Relationship between pubertal testicular ultrasonographic evaluation and future reproductive performance potential in Piétrain boars.

New ways of predicting sperm quality and output performance in young artificial insemination (AI) boars are important for breeding companies to ensure that the pubertal boars delivered to the AI studs have a high chance of meeting minimum quality standards to be used for insemination and therewith dissemination of desirable characteristics. The aim of the current study was to characterize the testicular development of 218 pubertal Piétrain boars (Line 408, Pig Improvement Company) to identify traits with predictable characteristics relative to their sperm quality as an adult AI boar. Scrotum, testes and epididymis were examined ultrasonographically at day (d) 100 (on-test) and 170 (off-test) followed by a computer-assisted grayscale analysis (GSA). Over the test period, paired testicular volume increased 7.3-fold from 22.7 ± 10.8 cm3 to 166.6 ± 62.2 cm3. The right testis was significantly (P = 0.014) larger than the left one at the off-test. Based on the sperm quality (ejaculate volume, sperm concentration, total sperm number, morphologically abnormal sperm and total sperm motility at day 3 of semen storage), 82.11% (n = 179) of the boars were classified as "productive" boars. These boars had a significantly (P = 0.039) larger paired testicular volume than "non-productive" boars (45.9 ± 19.9 cm3vs. 38.5 ± 12.6 cm3) at the on-test. For the right testis at on-test, significant differences for the standard deviation of mean gray value (P = 0.022), area under the curve (P = 0.004) and mean gradient value (GRAD, P = 0.030) regarding the future sperm production capacity (SPC) were shown. At off-test, there was a significant difference for minimum gray value (MIN GV, P = 0.003) and mean gray value (P = 0.001) related to SPC. To find SPC related cut-off values for GSA data, a two segmental non-linear regression analysis was carried out indicating breakpoints for GRAD ≥12 and MIN GV ≥ 40 for boars with low SPC. Off-test boars with MIN GV ≥ 40 showed a 2.4 higher risk to display low SPC (Odds ratio = 2.4 [1.1, 5.4]; P = 0.024). The results may enable breeding companies to include new sperm quality associated traits in their boar testing and selection programs.

Dose rates of antimicrobial substances in boar semen preservation-time to establish new protocols.

To achieve a standardized number of spermatozoa in the final AI dose, varying amounts of extender fluid with a fixed concentration of antimicrobial substances are currently added to boar ejaculates. This practice ignores the different degrees of dilution of the antimicrobials in the end product. In calculating the final concentration of gentamicin in AI doses from 27,538 processed boar ejaculates, we demonstrated varying gentamicin concentrations in the resultant extended boar semen samples. The median concentration was 220.37 mg/L. In 25 of the samples (0.09%), the gentamicin concentration fell below 5 mg/L, which is close to or below the epidemiological cut-off value for many bacteria. We calculated the minimum inhibitory concentration of gentamicin for bacteria isolated from raw and extended ejaculates. Five of the isolates from extended ejaculates exceeded the maximum test concentration of 512 mg/L. As a result, we are presenting an alternative method of boar semen preservation whereby a particular combination of gentamicin concentrate and antibiotic-free extender is incorporated that standardizes the antibiotic concentration in the diluted semen. The addition of standardized antibiotic concentrations did not negatively affect sperm quality when compared to the use of ready-to-use extenders. In conclusion, an end volume-based and standardized addition of gentamicin to boar ejaculates can be a helpful alternative to prevent insufficient dosage of antibiotics in liquid preserved boar semen without affecting semen quality.

Quality Control of Boar Sperm Processing: Implications from European AI Centres and Two Spermatology Reference Laboratories.

In recent years, increased automatization has resulted in a higher efficiency of boar semen processing in AI laboratories. Sophisticated laboratory management and efficient quality control programmes are needed for current tendencies in major pork-producing countries to reduce the sperm number per AI dose, to lengthen semen storage times and to adopt responsible methods for bacterial control and prevention of the development of multiresistant bacteria. The objective of the present review was to outline current trends in boar semen production and the critical steps in semen processing which affect sperm quality. In addition, integrated elements of a quality assurance programme in use by thirty European AI centres in association with the two German spermatology reference laboratories are described.

