Factors affecting fertility in frozen-thawed boar sperm

Frozen-thawed boar sperm holds the potential to have an impact on the future of the swine industry. Utilization of this technology could improve a swine producer’s ability to access top-tier genetics from around the world, to improve efficiency, profitability, and the quality of product to meet consumer demands. Effective application of frozen-thawed sperm can help reduce the potential risk associated with devastating economic loss due to the spread of disease. Frozen storage of boar sperm also provides a safeguard in the event of disease outbreaks, as genetic material from paternal lines can be preserved and banked for repopulation purposes. Historically these benefits have been masked by reduction in fertility measures such as litter size. The reduced fertility results from the damage sustained by the sperm cell during cryopreservation. However, increased understanding of this damage has lead to improved cryopreservation methods, ultimately increasing post-thaw viability and fertility. Enhancements in breeding technology have also resulted in a better understanding of the AI methods required to achieve acceptable farrowing rates and litter size. Fertility following AI with frozen-thawed sperm is approaching that of liquid stored sperm, and producers may soon reap the benefits of this technology. This thesis will outline the current swine industry, opportunities for utilizing frozen-thawed sperm, the main components of sperm, why they are susceptible to damage, and current freezing and breeding practices. Objective 1 was to develop a cryopreservation protocol for our lab that resulted in consistent post-thaw motility ( ≥ 40%) that would eventually be used by Illinois boar studs for domestic and international sale of frozen sperm. Evaluation with both manual microscopy and CASA methods were conducted to verify quality. A preliminary breeding trial was then conducted to test the fertility of sperm frozen with this method. There were 41 ejaculates from 23 boars used for freezing. Sperm were frozen at 1.4x109 sperm/mL, averaging 55.61.1% (meanSE) motility, following thaw. The samples assessed were not different (P>0.05) in motility when compared with manual or CASA systems, and results were most reliable at a 1:40 sperm dilution. In the preliminary breeding trial, gilts (n=14) were inseminated with either a single (n=10) or double (n=4) AI using 1, 2, or 4x109 motile, frozen-thawed sperm. Overall, the resulting pregnancy rates averaged 71.4% and numbers of normal fetuses per litter averaged 15.51.3 per litter. A feasibility study for freezing cost per ejaculate was estimated at $275/ejaculate or $11/dose of frozen-thawed semen at standard doses of 5x109 total frozen-thawed sperm. This cost estimate did not include genetic value, fixed equipment costs, depreciation, or variable lab space fees. Objective 2 focused on the proper methods for breeding with frozen-thawed boar sperm to achieve fertility. Our hypothesis was that increased numbers of inseminations and increased numbers of motile frozen-thawed sperm would improve pregnancy rate and litter size. Results showed acceptable fertility at high sperm numbers, but also the optimal method for insemination with the lowest dose tested. Gilts (n=111) responded to synchronization methods and were bred with 1, 2, or 4x109 motile frozen-thawed sperm from six boars using a single AI at 32 h, or a double AI, with the first AI at 24 and 32 h following estrus. Ultrasound was conducted at 12 h intervals to estimate the time of ovulation. On day 32 of gestation, overall pregnancy rate (73%) and number of normal fetuses per litter (10.80.5) across all treatments did not differ, and were not affected by number of motile sperm, or the interaction of number of motile sperm and number of inseminations. However, the number of inseminations tended to affect (P=0.14) the number of normal fetuses. Litter size increased with a double AI compared to single AI. Multiple inseminations helped to allow insemination to occur close to ovulation in response to variation in the time of ovulation. Both pregnancy rate and number of normal fetuses were greater when the time of the AI at 32 h occurred closer to the estimated time of ovulation (P<0.05). In addition, other factors such as presence of an abnormal ovary at day 30 decreased (P<0.001) pregnancy rate, while boar affected number of normal fetuses (P<0.01). Analysis of our data using a fertility index revealed doses of 2x109 motile sperm with multiple AI can achieve acceptable fertility with use of less sperm, when compared to AI using 4x109 motile sperm. The methods described here will investigate the potential for improved fertility when using frozen-thawed sperm, while accounting for variation in time of ovulation.

