Comparison of the genetic characteristics of directly measured and Fourier-transform mid-infrared-predicted bovine milk fatty acids and proteins.

Fourier-transform mid-infrared (FT-MIR) spectroscopy is a high-throughput and inexpensive methodology used to evaluate concentrations of fat and protein in dairy cattle milk samples. The objective of this study was to compare the genetic characteristics of FT-MIR predicted fatty acids and individual milk proteins with those that had been measured directly using gas and liquid chromatography methods. The data used in this study was based on 2,005 milk samples collected from 706 Holstein-Friesian × Jersey animals that were managed in a seasonal, pasture-based dairy system, with milk samples collected across 2 consecutive seasons. Concentrations of fatty acids and protein fractions in milk samples were directly determined by gas chromatography and high-performance liquid chromatography, respectively. Models to predict each directly measured trait based on FT-MIR spectra were developed using partial least squares regression, with spectra from a random selection of half the cows used to train the models, and predictions for the remaining cows used as validation. Variance parameters for each trait and genetic correlations for each pair of measured/predicted traits were estimated from pedigree-based bivariate models using REML procedures. A genome-wide association study was undertaken using imputed whole-genome sequence, and quantitative trait loci (QTL) from directly measured traits were compared with QTL from the corresponding FT-MIR predicted traits. Cross-validation prediction accuracies based on partial least squares for individual and grouped fatty acids ranged from 0.18 to 0.65. Trait prediction accuracies in cross-validation for protein fractions were 0.53, 0.19, and 0.48 for α-casein, ß-casein, and κ-casein, 0.31 for α-lactalbumin, 0.68 for ß-lactoglobulin, and 0.36 for lactoferrin. Heritability estimates for directly measured traits ranged from 0.07 to 0.55 for fatty acids; and from 0.14 to 0.63 for individual milk proteins. For FT-MIR predicted traits, heritability estimates were mostly higher than for the corresponding measured traits, ranging from 0.14 to 0.46 for fatty acids, and from 0.30 to 0.70 for individual proteins. Genetic correlations between directly measured and FT-MIR predicted protein fractions were consistently above 0.75, with the exceptions of C18:0 and C18:3 cis-3, which had genetic correlations of 0.72 and 0.74, respectively. The GWAS identified trait QTL for fatty acids with likely candidates in the DGAT1, CCDC57, SCD, and GPAT4 genes. Notably, QTL for SCD were largely absent in the FT-MIR predicted traits, and QTL for GPAT4 were absent in directly measured traits. Similarly, for directly measured individual proteins, we identified QTL with likely candidates in the CSN1S1, CSN3, PAEP, and LTF genes, but the QTL for CSN3 and LTF were absent in the FT-MIR predicted traits. Our study indicates that genetic correlations between directly measured and FT-MIR predicted fatty acid and protein fractions are typically high, but that phenotypic variation in these traits may be underpinned by differing genetic architecture.

Non-additive QTL mapping of lactation traits in 124,000 cattle reveals novel recessive loci.

BACKGROUND: Deleterious recessive conditions have been primarily studied in the context of Mendelian diseases. Recently, several deleterious recessive mutations with large effects were discovered via non-additive genome-wide association studies (GWAS) of quantitative growth and developmental traits in cattle, which showed that quantitative traits can be used as proxies of genetic disorders when such traits are indicative of whole-animal health status. We reasoned that lactation traits in cattle might also reflect genetic disorders, given the increased energy demands of lactation and the substantial stresses imposed on the animal. In this study, we screened more than 124,000 cows for recessive effects based on lactation traits. RESULTS: We discovered five novel quantitative trait loci (QTL) that are associated with large recessive impacts on three milk yield traits, with these loci presenting missense variants in the DOCK8, IL4R, KIAA0556, and SLC25A4 genes or premature stop variants in the ITGAL, LRCH4, and RBM34 genes, as candidate causal mutations. For two milk composition traits, we identified several previously reported additive QTL that display small dominance effects. By contrasting results from milk yield and milk composition phenotypes, we note differing genetic architectures. Compared to milk composition phenotypes, milk yield phenotypes had lower heritabilities and were associated with fewer additive QTL but had a higher non-additive genetic variance and were associated with a higher proportion of loci exhibiting dominance. CONCLUSIONS: We identified large-effect recessive QTL which are segregating at surprisingly high frequencies in cattle. We speculate that the differences in genetic architecture between milk yield and milk composition phenotypes derive from underlying dissimilarities in the cellular and molecular representation of these traits, with yield phenotypes acting as a better proxy of underlying biological disorders through presentation of a larger number of major recessive impacts.

