An investigation into the transmission and control of pestivirus in sheep in Australia.

Border disease virus (BDV) is a member of the pestivirus genus that primarily affects sheep, causing reproductive losses through abortion, still births and the birth of weak lambs. The key characteristic of this disease is the birth of persistently infected (PI) lambs which, after surviving transplacental infection, are born antibody negative, yet virus positive, and thus shed the virus for their entire life and are the primary source of spread within a flock. The cornerstones of BDV control are detection and elimination of PI animals, biosecurity measures to prevent re-infection, and surveillance programs. Recommendations for the control of BDV in sheep are centred around the approach to bovine viral diarrhoea virus (BVDV), the prominent cattle pestivirus species, due to a lack of specific research into BDV control and elimination. In this study, two aspects of a BDV control program were investigated: the effectiveness of the BVDV vaccine, Pestigard®, and the rate of seroconversion in a flock deliberately exposed to known PI lambs. The vaccine appeared to be safe, and the optimal dose was the full cattle dose (2 mL). While vaccination induced high virus neutralising titres to BVDV when administered as either a quarter, half or full dose registered for cattle, the BDV titres achieved were low and unlikely to prevent transplacental infection. In a second study, after exposure of between 2 and 15 days exposure to two PI lambs in confined conditions, only 3 of 66 previously naïve sheep demonstrated seroconversion. This demonstrated a very low rate of transmission and suggested that deliberate exposure to PI lambs at low-risk times for less than 15 days was not likely to be an effective means of achieving seroconversion throughout a flock and, therefore, not provide protection against BDV challenge during gestation.

Evaluating the effect of calcium, magnesium and sodium supplementation of Merino ewes on their lambs' growth.

Our study aimed to investigate responses to supplementation of calcium (Ca), magnesium (Mg) and sodium (Na) to lactating ewes and lambs grazing barley forage from lamb marking to weaning. A 10-ha paddock sown to barley was subdivided into eight plots as four replicates of two treatments. Merino ewes (n = 104) with lambs at foot were stratified to the eight plots (13 ewes and 21 lambs/plot) based on number of lambs (twin or single) and ewes' weight. Supplemented groups had access to mineral supplements (30 g/ewe/day) supplying 12 g/ewe/day ground limestone, 12 g/ewe/day Causmag® and 6 g/ewe/day coarse salt in a ratio of 2: 2: 1 by weight (as fed) from day 0 (a day before lamb marking) after sample collection. Control groups were not supplemented with minerals. Blood, milk and urine samples from ewes and blood from lambs were collected at different time points, namely, a day prior to lamb marking (day 0), 14 days after the commencement of study (day 14), and 28 days after the commencement of study (day 28). Weight of the lambs was also recorded at each time point. We found that the concentration of the forage minerals (Ca, Mg and potassium (K)) was lower on day 28 than on day 0 (P < 0.025). Liveweight gain was greater in the first 14 days compared with the second weight gain period (P < 0.001). The interaction of time and treatment was significant for liveweight (P < 0.001). Due to the improvement in weight gain of supplemented lambs, we recommend that mineral supplementation during late lactation is beneficial considering the low cost of minerals, even though the mineral content of the forage was not deficient.

Assessment of cannabidiol use in pets according to a national survey in the USA.

OBJECTIVES: Conduct an online questionnaire to understand the motivations and perceptions about cannabidiol use in companion animals in the USA. MATERIALS AND METHODS: Data from a USA population sample who owned a pet were gathered using an online questionnaire. Perception of cannabidiol efficacy was analysed for independence against explanatory variables using the Pearson chi-square test, followed by a binary logistic regression. RESULTS: A total of 1238 participants completed the survey; 356 had administered cannabidiol to their pet before. Dogs were the most prevalent pet, followed by cats (75.8 and 22.2%, respectively). Treats/chews and oils were the most common forms of cannabidiol (44.6 and 42.9%, respectively). The most prevalent condition for treating with cannabidiol was anxiety and stress (67.4%), followed by joint pain and inflammation (23%). Doses and frequency of cannabidiol used by many pet owners were inconsistent, and yet many participants perceived an improvement of their pets' condition with supplementation, with mild to no side effects. Most respondents had not given cannabidiol to their pets before due to uncertainty about its efficacy and safety. The frequency of cannabidiol administration and length of time administered were both significant for whether participants found it efficacious in treating a given condition, and this was more evident when supplementing cannabidiol for a longer time. CLINICAL SIGNIFICANCE: We found heterogeneity regarding cannabidiol dosage and dosing frequency. Cannabidiol was mostly perceived as safe and effective, but there is a need to conduct further research on cannabidiol long-term tolerability and therapeutic efficacy for treating various conditions.

