Validation of a portable device (iSperm® ) for the assessment of stallion sperm motility and concentration.

The objective of this study was to evaluate the accuracy of a novel, portable device (iSperm® Equine for assessing concentration and motility of stallion semen). In the first experiment, semen concentration was determined by the iSperm® Equine (Aidmics Biotechnology), Androvision® (Minitube) and NucleoCounter® SP-100™ (ChemoMetec). The total motility and progressive motility were determined by the iSperm® Equine and the Androvision® using the manufacturer's guidelines. Frozen/thawed semen samples (n = 33) at various dilutions were analysed for concentration and motility with the above-mentioned devices. There was a significant correlation between the concentrations measured with iSperm® and NucleoCounter® at all the measured dilutions. Moreover, <10% difference in concentrations was observed between the iSperm® and NucleoCounter® using the Bland-Altman test. There was also a significant correlation between iSperm® and Androvision® for total and progressive motility. In the second experiment, the parameters used in the Androvision® were modified to match those of the iSperm® . Total motility and progressive motility of frozen/thawed semen samples (n = 10) were determined, and the similarity between the Androvision® and iSperm® was confirmed by correlation studies and Bland-Altman test. The results of these experiments demonstrate that the iSperm® offers a reliable and practical alternative for the semi-automated measurement of concentration and motility of stallion semen in the field. The iSperm® enables the practitioner to obtain objective and repeatable measurements on a variety of semen types (fresh, cooled and frozen) in the field at the time of insemination and thus acquire more insight into the quantity and quality of the provided insemination doses. This mare-side diagnostic tool may help practitioners in identifying presumed subfertility problems more rapidly and act accordingly.

Effect of environmental factors and changes in the body condition score on the onset of the breeding season in mares.

Several methods have been proposed to advance the onset of the breeding season in horses. Most of them are based on the exposure to an artificial lighting period combined with hormonal treatments. Mares exposed to an artificial photoperiod are most often housed indoors where the ambient temperature is often higher than the outside temperature. Mares held in barns are also exposed to different daylight intensities than horses kept outside, depending on the architecture. In the current study, we evaluated the impact of ambient temperature, daylight intensity and changes in body condition score (BCS) on the timing of first ovulation after winter anestrus in mares exposed to an artificial photoperiod. Mares (n = 211) were housed in barns with different ambient temperature and daylight exposure but with the same artificial photoperiod exposure (except for a natural photoperiod control group). Artificial photoperiod as well as an increase in BCS over the winter significantly advanced the first spring ovulation. The BCS at the start and end of the anestrus period did not have an effect on the interval to first ovulation and neither did the modest increase in ambient temperature in the barn. However, a higher light intensity during the daytime significantly advanced the first spring ovulation. The results of this study suggest that exposure to more sunlight advances the onset of the breeding season. This effect is likely mediated through the biological effect of short wavelength blue light and its impact on melatonin suppression and biological rhythms. We suggest that greater/direct exposure to the blue light component of daylight improves the response to the artificial photoperiod. The results of the present study can further assist to optimize the conditions that lead to an efficient spring transition of breeding mares.

Suppression of keratin 18 gene expression in bovine blastocysts by RNA interference.

The expression of the cytoskeleton protein Keratin 18 (KRT18) starts at the onset of bovine blastocyst formation. KRT18 is solely expressed in the trophectoderm and can therefore be used as a marker for trophectodermal differentiation. In the present study, the expression of KRT18 was suppressed by RNA interference to probe its functional importance in bovine blastocyst formation. Microinjection of KRT18 double-stranded RNA into the cytoplasm of zygotes resulted in reduced KRT18 mRNA (76% reduction) and protein expression at the blastocyst stage and a lower developmental competence (41% reduction in the percentage of blastocyst formation) compared with non-injected and phosphate-buffered saline (PBS)-injected controls. KRT18 downregulation was associated with reduced mRNA expression of KRT8, the binding partner of KRT18, but had no effect on the expression of KRT19, CDH1 and DSP, other genes involved in intermediate filament and cytoskeleton formation. The results of the present study demonstrated that KRT18 knockdown in preimplantation embryos results in reduced blastocyst formation, but no further morphological aberrations were observed with regard to the biological function of KRT18. These observations could be due to the function of KRT18 being replaced by that of another gene, the surviving blastocysts expressing the minimum level of KRT18 required for normal blastocyst development or the possibility that further aberrations may occur later in development.