Role of the CTCF Binding Site in Human T-Cell Leukemia Virus-1 Pathogenesis.

During HTLV-1 infection, the virus integrates into the host cell genome as a provirus with a single CCCTC binding protein (CTCF) binding site (vCTCF-BS), which acts as an insulator between transcriptionally active and inactive regions. Previous studies have shown that the vCTCF-BS is important for maintenance of chromatin structure, regulation of viral expression, and DNA and histone methylation. Here, we show that the vCTCF-BS also regulates viral infection and pathogenesis in vivo in a humanized (Hu) mouse model of adult T-cell leukemia/lymphoma. Three cell lines were used to initiate infection of the Hu-mice, i) HTLV-1-WT which carries an intact HTLV-1 provirus genome, ii) HTLV-1-CTCF, which contains a provirus with a mutated vCTCF-BS which abolishes CTCF binding, and a stop codon immediate upstream of the mutated vCTCF-BS which deletes the last 23 amino acids of p12, and iii) HTLV-1-p12stop that contains the intact vCTCF-BS, but retains the same stop codon in p12 as in the HTLV-1-CTCF cell line. Hu-mice were infected with mitomycin treated or irradiated HTLV-1 producing cell lines. There was a delay in pathogenicity when Hu-mice were infected with the CTCF virus compared to mice infected with either p12 stop or WT virus. Proviral load (PVL), spleen weights, and CD4 T cell counts were significantly lower in HTLV-1-CTCF infected mice compared to HTLV-1-p12stop infected mice. Furthermore, we found a direct correlation between the PVL in peripheral blood and death of HTLV-1-CTCF infected mice. In cell lines, we found that the vCTCF-BS regulates Tax expression in a time-dependent manner. The scRNAseq analysis of splenocytes from infected mice suggests that the vCTCF-BS plays an important role in activation and expansion of T lymphocytes in vivo . Overall, these findings indicate that the vCTCF-BS regulates Tax expression, proviral load, and HTLV pathogenicity in vivo . Author Summary: Human T-cell leukemia virus type 1 (HTLV-1) is a cause of leukemia and lymphoma, and several inflammatory medical disorders. The virus integrates in the host cell DNA, and it includes a single binding site for a cellular protein designated CTCF. This protein is important in regulation of many viruses, as well as properties of normal and malignant cells. In order to define the role of CTCF in HTLV-1 pathogenesis in vivo , we analyzed a mutant virus lacking the binding site in humanized mice. We found that this mutation slowed virus spread and attenuated the development of disease. Gene expression studies demonstrated a dynamic role of CTCF in regulating viral gene expression and T lymphocyte activation.

A DIO2 missense mutation and its impact on fetal response to PRRSV infection.

BACKGROUND: Porcine reproductive and respiratory syndrome virus 2 (PRRSV-2) infection during late gestation substantially lowers fetal viability and survival. In a previous genome-wide association study, a single nucleotide polymorphism on chromosome 7 was significantly associated with probability of fetuses being viable in response to maternal PRRSV-2 infection at 21 days post maternal inoculation. The iodothyronine deiodinase 2 (DIO2) gene, located ~ 14 Kilobase downstream of this SNP, was selected as a priority candidate related to fetal susceptibility following maternal PRRSV-2 infection. Our objectives were to identify mutation(s) within the porcine DIO2 gene and to determine if they were associated with fetal outcomes after PRRSV-2 challenge. Sequencing of the DIO2, genotyping identified variants, and association of DIO2 genotypes with fetal phenotypes including DIO2 mRNA levels, viability, survival, viral loads, cortisol and thyroid hormone levels, and growth measurements were conducted. RESULTS: A missense variant (p.Asn91Ser) was identified in the parental populations from two independent PRRSV-2 challenge trials. This variant was further genotyped to determine association with fetal PRRS outcomes. DIO2 mRNA levels in fetal heart and kidney differed by the genotypes of Asn91Ser substitution with significantly greater DIO2 mRNA expression in heterozygotes compared with wild-type homozygotes (P < 0.001 for heart, P = 0.002 for kidney). While Asn91Ser did not significantly alter fetal viability and growth measurements, interaction effects of the variant with fetal sex or trial were identified for fetal viability or crown rump length, respectively. However, this mutation was not related to dysregulation of the hypothalamic-pituitary-adrenal and thyroid axis, indicated by no differences in circulating cortisol, T4, and T3 levels in fetuses of the opposing genotypes following PRRSV-2 infection. CONCLUSIONS: The present study suggests that a complex relationship among DIO2 genotype, DIO2 expression, fetal sex, and fetal viability may exist during the course of fetal PRRSV infection. Our study also proposes the increase in cortisol levels, indicative of fetal stress response, may lead to fetal complications, such as fetal compromise, fetal death, or premature farrowing, during PRRSV infection.

