In-vivo analysis of Protec™ and ß-glucan supplementation on innate immune performance and intestinal health of rainbow trout.

Functional diets are often given to fish during key stages to improve health through the interaction of the feed components with the host intestine. The additional factors added in these diets are known to modulate the immune response and as such may also offer protection against pathogenic challenges. The present study was undertaken to evaluate whether ß-glucan supplementation for 6 weeks can alter the magnitude of immune response to immunological challenges and subsequently offer an improved innate immune response to bacterial challenge in rainbow trout. Two experimental diets were used to study these effects: a basic commercial diet supplemented with ß-glucan and a commercially available functional diet (Protec™) that has ß-glucan as a functional component in addition to other components were compared to a basic commercial control diet. No significant differences were observed in biometric data. Histological analysis revealed a significantly greater number of goblet cells in the fish fed Protec™ and ß-glucan diets compared to those fed a control diet. Cell marker gene expression of distal intestine leucocytes indicated higher expression of T- and B-cells marker genes to both the ß-glucan containing diets in comparison to control. The Protec™ diet demonstrated modulation of innate immune markers after 6 weeks of feeding with key antimicrobial genes (SAA, HAMP, IL-1ß and TNFα) showing significant increases compared to the other diets. After stimulation with both PAMPs and an immune challenge with A. salmonicida fish fed the ß-glucan diet and the Protec™ exhibited modulation of the innate immune response. An immune challenge with A. salmonicida was carried out to identify if dietary composition led to differences in the innate immune response of rainbow trout. Modulation of the magnitude of response in some immune genes (SAA, IL-1ß and HAMP) was observed in both the distal intestine and head kidney in the Protec™ and ß-glucan fed fish compared to those fed the control diet.

Response of soil health indicators to dung, urine and mineral fertilizer application in temperate pastures.

Healthy soils are key to sustainability and food security. In temperate grasslands, not many studies have focused on soil health comparisons between contrasting pasture systems under different management strategies and treatment applications (e.g. manures and inorganic fertilisers). The aim of this study was to assess the responses of soil health indicators to dung, urine and inorganic N fertiliser in three temperate swards: permanent pasture not ploughed for at least 20 years (PP), high sugar ryegrass with white clover targeted at 30% coverage reseeded in 2013 (WC), and high sugar ryegrass reseeded in 2014 (HG). This study was conducted on the North Wyke Farm Platform (UK) from April 2017 to October 2017. Soil health indicators including soil organic carbon (SOC, measured by loss of ignition and elemental analyser), dissolved organic carbon (DOC), total nitrogen (TN), C:N ratio, soil C and N bulk isotopes, pH, bulk density (BD), aggregate stability, ergosterol concentration (as a proxy for fungi biomass), and earthworms (abundance, mass and density) were measured and analysed before and after application of dung and N fertilizer, urine and N fertiliser, and only N fertiliser. The highest SOC, TN, DOC, ergosterol concentration and earthworms as well as the lowest BD were found in PP, likely due to the lack of ploughing. Differences among treatments were observed due to the application of dung, resulting in an improvement in chemical indicators of soil health after 50 days of its application. Ergosterol concentration was significantly higher before treatment applications than at the end of the experiment. No changes were detected in BD and aggregate stability after treatment applications. We conclude that not enough time had passed for the soil to recover after the ploughing and reseeding of the permanent pasture, independently of the sward composition (HG or WC). Our results highlight the strong influence of the soil management legacy in temperate pasture and the positive effects of dung application on soil health over the short term. In addition, we point out the relevance of using standardised methods to report soil health indicators and some methodological limitations.

Immune responses to prebiotics in farmed salmonid fish: How transcriptomic approaches help interpret responses.

Within aquaculture, prebiotics are composed of complex carbohydrate molecules that cannot be digested by the fish directly but are metabolised by the microbial communities within the host gut, with the desire that "healthy" bacterial species are promoted with subsequently improved performance of the fish, there are likely some direct responses of intestinal cells to these dietary components. The sources and processing of prebiotics, which fall under the overarching theme of "functional feeds" are highly varied between species and types of prebiotics administered. How these feeds exert their effect, and the host responses are hard to determine, but new technologies and the development of high-throughput technologies (omics) are enabling the mechanisms and methods of action to be further understood. The recent advances in the availability of 'omics' technologies with the transition from single gene assays to microarray and RNA-seq in fish health have enabled novel functional ingredients to be analysed. This review will focus on recent studies on targeted gene expression and 'omics' technologies to characterize immune responses. Comparisons between the immunomodulatory effect of different prebiotics have been made and specific examples of how transcriptomics techniques have been used to identify immune responses to prebiotics are given.

