First detection, isolation and molecular characterization of infectious salmon anaemia virus associated with clinical disease in farmed Atlantic salmon (Salmo salar) in Chile

Background  

Several forms of progressive retinal atrophy (PRA) segregate in more than 100 breeds of dog with each PRA segregating in one or a few breeds. This breed specificity may be accounted for by founder effects and genetic drift, which have reduced the genetic heterogeneity of each breed, thereby facilitating the identification of causal mutations. We report here a new form of PRA segregating in the Border Collie breed. The clinical signs, including the loss of night vision and a progressive loss of day vision, resulting in complete blindness, occur at the age of three to four years and may be detected earlier through systematic ocular fundus examination and electroretinography (ERG).     

Changes in invertase activities precede ovary growth induced by gibberellic acid in

Developing pods of pea ( Pisum sativum L. cv. Alaska no 7) were used to study the enzymes of sucrose metabolism. Acid and neutral invertase (EC 3.2.1.26). sucrose synthase (SS, EC 2.4.1.13) and sucrose phosphate synthase (SPS, EC 2.4.1.14) have been localized in the soluble fraction. Acid invertase activity was also present in the insoluble fraction and in pea ovary apoplast. In pea pods, sucrose breakdown was dominated by the invertase pathway. SS specific activity only increased at late stages of parthenocarpic pod development, while SPS did so in pods obtained by pollination. Changes in time course of invertase activities have been correlated with the growth rate of fruits induced to develop either by fertilization or by exogenous application of giberellic acid (GA), 2,4-dichloro-phenoxy acetic acid (2,4-D) or 6-benzylaminopurine (6-BAP). The soluble neutral activities might be associated with pod elongation, while the acid ones were rather related to assimilate import by the induced fruits. Application of gibberellic acid to non-pollinated ovaries significantly enhanced the soluble neutral invertase activity before any ovary outgrowth was detected (up to 2 h after treatment). Within the same period of time. GA-treated ovaries showed a decrease in the acid invertase activity of the soluble fraction and an increase of the acid invertase activity in the apopiast. preceding in time the increment of the acid invertase activity associated with the insoluble fraction. Our results suggest that the early GA response may be mediated through a promotion of processes of protein secretion.     

Investigating Protein-protein Interactions in Live Cells Using Bioluminescence Resonance Energy Transfer

Assays based on Bioluminescence Resonance Energy Transfer (BRET) provide a sensitive and reliable means to monitor protein-protein interactions in live cells. BRET is the non-radiative transfer of energy from a ''donor'' luciferase enzyme to an ''acceptor'' fluorescent protein. In the most common configuration of this assay, the donor is Renilla reniformis luciferase and the acceptor is Yellow Fluorescent Protein (YFP). Because the efficiency of energy transfer is strongly distance-dependent, observation of the BRET phenomenon requires that the donor and acceptor be in close proximity. To test for an interaction between two proteins of interest in cultured mammalian cells, one protein is expressed as a fusion with luciferase and the second as a fusion with YFP. An interaction between the two proteins of interest may bring the donor and acceptor sufficiently close for energy transfer to occur. Compared to other techniques for investigating protein-protein interactions, the BRET assay is sensitive, requires little hands-on time and few reagents, and is able to detect interactions which are weak, transient, or dependent on the biochemical environment found within a live cell. It is therefore an ideal approach for confirming putative interactions suggested by yeast two-hybrid or mass spectrometry proteomics studies, and in addition it is well-suited for mapping interacting regions, assessing the effect of post-translational modifications on protein-protein interactions, and evaluating the impact of mutations identified in patient DNA.     