Autonomic Effects of Pulsed Field vs Thermal Ablation for Treating Atrial Fibrillation: Sub-analysis of ADVENT.

BACKGROUND: Autonomic denervation is an ancillary phenomenon during thermal ablation of atrial fibrillation (AF), that may have synergistic effects on symptomatic improvement and long-term freedom from AF. Pulsed field ablation (PFA), a non-thermal ablation modality, was non-inferior to thermal ablation in treating AF; however, PFA's relative myocardial selectivity may minimize autonomic effects. OBJECTIVES: Compare heart rate (HR) and heart rate variability (HRV) metrics as markers of autonomic function after ablation using PFA versus thermal ablation. METHODS: ADVENT was a randomized pivotal study comparing PFA (pentaspline catheter) with thermal ablation (radiofrequency [RF] or cryoballoon [CB]) for treating paroxysmal AF. Baseline HR was acquired from a pre-ablation 12-lead ECG, while follow-up HRs, as well as HRV (SDNN, SDANN) metrics, were derived from 72-hour Holter monitors at 6 and 12-months. RESULTS: This study included 379 paroxysmal AF patients undergoing PFA (n=194) or thermal ablation (n=185; n=102 RF, n=83 CB) completing 6 and 12-month Holter monitors. Compared to PFA, thermal patients had significantly greater increases in HR from baseline to 6 months (ΔHR; 10.1 vs. 5.9bpm; p=0.02) and 12 months (ΔHR; 8.8 vs. 5.2bpm; p=0.03). This increase in HR at 6 and 12 months was similar between CB and RF (p=0.94 and 0.83, respectively). HRV, both SDNN and SDANN, were significantly lower at both 6- and 12-month after thermal ablation compared to PFA (p<0.01). CONCLUSIONS: PFA's effect on the autonomic nervous system was attenuated compared to thermal ablation. Whether this affects long-term freedom from AF or symptomatic bradycardia/pauses after AF ablation requires further study.

Pulmonary vein narrowing after pulsed field versus thermal ablation.

AIMS: When it occurs, pulmonary vein (PV) stenosis after atrial fibrillation (AF) ablation is associated with significant morbidity. Even mild-to-moderate PV narrowing may have long-term implications. Unlike thermal ablation energies, such as radiofrequency (RF) or cryothermy, pulsed field ablation (PFA) is a non-thermal modality associated with less fibrotic proliferation. Herein, we compared the effects of PFA vs. thermal ablation on PV narrowing after AF ablation. METHODS AND RESULTS: ADVENT was a multi-centre, randomized, single-blind study comparing PFA (pentaspline catheter) with thermal ablation-force-sensing RF or cryoballoon (CB)-to treat drug-refractory paroxysmal AF. Pulmonary vein diameter and aggregate cross-sectional area were obtained by baseline and 3-month imaging. The pre-specified, formally tested, secondary safety endpoint compared a measure of PV narrowing between PFA vs. thermal groups, with superiority defined by posterior probability > 0.975. Among subjects randomized to PFA (n = 305) or thermal ablation (n = 302), 259 PFA and 255 thermal ablation (137 RF and 118 CB) subjects had complete baseline and 3-month PV imaging. No subject had significant (≥70%) PV stenosis. Change in aggregate PV cross-sectional area was less with PFA (-0.9%) than thermal ablation (-12%, posterior probability > 0.999)-primarily driven by the RF sub-cohort (-19.5%) vs. CB sub-cohort (-3.3%). Almost half of all PFA PV diameters did not decrease, but the majority (80%) of RF PVs decreased, regardless of PV anatomic location. CONCLUSION: In this first randomized comparison of PFA vs. thermal ablation, PFA resulted in less PV narrowing-thereby underscoring the qualitatively differential and favourable impact of PFA on PV tissue.

Genetic analysis of the blood transcriptome of young healthy pigs to improve disease resilience.

