Bovine Follicular Dynamics, Oocyte Recovery,and Development of Oocytes Microinjected with a Green Fluorescent Protein Construct

The present study was carried out to 1) evaluate the viability of in vitro fertilized zygotes after microinjection of DNA, 2) assess the influence of oocyte quality upon the development rate of embryos when injected with DNA, and 3) determine the integration frequency of green fluorescent protein DNA into microinjected embryos. Oocytes were aspirated from ovaries of nine nonlactating Holsteins and were categorized into grades A, B, C, and D. At 16 h after in vitro fertilization, approximately half of the pronuclear stage presumptive zygotes were classified as having 1 pronucleus or 2 pronuclei, and they were microinjected with DNA constructs. A potential predictor of DNA integration frequency at d 10 was assessment of the incidence of green fluorescing embryos. The proportion of cleaved embryos that developed to morulae or blastocysts was not different between groups with 1 pronucleus injected (45%), 1 pronucleus uninjected (64%), or 2 pronuclei injected (49%). However, the development of morulae or blastocysts was higher in the group with 2 pronuclei uninjected (69%). The overall developmental score of green fluorescent protein-positive embryos was higher for grade A oocytes (1.3 &#;&#;0.1) than for grade B (0.8 &#; 0.1), C (0.6 &#;&#;0.1), or D (0.3 &#;&#;0.1) oocytes. The results show that production of transgenic bovine blastocysts can occur from the microinjection of a presumptive zygote having only one visible pronucleus. Initial oocyte quality is an important factor in selection of oocytes suitable for microinjection of DNA and for preimplantation development to produce bovine transgenic embryos

Reflections on a 'virtual' practice development unit: changing practice through identity development

Aims. This paper draws together the personal thoughts and critical reflections of key people involved in the establishment of a ‘virtual’ practice development unit of clinical nurse specialists in the south of England. Background. This practice development unit is ‘virtual’ in that it is not constrained by physical or specialty boundaries. It became the first group of Trust-wide clinical nurse specialists to be accredited in the UK as a practice development unit in 2004. Design and methods. The local university was asked to facilitate the accreditation process via 11 two-hour audio-recorded learning sessions. Critical reflections from practice development unit members, leaders and university staff were written 12 months after successful accreditation, and the framework of their content analysed. Findings and discussion. Practice development was seen as a way for the clinical nurse specialists to realize their potential for improving patient care by transforming care practice in a collaborative, interprofessional and evolutionary manner. The practice development unit provided a means for these nurses to analyse their role and function within the Trust. Roberts’ identity development model for nursing serves as a useful theoretical underpinning for the reflections contained in this paper. Conclusions. These narratives provide another example of nurses making the effort to shape and contribute to patient care through organizational redesign. This group of nurses began to realize that the structure of the practice development unit process provided them with the means to analyse their role and function within the organization and, as they reflected on this structure, their behaviour began to change. Relevance to clinical practice. Evidence from these reflections supports the view that practice development unit participants have secured a positive and professional identity and are, therefore, better able to improve the patient experience

Association between FAMACHA scores and fecal egg counts in katahdin lambs

© 2017 American Society of Animal Science. All rights reserved. The FAMACHA system was introduced to the U.S. just over 10 yr ago to allow selective deworming of lambs with anemia associated with Haemonchus contortus and retard the development of anthelmintic resistance. The FAMACHA system was initially developed as a predictor of packed cell volume (PCV), but correlations between FAMACHA and fecal egg counts (FEC) have also been reported. It is important to understand factors that influence FAMACHA scores among farms to improve management of gastrointestinal nematodes. The objectives of this study were therefore to quantify associations between FAMACHA scores, FEC, BW, and age in Katahdin lambs at 2 different measurement times in 8 flocks in the eastern U.S., and to assess consistency of relationships between FAMACHA and FEC among flocks. Data came from 1,644 Katahdin lambs from 7 flocks sampled at approximately 90 d of age, and 1,295 lambs from 6 flocks sampled at approximately 120 d of age over a 5 yr period. Residual correlations among log-transformed FEC (LFEC), FAMACHA scores, BW, and lamb ages at each measurement time were determined. Repeatability of each variable was also determined as residual correlations among repeated measures. At both 90 and 120 d of age, correlations of FAMACHA scores with LFEC and BW were significant (P \u3c 0.001), but numerically modest (0.25 and -0.16, respectively at 90 d; 0.31 and -0.16, respectively at 120 d), demonstrating that higher FAMACHA scores were associated with higher FEC and more likely to be observed in lighter lambs. A small negative correlation was observed between FAMACHA score and lamb age (r = -0.05, P = 0.05, 90 d; r = -0.11, P \u3c 0.001, 120 d) indicating that younger lambs were more likely to have elevated FAMACHA scores. Thus, younger and lighter lambs will likely be more susceptible to parasitism and may need to be managed more diligently than older or heavier lambs. In addition, FAMACHA scores have potential to improve breeding value estimates in programs designed to genetically improve parasite resistance

