Influence of sperm-oocyte coincubation period on porcine in vitro fertilization (IVF) efficiency

A major obstacle for successful in vitro production of porcine embryos is the polyspermic fertilization. One possibility to reduce polyspermic penetration is decreasing the number of spermatozoa added to the fertilization medium. Unfortunately, the lower rate of polyspermy is accompanied by a reduced penetration rate. A short gamete coincubation period of 10 min has been described to obtain fertilization rates similar to 6 h of coincubation and may improve IVF efficiency (number of monospermic fertilized oocytes/total number inseminated) depending on sperm-oocyte ratio (Gil, 2007, Theriogenology, 67(3), 620–626). Here we demonstrate that the optimal coincubation period in our IVF conditions is between 10 min and 6 h. In vitro matured oocytes (n = 600) were inseminated with frozen-thawed epididymal semen with 600 spermatozoa per oocyte and coincubated for 2, 4 and 6 h. At 2 and 4 h post insemination (hpi), oocytes were vortexed and transferred to fertilization medium without spermatozoa. At 6 hpi, presumed zygotes of all groups were washed three times in culture medium and cultured. At 22 hpi, zygotes were fixed overnight and stained with Hoechst 33,342 for the assessment of fertilization and polyspermy. The IVF efficiency was higher for the 4 h group (40 ± 5%) than the 2 and 6 h group (19 ± 8% and 17 ± 5%). Between 4 and 6 h of gamete coincubation, the increase in the number of polyspermic oocytes was relatively higher than the increase in penetration rate (+39% vs. +15%), resulting in a decline in efficiency. (This study was supported by Research Foundation-Flanders)

Ovariële cysten bij de zeug: een multifactoriële aandoening met een weerslag op de vruchtbaarheid

Ovarian cysts are an important cause of reduced reproductive performance in gilts and sows. The formation of follicular cysts, although not completely elucidated yet, is associated with a deficiency in LH release and the continuous growth of follicles. Luteal cysts might arise from ovulated follicles due to premature closure of the ovulation site. The formation of ovarian cysts is probably multifactorial. Genetic factors may affect the susceptibility to this fertility disorder. Risk factors for the development of ovarian cysts are stress, mycotoxins and hormonal treatment over a long period of time, or at an improper phase of the reproductive cycle. Other, still unknown risk factors may be involved. Ovarian cysts can cause anestrus, irregular estrus cycles, behavioral changes, lower conception rates and a reduced litter size. However, not all ovarian cysts are clinically apparent. The diagnosis of ovarian cysts is based on ultrasonography. Various treatments are possible (GnRH, hCG or PGF(2a)) but the effectiveness is unsatisfactory. Therefore, in most cases the sow is culled

Risicofactoren voor oorhematomen bij biggen

In this study, the incidence of auricular hematomas in weaned piglets was examined, and it was investigated whether the incidence was linked with certain risk factors. In total, five successive batches of weaned pigs (n=10,657) were investigated from weaning (three weeks of age) until the end of the nursery period (ten weeks of age). The mean incidence was 2.3%, ranging from 1.3% to 2.9% for the different weaned batches. The incidence in barrows was 2.1% and in female piglets 2.5% (P > 0.05). The incidence in piglets from primiparous sows was 0.92% and in piglets of multiparous sows 2.45% (P < 0.001). The highest incidence was observed four to five weeks after weaning. There was a two-times-higher risk to develop an auricular hematoma at the ear with the ear tag (RR=2.28 [1.74 - 2.98]). Further research, preferably in more herds, is warranted to confirm and/or explain the present results and to further optimize control measures

Associations of faecal microbiota with influenza-like illness in participants aged 60 years or older:an observational study

Background People aged 60 years or older are at high risk for respiratory infections, one of the leading causes of mortality worldwide. Vaccination is the main way to protect against these infections; however, vaccination is less effective in older adults than in younger adults due to ageing of the immune system, so innovative strategies that improve vaccine responses could provide a major public health benefit. The gut microbiota regulates host immune homoeostasis and response against pathogens, but human studies showing the effects of the gut microbiota on respiratory infections in older adults are sparse. We aimed to investigate the composition of the microbiota in relation to respiratory infections and local and systemic immune markers in older adults during an influenza season. Methods In this observational study, participants were selected from an influenza-like illness (ILI) prospective surveillance cohort in which community-dwelling adults aged 60 years and older in the Netherlands were recruited through their general practitioner or the Civil Registry. Inclusion criteria have been described elsewhere. Participants completed questionnaires and self-reported symptoms. To measure microbiota composition, faecal samples were collected from participants registering an ILI event, with a follow-up (recovery) sample collected 7-9 weeks after the ILI event, and from asymptomatic participants not reporting any event throughout the season. We tested associations between microbiota profiles and a set of health-related variables, patient characteristics, and local and systemic immune markers. We cultured identified bacterial biomarkers for ILI with CaCo-2 cells in an in vitro intestinal epithelial model and measured the induced immune response. This study is registered with http://www.trialregister.nl, NL4666. Findings Between Oct 1, 2014, and April 30, 2015, 2425 older adults were recruited into the ILI surveillance cohort. From Oct 1, 2014, to June 15, 2015, faecal samples were collected from 397 participants, of whom 213 (54%) reported an ILI event once throughout the season and 184 (46%) did not. 192 ILI participants recovered and provided follow-up samples. Microbiota composition was altered during an ILI event. The Bacteroidetes (mean relative abundance 17.51% [SD 11.41] in the ILI group and 14.19% [10.02] in the control group; adjusted p=0.014) and the Proteobacteria (3.40% [8.10] in the ILI group and 1.57% [3.69] in the control group; adjusted p=0.015) were more abundant in the ILI group than in the control group. The abundance of Ruminococcus torques was positively associated with ILI and the abundance of Escherichia/Shigella, negatively correlated with alpha diversity, and negatively co-occurred with beneficial taxa, including butyrate producers. R torques was associated with pro-inflammatory profiles, both locally in faeces and systemically in blood. ILI-associated taxa (R torques and Escherichia coli) had symbiotic effects on the cellular immune response when cultured together in an in vitro model. Interpretation The abundances of specific bacteria could be used as potential biomarkers for susceptibility to respiratory infections and as targets for intervention in the ageing population. Copyright (C) 2020 The Author(s). Published by Elsevier Ltd