BACKGROUND: Disease resilience is the ability of an animal to maintain productive performance under disease conditions and is an important selection target. In pig breeding programs, disease resilience must be evaluated on selection candidates without exposing them to disease. To identify potential genetic indicators for disease resilience that can be measured on selection candidates, we focused on the blood transcriptome of 1594 young healthy pigs with subsequent records on disease resilience. Transcriptome data were obtained by 3'mRNA sequencing and genotype data were from a 650 K genotyping array. RESULTS: Heritabilities of the expression of 16,545 genes were estimated, of which 5665 genes showed significant estimates of heritability (p < 0.05), ranging from 0.05 to 0.90, with or without accounting for white blood cell composition. Genes with heritable expression levels were spread across chromosomes, but were enriched in the swine leukocyte antigen region (average estimate > 0.2). The correlation of heritability estimates with the corresponding estimates obtained for genes expressed in human blood was weak but a sizable number of genes with heritable expression levels overlapped. Genes with heritable expression levels were significantly enriched for biological processes such as cell activation, immune system process, stress response, and leukocyte activation, and were involved in various disease annotations such as RNA virus infection, including SARS-Cov2, as well as liver disease, and inflammation. To estimate genetic correlations with disease resilience, 3205 genotyped pigs, including the 1594 pigs with transcriptome data, were evaluated for disease resilience following their exposure to a natural polymicrobial disease challenge. Significant genetic correlations (p < 0.05) were observed with all resilience phenotypes, although few exceeded expected false discovery rates. Enrichment analysis of genes ranked by estimates of genetic correlations with resilience phenotypes revealed significance for biological processes such as regulation of cytokines, including interleukins and interferons, and chaperone mediated protein folding. CONCLUSIONS: These results suggest that expression levels in the blood of young healthy pigs for genes in biological pathways related to immunity and endoplasmic reticulum stress have potential to be used as genetic indicator traits to select for disease resilience.

Characterization of the bacterial fecal microbiota composition of pigs preceding the clinical signs of swine dysentery.

Swine dysentery (SD) is a worldwide production-limiting disease of growing-finishing pigs in commercial farms. The importance of the large intestinal microbiota in the swine dysentery pathogenesis has been established, but not well characterized. The objective of this study was to characterize the fecal bacterial microbiota of pigs immediately prior to developing clinical signs of swine dysentery. A total of 60 fecal samples were collected from 15 pigs with SD. Sampling times included a time point prior to SD (d0, n=15), 2 days before mucohaemorrhagic diarrhea was observed (d-2SD, n=15), 1 day before mucohaemorrhagic diarrhea was observed (d-1SD, n=15), and the day when pigs developed mucohemorragic diarrhea (MHD, n=15). Sequencing of cpn60 amplicons was used to profile the microbiome, and analyses were performed on QIIME2. Increased Chao1 index in d-1SD and MHD samples when compared to the d0 was the only change observed in alpha diversity. No differences between sampling times on beta diversity (Bray-Curtis dissimilarity) were found. Although a small sample size was investigated, differential abundance analysis revealed that Alistipes dispar and Parabacteroides gordonii were increased in MHD fecal samples when compared to d-2SD and d-1SD. It is suggested that these taxa may play a role in the pathogenesis of SD, which is known to require the presence of Brachyspira spp. and an anaerobe for severe disease development.

Pulsed Field or Conventional Thermal Ablation for Paroxysmal Atrial Fibrillation.

BACKGROUND: Catheter-based pulmonary vein isolation is an effective treatment for paroxysmal atrial fibrillation. Pulsed field ablation, which delivers microsecond high-voltage electrical fields, may limit damage to tissues outside the myocardium. The efficacy and safety of pulsed field ablation as compared with conventional thermal ablation are not known. METHODS: In this randomized, single-blind, noninferiority trial, we assigned patients with drug-refractory paroxysmal atrial fibrillation in a 1:1 ratio to undergo pulsed field ablation or conventional radiofrequency or cryoballoon ablation. The primary efficacy end point was freedom from a composite of initial procedural failure, documented atrial tachyarrhythmia after a 3-month blanking period, antiarrhythmic drug use, cardioversion, or repeat ablation. The primary safety end point included acute and chronic device- and procedure-related serious adverse events. RESULTS: A total of 305 patients were assigned to undergo pulsed field ablation, and 302 were assigned to undergo thermal ablation. At 1 year, the primary efficacy end point was met (i.e., no events occurred) in 204 patients (estimated probability, 73.3%) who underwent pulsed field ablation and 194 patients (estimated probability, 71.3%) who underwent thermal ablation (between-group difference, 2.0 percentage points; 95% Bayesian credible interval, -5.2 to 9.2; posterior probability of noninferiority, >0.999). Primary safety end-point events occurred in 6 patients (estimated incidence, 2.1%) who underwent pulsed field ablation and 4 patients (estimated incidence, 1.5%) who underwent thermal ablation (between-group difference, 0.6 percentage points; 95% Bayesian credible interval, -1.5 to 2.8; posterior probability of noninferiority, >0.999). CONCLUSIONS: Among patients with paroxysmal atrial fibrillation receiving a catheter-based therapy, pulsed field ablation was noninferior to conventional thermal ablation with respect to freedom from a composite of initial procedural failure, documented atrial tachyarrhythmia after a 3-month blanking period, antiarrhythmic drug use, cardioversion, or repeat ablation and with respect to device- and procedure-related serious adverse events at 1 year. (Funded by Farapulse-Boston Scientific; ADVENT ClinicalTrials.gov number, NCT04612244.).