Non-additive association analysis using proxy phenotypes identifies novel cattle syndromes.

Mammalian species carry ~100 loss-of-function variants per individual1,2, where ~1-5 of these impact essential genes and cause embryonic lethality or severe disease when homozygous3. The functions of the remainder are more difficult to resolve, although the assumption is that these variants impact fitness in less manifest ways. Here we report one of the largest sequence-resolution screens of cattle to date, targeting discovery and validation of non-additive effects in 130,725 animals. We highlight six novel recessive loci with impacts generally exceeding the largest-effect variants identified from additive genome-wide association studies, presenting analogs of human diseases and hitherto-unrecognized disorders. These loci present compelling missense (PLCD4, MTRF1 and DPF2), premature stop (MUS81) and splice-disrupting (GALNT2 and FGD4) mutations, together explaining substantial proportions of inbreeding depression. These results demonstrate that the frequency distribution of deleterious alleles segregating in selected species can afford sufficient power to directly map novel disorders, presenting selection opportunities to minimize the incidence of genetic disease.

A new mechanism for a familiar mutation - bovine DGAT1 K232A modulates gene expression through multi-junction exon splice enhancement.

BACKGROUND: The DGAT1 gene encodes an enzyme responsible for catalysing the terminal reaction in mammary triglyceride synthesis, and underpins a well-known pleiotropic quantitative trait locus (QTL) with a large influence on milk composition phenotypes. Since first described over 15 years ago, a protein-coding variant K232A has been assumed as the causative variant underlying these effects, following in-vitro studies that demonstrated differing levels of triglyceride synthesis between the two protein isoforms. RESULTS: We used a large RNAseq dataset to re-examine the underlying mechanisms of this large milk production QTL, and hereby report novel expression-based functions of the chr14 g.1802265AA > GC variant that encodes the DGAT1 K232A substitution. Using expression QTL (eQTL) mapping, we demonstrate a highly-significant mammary eQTL for DGAT1, where the K232A mutation appears as one of the top associated variants for this effect. By conducting in vitro expression and splicing experiments in bovine mammary cell culture, we further show modulation of splicing efficiency by this mutation, likely through disruption of an exon splice enhancer as a consequence of the allele encoding the 232A variant. CONCLUSIONS: The relative contributions of the enzymatic and transcription-based mechanisms now attributed to K232A remain unclear; however, these results suggest that transcriptional impacts contribute to the diversity of lactation effects observed at the DGAT1 locus.

Inflammatory Effects of Blood-Air Interface in a Porcine Cardiopulmonary Bypass Model.

Cardiopulmonary bypass (CPB) causes a systemic inflammatory response syndrome (SIRS) associated with multiorgan injury. A model was developed to test whether a blood-air interface (BAI) in the CPB circuit causes blood element activation and inflammation. Ten healthy swine were placed on partial CPB for 2 hours via the cervical vessels and monitored for 96 hours postoperatively. Five pigs (control group) had minimal air exposure in the circuit, while five were exposed to a BAI simulating cardiotomy suction. There were no significant differences in bypass flow or hemodynamics between the groups. In the BAI group, there was an increase in hemolysis after bypass (plasma-free hemoglobin 5.27 ± 1.2 vs. 0.94 ± 0.8 mg/dl; p = 0.01), more aggressive platelet consumption (28% vs. 83% of baseline; p = 0.009), leukocyte consumption (71% vs. 107% of baseline; p = 0.02), and increased granulocyte CD11b expression (409% vs. 106% of baseline; p = 0.009). These data suggest the inflammatory pattern responsible for the CPB-SIRS phenomenon may be driven by blood-air interaction. Future efforts should focus on BAI-associated mechanisms for minimizing blood trauma and inflammation during CPB.

