Antibiotics in semen extenders - a multiplicity of paradoxes.

Addition of antibiotics to semen extenders was taken for granted for many years, from the time that commercial artificial insemination in livestock first began many decades ago. However, there is now a growing realisation that this non-therapeutic utilisation of antibacterial agents is contrary to current recommendations for prudent use that medical and veterinary professionals are advised to follow. Furthermore, antibiotics are not benign, having negative effects on sperm samples, the inseminated female, personnel and potentially the environment. The purpose of this review is three-fold: to highlight the fact that antibiotics are used in semen extenders, with the result that considerable amounts are used globally in animal breeding, to review recent studies on the negative aspects of using antibiotics for this purpose, and to look at possible alternatives. Recent changes in the legislation regarding semen extenders occurred in some, but not all, countries, leaving question marks for semen producers as to whether antibiotics should be added to semen extenders or not.

Reduced bacterial load in stallion semen by modified single layer centrifugation or sperm washing.

The presence of bacteria poses a significant challenge to the quality of stallion semen used in artificial insemination. The bacterial content of insemination doses arises from various sources, such as the healthy stallion, environment, and collection equipment, and is implicated in fertility problems as well as reduced sperm quality during storage. The conventional approach of adding antibiotics to semen extenders raises concerns about antimicrobial resistance and potential negative effects on sperm characteristics, and may not be effective in inhibiting all bacteria. The objective of this study was to determine whether an innovative alternative to antibiotic usage - centrifugation through a single layer of a low density colloid (SLC) - could reduce the bacterial load in stallion semen, and to compare sperm characteristics in samples arising from this procedure, or simple extension of the ejaculate in semen extender, or from sperm washing, i.e. adding extender and then centrifuging the sample to allow the removal of most of the seminal plasma and extender. Eighteen semen samples were collected from six stallions. The semen samples were split and extended prior to washing or SLC, or received no further treatment other than extension. After preparation aliquots from each type of sample were sent for bacteriological examination; the remaining samples were stored for up to 72 h, with daily checks on sperm quality. The low density colloid SLC outperformed sperm washing or extension for bacterial reduction, effectively removing several bacterial species. The bacterial load in the samples was as follows: extended semen, 16 ± 6.7 × 105; washed, 5.8 ± 2.0 × 105; SLC, 2.3 ± 0.88 × 105, p < 0.0001. In addition, SLC completely removed some bacterial species, such as Staphylococcus xylosus. Although there is no selection for robust spermatozoa with the low density colloid, sperm motility, membrane integrity, and DNA fragmentation were not different to washed sperm samples. These findings suggest that SLC with a low density colloid offers a promising method for reducing bacterial contamination in stallion semen without resorting to antibiotics.

Single layer centrifugation as a method for bacterial reduction in bull semen for assisted reproduction.

Semen samples contain bacteria originating from the animal urogenital tract, environment, and/or contamination during semen processing, negatively affecting sperm quality by producing toxins and/or competing for nutrients in extenders. The aims of this study were to evaluate two methods of Single-layer centrifuges (SLC), high and low density colloid, as a method for bacterial removal from bull semen, and to evaluate sperm quality after treatment. In total, semen samples from 20 bulls (3 ejaculates per bull) were used in this study. Bacterial reduction was evaluated by bacterial quantification (colony forming unit - CFU/mL) while bacterial identification was performed by matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) after culturing bacteria on blood agar. Sperm motility parameters were evaluated by Computer Assisted Sperm Analyses (CASA), and sperm chromatin structure assay (SCSA) by Flow cytometry. Both, High and Low density SLC reduced number of bacteria significantly (p < 0.001) compared with control. The difference in bacterial count between High and Low SLC was also significant (p < 0.001). Furthermore, High density SLC was successful in removing almost all Bacillus and Proteus spp. Most CASA parameters were significantly improved after both treatments (p < 0.001, p < 0.01, p < 0.05). The Deoxyribonucleic acid (DNA) fragmentation index evaluated by SCSA in High (p < 0.01) and Low (p < 0.05) SLC group differed significantly compared with control. Single-layer centrifugation (SLC) with either a high or a low density colloid is a suitable method for bacterial removal in bull semen.

Animal Welfare Assessment Protocols for Bulls in Artificial Insemination Centers: Requirements, Principles, and Criteria.