Comparison of the efficacy of macrocyclic lactone anthelmintics, either singly or in combination with other anthelmintic(s), in nine beef herds in southern NSW.

Anthelmintic resistance (AR) is a well-recognized challenge in farmed ruminants. The use of anthelmintics in combination is one of the strategies recommended to slow the rate of AR development. Two studies were undertaken in 2017 and 2019 to assess the efficacy of single-dose macrocyclic lactone (ML) anthelmintic and ML combination drenches. In total, 11 Faecal Egg Count Reduction Trials (FECRTs) were set up in 10 different beef herds, with results available from 10 of those FECRTs (9 herds). AR to a single ML anthelmintic was detected in all 9 herds, with resistance to Cooperia and Haemonchus spp on 9 farms, and resistance to Ostertagia and Trichostrongylus spp on 2 farms. In contrast, for the ML combination anthelmintics, all FECRTs resulted in efficacies of 99%-100%. The results suggest that cattle producers should strongly consider using combination drenches in their herds in preference to single actives.

The impact of feeding supplemental minerals to sheep on the return of micronutrients to pasture via urine and faeces.

The form (organic versus inorganic) of minerals (Se, Zn, Cu and Mn), supplemented to sheep (Charolais × Suffolk-Mule (mean weight = 57 ± 2.9 kg) at two European industrial doses, on the return of micronutrients to pasture via nutrient partitioning and composition in sheep urine and faeces was investigated. This gave four treatments in total with 6 animals per treatment (n = 24). The form of the supplemented minerals did not influence the excretory partitioning of micronutrients (Se, Zn, Cu and Mn) between urine and faeces, nor on their concentrations in the excreta. The two doses trialed however, may influence the Se flux in the environment through altering the ratios of Se:P and Se:S ratios in the faeces and Se:S ratio in the urine. Administration of the mineral supplements also improved the retention of P in sheep reducing its excretion via urine. Although the concentrations of readily bioavailable micronutrients in the faeces were not affected by the mineral forms, there were differences in the more recalcitrant fractions of Se, Zn and Cu (as inferred via a sequential extraction) in faeces when different forms of supplemental minerals were offered. The potential impact of these differences on micronutrient flux in pasture requires further investigation.

A survey on bloat in southern Australian beef production systems.

Frothy bloat, associated predominantly with grazing legume-based pastures, is considered the second most costly disease in beef cattle in southern Australia, costing $84.4 M annually. It frequently results in the sudden death of cattle. In response to concerns from cattle producers, an online survey was conducted in southern Australia in late 2020 to determine the impact of bloat, identify risk factors and determine the efficacy of current preventive measures. For 217 responses, over two-thirds (70%) of producers reported bloat occurring in the previous 12 months, with estimated morbidity and mortality rates of 3.7% and 5.0% respectively. Bloat was associated with clover or clover-dominant paddocks (79%) and was not associated with grass or grass-dominant pastures or low clover pastures (92%) nor grazing crops (27%). For bloat that occurred in the past 12 months, cattle were very commonly grazing on clover or clover-dominant paddocks (90%) and occasionally lucerne-dominated paddocks (7%). Two-thirds of producers reported having preventive measures in place when losses occurred. Bayesian Network analysis confirmed that grazing clover-based pastures for more than 7 days, yearling cattle and the months of July-September were the main risk factors for bloat occurrence, with pasture type (clover) being the most important. Conversely, no clear relationship between weather conditions and bloat occurrence was evident. This survey highlights the known risk of clover-based pastures for causing bloat in cattle, and that losses occur in many cases despite preventive measures being used. This suggests that current methods for preventing bloat in cattle are suboptimal.

Moderate exercise protects against joint disease in a murine model of osteoarthritis.

