Inclusion of supplemental antibiotics (amikacin - penicillin) in a commercial extender for stallion semen: Effects on sperm quality, bacterial growth, and fertility following cooled storage.

In this study, the effectiveness of supplementing INRA-96® extender (INRA-Control; original antibiotic formulation: potassium penicillin G = 38 µg/mL; gentamicin sulfate = 105 µg/mL; amphotericin B = 0.315 µg/mL) with amikacin sulfate and potassium penicillin G (AP) was determined. In Exp. 1, two sources of amikacin (INRA-AP-Sigma or INRA-AP-GoldBio) in combination with penicillin G were compared with ticarcillin/clavulanate (INRA-Tim) or no-supplemental antibiotics (INRA-Control) to examine effects on sperm quality and commensal bacterial growth. No differences were detected in semen quality among treatments after 30 min of exposure (Time 30min) or 24 h of cooled storage (Time 24 h; P > 0.05). At both time periods, commensal bacterial growth was significantly lower in Groups INRA-AP-GoldBio and INRA-AP-Sigma than in INRA-Tim or INRA-Control (P < 0.05). In Exp. 2, increasing doses of amikacin sulfate (GoldBio) plus potassium penicillin G (Sigma) - AP (AP-1000, 2000, 3000, 4000 or 5000 µg-IU/mL, respectively) were added to INRA-96® extender and their effects on sperm quality and commensal bacterial growth were evaluated at Time 30min and Time 24 h. Slight reductions in progressive motility and viability were observed at Time 30min in Groups AP-4000 and AP-5000 as compared to other treatment groups (P < 0.05); however, no differences in sperm quality were detected among treatment groups at Time 24 h (P > 0.05). At both time periods, commensal bacterial growth was significantly lower in Groups AP-3000, AP-4000 and AP-5000 than in AP-1000 and AP-2000 (P < 0.05). In Exp. 3, a breeding trial was conducted to determine the effect of adding a high dose of AP (AP-5000) to INRA-96® extender on resulting pregnancy rates of mares bred with cool-stored semen (Time 24 h). Numerical, but not statistical differences, were observed in pregnancy rates between the mares bred with INRA-Control (6/11; 55%) or INRA-AP-5000 (9/11; 82%; P > 0.05). Supplementation of INRA-96® extender with two different concentrations of AP (AP-1000 or AP-5000) was tested in two clinical cases of stallions where semen was moderately to heavily contaminated with Pseudomonas aeruginosa, or both Klebsiella pneumoniae and Pseudomonas aeruginosa. In both cases, addition of AP resulted in a considerable decrease on bacterial growth in cool-stored semen when compared to the use of the original INRA-96® extender without supplemental antibiotics. In conclusion, the addition of amikacin sulfate and potassium penicillin G to INRA-96® extender allowed for effective control of commensal bacteria without affecting sperm quality. Higher doses of amikacin and penicillin can be safely added to INRA-96® extender to improve the antibacterial activity of this extender against commensal, and potentially pathogenic bacteria, while sperm quality and fertility of cooled semen remains unaffected. Based on the results of the present study, we currently recommend that INRA-96® extender can be safely supplemented with amikacin/penicillin by using a conventional dose of 1000 µg/mL - 1000 IU/mL as a prophylactic measure in cases where contamination of the ejaculates with commensal bacteria is evident. Alternatively, a high dose (5000 µg/mL - 5000 IU/mL) can be used as a control method for potentially pathogenic bacteria.

Effect of feeding a DHA-enriched nutriceutical on the quality of fresh, cooled and frozen stallion semen.

Eight stallions were used in 2 x 2 crossover study to determine if feeding a nutriceutical rich in docosahexaenoic acid (DHA) would improve semen quality. Stallions were randomly assigned to one of two treatment groups (n = 4 per group). Stallions were fed their normal diet (control) or their normal diet top-dressed with 250 g of a DHA-enriched nutriceutical. Feeding trials lasted for 14 week, after which a 14-week washout period was allowed and the treatment groups were reversed for another 14 week feeding trial. Feeding the nutriceutical resulted in a three-fold increase in semen DHA levels and 50% increase in the ratio of DHA to DPA in semen. Sperm motion characteristics in fresh semen were unaffected by treatment. After 24 h of cooled semen storage in an Equitainer, total and progressive motility did not differ between treatment groups, but sperm from stallions fed the nutriceutical exhibited higher velocity and straighter projectory (P < 0.05). After 48 h of cooled storage, increases in the percentages of sperm exhibiting total motility (P = 0.07), progressive motility (P = 0.06) and rapid motility (P = 0.04), were observed when stallions were being fed the nutriceutical. For a subset of four stallions, whose progressive sperm motility was <40% after 24 h of cooled storage when fed the control diet, feeding the nutriceutical resulted in improvements in mean progressive motility of sperm after 24 h (P = 0.10) and 48 h (P = 0.03) of storage. Feeding the nutriceutical resulted in similar improvements in motion characteristics being observed in frozen-thawed semen. While it appears that feeding the nutriceutical may improve the motion characteristics of cool-stored stallion semen, it may be most beneficial for stallions of marginal fertility whose sperm do not tolerate the rigors of cooling and storage. The nutriceutical also appeared to improve the freezability of semen. More dramatic improvements in semen quality may be observed if modifications in the main fat content of the diet are incorporated with the DHA supplement.