Review on international trade with boar semen.

For more than 40 years, AI (artificial insemination) has been carried out with pigs. In some countries, it constitutes since the 1990 s the dominant procedure with piglet production to fertilize the sow. This procedure of insemination with fresh semen has become prevalent in all countries on a worldwide basis with an important pig meat production, with the exception of China. Meanwhile, up to 90% of the sows have been artificially inseminated. The trend is still upwards. As the need of pig meat and thus the production continues to increase, one can proceed on the assumption that the number of semen doses, which is necessary for this procedure, will likewise increase correspondingly. Until now, the trade beyond borders has only been marginal. An improvement in the sense of a longer shelf life for semen doses is indispensable for the trade over longer distances.

Sperm chromatin structural integrity in normospermic boars is not related to semen storage and fertility after routine AI.

Standard semen parameters are limited in their capacity to distinguish subfertile boars and to assess storage influences on liquid preserved boar semen. The evaluation of sperm chromatin structural integrity could have potential as a diagnostic tool in AI practice. This study assessed whether the determination of sperm DNA integrity adds a useful diagnostic tool for selection of boar ejaculates in routine AI procedure and assessment of storage effects in diluted semen. Special emphasis was laid on the standard spermatological characterization of semen samples in parallel with the determination of the DNA fragmentation index (DFI) through the sperm chromatin structure assay (SCSA). Six hundred ninety two (692) ejaculates from 79 Piétrain boars in an AI center were analyzed for motility, morphology and DFI over a period of 24 weeks. 95.5% of the semen samples had a DFI < 5% with low distribution of variation for DFI due to boar and ejaculate (< 5%). 61.3% of ejaculates with DFI > 5% showed values below thresholds for sperm motility or morphology. Based on field data from 13,239 inseminations, fertility of boars with temporarily elevated DFI was not impaired (P > 0.05). 24 randomly selected diluted semen samples did not show a significant increase of DFI during 168 h storage (P > 0.05). In a further experiment, 42 diluted semen samples from 14 normospermic boars were assessed for motility, membrane integrity (PI, FITC-PNA) and SCSA parameters. Three single ejaculates showed an increase of DFI at 120 and 168 h storage time. This was accompanied by a pronounced loss of both motility and membrane integrity. In conclusion, the evaluation of sperm chromatin structural integrity by the SCSA has only limited value for identifying sperm deficiencies in normospermic fresh or stored boar semen. Temporarily elevated DFIs seem not to be indicative of subfertility in normospermic boars.

Age-dependent expression and localization of the progesterone receptor in the boar testis.

Expression of the progesterone receptor (PR) was monitored in testes of groups of five boars aged 50, 100, 150, 200 and 250 days. The primary monoclonal antibody used for immunohistochemistry (IHC) was raised against a peptide mapping the amino acids 922-933 of the carboxy-terminus of the human PR, negative controls were set up using an irrelevant monoclonal isotype-specific antibody, porcine endometrium served as positive control tissue. In parallel, qualitative reverse transcription-polymerase chain reaction was applied. Based on the developmental status of spermatogenesis the 50- and 100-day-old boars were considered as immature, the boars aged 200 and 250 days as mature. Positive and negative controls confirmed specificity of IHC. In the 50-day-old boars 85.1% of the prespermatogonia that had reached the basal lamina and 18.2% of the centrally located prespermatogonia stained positive, while it was 92.1% respectively 2.1% in the 100-day-old boars. The effect of time and location was highly significant (p < 0.005 resp. 0.0001). In mature boars between 77 and 80% of the A and B spermatogonia stained positive, there was no effect of boar age and stage of spermatogenesis. In both groups also few peritubular myoid cells stained positive. It is hypothesized that Leydig cell-derived progesterone plays a functional role in spermatogoniogenesis in a synergistic manner with Leydig cell-derived oestrogens.