The WUR0000125 PRRS resilience SNP had no apparent effect on pigs' infectivity and susceptibility in a novel transmission trial.

BACKGROUND: Porcine reproductive and respiratory syndrome (PRRS) remains one of the most important infectious diseases for the pig industry. A novel small-scale transmission experiment was designed to assess whether the WUR0000125 (WUR for Wageningen University and Research) PRRS resilience single nucleotide polymorphism (SNP) confers lower susceptibility and infectivity to pigs under natural porcine reproductive and respiratory syndrome virus (PRRSV-2) transmission. METHODS: Commercial full- and half-sib piglets (n = 164) were assigned as either Inoculation, Shedder, or Contact pigs. Pigs were grouped according to their relatedness structure and WUR genotype, with R- and R+ referring to pigs with zero and one copy of the dominant WUR resilience allele, respectively. Barcoding of the PRRSV-2 strain (SD09-200) was applied to track pig genotype-specific transmission. Blood and nasal swab samples were collected and concentrations of PRRSV-2 were determined by quantitative (q)-PCR and cell culture and expressed in units of median tissue culture infectious dose (TCID50). The Log10TCID50 at each sampling event, derived infection status, and area under the curve (AUC) were response variables in linear and generalized linear mixed models to infer WUR genotype differences in Contact pig susceptibility and Shedder pig infectivity. RESULTS: All Shedder and Contact pigs, except one, became infected through natural transmission. There was no significant (p > 0.05) effect of Contact pig genotype on any virus measures that would indicate WUR genotype differences in susceptibility. Contact pigs tended to have higher serum AUC (p = 0.017) and log10TCID50 (p = 0.034) when infected by an R+ shedder, potentially due to more infectious R+ shedders at the early stages of the transmission trial. However, no significant Shedder genotype effect was found in serum (p = 0.274) or nasal secretion (p = 0.951) that would indicate genotype differences in infectivity. CONCLUSIONS: The novel design demonstrated that it is possible to estimate genotype effects on Shedder pig infectivity and Contact pig susceptibility that are not confounded by family effects. The study, however, provided no supportive evidence that genetic selection on WUR genotype would affect PRRSV-2 transmission. The results of this study need to be independently validated in a larger trial using different PRRSV strains before dismissing the effects of the WUR marker or the previously detected GBP5 gene on PRRSV transmission.

Antimicrobial susceptibility of western Canadian Brachyspira isolates: Development and standardization of an agar dilution susceptibility test method.

The re-emergence of Brachyspira-associated disease in pigs since the late 2000s has illuminated some of the diagnostic challenges associated with this genus; notably, the lack of standardized antimicrobial susceptibility testing (AST) methods and interpretive criteria. Consequently, laboratories have relied heavily on highly variable in-house developed methods. There are currently no published investigations describing the antimicrobial susceptibility of Brachyspira isolates collected from pigs in Canada. The first objective of this study was therefore to develop a standardized protocol for conducting agar dilution susceptibility testing of Brachyspira spp., including determining the optimal standardized inoculum density, a key test variable that impacts test performance. The second objective was to determine the susceptibility of a collection of western Canadian Brachyspira isolates using the standardized methodology. After assessing multiple media, an agar dilution test was standardized in terms of starting inoculum (1-2 × 108 CFU/ml), incubation temperature and time, and assessed for repeatability. The antimicrobial susceptibility of a collection of clinical porcine Brachyspira isolates (n = 87) collected between 2009-2016 was then determined. This method was highly reproducible; repeat susceptibility testing yielded identical results 92% of the time. Although most of the isolates had very low MICs to the commonly used antimicrobials to treat Brachyspira-associated infections, several isolates with elevated MICs (>32 µg/ml) for tiamulin, valnemulin, tylosin, tylvalosin, and lincomycin were identified. Overall, this study underscores the importance of establishing CLSI approved clinical breakpoints for Brachyspira to facilitate the interpretation of test results and support the evidence-based selection of antimicrobials in swine industry.