Zika : how to communicate effectively

Communicating Zika virus preventive behaviors, transmission paths, and the underlying science are challenging. There are many scientific unknowns, an uncertainty of the risk, and tragic consequences for pregnant women and their fetuses. Clear, consistent, and frequent communication from trusted sources and channels makes it easier for people to understand Zika information which can help them to make decisions. Effective communication can support attitudes, norms, and behaviors to prevent the spread of Zika and protect pregnant women.

The viability of a tool for fetal health monitoring

Health monitoring has become widespread these past few years. Such applications include from exercise, food intake and weight watching, to specific scenarios like monitoring people who suffer from chronic diseases. More and more we see the need to also monitor the health of new-born babies and even fetuses. Congenital Heart Defects (CHDs) are the main cause of deaths among babies and doctors do not know most of these defects. Hence, there is a need to study what causes these anomalies, and by monitoring the fetus daily there will be a better chance of identifying the defects in earlier stages. By analyzing the data collected, doctors can find patterns and come up with solutions, thus saving peoples’ lives. In many countries, the most common fetal monitor is the ultrasound and the use of it is regulated. In Sweden for normal pregnancies, there is only one ultrasound scan during the pregnancy period. There is no great evidence that ultrasound can harm the fetus, but many doctors suggest to use it as little as possible. Therefore, there is a demand for a new non-ultrasound device that can be as accurate, or even better, on detecting the FHR and not harming the baby. The problems that are discussed in this thesis include how can accurate fetus health be monitored non-invasively at home and how could a fetus health monitoring system for home use be designed. The first part of the research investigates different technologies that are currently being used on fetal monitoring, and techniques and parameters to monitor the fetus. The second part is a qualitative study held in Sweden between April and May 2016. The data for the qualitative study was collected through interviews with 21 people, 10 mothers/mothers-to-be and 11 obstetricians/gynecologists/midwives. The questions were related to the Swedish pregnancy protocol, the use of technology in medicine and in particular during the pregnancy process, and the use of an ECG based monitoring device. The results show that there is still room for improvements on the algorithms to extract the fetal ECG and the survey was very helpful in understanding the need for a fetal home monitor. Parents are open to new technologies especially if it doesn't affect the baby's growth. Doctors are open to use ECG as a great alternative to ultrasound; on the other hand, midwives are happy with the current system. The remote monitoring feature is very desirable to everyone, if such system will be used in the future.

Ontogeny and nutritional programming of mitochondrial proteins in the ovine kidney, liver and lung

This study investigated the developmental and nutritional programming of two important mitochondrial proteins, namely voltage dependent anion channel (VDAC) and cytochrome c in the sheep kidney, liver and lung. The effect of maternal nutrient restriction between early to mid gestation (i.e. 28 to 80 days gestation, the period of maximal placental growth) on the abundance of these proteins was also examined in fetal and juvenile offspring. Fetuses were sampled at 80 and 140 days gestation (term ~147 days), and postnatal animals at 1 and 30 days and 6 months of age. The abundance of VDAC peaked at 140 days gestation in the lung, compared with 1 day after birth in the kidney and liver, whereas cytochrome c abundance was greatest at 140 days gestation in the liver, 1 day after birth in the kidney and 6 months of age in lungs. This differential ontogeny in mitochondrial protein abundance between tissues was accompanied with very different tissue specific responses to changes in maternal food intake. In the liver, maternal nutrient restriction only increased mitochondrial protein abundance at 80 days gestation, compared with no effect in the kidney. In contrast, in the lung mitochondrial protein abundance was raised near to term, whereas VDAC abundance was decreased by 6 months of age. These findings demonstrate the tissue specific nature of mitochondrial protein development that reflects differences in functional adaptation after birth. The divergence in mitochondrial response between tissues to maternal nutrient restriction early in pregnancy further reflects these differential ontogeny’s.

Select toxic metals status of pregnant women with history of pregnancy complications in Benin City, South-South Nigeria.