Sex influence on genetic expressions of early growth in Afshari lambs

Very little is known about the genetic aspects of sexual dimorphism of body weight in domestic sheep, and therefore this study was conducted to quantify the genetic basis of sexual dimorphism for early-growth-related traits in Afshari lambs. Traits evaluated included birth weight (BW), weaning weight (WW), and growth rate (GR) in male and female lambs. Male lambs were 6.6 % heavier at birth, had 14.4 % higher preweaning growth rates and were 16.0 % heavier at weaning compared to female lambs. Levels of sexual-size dimorphism (SSD), expressed as the ratio of male to female means, for BW, WW and GR were 1.07, 1.14 and 1.15, respectively, which indicated low levels of SSD in the traits studied. Fixed effects of year of birth and type of birth interacted with sex effects, with greater variability in birth and weaning weights among years and birth types in male lambs, suggesting greater environmental sensitivity in the males. Bivariate animal models and restricted maximum likelihood (REML) procedures were used to estimate phenotypic variances and their genetic and non-genetic components in male and female lambs. Estimates of the direct heritability (h2) and additive coefficient of variation (CVA) for BW were higher in males. However, for WW and GR, heritability estimates were higher in females. In contrast, whereas the contribution of maternal permanent environmental effects (c2) to variation of BW was higher in females, for WW and GR higher estimates of c2 were observed in males. Respective genetic and maternal permanent environmental correlations between records on males and females were 0.986 and 0.723 for BW, 0.995 and 0.983 for WW, and 0.995 and 0.966 for GR, indicating possible sexual dimorphism only for maternal effects on BW. Based on an approximate 95 % confidence interval, none of the observed differences in variance components between sexes differed from zero and none of the observed genetic or maternal correlations differed from 1.0, indicating no need or opportunity for sex-specific selection strategies

Genetic parameters for fecal egg counts and their relationship with body weights in katahdin lambs

© The Author(s) 2018. Reliance on anthelmintic drugs to control internal parasites in sheep is no longer sustainable because of the development of resistance to these drugs in parasite populations. Genetic selection may offer an alternative long-term solution, as differences in parasite resistance exist both within and among sheep breeds. However, selection for parasite resistance may have correlated effects on other production traits. The objectives of this study were to estimate genetic parameters for weaning (WFEC) and postweaning (PWFEC) fecal egg counts (FEC) and assess their relationship with birth (BWT), weaning (WWT), and postweaning (PWWT) BW in Katahdin lambs. The study used WFEC (n = 2,537), PWFEC (n = 3.421), BWT (n = 12,869), WWT (n = 10,961), and PWWT (n = 7,812) from 12,869 lambs measured between 2003 and 2015 in 13 flocks enrolled in the U.S. National Sheep Improvement Program. Animal and sire models were fitted to the data using the ASReml statistical package. Records were corrected for fixed effects of dam age, joint effect of type of birth and rearing, and management group (defined by joint effects of flock, sex, and birth year and season); lamb age in days at each measurement time was fitted as a covari-ate. Maternal additive and maternal permanent environmental effects were not significant (P \u3e 0.05), but litter effects influenced (P \u3c 0.01) both WFEC and PWFEC. Heritability estimates ranged from 0.18 to 0.26 for WFEC and 0.23 to 0.46 for PWFEC, depending on the model used. Heritability estimates from sire models were higher than estimates from animal models. Direct additive, litter, residual, and phenotypic correlations between WFEC and PWFEC were 0.82, 0.25, 0.15, and 0.29, respectively. Bivariate analyses revealed low to moderate correlations between BW and FEC. Moderate heritabilities for FEC in this study indicated that genetic progress for this trait can be achieved in Katahdin lambs and that selection for low FEC should have little or no effect on BW