A true congenital pancreatic cyst in a dog.

BACKGROUND: True congenital pancreatic cysts are a rare pathological process reported within feline and human literature. To date there has been no documented case of a true congenital cyst affecting a canine patient. The objective of this case report is to document the clinical findings, diagnostic investigations, surgical treatment, histopathological diagnosis and long-term outcome of a dog with a true pancreatic cyst. CASE PRESENTATION: A 5-month-old crossbreed dog was presented with a six-week history of abdominal pain, apparent bilateral pelvic limb weakness, reluctance to walk and intermittent vomiting and diarrhoea. An abdominal ultrasound examination performed by the dog's primary care veterinarian identified a large intra-abdominal structure of unclear origin. A computed tomographic examination identified a large ovoid structure measuring 156 mm in length, 95 mm in height and 89 mm in width and apparently originating from the left limb of the pancreas. An exploratory coeliotomy was performed and a partial pancreatectomy was performed to allow complete removal of the cystic structure. Histopathological analysis of sections of the wall of the large fluid-filled cyst identified a thick fibromuscular wall lined by a well regimented hyperplastic tall columnar epithelium with basally located round to ovoid nuclei featuring fine chromatin stippling and abundant apically located and surface mucin, concurrent with a true congenital pancreatic cyst. A long-term follow-up of twenty-nine months identified no clinical signs of recurrence. CONCLUSION: A partial pancreatectomy and en bloc excision of a true pancreatic cyst provided an excellent long-term outcome in a dog.

Exploring genetic resistance to infectious salmon anaemia virus in Atlantic salmon by genome-wide association and RNA sequencing.

BACKGROUND: Infectious Salmonid Anaemia Virus (ISAV) causes a notifiable disease that poses a large threat for Atlantic salmon (Salmo salar) aquaculture worldwide. There is no fully effective treatment or vaccine, and therefore selective breeding to increase resistance to ISAV is a promising avenue for disease prevention. Genomic selection and potentially genome editing can be applied to enhance host resistance, and these approaches benefit from improved knowledge of the genetic and functional basis of the target trait. The aim of this study was to characterise the genetic architecture of resistance to ISAV in a commercial Atlantic salmon population and study its underlying functional genomic basis using RNA Sequencing. RESULTS: A total of 2833 Atlantic salmon parr belonging to 194 families were exposed to ISAV in a cohabitation challenge in which cumulative mortality reached 63% over 55 days. A total of 1353 animals were genotyped using a 55 K SNP array, and the estimate of heritability for the trait of binary survival was 0.13-0.33 (pedigree-genomic). A genome-wide association analysis confirmed that resistance to ISAV was a polygenic trait, albeit a genomic region in chromosome Ssa13 was significantly associated with resistance and explained 3% of the genetic variance. RNA sequencing of the heart of 16 infected (7 and 14 days post infection) and 8 control fish highlighted 4927 and 2437 differentially expressed genes at 7 and 14 days post infection respectively. The complement and coagulation pathway was down-regulated in infected fish, while several metabolic pathways were up-regulated. The interferon pathway showed little evidence of up-regulation at 7 days post infection but was mildly activated at 14 days, suggesting a potential crosstalk between host and virus. Comparison of the transcriptomic response of fish with high and low breeding values for resistance highlighted TRIM25 as being up-regulated in resistant fish. CONCLUSIONS: ISAV resistance shows moderate heritability with a polygenic architecture, but a significant QTL was detected on chromosome 13. A mild up-regulation of the interferon pathway characterises the response to the virus in heart samples from this population of Atlantic salmon, and candidate genes showing differential expression between samples with high and low breeding values for resistance were identified.

A rapid ammonium fluoride method to determine the oxygen isotope ratio of available phosphorus in tropical soils.