Toxic metals are part of the most important groups of environmental pollutants that can bind to vital cellular components and interfere with their functions via inhalation, foods, water etc. The serum levels of toxic metals (lead, mercury, cadmium and arsenic) in pregnant women with history of pregnancy complications, pregnant women without history of pregnancy complication and non-pregnant women in Benin City, South – South Nigeria was investigated in this paper, with total of 135 healthy women comprising of 45 pregnant women with history of previous pregnancy complications, 45 pregnant women without history of pregnancy complications and 45 non-pregnant women without history of pregnancy complications (controls). Some demographic characteristics and 4ml of blood samples were obtained from each subject for the analysis of lead, mercury, cadmium and arsenic by standard methods. Pregnant women with history of pregnancy complications recorded a highly significant increase in the toxic metal (lead) mean value of 25.81μg/dl as against 23.70μg/dl for pregnant women without history of pregnancy complications and 11.23μg/dl for non-pregnant (control) women without history of pregnancy complications as well as significant increases in the mean values of other toxic metals (mercury, cadmium and arsenic) compared with controls (p<0.001). The selected toxic metals (Pb, Hg, Cd and As) may be involved in the development of pregnancy complications among pregnant women in Benin City, South– South Nigeria. Lead in particular may pose threat to mothers and fetuses as its mean values in the two groups of pregnant women were well above normal.

Short Term Outcome of Neonates Born With Abnormal Umbilical Cord Arterial Blood Gases

Background: Umbilical arterial blood gas (UABG) analysis is more objective than other methods for predicting neonatal outcome. Acidemic neonates may be at risk for unfavorable outcome after birth, but all neonates with abnormal arterial blood gas (ABG) analysis do not always have poor outcome. Objectives: This study was carried out to determine the short term outcome of the neonates born with an abnormal ABG. Patients and Methods: In a cohort prospective study 120 high risk mother-neonate pairs were enrolled and UABG was taken immediately after birth. All neonates with an umbilical cord pH less than 7.2 were considered as case group and more than 7.2 as controls. Outcomes like need to resuscitation, admission to newborn services and/or NICU), seizure occurrence, hypoxic ischemic encephalopathy (HIE), delayed initiation of oral feeding and length of hospital stay were recorded and compared between the two groups. P value less than 0.05 was considered as being significant. Results: Comparison of short term outcomes between normal and abnormal ABG groups were as the fallowing: need for advanced resuscitation 4 vs. 0 (P = 0.001), NICU admission 16 vs. 4 (P = 0.001), convulsion 2 vs. 0 (P = 0.496), HIE 17 vs. 4 (P = 0.002), delay to start oral feeding 16 vs. 4 (P = 0.001), mean hospital stay 4 vs. 3 days (P = 0.001). None of the neonates died in study groups. Conclusions: An umbilical cord PH less than 7.2 immediately after birth can be used as a prognostic factor for unfavorable short term outcome in newborns.

Estudo sobre a migração transuterina e a razão sexual de conceptos em bovinos da raça Nelore (Bos taurus indicus)

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Agronomia e Medicina Veterinária, 2016.

Conjoined twins in a monochorionic triplet pregnancy after in vitro fertilization: a case report

Background: Monozygotic monochorionic triplet pregnancy with conjoined twins is a very rare condition and is associated with many complications. Case: In this study, we describe a monochorionic–diamniotic triplet pregnancy after in vitro fertilization with an intracytoplasmic sperm injection. At a gestational age of 6 weeks and 4 days of pregnancy one gestational sac was observed, and at a gestational age of 12 weeks and 2 days, triplets with conjoined twins were diagnosed. After consulting with the parents, they chose fetal reduction of the conjoined twins. Selective feticide was successfully performed by radiofrequency ablation at 16 weeks of pregnancy. Unfortunately, the day after the procedure, the membrane ruptured, and 1 week later, all fetuses and placenta were spontaneously aborted. Conclusion: Monochorionic triplet pregnancy with conjoined twins is very rare. These pregnancies are associated with very serious complications. Intra cytoplasmic sperm injection increases the rate of monozygotic twinning and conjoined twins. Counseling with parents before IVF is very important.

