Assessing periparturient ewe characteristics and nemabiome composition to guide targeted selective treatment for sustainable gastrointestinal nematodes control in sheep.

Gastrointestinal nematodes (GINs) are a significant threat to the sustainability of global sheep production. Periparturient ewes play a key role in GIN epidemiology, with increased GIN faecal egg counts (FECs) in these ewes resulting in heavy pasture contamination that facilitates parasitic gastroenteritis in immunologically naïve lambs later during the grazing period. Traditionally, blanket anthelmintic treatment would suppress GIN egg outputs in these ewes and subsequent pasture contamination. However, farmers are now advised to implement targeted selective treatment (TST) to reduce anthelmintic use and subsequent anthelmintic resistance development, yet, there is currently limited evidence to determine optimal TST strategies in ewes. In this study, the characteristics of 226 ewes on seven Welsh farms were assessed postlambing to identify factors associated with their individual strongyle FECs using negative binomial mixed model analysis. Nemabiome analysis was conducted on 34 ewes across two study farms using the Oxford Nanopore MinIon platform with an aim of identifying factors associated with variations in ewe nemabiome composition within flocks. The best-fitted model of ewe FEC incorporated ewe body condition score, dag score, breed, and an interaction effect between ewe age and litter size as fixed factors. The addition of a mean FEC value for ewes of a specific litter size on each farm further improved model fit and reduced between-farm variance in the model. Nemabiome analysis revealed significant variation in within flock nemabiome diversity on individual farms, with significantly reduced nemabiome diversity recorded in ewes exhibiting dags and in twin-bearing ewes on respective farms, whilst T. circumcincta was present as a significantly higher proportion of the nemabiome in Suffolk ewes and twin bearing ewes (P < 0.05) in respective flocks. Our data demonstrate that commonly recorded ewe characteristics can be exploited to predict individual periparturient ewe FEC and subsequently may be used as a guide for TST strategies on sheep farms once specific TST thresholds are identified to deliver the optimal balance between minimal pasture contamination and maximal GIN refugia. This study is the first to utilise Oxford Nanopore MinIon sequencing to evaluate the nemabiome of sheep, and to molecularly assess the nemabiome of individual ruminants within a flock/herd, with results indicating that significant within flock variations in nemabiome composition which may have implications for TST and flock management strategies.

Effects of photobiomodulation and caffeine treatment on acute kidney injury in a hypoxic ischemic neonatal rat model.

Hypoxic ischemic encephalopathy (HIE) occurs in 2-5/1000 births, with acute kidney injury (AKI) occurring in 40%. AKI increases morbidity and mortality. Caffeine, an adenosine receptor antagonist, and photobiomodulation (PBM), working on cytochrome c oxidase, are potential treatments for AKI. To examine effects of caffeine and PBM on AKI in rats, Day 7 pups underwent a HIE intervention (Modified Rice-Vannucci model) replicating pathology observed in humans. Caffeine was administered for 3 days and/or PBM for 5 days following HIE. Weights and urine for biomarkers (NGAL, albumin, KIM-1, osteopontin) were collected prior to HIE, daily post intervention and at sacrifice. Both treatments reduced kidney injury seen on electron microscopy, but not when combined. HIE elevated urinary NGAL and albumin on Days 1-3 post-HIE, before returning to control levels. This elevation was significantly reduced by PBM or caffeine. KIM-1 was significantly elevated for 7 days post-HIE and was reduced by both treatments. Osteopontin was not altered by HIE or the treatments. Treatments, individually but not in combination, improved HIE-induced reductions in the enzymatic activity of mitochondrial complexes II-III. PBM and caffeine also improved weight gain. PBM and caffeine reduces AKI diagnosed by urinary biomarkers and confirmed by EM findings.

Moving the Needle Toward Fair Compensation in Pediatric Nephrology.

