Three-dimensional live imaging of bovine embryos by optical coherence tomography

While embryo transfer (ET) is widely practiced, many of the transferred embryos fail to develop in cattle. To establish a more effective method for selecting bovine embryos for ET, here we quantified morphological parameters of living embryos using three-dimensional (3D) images non-invasively captured by optical coherence tomography (OCT). Seven Japanese Black embryos produced by in vitro fertilization that had reached the expanded blastocyst stage after 7 days of culture were transferred after imaged by OCT. Twenty-two parameters, including thickness and volumes of the inner cell mass, trophectoderm, and zona pellucida, and volumes of blastocoel and whole embryo, were quantified from 3D images. Four of the seven recipients became pregnant. We suggest that these 22 parameters can be potentially employed to evaluate the quality of bovine embryos before ET.     

Differentially methylated CpG sites related to fertility in Japanese Black bull spermatozoa: epigenetic biomarker candidates to predict sire conception rate

For semen suppliers, predicting the low fertility of service bull candidates before artificial insemination would help prevent economic loss; however, predicting bull fertility through in vitro assessment of semen is yet to be established. In the present study, we focused on the methylated CpG sites of sperm nuclear DNA and examined methylation levels to screen new biomarkers for predicting bull fertility. In frozen-thawed semen samples collected from Japanese Black bulls, for which the sire conception rate (SCR) was recorded, the methylation level of each CpG site was analyzed using human methylation microarray. According to regression analysis, 143 CpG sites related to SCR were significantly differentially methylated. Whole genome bisulfite sequence data were obtained from three semen samples and the differentially methylated regions (DMRs) that included the target CpG sites selected by human methylation microarray were confirmed. Using combined bisulfite restriction analysis, fertility-related methylation changes were detected in 10 DMRs. With the exception of one DMR, the methylation levels of these DMRs were significantly different between groups with high fertility (> 50%) and low fertility (< 40%). From multiple regression analysis of methylation levels and SCR, three DMRs were selected that could effectively predict bull fertility. We suggest that these fertility-related differences in spermatozoal methylation levels could be new epigenetic biomarkers for predicting bull fertility.     

Fine mapping the number of corpora lutea quantitative trait loci on SSC3: Analysis of the porcine follicle‐stimulating hormone receptor gene

In females, follicle‐stimulating hormone (FSH) targets a FSH receptor (FSHR) expressed only on granulose cells, inducing maturation of the ovarian follicles. We hypothesized that genetic variants in the FSHR gene influence litter size by affecting the number of corpora lutea. We fine‐mapped a region of Sus Scrofa chromosome 3 that contains quantitative trait loci for corpora lutea. Polymorphisms were detected in the exons and 5′ flanking region of the porcine FSHR gene, a positional candidate for the statistically most significant of the quantitative trait loci. Finally, 248 F₂ animals from a Duroc and Meishan cross were genotyped for three FSHR SNPs at positions 74, 532 and 1166, and these were correlated with the phenotypes of litter size and corpus luteum number. Three haplotypes were identified: M1 (G/G/C), M2 (C/A/T) and D (C/A/C). In the F₂ population, the M1 haplotype was associated with a greater number of corpora lutea (P < 0.01) and also seemed to be associated with increased litter size, although the association was not significant (P = 0.2571). Some polymorphisms resulting in amino acid substitutions in these genes were excluded from the polymorphisms possibly responsible for the number of corpora lutea.     

Approachability and contact behavior of commercial dairy calves to humans

Calves (n = 106) on four dairy farms were observed for their approachability to humans. All calves experienced similar rearing conditions: Beginning individual pen, after birth until weaning at about 2 months, where they were housed individually and fed milk and a milk replacement; Late individual pen, after weaning until grouping at about 3.5 months, where they were housed individually and fed hay, silage and concentrate feed; Beginning group pen, after grouping until 5 months, where they were housed in groups of 2–5 animals and fed hay, silage and concentrate feed; later group pen, from 5 to 7 months. The number of calves that contacted an experimenter who stood in front of their pens for 10 min was recorded on 6 separate days over 3 months. Latency to touch and time spent in activities during touching such as sucking, licking, biting and rubbing were also measured. There were no significant differences in the latency to touch and the ratio of touch to non‐touch calves between the rearing conditions and the farms. The time spent touching was significantly affected by the interaction between the rearing condition and the farm (P < 0.01). In detail, the time spent sucking (P < 0.001) and licking (P < 0.01) was different between the rearing condition × farm variables. The proportion of calves that approached and touched the experimenter tended to be higher in the farms in which a stockperson worked longer inside and outside their pens (both ρ = 0.95, P = 0.051). These results were interpreted according to the perspectives of early positive reinforcement with food and the habituation process to humans existing nearby.     

Isolation and characterization of the interspecific fusants from Streptomycetes obtained using a liquid regeneration method

ABSTRACT: Protoplast fusion between different species of Streptomyces was performed using a liquid regeneration method developed for a rapid and simple preparation of the fusants. Consequently, new clones, which could not be obtained using the conventional agar regeneration method, were obtained. In the crosses between S. griseus and S. durhamensis , and between S. californicus and S. catenulae , eight and two recombinants, respectively, were obtained using the liquid regeneration method. Conversely, in the case of crosses between S. ornatus and S. catenulae , and between S. ornatus and S. vendargensis , seven recombinants each were obtained using only the agar method. The physiological characteristics, such as the assimilation of carbohydrate and antibiotic resistance, of these fusants differed considerably from those of their parental strains. Using the proposed liquid regeneration method, a simpler and quicker procedure for protoplast fusion is described.     

Glomus tumor of the liver in a cow

An 11-year-old Holstein-Friesian cow exhibited anorexia and jaundice. A large mass was found in the liver during necropsy. Macroscopically, the mass was composed of dark red multilobular tissue and a centrally located abscess, which was connected to the hepatic duct. Histologically, the mass consisted of proliferation of small neoplastic cells and was demarcated from the hepatic parenchyma by a thick region of granulation tissue. The neoplastic cells were predominantly arranged in solid sheets, but they also formed blood-filled cancellous structures, and proliferating foci were seen around blood vessels. Periodic acid-Schiff reaction demonstrated that a fine basement membrane-like structure surrounded the neoplastic cells. Immunohistochemically, the neoplastic cells were positive for vimentin and alpha smooth muscle actin and negative for cytokeratin, factor VIII-related antigen, chromogranin and desmin. Based on its histopathological features, the hepatic neoplasm was diagnosed as a primary glomus tumor. This is the first report about a primary glomus tumor of the liver in a cow.