Validation of Live Animal Ultrasonic Measurements of Body Composition in Market Lambs

Market lambs from the state fair of Virginia (n = 172) were ultrasonically evaluated by 4 scan technicians and 3 image interpreters to determine accuracy of ultrasonic estimates of loin muscle area (ULMA), backfat thickness (UBF), and body wall thickness (UBW). Lambs were initially scanned at the preferred magnification setting of each technician; 2 chose 1.5× and 2 chose 2.0×. Lambs were then scanned a second time for ULMA and UBF with machine magnification settings changed from 1.5 to 2.0×, or vice versa, midway through the second scan. Lambs were then slaughtered, and analogous measurements [carcass loin muscle area, carcass backfat thickness, and carcass body wall thickness (CBW)] were recorded on chilled carcasses. Pooled, residual correlation coefficients within technicians and interpreters between ultrasonic measurements from the first scan and carcass measurements were 0.66 for loin muscle area, 0.78 for backfat thickness, and 0.73 for body wall thickness, but were reduced to 0.43, 0.69, and 0.50, respectively, by inclusion of linear effects of carcass weight in the model. Mean bias for technicians and interpreters ranged from −1.30 to −2.66 cm2 for loin muscle area, −0.12 to −0.17 cm for backfat thickness, and 0.14 to −0.03 cm for body wall thickness; prediction errors ranged from 1.86 to 2.22 cm2, 0.12 to 0.14 cm, and 0.35 to 0.38 cm, respectively. Pooled correlations between repeated measures were 0.67 for ULMA, 0.79 for UBF, and 0.68 for UBW at the same magnification and 0.73 for ULMA and 0.76 for UBF across different magnification settings. Mean differences between repeated measures were more variable among technicians and interpreters than statistics comparing ultrasound to carcass measures. Standard errors of repeatability ranged from 1.61 to 2.45 cm2 for ULMA, 0.07 to 0.11 cm for UBF, and 0.36 to 0.42 cm for UBW. The effect of changing magnification setting on technician and interpreter repeatability was small for UBF and ULMA. The accuracy of prediction of CBW from UBW was similar to that achieved for backfat thickness; further assessment of the value of ultrasonic measurements of body wall thickness in lambs is warranted. These results indicate that ultrasound scanning can reliably predict carcass loin muscle area and backfat thickness in live lambs and, accordingly, has value in selection programs to improve composition. Development of certification standards for US lamb ultrasound technicians based on results of this study and others is proposed

The Impact of Nonequilibrium Conditions in Lung Surfactant: Structure and Composition Gradients in Multilamellar Films

The lipid–protein mixture that covers the lung alveoli, lung surfactant, ensures mechanical robustness and controls gas transport during breathing. Lung surfactant is located at an interface between water-rich tissue and humid, but not fully saturated, air. The resulting humidity difference places the lung surfactant film out of thermodynamic equilibrium, which triggers the buildup of a water gradient. Here, we present a millifluidic method to assemble multilamellar interfacial films from vesicular dispersions of a clinical lung surfactant extract used in replacement therapy. Using small-angle X-ray scattering, infrared, Raman, and optical microscopies, we show that the interfacial film consists of several coexisting lamellar phases displaying a substantial variation in water swelling. This complex phase behavior contrasts to observations made under equilibrium conditions. We demonstrate that this disparity stems from additional lipid and protein gradients originating from differences in their transport properties. Supplementing the extract with cholesterol, to levels similar to the endogenous lung surfactant, dispels this complexity. We observed a homogeneous multilayer structure consisting of a single lamellar phase exhibiting negligible variations in swelling in the water gradient. Our results demonstrate the necessity of considering nonequilibrium thermodynamic conditions to study the structure of lung surfactant multilayer films, which is not accessible in bulk or monolayer studies. Our reconstitution methodology also opens avenues for lung surfactant pharmaceuticals and the understanding of composition, structure, and property relationships at biological air–liquid interfaces

Administration of intrapulmonary sodium polyacrylate to induce lung injury for the development of a porcine model of early acute respiratory distress syndrome.