Remuneration issues are a substantial threat to the long-term stability of the pediatric nephrology workforce. It is uncertain whether the pediatric nephrology workforce will meet the growing needs of children with kidney disease without a substantial overhaul of the current reimbursement policies. In contrast to adult nephrology, the majority of pediatric nephrologists practice in an academic setting affiliated with a university and/or children's hospital. The pediatric nephrology service line is crucial to maintaining the financial health and wellness of a comprehensive children's hospital. However, in the current fee-for-service system, the clinical care for children with kidney disease is neither sufficiently valued, nor appropriately compensated. Current compensation models derived from the relative value unit (RVU) system contribute to the structural biases inherent in the current inequitable payment system. The perceived negative financial compensation is a significant driver of waning trainee interest in the field which is one of the least attractive specialties for students, with a significant proportion of training spots going unfilled each year and relatively stagnant growth rate as compared to the other pediatric subspecialties. This article reviews the current state of financial compensation issues plaguing the pediatric nephrology subspecialty. We further outline strategies for pediatric nephrologists, hospital administrators, and policy-makers to improve the landscape of financial reimbursement to pediatric subspecialists. A physician compensation model is proposed which aligns clinical activity with alternate metrics for current non-RVU producing activities that harmonizes hospital and personal mission statements.

In vitro screening as an anthelmintic discovery pipeline for Calicophoron daubneyi: nutritive media and rumen environment-based approaches.

Paramphistomosis can lead to morbidity and mortality of ruminant livestock within tropical and sub-tropical climates. In recent decades, rumen fluke has become an emerging infection in temperate climates across Western Europe, with Calicophoron daubneyi, the primary species present. Clinical outbreaks with C. daubneyi larvae are reported and adults might be responsible for production losses. There is not currently a widely licensed anthelmintic product available to control C. daubneyi. In this study, three existing flukicide anthelmintics were tested for efficacy against mature C. daubneyi, comparing a standard in vitro culturing assay and a new more relevant rumen fluid based in vitro compound screening protocol. The new rumen based screen confirmed that oxyclozanide was active against adult C. daubneyi and identified activity with praziquantel. The study highlighted the downstream value of incorporating relevant in vitro screening for anthelmintic discovery pipelines.

Development and validation of a liquid chromatography-mass spectrometric assay for simultaneous determination of tacrolimus and 13-O-desmethyl tacrolimus in rat kidney tissue.

A sensitive and robust LC-MS/MS method has been developed and validated to determine the concentrations of tacrolimus and its major metabolite 13-O-desmethyl tacrolimus (13-ODMT) in kidney tissue from rats who received tacrolimus intra-peritoneally at doses of 0.5mg/kg and 2mg/kg. The samples were prepared by a liquid-liquid extraction procedure using ethyl ether as the extraction solvent and ascomycin as the internal standard. Chromatographic separation was achieved using Phenomenex Kinetex column (2.6µm C18 100Å, 100×2.1mm, Phenomenex, Torrance CA) and a gradient mobile phase of water and methanol-acetonitrile (50:50, v/v) both containing 0.1% formic acid. The limit of quantification was 0.25ng/ml and the calibration curves covered a concentration range from 0.25 to 50ng/ml. Intra-and inter-assay precision and accuracy for both tacrolimus and 13-ODMT were all within FDA guidelines for bioanalysis. Extraction efficiency for tacrolimus ranged from 67.00 to 74.90% and from 66.70 to 78.40% for 13-ODMT. Several challenges interfering with the performance of the method such as phospholipid build-up have also been addressed. Kidney tissue samples from six rats receiving either 0.5 or 2mg/kg dose were analyzed and resulted in a median concentration of 11.54 and 0.72ng/ml for tacrolimus and 13-ODMT, respectively, for the lower dose level, and a median concentration of 8.89ng/ml and 1.50ng/ml for tacrolimus and 13-ODMT, respectively, at the higher dose level.

Early hepatic and peritoneal changes and immune response in goats vaccinated with a recombinant glutathione transferase sigma class and challenged with Fasciola hepatica.