RATIONALE: The isotopic composition of oxygen bound to phosphorus (δ18 OP value) offers an opportunity to gain insight into P cycling mechanisms. However, there is little information for tropical forest soils, which presents a challenge for δ18 OP measurements due to low available P concentrations. Here we report the use of a rapid ammonium fluoride extraction method (Bray-1) as an alternative to the widely used anion-exchange membrane (AEM) method for quantification of δ18 OP values of available P in tropical forest soils. METHODS: We compared P concentrations and δ18 OP values of available and microbial P determined by AEM and Bray-1 extraction for a series of tropical forest soils from Panama spanning a steep P gradient. This involved an assessment of the influence of extraction conditions, including temperature, extraction time, fumigation time and solution-to-soil ratio, on P concentrations and isotope ratios. RESULTS: Depending on the extraction conditions, Bray-1 P concentrations ranged from 0.2 to 66.3 mg P kg-1 across the soils. Extraction time and temperature had only minor effects on Bray-1 P, but concentrations increased markedly as the solution-to-soil ratio increased. In contrast, extraction conditions did not affect Bray-1 δ18 OP values, indicating that Bray-1 provides a robust measure of the isotopic composition of available soil P. For a relatively high P soil, available and fumigation-released (microbial) δ18 OP values determined by Bray-1 extraction (20‰ and 16‰, respectively) were higher than those determined by the AEM method (18‰ and 12‰, respectively), which we attribute to slightly different P pools extracted by the two methods and/or differences resulting from the longer extraction time needed for the AEM method. CONCLUSIONS: The short extraction time, insensitivity to extraction conditions and smaller mass of soil required to extract sufficient P for isotopic analysis make Bray-1extraction a suitable alternative to the AEM method for the determination of δ18 OP values of available P in tropical soils.

Arginine, ornithine and citrulline supplementation in rainbow trout: Free amino acid dynamics and gene expression responses to bacterial infection.

Supplementing the diet with functional ingredients is a key strategy to improve fish performance and health in aquaculture. The amino acids of the urea and nitric oxide (NO) cycles - arginine, ornithine and citrulline - perform crucial roles in the immune response through the generation of NO and the synthesis of polyamine used for tissue repair. We previously found that citrulline supplementation improves and maintains circulating free arginine levels in rainbow trout more effectively than arginine supplementation. Here, to test whether supplementation of urea cycle amino acids modulates the immune response in rainbow trout (Oncorhynchus mykiss), we supplemented a commercial diet with high levels (2% of total diet) of either arginine, ornithine or citrulline during a 7-week feeding trial, before challenging fish with the bacterium Aeromonas salmonicida. We carried out two separate experiments to investigate fish survival and 24 h post-infection to investigate the immediate response of free amino acid levels, and transcriptional changes in genes encoding urea cycle, NO cycle and polyamine synthesis enzymes. There were no differences in percentage fish mortality between diets, however there were numerous highly significant changes in free amino acid levels and gene expression to both dietary supplementation and infection. Out of 26 amino acids detected in blood plasma, 8 were significantly changed by infection and 9 by dietary supplementation of either arginine, ornithine or citrulline. Taurine, glycine and aspartic acid displayed the largest decreases in circulating levels in infected fish, while ornithine and isoleucine were the only amino acids that increased in concentration. We investigated transcriptional responses of the enzymes involved in arginine metabolism in liver and head kidney; transcripts for polyamine synthesis enzymes showed highly significant increases in both tissues across all diets following infection. The paralogous arginase-encoding genes, Arg1a, Arg1b, Arg2a and Arg2b, displayed complex responses across tissues and also due to diet and infection. Overall, these findings improve our understanding of amino acid metabolism following infection and suggests new potential amino acid targets for improving the immune response in salmonids.

Supplementation of arginine, ornithine and citrulline in rainbow trout (Oncorhynchus mykiss): Effects on growth, amino acid levels in plasma and gene expression responses in liver tissue.