Exercise is recommended as a non-pharmacological therapy for osteoarthritis (OA). Various exercise regimes, with differing intensities and duration, have been used in a range of OA rodent models. These studies show gentle or moderate exercise reduces the severity of OA parameters while high intensity load bearing exercise is detrimental. However, these studies were largely conducted in rats or in mouse models induced by severe injury, age or obesity, whilst destabilization of the medial meniscus (DMM) in mice has become a widely accepted model due to its lower variability, moderate progression and timescale. The present study was undertaken to provide insight into the effect of moderate exercise on early joint pathology in the DMM mouse model. Exercise was induced a week after induction by forced wheel walking for three or 7 weeks. Joints were analyzed by microcomputed tomography and histology. Assessment of skeletal parameters revealed that exercise offered protection against cartilage damage after 7 weeks of exercise, and a temporary protection against osteosclerosis was displayed after 3 weeks of exercise. Furthermore, exercise modified the metaphyseal trabecular microarchitecture of the osteoarthritic leg in both time points examined. Collectively, our findings corroborate previous studies showing that exercise has an important effect on bone in OA, which subsequently, at 8 weeks post-induction, translates into less cartilage damage. Thus, providing an exercise protocol in a surgical mouse model of OA, which can be used in the future to further dissect the mechanisms by which moderate exercise ameliorates OA.

Cereal grain mineral micronutrient and soil chemistry data from GeoNutrition surveys in Ethiopia and Malawi.

The dataset comprises primary data for the concentration of 29 mineral micronutrients in cereal grains and up to 84 soil chemistry properties from GeoNutrition project surveys in Ethiopia and Malawi. The work provided insights on geospatial variation in the micronutrient concentration in staple crops, and the potential influencing soil factors. In Ethiopia, sampling was conducted in Amhara, Oromia, and Tigray regions, during the late-2017 and late-2018 harvest seasons. In Malawi, national-scale sampling was conducted during the April-June 2018 harvest season. The concentrations of micronutrients in grain were measured using inductively coupled plasma mass spectrometry (ICP-MS). Soil chemistry properties reported include soil pH; total soil nitrogen; total soil carbon (C); soil organic C; effective cation exchange capacity and exchangeable cations; a three-step sequential extraction scheme for the fractionation of sulfur and selenium; available phosphate; diethylenetriaminepentaacetic acid (DTPA)-extractable trace elements; extractable trace elements using 0.01 M Ca(NO3)2 and 0.01 M CaCl2; and isotopically exchangeable Zn. These data are reported here according to FAIR data principles to enable users to further explore agriculture-nutrition linkages.

Soil and landscape factors influence geospatial variation in maize grain zinc concentration in Malawi.

Dietary zinc (Zn) deficiency is widespread globally, and in particular among people in sub-Saharan Africa (SSA). In Malawi, dietary sources of Zn are dominated by maize and spatially dependent variation in grain Zn concentration, which will affect dietary Zn intake, has been reported at distances of up to ~ 100 km. The aim of this study was to identify potential soil properties and environmental covariates which might explain this longer-range spatial variation in maize grain Zn concentration. Data for maize grain Zn concentrations, soil properties, and environmental covariates were obtained from a spatially representative survey in Malawi (n = 1600 locations). Labile and non-labile soil Zn forms were determined using isotopic dilution methods, alongside conventional agronomic soil analyses. Soil properties and environmental covariates as potential predictors of the concentration of Zn in maize grain were tested using a priori expert rankings and false discovery rate (FDR) controls within the linear mixed model (LMM) framework that informed the original survey design. Mean and median grain Zn concentrations were 21.8 and 21.5 mg kg-1, respectively (standard deviation 4.5; range 10.0-48.1). A LMM for grain Zn concentration was constructed for which the independent variables: soil pH(water), isotopically exchangeable Zn (ZnE), and diethylenetriaminepentaacetic acid (DTPA) extractable Zn (ZnDTPA) had predictive value (p < 0.01 in all cases, with FDR controlled at < 0.05). Downscaled mean annual temperature also explained a proportion of the spatial variation in grain Zn concentration. Evidence for spatially dependent variation in maize grain Zn concentrations in Malawi is robust within the LMM framework used in this study, at distances of up to ~ 100 km. Spatial predictions from this LMM provide a basis for further investigation of variations in the contribution of staple foods to Zn nutrition, and where interventions to increase dietary Zn intake (e.g. biofortification) might be most effective. Other soil and landscape factors influencing spatially dependent variation in maize grain Zn concentration, along with factors operating over shorter distances such as choice of crop variety and agronomic practices, require further exploration beyond the scope of the design of this survey.