Improving the consistency of experimental swine dysentery inoculation strategies.

Swine dysentery (SD) caused by pathogenic Brachyspira spp. is an economic challenge for the swine industry. In research settings, experimental reproduction of swine dysentery typically relies on intragastric inoculation which has shown variable success. This project aimed to improve the consistency of the experimental inoculation protocol used for swine dysentery in our laboratory. Over six experiments, we evaluated the influence of group housing in inoculated pigs using a frozen-thawed broth culture of strongly hemolytic B. hyodysenteriae strain D19 (Trial A), compared the relative virulence of B. hyodysenteriae strains D19 and G44 (Trial B), compared inoculum volumes (50 mL vs 100 mL) for G44 and B. hampsonii 30446 (Trial C), and performed three independent trials evaluating intragastric inoculation using different oral inoculation methods: oral feed balls (Trial D), and oral syringe bolus of 100 mL (Trial E) or 300 mL (Trial F). Intragastric inoculation with a fresh broth culture of B. hyodysenteriae strain G44 resulted in a shorter incubation period and a higher proportionate duration of mucohemorrhagic diarrhea (MMHD) compared to D19. Intragastric inoculation with either 50 or 100 mL of B. hampsonii 30446 or B. hyodysenteriae (G44) were statistically equivalent. Oral inoculation with 100 mL or 300 mL also yielded similar results to intragastric inoculation but was more expensive due to the additional work and supplies associated with syringe training. Our future research will use intragastric inoculation with 100 mL of a fresh broth culture containing B. hyodysenteriae strain G44 as it yields a high incidence of mucohaemorrhagic diarrhea with a reasonable cost.

Electron transparent nanotubes reveal crystallization pathways in confinement.

The cylindrical pores of track-etched membranes offer excellent environments for studying the effects of confinement on crystallization as the pore diameter is readily varied and the anisotropic morphologies can direct crystal orientation. However, the inability to image individual crystals in situ within the pores in this system has prevented many of the underlying mechanisms from being characterized. Here, we study the crystallization of calcium sulfate within track-etched membranes and reveal that oriented gypsum forms in 200 nm diameter pores, bassanite in 25-100 nm pores and anhydrite in 10 nm pores. The crystallization pathways are then studied by coating the membranes with an amorphous titania layer prior to mineralization to create electron transparent nanotubes that protect fragile precursor materials. By visualizing the evolutionary pathways of the crystals within the pores we show that the product single crystals derive from multiple nucleation events and that orientation is determined at early reaction times. Finally, the transformation of bassanite to gypsum within the membrane pores is studied using experiment and potential mean force calculations and is shown to proceed by localized dissolution/reprecipitation. This work provides insight into the effects of confinement on crystallization processes, which is relevant to mineral formation in many real-world environments.

Dynamic tilting in perovskites.

A new computational analysis of tilt behaviour in perovskites is presented. This includes the development of a computational program - PALAMEDES - to extract tilt angles and the tilt phase from molecular dynamics simulations. The results are used to generate simulated selected-area electron and neutron diffraction patterns which are compared with experimental patterns for CaTiO3. The simulations not only reproduced all symmetrically allowed superlattice reflections associated with tilt but also showed local correlations that give rise to symmetrically forbidden reflections and the kinematic origin of diffuse scattering.

Genome wide association study of thyroid hormone levels following challenge with porcine reproductive and respiratory syndrome virus.

