Cryopreservation of Ram Semen in Extenders Containing Soybean Lecithin as Cryoprotectant and Hyaluronic Acid as Antioxidant

A soybean lecithin‐based extender supplemented with hyaluronic acid (HA) was assayed for effectiveness to improve the quality of frozen–thawed ram semen. HA has not been tested yet in an extender containing soybean lecithin for freezing ram semen. Thus, the aim of this study was to analyse the effects of soybean lecithin at 1% or 1.5% along with HA at 0, 0.5 and 1 mg ml^‐1 in a Tris‐based extender on the motion characteristics, membrane integrity (HOST), viability, GSH peroxidase (GSH‐PX) activity, lipid peroxidation and acrosomal status after freezing–thawing. Semen was collected from four Mehraban rams during the breeding season and frozen in the six lecithin×HA extenders. The extender containing 1.5% lecithin supplemented with no HA yielded higher total motility (52.5%±1.6), viability (55.8%±1.6) and membrane integrity (44.5%±1.7), but the effects of the lecithin concentration did not reach signification. Linearity‐related parameters, ALH, BCF, lipid peroxidation, GSH‐PX activity, morphology and acrosomal status were not affected by the extender composition. In general, adding HA significantly decreased sperm velocity (1 mg ml^‐1 HA), total motility (only with 1.5% lecithin), viability (1 mg ml^‐1 HA for 1% lecithin; both concentrations for 1.5% lecithin) and membrane integrity. In conclusion, adding HA to the freezing extender supplemented with soybean lecithin failed to improve quality‐related variables in ram semen. Increasing the lecithin content could have a positive effect, but further studies are needed.     

Determination of Feeder Cell‐Based Cellular Niches Supporting the Colonization and Maintenance of Spermatogonial Stem Cells from Prepubertal Domestic Cat Testes

Recently, isolation and in vitro culture of putative spermatogonial stem cells (SSCs) in the domestic cat have been conducted. However, the cellular niche conditions that facilitate the establishment and long‐term maintenance of feline SSCs (FSSCs) have not been described. Therefore, we investigated the type of feeder cells used to stimulate colony formation and growth of FSSCs among the various factors in the FSSC niche. Spermatogonial stem cells isolated from feline testes were cultured on mitotically inactivated testicular stromal cells (TSCs) derived from cats, dogs and mice, and mouse embryonic fibroblasts (MEFs). The formation and growth of colonies derived from SSCs cultured on each type of feeder cell were identified at passage 0, and the morphology, alkaline phosphatase (AP) activity and expression of SSC‐specific genes in surviving colonies were investigated at passage 4. Among these diverse feeder cells, TSCs from cat showed the greatest colony formation, growth and maintenance of FSSCs, and SSC colonies cultured by passage 4 showed a typical dome‐shaped morphology, AP activity and expression of SSC‐specific genes (NANOG, OCT4, SOX2 and CD9). Accordingly, these results demonstrate that feline TSCs could be used as feeder cells to support the establishment and maintenance of SSCs from domestic cats.     

Superradiance in a torus magnetosphere around a black hole

The coalescence of a neutron star and a black hole in a binary system is believed to form a torus around a Kerr black hole. A similarly shaped magnetosphere then results from the remnant magnetic field of the neutron star. In the strong-field case, it contains a cavity for plasma waves located between the barrier of the gravitational potential and the surrounding torus. This cavity may be unstable to superradiance of electromagnetic waves. Superradiant amplification of such waves, initially excited by turbulence in the torus, should inflate into a bubble in a time as short as approximately 0.75 (1 percent/&cjs3539;epsilon&cjs3539;2)(M/7M middle dot in circle) seconds approximately 0.15 to 1.5 seconds, assuming an efficiency &cjs3539;epsilon&cjs3539;2 = 0.5 to 5 percent and a mass M = 7M middle dot in circle. These bubbles may burst and repeat, of possible relevance to intermittency in cosmological gamma-ray bursts. The model predicts gamma-ray bursts to be anticorrelated with their gravitational wave emissions.     