The Opioid Crisis: Prevalence and Markets of Opioids.

The US opioid crisis came in three waves - prescription opioids, heroin, and illicitly manufactured fentanyls - Centers for Disease Control and Prevention Centers for Disease Control and Prevention resulting in the deaths of nearly 500,000 people from 1996 to 2019. In 2009, drug overdose deaths exceeded those involving automobiles. Opioid overdose deaths contributed to the decrease in life expectancy for Americans from 78.8 to 78.5 during 2014 to 2017. The overprescribing of a schedule II prescription opioid was escalated by pharmaceutical companies promoting a growing belief that pain was an undertreated condition. In 2012, the number of opioid prescriptions peaked at 255 million and deaths exceeded 11,000 per year. The typical prescription opioid abuser was white, male, and 45-55 years of age. The hardest-hit states were in Appalachia and the Northeast. When an abuse-resistant formulation was introduced for OxyContin, the most prevalent prescription opioid, users turned to heroin. From the early 1980s, a new pizza delivery style of Mexican trafficking in black tar heroin infiltrated many of the same states hit hardest by prescription opioids. Heroin overdose deaths reached 14,495 in 2017. As heroin abuse increased in states supplied with black tar heroin, fentanyl-contaminated white powder heroin began to appear in the Northeast. Fentanyl was quickly followed by fentanyl analogs. While heroin deaths continued to escalate through 2017, they were soon overshadowed by fentanyl overdose deaths. Finally, prescription opioid and heroin overdose deaths started to decline in 2017, though fentanyl deaths continued to increase. In late 2019, it appeared that restrictions on transportation and travel due to the COVID-19 pandemic had resulted in decreased availability of illicit drugs, but by 2020 drug abuse had escalated in many countries. Globally, heroin was the primary opioid of abuse and only a few countries - including Canada, Germany, Austria, and Belgium - have experienced a significant increase in prescription opioids. However, illicitly manufactured fentanyls are a growing problem in North America, Europe, Australia, and Asia.

Liming impacts barley yield over a wide concentration range of soil exchangeable cations.

Liming has widespread and significant impacts on soil processes and crop responses. The aim of this study was to describe the relationships between exchangeable cation concentrations in soil and the relative yield of spring barley. The hypothesis was that yield is restricted by the concentration of a single exchangeable cation in the soil. For simplicity, we focused on spring barley which was grown in nine years of a long-term experiment at two sites (Rothamsted and Woburn). Four liming rates were applied and in each year the relative yield (RY) and the concentrations of exchangeable cations were assessed. Liming had highly significant effects on the concentrations of most exchangeable cations, except for Cu and K. There were significant negative relationships (either linear or exponential) between the exchangeable concentrations of Mn, Cd, Cr, Al, Fe, Cu, Co, Zn and Ni in soil and soil pH. The relationships between RY and the concentrations of selected exchangeable cations (Mn, Ca and Al) were described well using log-logistic relationships. For these cations a significant site effect was probably due to fundamental differences in soil properties. At both sites the concentrations of exchangeable soil Al were excessive (> 7.5 mg kg-1) and were most likely responsible for reduced barley yields (where RY ≤ 0.5) with soil acidification. At Rothamsted barley yield was non-limited (where RY ≥ 1) at soil exchangeable Mn concentrations (up to 417 mg kg-1) greater than previously considered toxic, which requires further evaluation of critical Mn concentrations. SUPPLEMENTARY INFORMATION: The online version of this article (10.1007/s10705-020-10117-2) contains supplementary material, which is available to authorized users.

The nutritional quality of cereals varies geospatially in Ethiopia and Malawi.