Animal welfare is a complex subject; as such, it requires a multidimensional approach with the main aim of providing the animals with the "five freedoms". The violations of any one of these freedoms could have an influence on animal wellbeing on different levels. Over the years, many welfare quality protocols were developed in the EU thanks to the Welfare Quality® project. Unfortunately, there is a lack of such summarized information about bull welfare assessment in artificial insemination stations or about how disturbed welfare can be reflected in their productivity. Animal reproduction is the basis for the production of meat and milk; therefore, factors contributing to reduced fertility in bulls are not only indicators of animal welfare but also have implications for human health and the environment. Optimizing the reproductive efficiency of bulls at an early age can help to reduce greenhouse gas emissions. In this review, welfare quality assessment will be evaluated for these production animals using reproduction efficiency as a key area, focusing on stress as a main effect of poor animal welfare and, thereby, reduced fertility. We will address various welfare aspects and possible changes in resources or management to improve outcomes.

Effect of Some Plant-Based Substances on Microbial Content and Sperm Quality Parameters of Bull Semen.

The rapid emergence of antibacterial resistance requires alternatives to antibiotics to be found, including for semen preservation. One of the possible alternatives would be to use plant-based substances with known antimicrobial effects. The objective of this study was to test the antimicrobial effect of pomegranate powder, ginger, and curcumin extract in two concentrations on bull semen microbiota after exposure for <2 h and 24 h. An additional aim was to evaluate the effect of these substances on sperm quality parameters. The bacterial count in semen was low from the beginning; however, a reduction was present for all tested substances compared with control. A reduction in bacterial count in control samples was also observed with time. Curcumin at a concentration of 5%, reduced bacterial count by 32% and was the only substance that had a slight positive effect on sperm kinematics. The other substances were associated with a decline in sperm kinematics and viability. Neither concentration of curcumin had a deleterious effect on sperm viability parameters measured by flow cytometry. The results of this study indicate that curcumin extract at a concentration of 5% can reduce the bacterial count and does not have a negative influence on bull sperm quality.

Identification of Bull Semen Microbiome by 16S Sequencing and Possible Relationships with Fertility.

Reports on the use of 16S sequencing for the identification of bacteria in healthy animals are lacking. Bacterial contamination of bull semen can have a negative effect on the sperm quality. The aims of this study were threefold: to identify bacteria in the semen of healthy bulls using 16S sequencing; to investigate the differences in the bacterial community between individual bulls; and to establish if there was a relationship between the bacteria isolated and bull fertility. Semen from 18 bulls of known fertility was used for the DNA extraction and 16S sequencing; 107 bacterial genera were identified. The differences in the amplicon sequence variants (ASVs) and the numbers of genera between bulls were noted. Negative correlations (p < 0.05) between several bacterial genera with Curvibacter, Rikenellaceae RC9-gut-group and Dyella spp. were seen. Other negatively correlated bacteria were Cutibacterium, Ruminococcaceae UCG-005, Ruminococcaceae UCG-010 and Staphylococcus, all within the top 20 genera. Two genera, W5053 and Lawsonella, were enriched in bulls of low fertility; this is the first time that these bacteria have been reported in bull semen samples. The majority of the bacteria were environmental organisms or were species originating from the mucous membranes of animals and humans. The results of this study indicate that differences in the seminal microbiota of healthy bulls occur and might be correlated with fertility.

Transabdominal and transrectal ultrasonography of fetuses in Württemberg ewes: Correlation with gestational age.

It is useful to determine the gestational age in sheep to provide essential information for effective flock management practices. The aims of this study were to evaluate the efficacy and practical aspects for using two ultrasound techniques, transabdominal and transrectal, in determining the gestational age in Württemberg ewes. Monitoring of embryo and fetus developmental stages during the ewes' gestation was carried out with real time ultrasound using a transabdominal convex probe, frequency 3.5 MHz, and a transrectal linear probe, frequency 7.5 MHz. The size of the embryonic vesicle during the period from the 23rd to the 38th day of gestation can be used as a confirmational indicator of gestational age when the transrectal probe is used. The occipital nasal diameter correlated with the gestational age with both transabdominal (P < 0.05) and transrectal probes (P < 0.01) from the 46th to the 63rd day of gestation. The biparietal diameter of the fetal head measured by transabdominal probe during the period from the 46th to the 63rd day of gestation correlates with gestation age (P < 0.05). The diameter of the fetal eye orbit monitored by transrectal probe from the 46th to the 63rd day of gestation also correlated well with gestational age (P < 0.05).