Changes and local immune response were evaluated in the peritoneal cell populations, duodenal lamina propria and liver from goats immunized with recombinant glutathione transferase sigma class (rFhGST-S1) during early stages of infection with Fasciola hepatica. Group 1 (n=7) was unimmunized and uninfected; group 2 (n=10) was immunized with adjuvant Quil A and infected; group 3 (n=10) was immunised with rFhGST-S1 and infected. Three goats from each group were killed at 7-9 days post-infection (dpi) to evaluate early changes and immune response. The remaining goats were killed at 15 weeks post-infection (wpi). rFhGST-S1 vaccination induced variable response: three goats showed low fluke burden at 15 wpi and two goats showed low hepatic damage at early infection stages. This response was associated to a severe infiltrate of eosinophils in peritoneal fluid and hepatic necrotic foci, high iNOS expression in peritoneal cells and abundant infiltrate of eosinophils surrounding hepatic migrating flukes. T lymphocyte subsets were found in the vicinity of necrotic areas but they were absent in the vicinity of migrating larvae. No significant variation for T cell subsets, except for CD4 and γδ T lymphocytes, that were higher in the Quil A group compared to the rFhGST-S1 group. Expression of IL4 and IFN-γ in the hepatic inflammatory infiltrates was very occasional.

Chronic hemodialysis in pediatric patients: technical and practical aspects of use.

Chronic kidney disease (CKD) is a continuum of progressive reduction in kidney function lasting for more than three months, due to either structural and/or functional renal abnormalities that may lead to irreversible kidney damage. The term "renal supportive therapy" (RST) generally characterizes the spectrum of dialysis therapies available to support existing renal function in patients with CKD during progression to end-stage renal disease (ESRD) and/or renal transplantation. Chronic RST modalities include conventional hemodialysis, peritoneal dialysis and home hemodialysis therapies. The modality chosen to deliver RST in the pediatric patient is often guided by a variety of factors including institutional resources, local expertise, patient characteristics, treatment goals, and physician preference. Chronic RST in a pediatric population requires the flexible utilization of multiple delivery modalities for effective care across infancy into adulthood and is not typically initiated until GFR declines to between 15-30 mL/min per 1.73 m2, although thresholds for initiation of RST will vary between patients. This review will provide an overview of current approaches to management and technical approaches to pediatric patients requiring chronic hemodialysis.

Case report: successful treatment of recurrent focal segmental glomerulosclerosis with a novel rituximab regimen.

Focal segmental glomerulosclerosis (FSGS) is the cause of renal failure in more than 10% of pediatric patients undergoing renal transplantation. Recurrent FSGS is a major cause of pediatric allograft failure, with the risk increasing for patients undergoing retransplantation. Standard therapy for recurrent posttransplantation FSGS includes the use of intensive plasmapheresis (PP) in conjunction with cyclophosphamide or high-dose cyclosporine. However, many patients exhibit refractory disease, with rapid progression to allograft loss despite these interventions. Prior studies have reported conflicting data on the efficacy of adding rituximab therapy to the standard treatment regimen for recurrent posttransplantation FSGS. Here we present a successful therapeutic protocol with rapid elimination of PP after initiation of rituximab therapy for an adolescent patient with recurrent FSGS in the immediate postoperative period. The patient has maintained excellent allograft function through 12 months posttransplantation.

Immune responses directed at egg proteins during experimental infection with the liver fluke Fasciola hepatica.

Fasciola hepatica is responsible for human disease and economic livestock loss on a global scale. Unlike the well characterized schistosomes, only the adult and juvenile stages of F. hepatica are implicated in disease, whereas the freely voided egg is not thought to contribute to host-parasite interactions. We investigated specific immune responses to soluble F. hepatica egg proteins (SFHEP), during a 14-week experimental infection, demonstrating significant increases in anti-SFHEP IgG1 (P = 0.001), transforming growth factor beta-1 (P = 0.008) and IL-10 (P < 0.001) titres at the onset of egg production. Western blot analysis of soluble SFHEP demonstrates that protein bands migrating at 61.6, 54.8 and 44 kDa become sero-reactive before the appearance of eggs within host faeces. Therefore, expression of some egg-associated proteins indicates progression to chronic disease. Antigenic bands were investigated through mass spectrometry, identifying a protein disulphide isomerase (PDI) (61.6 kDa), an enolase and ferritin-related proteins (54.8 kDa), and a cocktail of dehydrogenases (44 kDa). Biochemical analysis of egg secretions reveals proteolytic activity, which increases over time, indicating that proteases may be continually secreted during the course of egg maturation. The implications of egg-specific immune responses and proteolytic secretions are further discussed.