Introduction: Porcine reproductive and respiratory syndrome virus (PRRSV) causes respiratory disease in piglets and reproductive disease in sows. Piglet and fetal serum thyroid hormone (i.e., T3 and T4) levels decrease rapidly in response to Porcine reproductive and respiratory syndrome virus infection. However, the genetic control of T3 and T4 levels during infection is not completely understood. Our objective was to estimate genetic parameters and identify quantitative trait loci (QTL) for absolute T3 and/or T4 levels of piglets and fetuses challenged with Porcine reproductive and respiratory syndrome virus. Methods: Sera from 5-week-old pigs (N = 1792) at 11 days post inoculation (DPI) with Porcine reproductive and respiratory syndrome virus were assayed for T3 levels (piglet_T3). Sera from fetuses (N = 1,267) at 12 or 21 days post maternal inoculation (DPMI) with Porcine reproductive and respiratory syndrome virus of sows (N = 145) in late gestation were assayed for T3 (fetal_T3) and T4 (fetal_T4) levels. Animals were genotyped using 60 K Illumina or 650 K Affymetrix single nucleotide polymorphism (SNP) panels. Heritabilities, phenotypic correlations, and genetic correlations were estimated using ASREML; genome wide association studies were performed for each trait separately using Julia for Whole-genome Analysis Software (JWAS). Results: All three traits were low to moderately heritable (10%-16%). Phenotypic and genetic correlations of piglet_T3 levels with weight gain (0-42 DPI) were 0.26 ± 0.03 and 0.67 ± 0.14, respectively. Nine significant quantitative trait loci were identified for piglet_T3, on Sus scrofa chromosomes (SSC) 3, 4, 5, 6, 7, 14, 15, and 17, and collectively explaining 30% of the genetic variation (GV), with the largest quantitative trait loci identified on SSC5, explaining 15% of the genetic variation. Three significant quantitative trait loci were identified for fetal_T3 on SSC1 and SSC4, which collectively explained 10% of the genetic variation. Five significant quantitative trait loci were identified for fetal_T4 on SSC1, 6, 10, 13, and 15, which collectively explained 14% of the genetic variation. Several putative immune-related candidate genes were identified, including CD247, IRF8, and MAPK8. Discussion: Thyroid hormone levels following Porcine reproductive and respiratory syndrome virus infection were heritable and had positive genetic correlations with growth rate. Multiple quantitative trait loci with moderate effects were identified for T3 and T4 levels during challenge with Porcine reproductive and respiratory syndrome virus and candidate genes were identified, including several immune-related genes. These results advance our understanding of growth effects of both piglet and fetal response to Porcine reproductive and respiratory syndrome virus infection, revealing factors associated with genomic control of host resilience.

Plasma protein levels of young healthy pigs as indicators of disease resilience.

Selection for disease resilience, which refers to the ability of an animal to maintain performance when exposed to disease, can reduce the impact of infectious diseases. However, direct selection for disease resilience is challenging because nucleus herds must maintain a high health status. A possible solution is indirect selection of indicators of disease resilience. To search for such indicators, we conducted phenotypic and genetic quantitative analyses of the abundances of 377 proteins in plasma samples from 912 young and visually healthy pigs and their relationships with performance and subsequent disease resilience after natural exposure to a polymicrobial disease challenge. Abundances of 100 proteins were significantly heritable (false discovery rate (FDR) <0.10). The abundance of some proteins was or tended to be genetically correlated (rg) with disease resilience, including complement system proteins (rg = -0.24, FDR = 0.001) and IgG heavy chain proteins (rg = -0.68, FDR = 0.22). Gene set enrichment analyses (FDR < 0.2) based on phenotypic and genetic associations of protein abundances with subsequent disease resilience revealed many pathways related to the immune system that were unfavorably associated with subsequent disease resilience, especially the innate immune system. It was not possible to determine whether the observed levels of these proteins reflected baseline levels in these young and visually healthy pigs or were the result of a response to environmental disturbances that the pigs were exposed to before sample collection. Nevertheless, results show that, under these conditions, the abundance of proteins in some immune-related pathways can be used as phenotypic and genetic predictors of disease resilience and have the potential for use in pig breeding and management.

A challenge of selection for disease resilience is that it is difficult to directly select pigs that have greater resilience to multiple diseases in the healthy nucleus herd environment which is essential for breeding programs. A possible alternative is to select an indicator trait or marker that can be measured in a healthy setting, is heritable, and is associated with the genetics of disease resilience. In this study, we investigated plasma protein levels measured on young healthy pigs as indicator traits to select for disease resilience. For this purpose, we used plasma proteome data collected prior to the natural exposure of nursery pigs to multiple diseases, performed phenotypic and genetic quantitative analyses, and investigated their relationships with disease resilience. Our results suggest that plasma protein levels of young healthy pigs have the potential as biomarkers to select for disease resistance.

Randomized prospective evaluation of same-day discharge after cryoballoon ablation of atrial fibrillation: results of the EASY PVI study.