Functional amino acids (FAA) regulate metabolic pathways directly linked to health, survival, growth and development. Arginine is a FAA with crucial roles in protein deposition and the immune response. In mammals, supplementation of arginine's precursor amino acid, citrulline, is known to increase circulating arginine to levels beyond direct arginine supplementation, however, citrulline supplementation is poorly studied in fish. To address this knowledge gap, we supplemented the diet of rainbow trout with arginine and its precursor amino acids, ornithine and citrulline, at 3 levels (0.5%, 1% and 2% of the total diet) during a 14-week experiment. We sampled fish at 3 h and 24 h post-feeding to investigate immediate and steady-state effects, respectively. There were no differences in fish growth for any of the diets across a range of indicators. In blood plasma, out of 26 amino acids detected, 11 and 6 displayed significant changes 24 h and 3 h post-prandial, respectively. Arginine, ornithine and citrulline levels were all significantly increased by the citrulline supplemented diets. In muscle, 8 amino acids were significantly altered by supplemented diets, while there were no significant changes in liver. Arginine was increased by 2% citrulline supplementation in muscle tissue. We also investigated the transcriptional responses of urea cycle, nitric oxide cycle and rate-limiting polyamine synthesis enzymes, related to arginine's metabolism, in liver. At both time points, only 2 enzymes were significantly altered by the supplemented diets, however several significant changes were observed comparing 3 h and 24 h post-prandial expression levels. Of these, the paralogous polyamine synthesis enzyme encoding genes ODC1 and ODC2 displayed the largest increases in 3 h post-prandial fish. These findings demonstrate that endogenous synthesis of arginine is possible from a citrulline supplemented diet and improve our understanding of arginine metabolism in fish.

Rainbow trout (Oncorhynchus mykiss) urea cycle and polyamine synthesis gene families show dynamic expression responses to inflammation.

The urea cycle is an endogenous source of arginine that also supports removal of nitrogenous waste following protein metabolism. This cycle is considered inefficient in salmonids, where only 10-15% of nitrogenous waste is excreted as urea. In rainbow trout, arginine is an essential amino acid that has attracted attention due to its many functional roles. These roles include the regulation of protein deposition, immune responses and polyamine synthesis; the latter is directly linked to the urea cycle and involved in tissue repair. The key enzymes used in the urea cycle, namely arginase, ornithine transcarbamylase, argininosuccinate synthase and argininosuccinate lyase, in addition to two rate limiting enzymes required for polyamine synthesis (ornithine decarboxylase and s-adenosylmethionine decarboxylase) are poorly studied in fishes, and their responses to inflammation remain unknown. To address this knowledge gap, we characterised these gene families using phylogenetics and comparative genomics, investigated their mRNA distribution among a panel of tissues and established their transcriptional responses to an acute inflammatory response caused by bacterial infection in liver and muscle. Gene duplicates (paralogues) were identified for arginase (ARG1a, 1b, 2a and 2b), ornithine decarboxylase (ODC1 and 2) and s-adenosylmethionine decarboxylase (SAMdc1 and 2), including paralogues retained from an ancestral salmonid-specific whole genome duplication. ARG2a and 2b were highly upregulated following bacterial infection in liver, whereas ARG1b was downregulated, while both paralogues of SAMdc and ODC were upregulated in liver and unchanged in muscle. Overall, these findings improve our understanding of the molecules supporting the urea cycle and polyamine synthesis in fish, highlighting major changes in the regulation of these systems during inflammation.

The stable oxygen isotope ratio of resin extractable phosphate derived from fresh cattle faeces.

RATIONALE: Phosphorus losses from agriculture pose an environmental threat to watercourses. A new approach using the stable oxygen isotope ratio of oxygen in phosphate (δ18 OPO4 value) may help elucidate some phosphorus sources and cycling. Accurately determined and isotopically distinct source values are essential for this process. The δ18 OPO4 values of animal wastes have, up to now, received little attention. METHODS: Phosphate (PO4 ) was extracted from cattle faeces using anion resins and the contribution of microbial PO4 was assessed. The δ18 OPO4 value of the extracted PO4 was measured by precipitating silver phosphate and subsequent analysis on a thermal conversion elemental analyser at 1400°C, with the resultant carbon monoxide being mixed with a helium carrier gas passed through a gas chromatography (GC) column into a mass spectrometer. Faecal water oxygen isotope ratios (δ18 OH2O values) were determined on a dual-inlet mass spectrometer through a process of headspace carbon dioxide equilibration with water samples. RESULTS: Microbiological results indicated that much of the extracted PO4 was not derived directly from the gut fauna lysed during the extraction of PO4 from the faeces. Assuming that the faecal δ18 OH2O values represented cattle body water, the predicted pyrophosphatase equilibrium δ18 OPO4 (Eδ18 OPO4 ) values ranged between +17.9 and +19.9‰, while using groundwater δ18 OH2O values gave a range of +13.1 to +14.0‰. The faecal δ18 OPO4 values ranged between +13.2 and +15.3‰. CONCLUSIONS: The fresh faecal δ18 OPO4 values were equivalent to those reported elsewhere for agricultural animal slurry. However, they were different from the Eδ18 OPO4 value calculated from the faecal δ18 OH2O value. Our results indicate that slurry PO4 is, in the main, derived from animal faeces although an explanation for the observed value range could not be determined.