Long-term exercise training in overweight adolescents improves plasma peptide YY and resistin.

The objective of this study was to investigate the effect of long-term exercise training on concentrations of five hormones related to appetite and insulin resistance in overweight adolescents. In addition, we were interested in the relationships of these hormones with each other and with anthropometric and/or cardiovascular disease marker changes. Participants were >or=the 85th percentile for BMI for age and sex and participated in an 8-month supervised aerobic training program. Anthropometrics, cardiovascular fitness assessment, and fasting blood samples were taken pre- and post-training. Glucose, insulin, total cholesterol (TC), high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, triglycerides, leptin, active ghrelin, total peptide YY (PYY), adiponectin, and resistin concentrations were measured. The participants increased their time to exhaustion on an incremental treadmill test and decreased both percent body fat and blood triglyceride concentrations. Total PYY concentration increased and resistin concentration decreased after long-term exercise training, which are favorable outcomes. Leptin concentrations were related to weight, percent body fat, waist circumference, and triglyceride concentrations pre- and post-training. The changes in resistin concentrations were related to the changes in triglyceride concentrations. We conclude that long-term exercise training has beneficial effects for overweight adolescents with respect to PYY and resistin, hormones related to appetite and insulin sensitivity.

A proteomics approach to quantify protein levels following RNA interference: case study with glutathione transferase superfamily from the model metazoan Caenorhabditis elegans.

Loss-of-function phenotypic analysis via interference RNA (RNAi) technology is a revolutionary approach to assigning gene function. While transcript-based methodologies commonly validate RNAi gene suppression investigations, protein-based validation is less developed. This report illustrates the potential for two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (2-DE) and gel analysis to quantify protein levels following RNAi. This case study involves three glutathione transferase (GST) genes targeted by RNAi from the model organism Caenorhabditis elegans.

Fluid and volume monitoring.

BACKGROUND: Fluid resuscitation is not only used to prevent acute kidney injury (AKI) but fluid management is also a cornerstone of treatment for patients with established AKI and renal failure. Ultrafiltration removes volume initially from the intravascular compartment inducing a relative degree of hypovolemia. Normal reflex mechanisms attempt to sustain blood pressure constant despite marked changes in blood volume and cardiac output. Thus, compensated shock with a normal blood pressure is a major cause of AKI or exacerbations of AKI during ultrafiltration. METHODS: We undertook a systematic review of the literature using MEDLINE, Google Scholar and PubMed searches. We determined a list of key questions and convened a 2-day consensus conference to develop summary statements via a series of alternating breakout and plenary sessions. In these sessions, we identified supporting evidence and generated clinical practice recommendations and/or directions for future research. RESULTS: We defined three aspects of fluid monitoring: i) normal and pathophysiological cardiovascular mechanisms; ii) measures of volume responsiveness and impending cardiovascular collapse during volume removal, and; iii) measured indices of each using non-invasive and minimally invasive continuous and intermittent monitoring techniques. The evidence documents that AKI can occur in the setting of normotensive hypovolemia and that under-resuscitation represents a major cause of both AKI and mortality ion critically ill patients. Traditional measures of intravascular volume and ventricular filling do not predict volume responsiveness whereas dynamic functional hemodynamic markers, such as pulse pressure or stroke volume variation during positive pressure breathing or mean flow changes with passive leg raising are highly predictive of volume responsiveness. Numerous commercially-available devices exist that can acquire these signals. CONCLUSIONS: Prospective clinical trials using functional hemodynamic markers in the diagnosis and management of AKI and volume status during ultrafiltration need to be performed. More traditional measure of preload be abandoned as marked of volume responsiveness though still useful to assess overall volume status.

