Mastitis is associated with decreased fertility in dairy cows. In the current study, we created an experimental model to simulate shortterm mastitis by a single intramammary administration of Gram-negative endotoxin of Escherichia coli origin (GK), or Gram-positive toxin of Staphylococcus aureus origin (GC), to examine the effect of mastitis on oocyte developmental competence. Healthy Holstein cows were synchronized, and follicular fluid (FF) of cows treated with GC or GK and of uninfected cows (controls) was aspirated from the preovulatory follicles by transvaginal ultrasound procedure. The aspirated FF was used as maturation medium for in vitro embryo production. The distribution of matured oocytes into different cortical granule classes and meiotic stages was affected by GK administration (P!0.05) but not by GC administration. The proportion of oocytes that cleaved to two-and four-cell stage embryos (44 h postfertilization) was lower in both GC and GK groups than in controls (P!0.05). Blastocyst formation rate (7-8 days postfertilization) was lower in the GK group (P!0.05) and numerically lower in the GC group compared with their uninfected counterparts. The total cell number in blastocysts did not differ among groups; however, the apoptotic index was higher in the GC group (P!0.05), but not in the GK group, relative to controls. Examining mRNA relative abundance in oocytes and early embryos revealed mastitis-induced alterations in PTGS2 (COX2), POU5F1, and HSF1 but not in SLC2A1 (GLUT1) or GDF9. Results indicate a differential disruptive effect of mastitis induced by GK and GC on oocyte developmental competence in association with alterations in maternal gene expression.

No abstract

In the last decade, potential exposure of humans and animals to industrial chemicals and pesticides has been a growing concern. In the present study, di-(2-ethylhexyl) phthalate (DEHP) and mono-(2-ethylhexyl) phthalate (MEHP) were used to model the effects of endocrine-disrupting compounds and their risk in relation to early embryonic losses. Exposure of cumulus oocyte complexes during maturation to 50 μM MEHP reduced the proportion of oocytes that underwent nuclear maturation (p < 0.05) and increased the proportion of apoptotic oocytes (p < 0.05). Furthermore, phthalates reduced cleavage rate in the MEHP-treated group (p < 0.05) and the proportion of embryos developing to the blastocyst stage in both DEHP- and MEHP-treated groups (p < 0.05). The total cell count for blastocysts developing from MEHP-treated oocytes was lower than in controls (p < 0.05). Exposure of oocytes to MEHP during maturation reduced (p < 0.05) the expression of ASAH1 (an anti-apoptotic factor), CCNA2 (involved in cell cycle control), and POU5F1 (responsible for pluripotency) in matured oocytes. Furthermore, the reduced mRNA expression of POU5F1 and ASAH1 lasted into two-cell stage embryos (p < 0.05). Phthalate-induced alterations in POU5F1, ASAH1, and CCNA2 expression might explain in part the reduced developmental competence of MEHP-treated oocytes.

We examined the effects of naturally occurring mastitis on bovine oocyte developmental competence in vitro. Specifically, we investigated the effects of intramammary infection on the ovarian pool of oocytes (i.e., follicle-enclosed oocytes) and their ability to undergo in vitro maturation, fertilization, and further development to the blastocyst stage. Culled Holstein cows (n=50) from 9 commercial dairy farms in Israel were allotted to 3 groups according to somatic cell count (SCC) records of the last 3 monthly milk tests as well as of quarter samples collected before slaughter: (1) low SCC (n=7), (2) medium SCC (n=16), or (3) high SCC (n=27). Means of SCC values differed among low-, medium-, and high-SCC groups: 148,000, 311,000 and 1,813,000 cell/mL milk, respectively. Milk yield and days in milk did not differ among the 3 groups. Bacterial isolates included coagulase-negative staphylococci, Escherichia coli, Streptococcus dysgalactiae, or no bacteria found. Ovaries were collected at the abattoir and brought to the laboratory. Cumulus oocyte complexes were recovered separately from each cow and subjected individually to in vitro maturation and fertilization, followed by 8d in culture. The number of aspirated oocytes did not differ among groups, with a range of 17 to 21 oocytes per cow. The proportion of oocytes that cleaved into 2- to 4-cell-stage embryos (86.1 ± 3.4%) did not differ among groups. In contrast, mean percentages of embryos developed to the blastocyst stage on d 7 and 8 after fertilization were less in both medium- and-high SCC groups than in the low-SCC group (5.6 ± 2.3 and 4.1 ± 1.8 vs. 18.1 ± 4.6%, respectively). Additional analysis indicated that cleavage and blastocyst-formation rates did not differ among the bacterial types in the low-, medium-, and high-SCC groups. These are the first results to demonstrate that naturally occurring mastitis disrupts the developmental competence of the ovarian pool of oocytes, (i.e., oocytes at the germinal vesicle stage). The disruption was associated with elevation of SCC rather than bacterial type. The results may provide a partial explanation for the low fertility of cows that have contracted mastitic pathogens before insemination.

No abstract

No abstract

We examined the association between progressive motility of spermatozoa and in vitro fertilization (IVF) competence of bovine ejaculates. Fresh semen was evaluated using a computerized sperm quality analyzer for bulls using progressive motility as the primary parameter. Ejaculates with high progressive motility (HPM; >81%) were compared with those with low progressive motility (LPM; 0.05). Examination of sperm morphology revealed a higher proportion of spermatozoa with abnormal morphology (P < 0.01) in LPM versus HPM ejaculates, the predominant abnormal feature being a bent tail (P < 0.05). Sperm viability, acrosome integrity and DNA fragmentation did not differ between HPM and LPM samples. Mitochondrial membrane potential was higher (P < 0.01) in HPM versus LPM semen. Zinc concentrations in the seminal plasma correlated with progressive motility (R2 = 0.463, P = 0.03). In addition, representative ejaculates from HPM and LPM groups were cryopreserved in straws and used for IVF. The proportions of embryos cleaved to 2- and 4-cell stages (88.1 ± 1.1 versus 80.5 ± 1.7, P = 0.001) and developed to blastocysts (33.5 ± 1.6 versus 23.5 ± 2.2, P = 0.026) were higher for HPM than LPM semen. The total cell number of embryos and blastocyst apoptotic index did not differ between groups. Although sperm progressive motility is associated with IVF competence, further examination is required to determine whether progressive motility can serve as a predictor of semen fertilization capacity in vivo.

This novel automatic vitrification device is capable to produce high survival rates of oocytes and embryos. We anticipate that as the demand for vitrification of gametes, embryos, and reproductive tissues increases worldwide, the availability of an automated vitrification device will become indispensable for standardization, simplification, and reproducibility of the entire process.