Phylogeny and expression analysis of C-reactive protein (CRP) and serum amyloid-P (SAP) like genes reveal two distinct groups in fish.

The acute phase response (APR) is an early innate immune function that is initiated by inflammatory signals, leading to the release of acute phase proteins to the bloodstream to re-establish homeostasis following microbial infection. In this study we analysed the Atlantic salmon (Salmo salar) whole-genome database and identified five C-reactive protein (CRP)/serum amyloid P component (SAP) like molecules namely CRP/SAP-1a, CRP/SAP-1b, CRP/SAP-1c, CRP/SAP-2 and CRP/SAP-3. These CRP/SAP genes formed two distinct sub-families, a universal group (group I) present in all vertebrates and a fish/amphibian specific group (group II). Salmon CRP/SAP-1a, CRP/SAP-1b and CRP/SAP-1c and CRP/SAP-2 belong to the group I family whilst salmon CRP/SAP-3 is a member of group II. Gene expression analysis showed that the salmon CRP/SAP-1a as well as serum amyloid A-5 (SAA-5), one of the major acute phase proteins, were significantly up-regulated by recombinant cytokines (rIL-1ß and rIFNγ) in primary head kidney cells whilst the other four CRP/SAPs remained refractory. Furthermore, SAA-5 was produced as the main acute phase protein (APP) in Atlantic salmon challenged with Aeromonas salmonicida (aroA(-) strain) whilst salmon CRP/SAPs remained unaltered. Overall, these data illustrate the potential different functions of expanded salmon CRP/SAPs to their mammalian homologues.

Phosphate stable oxygen isotope variability within a temperate agricultural soil.

In this study, we conduct a spatial analysis of soil total phosphorus (TP), acid extractable phosphate (PO4) and the stable oxygen (O) isotope ratio within the PO4 molecule (δ18OPO4 ) from an intensively managed agricultural grassland site. Total P in the soil was found to range from 736 to 1952 mg P kg- 1, of which between 12 and 48% was extractable using a 1 M HCl (HClPO4 ) solution with the two variables exhibiting a strong positive correlation. The δ18OPO4 of the extracted PO4 ranged from 17.0 to 21.6‰ with a mean of 18.8‰ (± 0.8). While the spatial variability of Total P has been researched at various scales, this is the first study to assess the variability of soil δ18OPO4 at a field-scale resolution. We investigate whether or not δ18OPO4 variability has any significant relationship with: (i) itself with respect to spatial autocorrelation effects; and (ii) HClPO4 , elevation and slope - both globally and locally. Results indicate that δ18OPO4 was not spatially autocorrelated; and that δ18OPO4 was only weakly related to HClPO4 , elevation and slope, when considering the study field as a whole. Interestingly, the latter relationships appear to vary in strength locally. In particular, the δ18OPO4 to HClPO4 relationship may depend on the underlying soil class and/or on different field managements that had operated across an historical north-south field division of the study field, a division that had been removed four years prior to this study.

Impact of selenium supplementation on fish antiviral responses: a whole transcriptomic analysis in rainbow trout (Oncorhynchus mykiss) fed supranutritional levels of Sel-Plex®.