Genome-wide association analysis reveals QTL and candidate mutations involved in white spotting in cattle.

BACKGROUND: White spotting of the coat is a characteristic trait of various domestic species including cattle and other mammals. It is a hallmark of Holstein-Friesian cattle, and several previous studies have detected genetic loci with major effects for white spotting in animals with Holstein-Friesian ancestry. Here, our aim was to better understand the underlying genetic and molecular mechanisms of white spotting, by conducting the largest mapping study for this trait in cattle, to date. RESULTS: Using imputed whole-genome sequence data, we conducted a genome-wide association analysis in 2973 mixed-breed cows and bulls. Highly significant quantitative trait loci (QTL) were found on chromosomes 6 and 22, highlighting the well-established coat color genes KIT and MITF as likely responsible for these effects. These results are in broad agreement with previous studies, although we also report a third significant QTL on chromosome 2 that appears to be novel. This signal maps immediately adjacent to the PAX3 gene, which encodes a known transcription factor that controls MITF expression and is the causal locus for white spotting in horses. More detailed examination of these loci revealed a candidate causal mutation in PAX3 (p.Thr424Met), and another candidate mutation (rs209784468) within a conserved element in intron 2 of MITF transcripts expressed in the skin. These analyses also revealed a mechanistic ambiguity at the chromosome 6 locus, where highly dispersed association signals suggested multiple or multiallelic QTL involving KIT and/or other genes in this region. CONCLUSIONS: Our findings extend those of previous studies that reported KIT as a likely causal gene for white spotting, and report novel associations between candidate causal mutations in both the MITF and PAX3 genes. The sizes of the effects of these QTL are substantial, and could be used to select animals with darker, or conversely whiter, coats depending on the desired characteristics.

Fixed-Dose Four-Factor Prothrombin Complex Concentrate for Vitamin K Antagonist Reversal: Does One Dose Fit All?

Four-factor prothrombin complex concentrate (4F-PCC) has emerged as the preferred option for emergent reversal of vitamin K antagonists (VKAs); however, the optimal dosing strategy is unknown. Although several studies have attempted to determine the optimal dose of 4F-PCC using a variety of dosing regimens, no dosing strategy has been found to be superior. Many of these studies have evaluated a low, fixed dose of 4F-PCC rather than individualized dosing as recommended in product labeling. The purpose of this review was to evaluate the efficacy and safety of various fixed-dose strategies of 4F-PCC for emergent VKA reversal and to assess limitations of the existing literature. A search of the PubMed database was performed from the earliest available date through 2018 for relevant articles describing fixed-dose 4F-PCC for VKA reversal. Reference lists of relevant articles were also manually reviewed. Most currently available studies are primarily observational and heterogeneous in design. A very low fixed dose of 500 IU is likely inadequate for successful VKA reversal, but increased fixed doses of 1000-1500 IU have found some degree of success and may be considered for VKA reversal. However, many of these studies consistently identified a trend toward international normalized ratio (INR) reversal failure in patients presenting with high baseline INR values or intracranial hemorrhage, suggesting that higher 4F-PCC doses are needed in these patients. Available studies are underpowered to determine whether a dose-dependent association with thrombotic risk exists. Additional large, randomized studies are needed to determine the optimal dosing strategy and ascertain the role for fixed-dose 4F-PCC.

Multiple QTL underlie milk phenotypes at the CSF2RB locus.