Adrenocortical tumor with left renal vein, vena cava and intrahepatic venous extension.

A 41 year old white female presented with upper respiratory distress and shortness of breath appeared on initial computed tomography (CT) scan to have a large left retroperitoneal mass with left renal vein extension including a mass up to the level of the atrium. This presentation suggested hypernephroma. She proved, however, to have an adrenal cortical carcinoma which displaced the kidney, exhibiting vascular invasion within the gland and non-adherent extension into the vena cava, atrium, common hepatic vein and left renal vein, where some adherence was present. This unusual tumor required extensive surgery for removal, including use of cardiopulmonary bypass, with good results.

Understanding triclabendazole resistance.

Triclabendazole (TCBZ) has been the drug of choice to treat liver fluke infections in livestock for >20 years, due to its high activity against both adult and juvenile flukes. More recently, it has been used successfully to treat human cases of fascioliasis. Resistance to TCBZ first appeared in the field in Australia in the mid-1990s. Since then, resistance has been reported from a number of countries throughout Europe: Ireland, Scotland, Wales, Spain and The Netherlands. The heavy reliance on a single drug puts treatment strategies for fascioliasis at risk. Should resistance develop further, the prospect is an alarming one. This review will present an overview of progress in understanding the mechanism of resistance to TCBZ, examining possible changes in the target molecule, in drug influx/efflux mechanisms and in the metabolism of TCBZ by the fluke. The review will also consider ways to deal with resistance, covering drug-oriented options such as: the use of alternative drugs, drug combinations and the search for new compounds.

The effect of vascular access location and size on circuit survival in pediatric continuous renal replacement therapy: a report from the PPCRRT registry.

PURPOSE: Well-functioning vascular access is essential for the provision of adequate CRRT. However, few data exist to describe the effect of catheter size or location on CRRT performance in the pediatric population. METHODS: Data for vascular access site, size, and location, as well as type of anticoagulant used and patient demographic data were gathered from the ppCRRT registry. Kaplan-Meier curves were generated and then analyzed by log-rank test or Cox Proportional Hazards model. RESULTS: Access diameter was found to significantly affect circuit survival. None of the 5 French catheters lasted longer than 20 hours. Seven and 9 French, but not 8 French, catheters fared worse than larger diameter catheters (p=0.002). Circuits associated with internal jugular access survived longer than subclavian or femoral access associated circuits (p<0.05). Circuit survival was also found to be favorably associated with the CVVHD modality (p<0.001). CONCLUSIONS: Functional CRRT circuit survival in children is favored by larger catheter diameter, internal jugular vein insertion site and CVVHD. For patients requiring catheter diameters less than 10 French, CRRT circuit survival might be optimized if internal jugular vein insertion is feasible. Conversely, when a vascular access site other than the internal jugular vein is most prudent, consideration should be given to using the largest diameter catheter appropriate for the size of the child. The CVVHD modality was associated with longer circuit survival, but the mechanism by which this occurs is unclear.

Disruption of neurofascin localization reveals early changes preceding demyelination and remyelination in multiple sclerosis.

