatment respond interval were estimated based on repeated ultrasound examinations. Blood serum P4 sampling was done on d -7, d-0 (start of the experiment) and on d 7 after ovulation. Resumption of cyclic activity occurred in 55.17% (32/58) of the treated cows, 56.52% in Group 1; 60.86% in Group 2 and 41.66% in the control group. Overall, the follicular growth rate was similar between the trials group and significant with regard to the cows in the control group (p<0.05). eCG or GnRH treated cows responded significantly faster 6.85±0.2 and 7.84±0.2 days, respectively, in comparison to the control group cows (17±0.7 days, p<0.001). Treatment with a single dose of GnRH or eCG caused resumption of follicular growth and ovulation following luteogenesis (forming CL) without significant changes in P4 concentrations on day 7 after ovulation (p>0.05). Cows in Group 2 had significantly higher incidence of multiple ovulations than cows in Group 1 (p<0.05). The eCG treatment resulted in a faster response and higher ovulation rate compared to GnRH treatment. In conclusion, both treatments have shown acceptable results in resumption of cyclicity in dairy cows with static ovaries.
Besir Jasari, Branko Atanasov, Irena Celeska, Miroslav Radeski, Martin Nikolovski and Toni Dovenski
Several reports indicated that a large proportion of dairy cows have not resumed cyclicity until day 60 after calving. These cows are traditionally classified as non-cycling (anoestrous or anovular cows). Static ovaries (SO, lack of luteal tissue and follicles >8 mm, and progesterone < 0.5 ng/mL) could be a possible underlying reason that contributes to a non-cycling status. Although SO affects both primiparous (PP) and multiparous (MP) cows, PP cows are more prone to be non-cycling than MP. Therefore, this study aims to compare the metabolic profiles and hormonal status between non-cycling PP and MP cows diagnosed with SO. One hundred and twenty one animals that did not express signs of oestrus until day 60 postpartum were grouped by parity (PP, n=58 and MP, n=63), then blood sampled and examined using transrectal ultrasonography. Blood samples were collected before the ultrasonographic examination. Out of those, 42 PP (72.4%) and 28 MP (44.4%) were diagnosed as non-cycling (bearing SO). Serum concentrations of triglycerides, cholesterol, total protein and albumin did not differ between parity groups. The glucose concentrations in PP cows (1.43 ± 0.59 mmol/L) and MP cows (1.69 ± 0.71 mmol/L) did not differ, however, they were less than the normal physiological concentration. In addition, no differences were detected between parity groups for concentrations of NEFA, β-HBA, progesterone and estradiol. In summary, we concluded that non-cycling PP and MP cows bearing SO have similar hormonal status and metabolic profiles.
Martin Nikolovski, Ljupco Mickov, Monika Dovenska, Vladimir Petkov, Branko Atanasov and Toni Dovenski
Motility patterns of spermatozoa are an indicator of their fertilizing capacity. Reduced glutathione (GSH) has been reported to induce positive effects on the biological quality of the frozen-thawed spermatozoa. The aim of this study was to investigate the effects of GSH addition to semen extender based on the following kinetic parameters: continuous line velocity (VSL), average path velocity (VAP), curvilinear velocity (VCL), amplitude of lateral head displacement (ALH), linearity (LIN), straightness (STR), beat cross frequency (BCF), total motility (tMOT) and progressive motility (pMOT), and to appoint them as indicators of its presence. A soybean-based extender was used for dilution of the semen samples, fractioned in two parts, one containing GSH (5 mMol/ml) and second without GSH. The ejaculates (n=48) were collected from two rams (January - May, 2013) which were classified in two groups according to the used extender: Group 1 (with GSH, n=24) and Group 2 (without GSH, n=24), and then frozen in liquid nitrogen on -196°C degrees in a programmable freezer. Assessment of the samples has been performed post-thawing (30 sec. at 37°C) on CASA equipment, acquiring kinetic parameters. Results showed that only VSL and BCF have a statistically significant difference between group1 and group 2 (102.98±15.13 vs. 88.47±20.63, t=2.77, p<0.01 and 32.01±2.68 vs. 89.47±2.92, t=3.13, p<0.01, respectively). The summary of the investigation concludes that none of the kinetic parameters could be appointed as indicators that confer the positive effect of GSH as an additive to soy-bean semen extender.