BACKGROUND: Selenium (Se) is required for the synthesis of proteins (selenoproteins) with essential biological functions. Selenoproteins have a crucial role in the maintenance of cellular redox homeostasis in nearly all tissues, and are also involved in thyroid hormone metabolism, inflammation and immunity. Several immune processes rely on Se status and can be compromised if this element is present below the required level. Previous work has supported the notion that when Se is delivered at levels above those deemed to be the minimal required but below toxic concentrations it can have a boosting effect on the organism's immune response. Based on this concept Se-enriched supplements may represent a valuable resource for functional feeds in animal farming, including aquaculture. RESULTS: In this study we tested the effects of Se supplemented as Sel-Plex during an immune challenge induced by polyinosinic:polycytidylic acid (poly(I:C)), a pathogen-associated molecular pattern (PAMP) that mimics viral infection. Trout were fed two diets enriched with 1 or 4 mg Se Kg(-1) of feed (dry weight) by Sel-Plex addition and a commercial formulation as control. The whole trout transcriptomic response was investigated by microarray and gene ontology analysis, the latter carried out to highlight the biological processes that were influenced by Sel-Plex supplementation in the head kidney (HK) and liver, the main immune and metabolic organs in fish. Overall, Sel-Plex enrichment up to 4 mg Se Kg(-1) induced an important response in the trout HK, eliciting an up-regulation of several genes involved in pathways connected with hematopoiesis and immunity. In contrast, a more constrained response was seen in the liver, with lipid metabolism being the main pathway altered by Se supplementation. Upon stimulation with poly(I:C), supplementation of 4 mg Se Kg(-1) increased the expression of principal mediators of the antiviral defences, especially IFN-γ, and down-stream molecules involved in the cell-mediated immune response. CONCLUSIONS: Supplementation of diets with 4 mg Se Kg(-1) using Sel-Plex remarkably improved the fish response to viral PAMP stimulation. Sel-Plex, being a highly bioavailable supplement of organic Se, might represent a suitable option for supplementation of fish feeds, to achieve the final aim of improving fish fitness and resistance against immune challenges.

Assessment of bioavailable organic phosphorus in tropical forest soils by organic acid extraction and phosphatase hydrolysis.

Soil organic phosphorus contributes to the nutrition of tropical trees, but is not accounted for in standard soil phosphorus tests. Plants and microbes can release organic anions to solubilize organic phosphorus from soil surfaces, and synthesize phosphatases to release inorganic phosphate from the solubilized compounds. We developed a procedure to estimate bioavailable organic phosphorus in tropical forest soils by simulating the secretion processes of organic acids and phosphatases. Five lowland tropical forest soils with contrasting properties (pH 4.4-6.1, total P 86-429 mg P kg- 1) were extracted with 2 mM citric acid (i.e., 10 µmol g- 1, approximating rhizosphere concentrations) adjusted to soil pH in a 4:1 solution to soil ratio for 1 h. Three phosphatase enzymes were then added to the soil extract to determine the forms of hydrolysable organic phosphorus. Total phosphorus extracted by the procedure ranged between 3.22 and 8.06 mg P kg- 1 (mean 5.55 ± 0.42 mg P kg- 1), of which on average three quarters was unreactive phosphorus (i.e., organic phosphorus plus inorganic polyphosphate). Of the enzyme-hydrolysable unreactive phosphorus, 28% was simple phosphomonoesters hydrolyzed by phosphomonoesterase from bovine intestinal mucosa, a further 18% was phosphodiesters hydrolyzed by a combination of nuclease from Penicillium citrinum and phosphomonoesterase, and the remaining 51% was hydrolyzed by a broad-spectrum phytase from wheat. We conclude that soil organic phosphorus can be solubilized and hydrolyzed by a combination of organic acids and phosphatase enzymes in lowland tropical forest soils, indicating that this pathway could make a significant contribution to biological phosphorus acquisition in tropical forests. Furthermore, we have developed a method that can be used to assess the bioavailability of this soil organic phosphorus.

Disrupted seasonal biology impacts health, food security and ecosystems.

The rhythm of life on earth is shaped by seasonal changes in the environment. Plants and animals show profound annual cycles in physiology, health, morphology, behaviour and demography in response to environmental cues. Seasonal biology impacts ecosystems and agriculture, with consequences for humans and biodiversity. Human populations show robust annual rhythms in health and well-being, and the birth month can have lasting effects that persist throughout life. This review emphasizes the need for a better understanding of seasonal biology against the backdrop of its rapidly progressing disruption through climate change, human lifestyles and other anthropogenic impact. Climate change is modifying annual rhythms to which numerous organisms have adapted, with potential consequences for industries relating to health, ecosystems and food security. Disconcertingly, human lifestyles under artificial conditions of eternal summer provide the most extreme example for disconnect from natural seasons, making humans vulnerable to increased morbidity and mortality. In this review, we introduce scenarios of seasonal disruption, highlight key aspects of seasonal biology and summarize from biomedical, anthropological, veterinary, agricultural and environmental perspectives the recent evidence for seasonal desynchronization between environmental factors and internal rhythms. Because annual rhythms are pervasive across biological systems, they provide a common framework for trans-disciplinary research.