Micronutrient deficiencies (MNDs) remain widespread among people in sub-Saharan Africa1-5, where access to sufficient food from plant and animal sources that is rich in micronutrients (vitamins and minerals) is limited due to socioeconomic and geographical reasons4-6. Here we report the micronutrient composition (calcium, iron, selenium and zinc) of staple cereal grains for most of the cereal production areas in Ethiopia and Malawi. We show that there is geospatial variation in the composition of micronutrients that is nutritionally important at subnational scales. Soil and environmental covariates of grain micronutrient concentrations included soil pH, soil organic matter, temperature, rainfall and topography, which were specific to micronutrient and crop type. For rural households consuming locally sourced food-including many smallholder farming communities-the location of residence can be the largest influencing factor in determining the dietary intake of micronutrients from cereals. Positive relationships between the concentration of selenium in grain and biomarkers of selenium dietary status occur in both countries. Surveillance of MNDs on the basis of biomarkers of status and dietary intakes from national- and regional-scale food-composition data1-7 could be improved using subnational data on the composition of grain micronutrients. Beyond dietary diversification, interventions to alleviate MNDs, such as food fortification8,9 and biofortification to increase the micronutrient concentrations in crops10,11, should account for geographical effects that can be larger in magnitude than intervention outcomes.

Spatial prediction of the concentration of selenium (Se) in grain across part of Amhara Region, Ethiopia.

Grain and soil were sampled across a large part of Amhara, Ethiopia in a study motivated by prior evidence of selenium (Se) deficiency in the Region's population. The grain samples (teff, Eragrostis tef, and wheat, Triticum aestivum) were analysed for concentration of Se and the soils were analysed for various properties, including Se concentration measured in different extractants. Predictive models for concentration of Se in the respective grains were developed, and the predicted values, along with observed concentrations in the two grains were represented by a multivariate linear mixed model in which selected covariates, derived from remote sensor observations and a digital elevation model, were included as fixed effects. In all modelling steps the selection of predictors was done using false discovery rate control, to avoid over-fitting, and using an α-investment procedure to maximize the statistical power to detect significant relationships by ordering the tests in a sequence based on scientific understanding of the underlying processes likely to control Se concentration in grain. Cross-validation indicated that uncertainties in the empirical best linear unbiased predictions of the Se concentration in both grains were well-characterized by the prediction error variances obtained from the model. The predictions were displayed as maps, and their uncertainty was characterized by computing the probability that the true concentration of Se in grain would be such that a standard serving would not provide the recommended daily allowance of Se. The spatial variation of grain Se was substantial, concentrations in wheat and teff differed but showed the same broad spatial pattern. Such information could be used to target effective interventions to address Se deficiency, and the general procedure used for mapping could be applied to other micronutrients and crops in similar settings.

Bacterial community structure and response to nitrogen amendments in Lake Shenandoah (VA, USA).

Microbial processes are critical to the function of freshwater ecosystems, yet we still do not fully understand the factors that shape freshwater microbial communities. Furthermore, freshwater ecosystems are particularly susceptible to effects of environmental change, including influx of exogenous nutrients such as nitrogen and phosphorus. To evaluate the impact of nitrogen loading on the microbial community structure of shallow freshwater lakes, water samples collected from Lake Shenandoah (Virginia, USA) were incubated with two concentrations of either ammonium, nitrate, or urea as a nitrogen source. The potential impact of these nitrogen compounds on the bacterial community structure was assessed via 16S rRNA amplicon sequencing. At the phylum level, the dominant taxa in Lake Shenandoah were comprised of Actinobacteria and Proteobacteria, which were not affected by exposure to the various nitrogen treatments. Overall, there was not a significant shift in the diversity of the bacterial community of Lake Shenandoah with the addition of nitrogen sources, indicating this shallow system may be constrained by other environmental factors.

Yield responses of arable crops to liming - An evaluation of relationships between yields and soil pH from a long-term liming experiment.