BACKGROUND: With advancements in technology and ablation techniques, catheter ablation for the treatment of atrial fibrillation (AF) has become safer with time. In the past, standard-of-care recommended overnight stay for outpatient procedures. As safety has improved and procedure times have shortened, some centers have allowed for same-day discharge. We report the results of a multi-center, randomized clinical trial investigating the safety of same-day discharge post-cryoballoon ablation. METHODS: Patients with paroxysmal atrial fibrillation underwent pulmonary vein isolation (PVI) with the Medtronic Arctic Advance cryoballoon at 3 US centers. Six hours after the procedure, patients were randomized to either stay overnight or be discharged same day. RESULTS: A total of 49 patients were enrolled. Two patients were withdrawn prior to randomization. One patient chose to withdraw after randomization. Of the 22 patients randomized to same-day discharge and the 23 patients randomized the overnight stay, no significant adverse outcomes were reported in either group. Occurrence of adverse events did not differ significantly between the two groups. Procedure time and fluoroscopy time did not significantly differ between groups. CONCLUSIONS: This is the first randomized trial examining the safety of same-day discharge post-cryoballoon ablation. Based on our results, same-day discharge following cryoballoon ablation for paroxysmal AF is a safe option following uncomplicated ablation for PVI. Operators should use their discretion in selecting patients for same-day discharge.

Exploration of plasma metabolite levels in healthy nursery pigs in response to environmental enrichment and disease resilience.

The purpose of this study was to explore plasma metabolite levels in young healthy pigs and their potential association with disease resilience and estimate genetic and phenotypic correlation with the change in lymphocyte concentration following disease challenge. Plasma samples were collected from 968 healthy nursery pigs over 15 batches at an average of 28 ± 3.23 d of age. Forty-four metabolites were identified and quantified by nuclear magnetic resonance. Pigs were then introduced into a natural disease challenge barn, and were classified into four groups based on the growth rate of each animal in the grow-to-finish phase (GFGR) and treatment rate (TR): resilient (RES), average (MID), susceptible (SUS), and dead (pigs that died before harvest). Blood samples were collected from all pigs before and 2 wk after disease challenge and complete blood count was determined. Environmental enrichment (inedible point source objects) was provided for half of the pigs in seven batches (N = 205) to evaluate its impact on resilience and metabolite concentrations. Concentration of all metabolites was affected by batch, while entry age affected the concentration of 16 metabolites. The concentration of creatinine was significantly lower for pigs classified as "dead" and "susceptible" when compared to "average" (P < 0.05). Pigs that received enrichment had significantly lower concentrations of six metabolites compared with pigs that did not receive enrichment (P ≤ 0.05). Both, group classification and enrichment affected metabolites that are involved in the same pathways of valine, leucine, and isoleucine biosynthesis and degradation. Resilient pigs had higher increase in lymphocyte concentration after disease challenge. The concentration of plasma l-α-aminobutyric acid was significantly negatively genetically correlated with the change in lymphocyte concentration following challenge. In conclusion, creatinine concentration in healthy nursery pigs was lower in pigs classified as susceptible or dead after disease challenge, whilst l-α-aminobutyric may be a genetic biomarker of lymphocyte response after pathogen exposure, and both deserve further investigation. Batch, entry age, and environmental enrichment were important factors affecting the concentration of metabolites and should be taken into consideration in future studies.

The focus of this study was to explore plasma metabolite levels in young healthy pigs and their potential association with health outcome classification following the exposure to a polymicrobial disease challenge. In addition, we explored the effect of the environmental enrichment on metabolite concentrations. Finally, we estimated genetic and phenotypic correlations between metabolites and the magnitude of change in lymphocytes levels following exposure to a polymicrobial disease challenge. We found that concentration of creatinine was lower in pigs that died before marketing, classified as "dead" and susceptible when compared to average group. This indicates that creatinine can be used as an early indicator of death and/or susceptibility of disease in pigs. Providing environmental enrichment affected the concentration of six metabolites and branched chain amino acids index. Such results would be very useful to design environmental enrichment strategies when pigs are challenged by disease in commercial farms. The magnitude of change in lymphocytes level was negatively genetically correlated with l-α-aminobutyric acid. This result indicates that l-αs-aminobutyric acid can be an early indicator of the magnitude of change in lymphocytes level. Such indicator can be collected from nucleus breeding herds in healthy animals and could provide an early biomarker of resilience.