The importance of soil drying and re-wetting in crop phytohormonal and nutritional responses to deficit irrigation.

Soil drying and re-wetting (DRW) occurs at varying frequencies and intensities during crop production, and is deliberately used in water-saving irrigation techniques that aim to enhance crop water use efficiency. Soil drying not only limits root water uptake which can (but not always) perturb shoot water status, but also alters root synthesis of phytohormones and their transport to shoots to regulate leaf growth and gas exchange. Re-wetting the soil rapidly restores leaf water potential and leaf growth (minutes to hours), but gas exchange recovers more slowly (hours to days), probably mediated by sustained changes in root to shoot phytohormonal signalling. Partial rootzone drying (PRD) deliberately irrigates only part of the rootzone, while the remainder is allowed to dry. Alternating these wet and dry zones (thus re-wetting dry soil) substantially improves crop yields compared with maintaining fixed wet and dry zones or conventional deficit irrigation, and modifies phytohormonal (especially abscisic acid) signalling. Alternate wetting and drying (AWD) of rice can also improve yield compared with paddy culture, and is correlated with altered phytohormonal (including cytokinin) signalling. Both PRD and AWD can improve crop nutrition, and re-wetting dry soil provokes both physical and biological changes which affect soil nutrient availability. Whether this alters crop nutrient uptake depends on competition between plant and microbes for nutrients, with the rate of re-wetting determining microbial dynamics. Nevertheless, studies that examine the effects of soil DRW on both crop nutritional and phytohormonal responses are relatively rare; thus, determining the cause(s) of enhanced crop yields under AWD and PRD remains challenging.

Comparison of insulin lispro protamine suspension versus insulin glargine once daily added to oral antihyperglycaemic medications and exenatide in type 2 diabetes: a prospective randomized open-label trial.

AIMS: To compare efficacy and safety of two, once-daily basal insulin formulations [insulin lispro protamine suspension (ILPS) vs. insulin glargine (glargine)] added to oral antihyperglycaemic medications (OAMs) and exenatide BID in suboptimally controlled type 2 diabetes (T2D) patients. METHODS: This 24-week, open-label, multicentre trial randomized patients to bedtime ILPS (n = 171) or glargine (n = 168). Non-inferiority of ILPS versus glargine was assessed by comparing the upper limit of 95% confidence intervals (CIs) for change in haemoglobin A1c (HbA1c) from baseline to week 24 (adjusted for baseline HbA1c) with non-inferiority margin 0.4%. RESULTS: Non-inferiority of ILPS versus glargine was demonstrated: least-squares mean between-treatment difference (ILPS minus glargine) (95% CI) was 0.22% (0.06, 0.38). Mean HbA1c reduction was less for ILPS- versus glargine-treated patients (-1.16 ± 0.84 vs. -1.40 ± 0.97%, p = 0.008). Endpoint HbA1c < 7.0% was achieved by 53.7% (ILPS) and 61.7% (glargine) (p = NS). Overall hypoglycaemia rates (p = NS) and severe hypoglycaemia incidence (p = NS) were similar. Nocturnal hypoglycaemia rate was higher in patients treated with ILPS versus glargine (p = 0.004). Weight gain was similar between groups (ILPS: 0.27 ± 3.38 kg; glargine: 0.66 ± 3.93 kg, p = NS). Endpoint total insulin doses were lower in patients treated with ILPS versus glargine (0.30 ± 0.17 vs. 0.37 ± 0.17 IU/kg/day, p < 0.001). CONCLUSIONS: ILPS was non-inferior to glargine for HbA1c change over 24 weeks, but was associated with less HbA1c reduction and more nocturnal hypoglycaemia. Treat-to-target basal insulin therapy improves glycaemic control and is associated with minimal weight gain when added to OAMs and exenatide BID for suboptimally controlled T2D.

Identification and characterisation of TLR18-21 genes in Atlantic salmon (Salmo salar).