The management of optimal soil pH is fundamental to sustainable crop production. Understanding the lime requirement for arable crops has developed gradually over the last several decades. The aim of this study was to examine the yield-pH relationship for a range of arable crops to understand their response to liming, based on the Long-Term Liming experiments established in 1962 at Rothamsted Research, UK. The main treatments of four different rates of lime and, therefore, four distinctly different soil pH levels were maintained for 35 years at two sites (Rothamsted and Woburn). The pH ranged from 4.4 to 8.0. The lime response was tested on the following crops: spring barley, spring oats, spring beans, spring lupins, winter lupins, potatoes, linseed, winter oilseed rape, winter triticale and winter wheat. Relative yield (RY) was used for non-linear regression analysis to detect site, year and phosphorus (P) fertiliser effects on the relationship with pH. Liming had a highly significant positive effect on soil pH, but overall there was no consistent increase or decrease in soil extractable P (Olsen) or exchangeable K. There were significant site effects detected for RY for most crops which reflect differences in the two soil types. Spring oats and potatoes had very weak responses to lime within the pH range tested. For spring barley, winter triticale, winter wheat and winter oilseed rape significant effects of P fertiliser on the yield-pH relationship were found, although the nature of effects differed between crops and sites. Findings from the Long-Term Liming experiment are invaluable in improving the fundamental understanding on the yield-pH relationship for important arable crops and this has significant implications on selecting crops for rotations. The pH at 90% RY was calculated for selected crops and the beneficial effect of fertiliser P was detected in significantly reducing the critical pH value.

Increasing body mass index and abdominal subcutaneous fat thickness are associated with increased skin-to-epidural space distance in pregnant women.

BACKGROUND: Body mass index does not indicate the distribution of adipose tissue. Central adiposity may be measured using ultrasound measurement of subcutaneous fat thickness. This study determined if the abdominal subcutaneous fat thickness measured correlated with skin-to-epidural space distance at delivery, and compared this with the booking body mass index. METHODS: We analysed a sub-set of participants from a single-centre, prospective cohort study that assessed the relationship between subcutaneous fat thickness and maternity outcomes. Abdominal subcutaneous fat thickness measurements were obtained during the routine fetal anomaly scan. The skin-to-epidural space distance was obtained in those having epidural or combined spinal-epidural analgesia. Linear regression was used to test for strength of association and adjusted R2 values calculated to determine if subcutaneous fat thickness or body mass index was more strongly associated with skin-to-epidural space distance. RESULTS: The 463 women had a median (IQR) booking body mass index of 25.0 kg/m2 (21.8-29.3) and subcutaneous fat thickness of 16.2 mm (13.0-21.0). The median (IQR) skin-to-epidural space distance was 5.0 cm (4.5-6.0). Both parameters significantly correlated with skin-to-epidural space distance (r=0.53 and 0.68 respectively, P <0.001). Adjusted linear regression coefficient (95% CI) for subcutaneous fat thickness was 0.09 (0.08 to 0.11), R2=0.30 and for body mass index 0.12 (0.11 to 0.13), R2=0.47. CONCLUSIONS: Booking body mass index had a stronger relationship with skin-to- epidural space distance at delivery than subcutaneous fat thickness, explaining 47% of the variation in the skin-to-epidural distance.

Challenges and controversies in the conservative management of uterine and ovarian cancer.

Uterine cancer is the fifth most common cancer in women worldwide with an estimated 320,000 annual diagnoses. Its most common form, endometrioid adenocarcinoma of the endometrium (endometrial adenocarcinoma [EAC]), is thought to develop through excessive proliferation of endometrial glands, and then increasing steadily in incidence. The current standard treatment for EAC is hysterectomy, which is often curative. However, it may be unacceptably expensive for women with severe medical comorbidities, those who are at risk of intra- and postoperative adverse events and those who desire fertility. Ovarian cancer is the most malignant of all gynaecological cancers, but patients with disease limited to one ovary and patients with non-epithelial tumours may expect a good prognosis. A selected group of young patients who desire fertility may be well treated with conservative surgery. This chapter reviews patient selection, diagnosis, pre-treatment evaluation, treatment options, surveillance and risk of relapse.

Metagenomic Investigation of Idiopathic Meningoencephalomyelitis in Dogs.