Isolation and characterization of porcine circovirus type 2 from pigs showing signs of post‐weaning multisystemic wasting syndrome in the Netherlands

Summary

Pigs with wasting syndrome were examined for macroscopic and histopathological lesions, and for porcine circovirus (PCV). Histopathological lesions were comparable to those previously documented for post‐weaning multisystemic wasting syndrome (PMWS). In addition, in seven out of ten examined PMWS‐affected pigs focal‐to‐slight mononuclear meningitis and focal cerebral mononuclear infiltrates (4 out of 10) were observed. A virus was isolated from organs and sera from pigs showing wasting syndrome. An immunoperoxidase monolayer assay and an indirect immunofluorescence assay were performed on the infected PK‐15 and Dulac cell cultures, respectively, and both assays indicated the presence of PCV type 2 (PCV2). The nested‐polymerase chain reaction (nPCR) technique, based on the use of PCV2 specific oligonucleotides, revealed specific amplified products of 481 bp. Nucleotide sequence analysis of the entire genome of the Dutch PCV isolate 24657 NL showed a homology with known nucleotide sequences of porcine PCV type 1 (PCV1) and PCV2 isolates of 77.1% and >96%, respectively. This is the first report of the isolation and characterization of PCV2 in PMWS‐affected pigs in the Netherlands.     

Oral transmission of porcine reproductive and respiratory syndrome virus by muscle of experimentally infected pigs

The current study was performed to determine if porcine reproductive and respiratory syndrome virus (PRRSV) could be transmitted to pigs by feeding muscle tissue obtained from recently infected pigs. Muscle obtained from pigs infected with either a European strain (EU donor pigs) or American strain (US donor pigs) of PRRSV was fed to PRRSV-free receiver pigs. The donor pigs were slaughtered 11 days post-infection (dpi). PRRSV was detected by conventional virus isolation in muscle at 11 dpi from 7 of 12 EU donor pigs and 5 of 12 US donor pigs. In contrast to conventional virus isolation, all muscle samples from infected pigs were positive for viral nucleic acid by PCR, except for muscle from one animal infected with the American strain of PRRSV. Five hundred grams of raw semimembranosus muscle from each of the donor pigs was fed over a 2 days period (250 g per day) to each of two receiver pigs (48 receiver pigs). The receiver pigs were housed separately in five groups. One of the five groups was fed muscle obtained from US donor pigs that was also spiked with the American strain of PRRSV. Sentinel pigs were placed in-contact with the group of receiver pigs fed spiked muscle. All receiver pigs became viraemic by 6 days post-feeding (dpf). There was evidence of horizontal transmission with sentinel pigs, in-contact with receiver pigs, becoming viraemic. The study demonstrates that PRRSV could be infectious through the oral route via the feeding of meat obtained from recently infected pigs.     

Stochastic simulation of dairy cattle nucleus breeding schemes: Influence of breeding strategy and biased breeding values in the population