A general method for calculating solid/liquid interfacial free energies from atomistic simulations: Application to CaSO4·xH2O.

We present a general method for computing interfacial free energies from atomistic simulations, which is particularly suitable for solid/liquid interfaces. Our method uses an Einstein crystal as a universal reference state and is more flexible than previous approaches. Surfaces with dipoles, complex reconstructions, and miscible species are all easily accommodated within the framework. It may also be extended to calculating the relative free energies of different phases and other types of defect. We have applied our method to interfaces of bassanite and gypsum with water and obtained interfacial free energies of the order of 0.12 J/m2, of which ∼45% is due to entropic contributions. Our calculations of the interfacial free energy of NaCl with water obtained a value of 0.13 J/m2, of which only 19% is from entropic contributions. We have also predicted equilibrium morphologies for bassanite and gypsum that compare well with experiments and previous calculations.

Effects of porcine reproductive and respiratory syndrome virus (PRRSV) on thyroid hormone metabolism in the late gestation fetus.

Porcine reproductive and respiratory syndrome virus (PRRSV) in late gestation causes a profound suppression of circulating maternal and fetal thyroid hormone during a critical window of development. To understand this relationship, we evaluated thyroid hormone metabolism at the maternal-fetal interface and within fetal tissues, along with hormone metabolite levels in serum. Fetuses were classified using an established model based on viral load in serum and thymus, and preservation status, including uninfected (UNIF), high-viral load viable (HV-VIA), and high-viral load meconium-stained (HV-MEC), with additional controls from sham-inoculated gilts (CON). Expression of three iodothyronine deiodinases, five sulfotransferases, sulfatase, and two solute carriers known to transport thyroid hormone were evaluated in maternal endometrium and fetal placenta, liver, and kidney. Serum thyroxin (T4), reverse triiodothyronine (rT3), and diiodothyronine (T2) were evaluated via liquid chromatography tandem mass spectrometry. Significant changes in gene expression were observed in all four tissues, with the liver being the most severely impacted. We observed local and fetal specific regulation of maternal tissues through significant upregulation of DIO2 and DIO3 expression in the endometrium corresponding to infected but viable fetuses relative to uninfected and control fetuses. Expression levels of DIO2 and DIO3 were significantly higher in the resilient (HV-VIA) fetuses relative to the susceptible (HV-MEC) fetuses. A substantial decrease in serum T4 was confirmed, with no corresponding increase in rT3 or T2. Collectively, these results show that thyroid hormone metabolism is altered at the maternal-fetal interface and within the PRRSV infected fetus and is associated with fetal viability.

Impact of SARS-CoV-2 lockdown on expansion of HIV transmission clusters among key populations: A retrospective phylogenetic analysis.

Background: Public health measures designed to reduce SARS-CoV-2 transmission led to reduced access to care and prevention services for people living with or at risk of acquiring HIV, particularly during the initial introduction of extensive restrictions. This reduction in access may have contributed to increases in HIV transmission not outweighed by decreases in transmission occurring as a result of reduced contact rates promoted by the same public health measures. Methods: We synthesize available province-wide HIV data in British Columbia, Canada, together with public mobility data to phylogenetically investigate the early impacts of SARS-CoV-2 on HIV transmission. Cluster growth, coalescent branching events and lineage-level diversification rates were assessed in "pre-lockdown" (January 22-March 21, 2020), "lockdown" (March 22-May 20, 2020) and "post-lockdown" (May 21-July 19, 2020) to facilitate comparison of transmission trends across key populations. Findings: Results reveal increased HIV transmission in a limited number of clusters in association with reduced access to health services during the initial introduction of SARS-CoV-2-related restrictions. In particular, clusters associated with people who inject drugs (PWID) show rapid growth, extensive branching events in phylogenetic trees during and following the lockdown period, and elevated median change in individuals' viral diversification rates during lockdown compared to clusters associated with men who have sex with men (MSM), consistent with increased transmission rates between PWID. Interpretation: Increased vigilance and innovative targeted solutions are critical to offset potential negative impacts of SARS-CoV-2 or future pandemic-related restrictions on HIV epidemic dynamics. Funding: Funding sources include Genome Canada and Genome BC, the Public Health Agency of Canada, the BC Centre for Excellence in HIV/AIDS, and the Canadian Institutes of Health Research Coronavirus Rapid Response Programme. Student funding includes a NSERC CREATE scholarship and a Canadian Institutes of Health Research graduate fellowship.