Whole-Inactivated Influenza Virus Is a Potent Adjuvant for Influenza Peptides Containing CD8+ T Cell Epitopes

Influenza peptide antigens coding for conserved T cell epitopes have the capacity to induce cross-protective influenza-specific immunity. Short peptide antigens used as a vaccine, however, often show poor immunogenicity. In this study, we demonstrate that whole-inactivated influenza virus (WIV) acts as an adjuvant for influenza peptide antigens, as shown by the induction of peptide-specific CD8+ T cells in HLA-A2.1 transgenic mice upon vaccination with the influenza-M1-derived GILGFVFTL peptide (GIL), formulated with WIV. By screening various concentrations of GIL and WIV, we found that both components contributed to the GIL-specific T cell response. Whereas co-localization of the peptide antigen and WIV adjuvant was found to be important, neither physical association between peptide and WIV nor fusogenic activity of WIV were relevant for the adjuvant effect of WIV. We furthermore show that WIV may adjuvate T cell responses to a variety of peptides, using pools of either conserved wild-type influenza peptides or chemically altered peptide ligands. This study shows the potential of WIV as an adjuvant for influenza peptides. The simple formulation process and the solid safety record of WIV make this an attractive adjuvant for T cell peptides, and may also be used for non-influenza antigens

Адаптация гидравлической модели водостока к бассейнам рек Дунай и Днестр

Гидравлическая модель водостока адаптирована к бассейну рек Дунай и Днестр. По данным орографии, атмосферных осадках или поверхностном стоке она позволяет рассчитывать объемы, расходы и уровни воды с пространственным разрешением 1 км. В модели возможно использование данные об экосистемах на земной поверхности, типах почвы. По данным наблюдений стока оценены среднемесячные величины расходов рек, которые соответствуют наблюдениям, что позволяет применять модель в дальнейших оценках стока, наносов и т.д.Hydraulic model of water inflow is adapted to the Danube and the Dniester rivers basin. According to the orography, precipitation and surface inflow data it permits to calculate water volumes, discharges and levels with spatial resolution 1 km. It is possible to use the data on ecosystems on the ground surface, types of soil in the model. According to the observations data of the inflow the average monthly values of river discharges corresponding to the observations are estimated. It permits to apply the model in the further estimations of inflow, alluvia e t.c

Inhibitors of zinc-dependent metalloproteases hinder sperm passage through the cumulus oophorus during porcine fertilization in vitro

In this study, we report for the first time on a possible contribution of metalloproteases in sperm passage through the cumulus matrix in pigs. The presence of 20 mu M 1,10-phenanthroline (1,10-PHEN), inhibitor of zinc-dependent metalloproteases, strongly inhibited the degree of sperm penetration in cumulus-intact (CI), but not in cumulus-free (CF), porcine oocytes during IVF. The inhibitory effect of 1,10-PHEN was due to the chelation of metal ions as a non-chelating analog (1,7-PHEN) did not affect IVF rates. Furthermore, incubation with 1,10-PHEN did not affect sperm binding to the zona pellucida nor sperm motility, membrane integrity, or acrosomal status. These findings led to the assumption that 1,10-PHEN interacts with a sperm-or cumulus-derived metalloprotease. Metalloproteases are key players in physiological processes involving degradation or remodeling of extracellular matrix. In vivo, their proteolytic activity is regulated by tissue inhibitors of metalloproteases (TIMP1-TIMP4). We tested the effect of TIMP3 on fertilization parameters after porcine IVF. Similar to 1,10-PHEN, TIMP3 inhibited total fertilization rate of CI but not CF oocytes and did not influence sperm quality parameters. Although the inhibitory effect was stronger in CI oocytes, TIMP3 also reduced the degree of sperm penetration in CF oocytes, suggesting the involvement of a metalloprotease in a subsequent step during fertilization. In conclusion, our results indicate the involvement of TIMP3-sensitive, zinc-dependent metalloprotease activity in sperm passage through the cumulus oophorus in pigs. The results should provide the basis for further biochemical research toward the localization and identification of the metalloprotease involved