Human congenital cytomegalovirus infection: characteristics and pathogenesis of fetal brain damage

Human cytomegalovirus (HCMV) causes congenital neurological lifelong disabilities. The study analyzed 10 HCMV-infected human fetuses at 21 weeks of gestation to evaluate the characteristics and pathogenesis of brain injury related to congenital human CMV (cCMV) infection. Specifically, tissues from cortical and white matter areas, subventricular zone, thalamus, hypothalamus, hippocampus, basal ganglia and cerebellum were analysed by: i) immunohistochemistry (IHC) to detect HCMV-infected cell distribution, ii) hematoxylin-eosin staining to evaluate histological damage and iii) real-time PCR to quantify tissue viral load (HCMV-DNA). Viral tropism was assessed by double IHC to detect HCMV-antigens and neural/neuronal markers: nestin (expressed in early differentiation stage), doublecortin (DCX, identifying neuronal precursor cells) and neuronal nuclei (NeuN, identifying mature neurons). HCMV-positive cells and viral DNA were found in the brain of 8/10 (80%) fetuses. For these cases, brain damage was classified in mild (n=4, 50%), moderate (n=3, 37.5%) and severe (n=1, 12.5%) based on presence of i) diffuse astrocytosis, microglial activation and vascular changes; ii) occasional (in mild) or multiple (in moderate/severe) microglial nodules and iii) necrosis (in severe). The highest median HCMV-DNA level was found in the hippocampus (212 copies/5ng of humanDNA [hDNA], range: 10-7,505) as well as the highest mean HCMV-infected cell value (2.9 cells, range: 0-23), followed by that detected in subventricular zone (1.8 cells, range: 0-19). This suggests a preferential HCMV tropism for immature neuronal cells, residing in these regions, confirmed by the detection of DCX and nestin in 94% and 63.3% of HCMV-positive cells, respectively. NeuN was not found among HCMV-positive cells and was nearly absent in the brain with severe damage, suggesting HCMV does not infect mature neurons and immature HCMV-infected neuronal cells do not differentiate into neurons. HCMV preferential tropism in immature neural/neuronal cells delays/inhibits their differentiation interfering with brain development processes that lead to structural and functional brain defects.

Role of second trimester ultrasound in prediction of newborns neurologic damage after maternal Cytomegalovirus infection

Objective To find a correlation between cerebral symptoms at birth and abnormalities found at anomaly scan, through the analysis of sensitivity of the anomaly scan in the prediction of severe CMV neonatal disease. Methods - Design, Setting, Population This was a retrospective collection of all cases of primary congenital CMV infection reported in our unit (Obstetrics and Perinatal Medicine, Policlinico di S Orsola, IRCSS, Bologna) over a period of 9 years (2013–2022). Only cases of fetal infection following confirmed maternal primary infection in the first trimester (MPI) and newborns with confirmed CMV infection on blood/saliva or urine were included. Results Between 2014 and 2022, 69 fetuses had an antenatal diagnosis of primary CMV infection. The infection occurred after MPI in the first, second, and third trimester in 63.7% (43/69), 27.5% (19/69), and 10% (7/69) of cases, respectively. Second-trimester assessment by anomaly scan was abnormal in 10/69 (15%) fetuses: 5/69 (7%) had an extracerebral STA and 5/69 (7%) had a cerebral STA. Normal anomaly scan was found in 59/69 (86%) fetuses. When looking at all fetuses infected in the first trimester, 12.5% (5/40) underwent TOP and 45% (18/40) had symptoms at birth. A mean follow-up of 22.4 months (range 12–48 months) was available for 68/69 (99%) live born neonates. Conclusion Anomaly scan results to have a predictive positive value of 67% fetuses infected in the first trimester. Serial assessment by ultrasound is necessary to predict the risk of sequelae occurring in 35% following fetal infection in the first trimester of pregnancy. This combined evaluation by US and trimester of infection should be useful when counselling on the prognosis of cCMV infection.