BACKGROUND: Meningoencephalomyelitis of unknown origin (MUO) is a common and life-threatening neuroinflammatory disease in dogs. Features of the disease are suggestive of an underlying immune-mediated process, but the association of this disease with a pathogen is still unknown. HYPOTHESIS/OBJECTIVES: To search for candidate etiologic agent associated with cases if MUO using next generation metagenomic sequencing. ANIMALS: Twenty-two dogs diagnosed with either MUO (11/22; 10 CSF and 3 brain), or noninflammatory CNS diseases inconsistent with MUO (11/22; 11 CSF and 2 brain) that served as negative controls. METHODS: A case control study was performed by identifying MUO and non-MUO cases. Samples were blindly processed and then unblinded for comparative analyses. Inclusion criteria for MUO cases included consistent MRI lesions and inflammatory CSF with a negative PCR panel for infectious agents or histopathologic diagnosis. Dogs with glucocorticoid therapy within 2 weeks of sample collection were excluded. Fresh-frozen cerebrospinal fluid (CSF; 21) and brain (5) samples were collected and RNA and DNA were extracted separately for shotgun metagenomic sequencing. Known positive samples were used as controls to validate our sequencing and analysis pipelines and to establish limits of detection. Sequencing results were analyzed at a nucleotide and protein level for broad comparison to known infectious organisms. RESULTS: No candidate etiologic agents were identified in dogs with MUO. CONCLUSIONS AND CLINICAL IMPORTANCE: These results support but do not prove the hypothesis that MUO is not associated with infectious agents and might be an autoimmune disease.

Evidence of persistence of Prevotella spp. in the cystic fibrosis lung.

Purpose. Prevotella spp. represent a diverse genus of bacteria, frequently identified by both culture and molecular methods in the lungs of patients with chronic respiratory infection. However, their role in the pathogenesis of chronic lung infection is unclear; therefore, a more complete understanding of their molecular epidemiology is required.Methodology. Pulsed Field Gel Electrophoresis (PFGE) and Random Amplified Polymorphic DNA (RAPD) assays were developed and used to determine the degree of similarity between sequential isolates (n=42) from cystic fibrosis (CF) patients during periods of clinical stability and exacerbation.Results. A wide diversity of PFGE and RAPD banding patterns were observed, demonstrating considerable within-genus heterogeneity. In 8/12 (66.7 %) cases, where the same species was identified at sequential time points, pre- and post-antibiotic treatment of an exacerbation, PFGE/RAPD profiles were highly similar or identical. Congruence was observed between PFGE and RAPD (adjusted Rand coefficient, 0.200; adjusted Wallace RAPD->PFGE 0.459, PFGE->RAPD 0.128). Furthermore, some isolates could not be adequately assigned a species name on the basis of 16S rRNA analysis: these isolates had identical PFGE/RAPD profiles to Prevotella histicola.Conclusion. The similarity in PFGE and RAPD banding patterns observed in sequential CF Prevotella isolates may be indicative of the persistence of this genus in the CF lung. Further work is required to determine the clinical significance of this finding, and to more accurately distinguish differences in pathogenicity between species.

A genomic case study of mixed fibrolamellar hepatocellular carcinoma.

BACKGROUND: Mixed fibrolamellar hepatocellular carcinoma (mFL-HCC) is a rare liver tumor defined by the presence of both pure FL-HCC and conventional HCC components, represents up to 25% of cases of FL-HCC, and has been associated with worse prognosis. Recent genomic characterization of pure FL-HCC identified a highly recurrent transcript fusion (DNAJB1:PRKACA) not found in conventional HCC. PATIENTS AND METHODS: We performed exome and transcriptome sequencing of a case of mFL-HCC. A novel BAC-capture approach was developed to identify a 400 kb deletion as the underlying genomic mechanism for a DNAJB1:PRKACA fusion in this case. A sensitive Nanostring Elements assay was used to screen for this transcript fusion in a second case of mFL-HCC, 112 additional HCC samples and 44 adjacent non-tumor liver samples. RESULTS: We report the first comprehensive genomic analysis of a case of mFL-HCC. No common HCC-associated mutations were identified. The very low mutation rate of this case, large number of mostly single-copy, long-range copy number variants, and high expression of ERBB2 were more consistent with previous reports of pure FL-HCC than conventional HCC. In particular, the DNAJB1:PRKACA fusion transcript specifically associated with pure FL-HCC was detected at very high expression levels. Subsequent analysis revealed the presence of this fusion in all primary and metastatic samples, including those with mixed or conventional HCC pathology. A second case of mFL-HCC confirmed our finding that the fusion was detectable in conventional components. An expanded screen identified a third case of fusion-positive HCC, which upon review, also had both conventional and fibrolamellar features. This screen confirmed the absence of the fusion in all conventional HCC and adjacent non-tumor liver samples. CONCLUSION: These results indicate that mFL-HCC is similar to pure FL-HCC at the genomic level and the DNAJB1:PRKACA fusion can be used as a diagnostic tool for both pure and mFL-HCC.