Teleost fish possess many types of toll-like receptor (TLR) some of which exist in other vertebrate groups and some that do not (ie so-called "fish-specific" TLRs). In this study, we identified in Atlantic salmon (Salmo salar) whole-genome shotgun (WGS) contigs seven TLRs that are not found in mammals, including six types of fish-specific TLRs (one TLR18, one TLR19, and four TLR20 members (two of which are putative soluble forms (s)) and one TLR21. Phylogenetic analysis revealed that teleost TLR19-21 are closely related with murine TLR11-TLR13, whilst teleost TLR18 groups with mammalian TLR1, 2, 6 and 10. A typical TLR protein domain structure was found in all these TLRs with the exception of TLR20b(s) and TLR20c(s). TLR-GFP expression plasmids transfected into SHK-1 cells showed that salmon TLR19, TLR20a and TLR20d were preferentially localised to the intracellular compartment. Real time PCR analysis suggested that salmon TLR19-TLR21 are mainly expressed in immune related organs, such as spleen, head kidney and gills, while TLR18 transcripts are more abundant in muscle. In vitro stimulation of primary head kidney cells with type I IFN, IFNγ and IL-1ß had no impact on TLR expression. Infectious salmon anaemia virus (ISAV) infection, in vivo, down-regulated TLR20a, TLR20b(s), TLR20d and TLR21 in infected salmon kidney tissue. In contrast, up-regulation of TLR19 and TLR20a expression was found in posterior kidney in rainbow trout with clinical proliferative kidney disease (PKD).

A neuroimaging measure to capture heterogeneous patterns of atrophy in Parkinson's disease and dementia with Lewy bodies.

INTRODUCTION: Parkinson's disease (PD) and Dementia with Lewy bodies (DLB) show heterogeneous brain atrophy patterns which group-average analyses fail to capture. Neuroanatomical normative modelling overcomes this by comparing individuals to a large reference cohort. Patient-specific atrophy patterns are measured objectively and summarised to index overall neurodegeneration (the 'total outlier count'). We aimed to quantify patterns of neurodegenerative dissimilarity in participants with PD and DLB and evaluate the potential clinical relevance of total outlier count by testing its association with key clinical measures in PD and DLB. MATERIALS AND METHODS: We included 108 participants with PD and 61 with DLB. PD participants were subclassified into high and low visual performers as this has previously been shown to stratify those at increased dementia risk. We generated z-scores from T1w-MRI scans for each participant relative to normative regional cortical thickness and subcortical volumes, modelled in a reference cohort (n = 58,836). Outliers (z < -1.96) were aggregated across 169 brain regions per participant. To measure dissimilarity, individuals' Hamming distance scores were calculated. We also examined total outlier counts between high versus low visual performance in PD; and PD versus DLB; and tested associations between these and cognition. RESULTS: There was significantly greater inter-individual dissimilarity in brain-outlier patterns in PD poor compared to high visual performers (W = 522.5; p < 0.01) and in DLB compared to PD (W = 5649; p < 0.01). PD poor visual performers had significantly greater total outlier counts compared to high (ß = -4.73 (SE = 1.30); t = -3.64; p < 0.01) whereas a conventional group-level GLM failed to identify differences. Higher total outlier counts were associated with poorer MoCA (ß = -0.55 (SE = 0.27), t = -2.04, p = 0.05) and composite cognitive scores (ß = -2.01 (SE = 0.79); t = -2.54; p = 0.02) in DLB, and visuoperception (ß = -0.67 (SE = 0.19); t = -3.59; p < 0.01), in PD. CONCLUSIONS: Neuroanatomical normative modelling shows promise as a clinically informative technique in PD and DLB, where patterns of atrophy are variable.

Cytosine-based nucleoside analogs are selectively lethal to DNA mismatch repair-deficient tumour cells by enhancing levels of intracellular oxidative stress.

BACKGROUND: DNA mismatch repair deficiency is present in a significant proportion of a number of solid tumours and is associated with distinct clinical behaviour. METHODS: To identify the therapeutic agents that might show selectivity for mismatch repair-deficient tumour cells, we screened a pair of isogenic MLH1-deficient and MLH1-proficient tumour cell lines with a library of clinically used drugs. To test the generality of hits in the screen, selective agents were retested in cells deficient in the MSH2 mismatch repair gene. RESULTS: We identified cytarabine and other related cytosine-based nucleoside analogues as being selectively toxic to MLH1 and MSH2-deficient tumour cells. The selective cytotoxicity we observed was likely caused by increased levels of cellular oxidative stress, as it could be abrogated by antioxidants. CONCLUSION: We propose that cytarabine-based chemotherapy regimens may represent a tumour-selective treatment strategy for mismatch repair-deficient cancers.