BACKGROUND: Over many years, artificial selection has substantially improved milk production by cows. However, the genes that underlie milk production quantitative trait loci (QTL) remain relatively poorly characterised. Here, we investigate a previously reported QTL located at the CSF2RB locus on chromosome 5, for several milk production phenotypes, to better understand its underlying genetic and molecular causes. RESULTS: Using a population of 29,350 taurine dairy cows, we conducted association analyses for milk yield and composition traits, and identified highly significant QTL for milk yield, milk fat concentration, and milk protein concentration. Strikingly, protein concentration and milk yield appear to show co-located yet genetically distinct QTL. To attempt to understand the molecular mechanisms that might be mediating these effects, gene expression data were used to investigate eQTL for 11 genes in the broader interval. This analysis highlighted genetic impacts on CSF2RB and NCF4 expression that share similar association signatures to those observed for lactation QTL, strongly implicating one or both of these genes as responsible for these effects. Using the same gene expression dataset representing 357 lactating cows, we also identified 38 novel RNA editing sites in the 3' UTR of CSF2RB transcripts. The extent to which two of these sites were edited also appears to be genetically co-regulated with lactation QTL, highlighting a further layer of regulatory complexity that involves the CSF2RB gene. CONCLUSIONS: This locus presents a diversity of molecular and lactation QTL, likely representing multiple overlapping effects that, at a minimum, highlight the CSF2RB gene as having a causal role in these processes.

Spreading Ebola Panic: Newspaper and Social Media Coverage of the 2014 Ebola Health Crisis.

During times of hot crises, traditional news organizations have historically contributed to public fear and panic by emphasizing risks and uncertainties. The degree to which digital and social media platforms contribute to this panic is essential to consider in the new media landscape. This research examines news coverage of the 2014 Ebola crisis, exploring differences in presentation between newspaper coverage and news shared on the social news platform Reddit. Results suggest that news shared on Reddit amplified panic and uncertainty surrounding Ebola, while traditional newspaper coverage was significantly less likely to produce panic-inducing coverage.

Role of Bedside Preparation in Reducing "Door-to-Needle" Tissue-Type Plasminogen Activator (Alteplase) Administration Times and Association with Patient Outcomes.

INTRODUCTION: The purpose of this study is to assess the benefit of bedside alteplase preparation as a component of the acute stroke process. METHODS: A retrospective, single center study, designed to evaluate the impact of a bedside alteplase preparation protocol. Stroke patients receiving intravenous (IV) alteplase prepared at bedside were compared to pre-bedside alteplase preparation patients. The primary outcome was to compare door-to-needle (DTN) times between the groups. The secondary outcomes included comparison of pre-bedside alteplase preparation to post-bedside alteplase preparation on the following variables: imaging-to-drug times, order entry to drug administration times, percentage of patients achieving the 60 minute DTN goal, rate of intracranial hemorrhage (ICH), and patient discharge disposition. RESULTS: Patients in the pre-bedside preparation group included those who received IV alteplase between Jan. 1, 2012 and Jan. 31, 2015 and post-bedside preparation patients between Feb. 1, 2015 and March 31, 2016. Thirty-one patients were enrolled in the study, 16 in the pre-bedside preparation group and 15 in the post-beside preparation group. The mean DTN time in the post-bedside alteplase preparation group was significantly reduced, as compared to the pre-bedside preparation group (66.6 minutes vs. 95.9 minutes, p=0.024). Percent of patients meeting the 60 minute DTN time goal was significantly improved when alteplase was prepared at bedside (53.3 percent vs. 18.8 percent) (p=0.044). Rates of ICH were not significantly different between the two populations. CONCLUSIONS: Bedside alteplase preparation significantly reduced DTN times in an academic hospital emergency department.

Functional Status Change Among Children With Extracorporeal Membrane Oxygenation to Support Cardiopulmonary Resuscitation in a Pediatric Cardiac ICU: A Single Institution Report.