Saltatory conduction in the nervous system is enabled through the intimate association between the leading edge of the myelin sheath and the axonal membrane to demarcate the node of Ranvier. The 186 kDa neuron specific isoform of the adhesion molecule neurofascin (Nfasc186) is required for the clustering of voltage gated Na+ channels at the node, whilst the 155 kDa glial specific isoform (Nfasc155) is required for the assembly of correct paranodal junctions. In order to understand the relationship between these vital structures and how they are affected in multiple sclerosis we have examined the expression of Nfasc155 and Nfasc186 in areas of inflammation, demyelination and remyelination from post-mortem brains. Fourteen cases of neuropathologically confirmed multiple sclerosis (8 female and 6 male; post-mortem delay 7-24 h; age 37-77 years; and disease duration 15-40 years), comprising 20 tissue blocks with 32 demyelinating or remyelinating lesions, were used in this study. A significant early alteration in Nfasc155+ paranodal structures occurs within and adjacent to actively demyelinating white matter lesions that are associated with damaged axons. Shaker-type Kv1.2 channels, normally located distally to the paranode, overlapped with the disrupted Nfasc155+ structures. In the absence of Nfasc155, Kv1.2 channels abutted normally clustered Nfasc186+ nodes, indicating that complete disruption of the paranodal structure and movement of Kv1.2 channels precede alterations at the node itself. Within areas of partial remyelination, a number of atypical triple-Nfasc155+ structures were noted that may represent transient oligodendrocyte-axonal contacts during the process of myelin repair or aberrant interactions. Within shadow plaques discretely clustered Na+v, Nfasc186+ and Nfasc155+ domains indicated the restoration of normal nodal architecture. The alterations in oligodendrocyte Nfasc155 expression that accompany inflammation and demyelination suggest an ongoing disruption to the axonal-oligodendrocyte complex within newly forming as well as established lesions in multiple sclerosis, resulting in destruction of the Nfasc186+/Na+v nodal complex vital to successful fast neurotransmission in the CNS.

Evidence for biogeographic patterning of mitochondrial DNA sequences in Eastern horse populations.

Equine mitochondrial DNA (mtDNA) phylogeny reconstruction reveals a complex pattern of variation unlike that seen in other large domesticates. It is likely that this pattern reflects a process of multiple and repeated, although not necessarily independent, domestication events. Until now, no clear geographic affiliation of clades has been apparent. In this study, amova analyses have revealed a significant non-random distribution of the diversity among equine populations when seven newly sequenced Eurasian populations were examined in the context of previously published sequences. The association of Eastern mtDNA types in haplogroup F was highly significant using Fisher's exact test of independence (P = 0.00000). For the first time, clear biogeographic partitioning has been detected in equine mtDNA sequence.

Mitochondrial DNA sequence diversity in extant Irish horse populations and in ancient horses.

Equine mitochondrial DNA sequence variation was investigated in three indigenous Irish horse populations (Irish Draught Horse, Kerry Bog Pony and Connemara Pony) and, for context, in 69 other horse populations. There was no evidence of Irish Draught Horse or Connemara Pony sequence clustering, although the majority of Irish Draught Horse sequences (47%) were assigned to haplogroup D. Conversely, 31% of the Kerry Bog Pony sequences were assigned to the rare haplogroup E. In addition to the extant population analyses, ancient DNA sequences were generated from three out of four Irish archaeological specimens, all of which were assigned to haplogroup A.

A novel nonsense mutation in the EYA1 gene associated with branchio-oto-renal/branchiootic syndrome in an Afrikaner kindred.

Branchio-oto-renal (BOR) syndrome is an autosomal dominant disorder characterized by the associations of hearing loss, branchial arch defects and renal anomalies. Branchiootic (BO) syndrome is a related disorder that presents without the highly variable characteristic renal anomalies of BOR syndrome. Dominant mutations in the human homologue of the Drosophila eyes absent gene (EYA1) are frequently the cause of both BOR and BO syndromes. We report a South African family of Afrikaner descent with affected individuals presenting with pre-auricular abnormalities and either hearing loss or bilateral absence of the kidneys. Genetic analysis of the pedigree detected a novel EYA1 heterozygous nonsense mutation in affected family members but not in unaffected family members or a random DNA panel. Through mutational analysis, we conclude that this particular mutation is the cause of BOR/BO syndrome in this family as a result of a truncation of the EYA1 protein that ablates the critical EYA homologous region. To the best of our knowledge, this is the first case of BOR/BO syndrome reported in Africa or in those of the Afrikaner descent.

Charcot-Marie-Tooth type 4F disease caused by S399fsx410 mutation in the PRX gene.

Charcot-Marie-Tooth type 4F disease (CMT4F) is an autosomal recessive neuropathy caused by mutations in the PRX gene. To date, only seven mutations have been identified in the PRX gene. In this study, the authors report a novel S399fsX410 mutation in the PRX gene and its effects at the protein level, which was identified in an 8-year-old patient with early-onset CMT disease.