BACKGROUND: The loss of alveolar epithelial and endothelial integrity is a central component in acute respiratory distress syndrome (ARDS); however, experimental models investigating the mechanisms of epithelial injury are lacking. The purpose of the present study was to design and develop an experimental porcine model of ARDS by inducing lung injury with intrapulmonary administration of sodium polyacrylate (SPA). METHODS: The present study was performed at the Centre for Comparative Medicine, University of British Columbia, Vancouver, British Columbia. Human alveolar epithelial cells were cultured with several different concentrations of SPA; a bioluminescence technique was used to assess cell death associated with each concentration. In the anesthetized pig model (female Yorkshire X pigs (n = 14)), lung injury was caused in 11 animals (SPA group) by injecting sequential aliquots (5 mL) of 1% SPA gel in aqueous solution into the distal airway via a rubber catheter through an endotracheal tube. The SPA was dispersed throughout the lungs by manual bag ventilation. Three control animals (CON group) underwent all experimental procedures and measurements with the exception of SPA administration. RESULTS: The mean (± SD) ATP concentration after incubation of human alveolar epithelial cells with 0.1% SPA (0.92 ± 0.27 μM/well) was approximately 15% of the value found for the background control (6.30 ± 0.37 μM/well; p < 0.001). Elastance of the respiratory system (E RS) and the lung (E L) increased in SPA-treated animals after injury (p = 0.003 and p < 0.001, respectively). Chest wall elastance (E CW) did not change in SPA-treated animals. There were no differences in E RS, E L, or E CW in the CON group when pre- and post-injury values were compared. Analysis of bronchoalveolar lavage fluid showed a significant shift toward neutrophil predominance from before to after injury in SPA-treated animals (p < 0.001) but not in the CON group (p = 0.38). Necropsy revealed marked consolidation and congestion of the dorsal lung lobes in SPA-treated animals, with light-microscopy evidence of bronchiolar and alveolar spaces filled with neutrophilic infiltrate, proteinaceous debris, and fibrin deposition. These findings were absent in animals in the CON group. Electron microscopy of lung tissue from SPA-treated animals revealed injury to the alveolar epithelium and basement membranes, including intra-alveolar neutrophils and fibrin on the alveolar surface and intravascular fibrin (microthrombosis). CONCLUSIONS: In this particular porcine model, the nonimmunogenic polymer SPA caused a rapid exudative lung injury. This model may be useful to study ARDS caused by epithelial injury and inflammation

Adding gastrointestinal parasite resistance to the breeding objective in hair sheep: initial step

The U.S. Maternal Hair Index was designed by the National Sheep Improvement Program (NSIP) to increase total weight of lamb weaned per ewe lambing (TW). Producers are interested in adding gastrointestinal nematode (GIN) resistance to this breeding objective since parasitism causes substantial economic losses. The NSIP provides producers with estimated breeding values (EBV) for postweaning fecal egg count (PFEC), an indicator of GIN resistance. Our objective was to gauge the effects of including PFEC as another selection criterion and goal trait. Selection index theory was used to construct 11 indexes. First was a benchmark index with TW as the goal. Weaning weight, and number of lambs born and weaned, were selection criteria. An index was then designed with PFEC added as a selection criterion. In 9 more indices, PFEC was also included as a goal trait, where the economic value of TW was scaled relative to that of PFEC. PFEC received a scaled economic value of −1 with that of TW increasing from +1 to +5 at +0.5 increments. Selection criteria were modeled as EBV or phenotypes. Annual genetic responses in goal traits were predicted. The top 3% of males and 26% of females were selected. Breeding values and phenotypes were simulated for 200 lambs by Cholesky decomposition and used to generate index scores, with 100 replicates run. Concordances with the animals selected in the benchmark scenario were determined. Using EBV as the selection criteria, TW increased by 1.46 kg/yr in the benchmark scenario. However, unfavorably, PFEC increased by 2.24%/yr. When using phenotypic criteria, TW increased by 0.52 kg/yr and PFEC by 0.28%/yr. Once added as a goal trait, PFEC decreased regardless of the scaled economic value of TW. However, responses in TW were also reduced, although less so as its scaled economic value increased. A scaled economic value of +3 for TW appeared reasonable over other choices with 79% of the emphasis placed on TW in the breeding objective. With EBV as selection criteria, PFEC declined by 7.96%/yr with 98% of the response in TW retained. Also, on average, 64% of males and 80% of females were chosen in common with the benchmark. With phenotypic selection criteria, PFEC declined by 5.13%/yr and 94% of the response in TW was retained; 61% of males and 80% of females were chosen in common with the benchmark. Implementing an index with scaled economic values of +3 for TW and −1 for PFEC would balance gains in TW with reductions in PFEC