OBJECTIVES: The purpose of this study is to describe the functional status of survivors from extracorporeal cardiopulmonary resuscitation instituted during in-hospital cardiac arrest using the Functional Status Scale. We aimed to determine risk factors leading to the development of new morbidity and unfavorable functional outcomes. DESIGN: This was a single-center retrospective chart review abstracting patient characteristics/demographic data, duration of cardiopulmonary resuscitation, duration of extracorporeal membrane oxygenation support, as well as maximum lactate levels within 2 hours before and after extracorporeal cardiopulmonary resuscitation. Cardiac arrest was defined as the administration of chest compressions for a nonperfusing cardiac rhythm. Extracorporeal cardiopulmonary resuscitation was defined by instituting extracorporeal membrane oxygenation during active chest compressions. Functional Status Scale scores were calculated at admission and on hospital discharge for patients who survived. SETTING: Patients admitted in the pediatric cardiac ICU at C.S. Mott Children's Hospital from January 1, 2005, to December 31, 2015. PATIENTS: Children less than 18 years who underwent extracorporeal cardiopulmonary resuscitation. INTERVENTIONS: Not applicable. MEASUREMENTS AND MAIN RESULTS: Of 608 extracorporeal membrane oxygenation events during the study period, 80 were extracorporeal cardiopulmonary resuscitation (14%). There were 40 female patients (50%). Median age was 40 days (interquartile range, 9-342 d). Survival to hospital discharge was 48% (38/80). Median Functional Status Scale score at admission was 6 (interquartile range, 6-6) and at hospital discharge 9 (interquartile range, 8-11). Out of 38 survivors, 19 (50%) had a change of Functional Status Scale score greater than or equal to 3, that is consistent with new morbidity, and 26 (68%) had favorable functional outcomes with a change in Functional Status Scale score of less than 5. CONCLUSIONS: This is the first extracorporeal cardiopulmonary resuscitation report to examine changes in Functional Status Scale from admission (baseline) to discharge as a measure of overall functional outcome. Half of surviving patients (19/38) had new morbidity, while 68% (26/38) had favorable outcomes. Lactate levels, duration of cardiopulmonary resuscitation, and duration of extracorporeal membrane oxygenation were not found to be risk factors for the development of new morbidity and poor functional outcomes. Functional Status Scale may be used as a metric to monitor improvement of extracorporeal cardiopulmonary resuscitation outcomes and help guide research initiatives to decrease morbidity in this patient population.

Development and innovation of system resources to optimize patient care.

PURPOSE: Various incremental and disruptive healthcare innovations that are occurring or may occur are discussed, with insights on how multihospital health systems can prepare for the future and optimize the continuity of patient care provided. SUMMARY: Innovation in patient care is occurring at an ever-increasing rate, and this is especially true relative to the transition of patients through the care continuum. Health systems must leverage their ability to standardize and develop electronic health record (EHR) systems and other infrastructure necessary to support patient care and optimize outcomes; examples include 3D printing of patient-specific medication dosage forms to enhance precision medicine, the use of drones for medication delivery, and the expansion of telehealth capabilities to improve patient access to the services of pharmacists and other healthcare team members. Disruptive innovations in pharmacy services and delivery will alter how medications are prescribed and delivered to patients now and in the future. Further, technology may also fundamentally alter how and where pharmacists and pharmacy technicians care for patients. This article explores the various innovations that are occurring and that will likely occur in the future, particularly as they apply to multihospital health systems and patient continuity of care. CONCLUSION: Pharmacy departments that anticipate and are prepared to adapt to incremental and disruptive innovations can demonstrate value in the multihospital health system through strategies such as optimizing the EHR, identifying telehealth opportunities, supporting infrastructure, and integrating services.

Mechanical Circulatory Support for the Failing Fontan: Conversion to Assisted Single Ventricle Circulation-Preliminary Observations.