SUMMARY: A stochastic simulation model of an open nucleus scheme was used to study the consequences of the breeding strategy and biased lactation records for population cows. Selection was for a single sex-limited trait with a heritability of 0.25 and based on animal model breeding value estimates. Selection of dams was across age classes while sires were required to have a progeny test before they could be selected as proven bull or bull sire. Dams to breed nucleus replacements and young bulls could be selected from the nucleus and the top population which contained 240 and 1600 replacement heifers annually. The first 15 years of the simulated period was used to reach a population with an equilibrium genetic progress for a progeny testing scheme. Comparisons were based on the 25 year period after an alternative breeding scheme was adopted. The annual genetic gain was calculated from the last 10 years of that period. The annual genetic gain in an open nucleus breeding scheme was .247 σ(a) . The annual genetic gain increased 5.4% when MOET was also used on cows selected to breed replacements for the top population. When, in addition the number of sires used on top population cows was reduced from 8 to 4, that being the number used in the nucleus, the annual genetic gain increased by another 2.8%. The reduction in annual genetic gain due to biased lactation records of top population cows ranged from 4.6 to 15.4%. The average bias in estimated breeding values of the top population dams selected to breed nucleus replacements ranged from 0.53 to 2,52 σ(a) . The regression coefficient of the EBV of the bull after progeny testing on the EBV of the dam at the time of selection was 0.55 without biased lactations and ranged from 0.10 to 0.27 with biased lactations. The reduction in genetic gain was especially related to the regression coefficient and to a lesser extent to the average bias. In practice, the expected reduction in annual genetic gain from biased lactation records of population cows is expected to be between 5 and 10 %. ZUSAMMENFASSUNG: Stochastische Simulation von Milchvieh-Nukleussystemen: Einflu? der Zuchtstrategie und verzerrter Zuchtwerte in der Population Eine stochastische Simulation eines offenen Nukleussystems wurde zur Untersuchung der Konsequenzen der Zuchtstrategie und verzerrter Laktationsabschlüsse für Populationskühe untersucht. Selektion bezog sich auf ein einzelnes weibliches Merkmal mit Heritabilit?t von 1/4 und gründete auf Tiermodell Zuchtwertsch?tzungen, Selektion von Muttertieren über Altersklassen, w?hrend Stiere vor der Selektion einen Nachkommenschaftstest haben mu?ten. Muttertiere für Nukleus- und Jungstiere kommen vom Nukleus und Spitzen der Population, die 240 und 1600 nachgestellte Kalbinnen umfa?ten. Die ersten 15 Jahre der simulierten Periode wurden zum Erreichen einer Population mit Gleichgewichtsfortschritt für ein Nachkommenschaftsprüfsystem verwendet. Vergleiche beruhten auf einer 25-Jahre-Periode nach Einrichtung des alternativen Zuchtsystems, und der j?hrliche Zuchtfortschritt wurde für die letzten 10 Jahre berechnet. Der j?hrliche Zuchtfortschritt im offenen Nukleussystem war 0,247 σ(a) und nahm um 5,4% zu, wenn MOETauch für Kühe zum Ersatz der Spitzenpopulation verwendet wurde. Wenn darüber hinaus die Zahl der Vatertiere in der Spitzenpopulation von 8 auf 4 reduziert wurde, die Zahl der im Nukleus verwendeten, konnte der j?hrliche genetische Fortschritt um weitere 2, 8% gesteigert werden. Die Verminderung des Zuchtfortschrittes auf Grund von verzerrten Laktationsabschlüssen der Spitzenkühe der Population variierte von 4,6 bis 15,4%. Die durchschnittliche Verzerrung der gesch?tzten Zuchtwerte der Populationsspitzenkühe für die Nukleusremonte bewegte sich von 0,53 bis 2,52 σ(a) . Der Regressionskoeffizient von EBV der Stiere auf Grund von Nachkommenschaftsprüfung auf EBV der Muttertiere beim Zeitpunkt der Selektion war 0,55 ohne verzerrte Laktationen und schwankte zwischen 0,10 und 0,27 bei verzerrten Laktationen. Die Verminderung des genetischen Fortschritts hing deutlich mit dem Regressionskoeffizient zusammen und weniger mit der durchschnittlichen Verzerrung. In der Praxis ist zu erwarten, da? die Reduktion des Zuchtfortschrittes durch verzerrte Laktationsabschlüsse der Population zwischen 5 und 10% liegt.     

The Adverse Effect of Phytoestrogens on the Synthesis and Secretion of Ovarian Oxytocin in Cattle

The current investigations were undertaken to study the mechanism of the adverse effect of phytoestrogens on the function of bovine granulosa (follicles >1< cm in diameter) and luteal cells from day 1–5, 6–10, 11–15, 16–19 of the oestrous cycle. The cells were incubated with genistein, daidzein or coumestrol (each at the dose of 1 × 10^?6 m ). The viability and secretion of estradiol (E2), progesterone (P4) and oxytocin (OT) were measured after 72 h of incubation. Moreover, the expression of mRNA for neurophysin‐I/OT (NP‐I/OT; precursor of OT) and peptidyl‐glycine‐α‐amidating monooxygenase (PGA, an enzyme responsible for post‐translational OT synthesis) was determined after 8 h of treatment. None of the phytoestrogens used affected the viability of cells except for coumestrol. The increased secretion of E2 and P4 was only obtained by coumestrol (p < 0.05) from granulosa cells from follicles <1 cm in diameter and decreased from luteal cells on days 11–15 of the oestrous cycle, respectively. All three phytoestrogens stimulated (p < 0.05) OT secretion from granulosa and luteal cells in all stages of the oestrous cycle and the expression of NP‐I/OT mRNA in the both types of cells. The expression of mRNA for PGA was stimulated (p < 0.05) by daidzein and coumestrol in granulosa cells, and by genistein and coumestrol in luteal cells. In conclusion, our results demonstrate that these phytoestrogens can impair the ovary function in cattle by adversely affecting the synthesis of OT in follicles and in corpus luteum. However, their influence on the ovarian steroids secretion was less evident.