BACKGROUND: Mechanical circulatory support (MCS) of a failing Fontan circulation remains challenging. We hypothesized that MCS can be provided by converting the Fontan circulation into a mechanically assisted single ventricle parallel circulation (MASVC). METHODS: A porcine model of functionally univentricular circulation was created under cardiopulmonary bypass (CPB) by performing an atrial septectomy, tricuspid valvectomy, and interrupting antegrade pulmonary blood flow. A centrifugal flow pump was placed with inflow from the common atrium. Eight millimeter Dacron grafts anastomosed to the ascending aorta and main pulmonary artery supplied systemic (Qs) and pulmonary (Qp) blood flow. Ultrasonic flow probes were used to measure Qs and Qp after weaning from CPB. The Qp/Qs ratio was regulated using an adjustable clamp. Hemodynamic and laboratory data were recorded. RESULTS: All four animals were successfully weaned from CPB onto the MASVC for a duration of two hours. Mechanically assisted single ventricle parallel circulation achieved satisfactory hemodynamics. As anticipated, the arterial oxygen saturation and partial pressure of oxygen in arterial blood were lower in the MASVC compared to baseline biventricular circulation. At the conclusion of the study, there was a trend towards a decrease in the mixed venous saturation with increasing oxygen extraction compared to the baseline. Serum lactate levels increased after weaning from CPB and did not return to baseline after two hours of support. CONCLUSION: Mechanically assisted single ventricle parallel circulation can be established in a single ventricle animal model. This strategy could potentially provide MCS of a single ventricle circulation. Studies with longer duration of support are required to assess adequacy of support and long-term sustainability.

Road Map for Research Training in the Residency Learning Experience.

The American Society of Health-System Pharmacists residency accreditation standards require all postgraduate residency training programs to teach and evaluate a resident's ability to advance practice through project development and presentation, underscoring the importance of conducting research in today's professional climate. Although many residents express strong interest in research participation and contributing to the medical literature, many obstacles to publication have been identified. We aim to illustrate a deliberate approach to teaching this material and structuring the longitudinal experience in a way that maximizes resources to overcome these barriers. Such efforts should aid residents, advisors, and program directors in establishing curriculum which leads to successful completion and publication of pharmacy resident's research projects.

DNA and RNA-sequence based GWAS highlights membrane-transport genes as key modulators of milk lactose content.

BACKGROUND: Lactose provides an easily-digested energy source for neonates, and is the primary carbohydrate in milk in most species. Bovine lactose is also a key component of many human food products. However, compared to analyses of other milk components, the genetic control of lactose has been little studied. Here we present the first GWAS focussed on analysis of milk lactose traits. RESULTS: Using a discovery population of 12,000 taurine dairy cattle, we detail 27 QTL for lactose concentration and yield, and subsequently validate the effects of 26 of these loci in a distinct population of 18,000 cows. We next present data implicating causative genes and variants for these QTL. Fine mapping of these regions using imputed, whole genome sequence-resolution genotypes reveals protein-coding candidate causative variants affecting the ABCG2, DGAT1, STAT5B, KCNH4, NPFFR2 and RNF214 genes. Eleven of the remaining QTL appear to be driven by regulatory effects, suggested by the presence of co-locating, co-segregating eQTL discovered using mammary RNA sequence data from a population of 357 lactating cows. Pathway analysis of genes representing all lactose-associated loci shows significant enrichment of genes located in the endoplasmic reticulum, with functions related to ion channel activity mediated through the LRRC8C, P2RX4, KCNJ2 and ANKH genes. A number of the validated QTL are also found to be associated with additional milk volume, fat and protein phenotypes. CONCLUSIONS: Overall, these findings highlight novel candidate genes and variants involved in milk lactose regulation, whose impacts on membrane transport mechanisms reinforce the key osmo-regulatory roles of lactose in milk.

Hyperlipidemia sink for anesthetic agents.

We present a case that involves anesthetic resistance during anesthesia for electroconvulsive therapy. Despite adequate dosing of both intravenous and inhalation anesthetics, our patient was resistant to induction of the state of general anesthesia. Subsequently, we noticed extreme hyperlipidemia. We hypothesized that the patient's extreme hyperlipidemia served as an anesthetic "sink" and prevented the full dose of intravenous agents from quickly reaching their intended site of action.