Tetrapod phylogeny inferred from 18S and 28S ribosomal RNA sequences and a review of the evidence for amniote relationships [published erratum appears in Mol Biol Evol 1991 May;8(3):398]

The 18S ribosomal RNAs of 21 tetrapods were sequenced and aligned with five published tetrapod sequences. When the coelacanth was used as an outgroup, Lissamphibia (living amphibians) and Amniota (amniotes) were found to be statistically significant monophyletic groups. Although little resolution was obtained among the lissamphibian taxa, the amniote sequences support a sister-group relationship between birds and mammals. Portions of the 28S ribosomal RNA (rRNA) molecule in 11 tetrapods also were sequenced, although the phylogenetic results were inconclusive. In contrast to previous studies, deletion or down- weighting of base-paired sites were found to have little effect on phylogenetic relationships. Molecular evidence for amniote relationships is reviewed, showing that three genes (beta-hemoglobin, myoglobin, and 18S rRNA) unambiguously support a bird-mammal relationship, compared with one gene (histone H2B) that favors a bird- crocodilian clade. Separate analyses of four other genes (alpha- crystallin A, alpha-hemoglobin, insulin, and 28S rRNA) and a combined analysis of all sequence data are inconclusive, in that different groups are defined in different analyses and none are strongly supported. It is suggested that until sequences become available from a broader array of taxa, the molecular evidence is best evaluated at the level of individual genes, with emphasis placed on those studies with the greatest number of taxa and sites. When this is done, a bird-mammal relationship is most strongly supported. When regarded in combination with the morphological evidence for this association, it must be considered at least as plausible as a bird-crocodilian relationship.      

Rilonacept in the treatment of acute gouty arthritis: a randomized,controlled clinical trial using indomethacin as the active comparator

Introduction

In phase-3 clinical trials, the interleukin (IL-1) blocker, rilonacept (IL-1 Trap), demonstrated efficacy for gout flare prevention during initiation of urate-lowering therapy. This trial evaluated rilonacept added to a standard-of-care, indomethacin, for treatment of acute gout flares.

Methods

Adults, aged 18-70 years, with gout presenting within 48 hours of flare onset and having at least moderate pain as well as swelling and tenderness in the index joint were randomized to subcutaneous (SC) rilonacept 320 mg at baseline plus oral indomethacin 50 mg TID for 3 days followed by 25 mg TID for up to 9 days (n = 74); SC placebo at baseline plus oral indomethacin as above (n = 76); or SC rilonacept 320 mg at baseline plus oral placebo (n = 75). The primary efficacy endpoint was change in pain in the index joint (patient-reported using a Likert scale (0 = none; 4 = extreme)) from baseline to the average of values at 24, 48 and 72 hours (composite time point) for rilonacept plus indomethacin versus indomethacin alone. Comparison of rilonacept monotherapy with indomethacin monotherapy was dependent on demonstration of significance for the primary endpoint. Safety evaluation included clinical laboratory and adverse event (AE) assessments.

Results

Patient characteristics were comparable among the groups; the population was predominantly male (94.1%), white (75.7%), with mean ± SD age of 50.3 ± 10.6 years. All treatment groups reported within-group pain reductions from baseline (P < 0.0001). Although primary endpoint pain reduction was greater with rilonacept plus indomethacin (-1.55 ± 0.92) relative to indomethacin alone (-1.40 ± 0.96), the difference was not statistically significant (P = 0.33), so formal comparison between monotherapy groups was not performed. Pain reduction over the 72-hour period with rilonacept alone (-0.69 ± 0.97) was less than that in the other groups, but pain reduction was similar among groups at 72 hours. Treatment with rilonacept was well-tolerated with no reported serious AEs related to rilonacept. Across all groups, the most frequent AEs were headache and dizziness.

Conclusions

Although generally well-tolerated, rilonacept in combination with indomethacin and rilonacept alone did not provide additional pain relief over 72 hours relative to indomethacin alone in patients with acute gout flare.

Trial registration

ClinicalTrials.gov registration number {"type":"clinical-trial","attrs":{"text":"NCT00855920","term_id":"NCT00855920"}}NCT00855920.     

Infectious salmon anaemia virus (ISAV) isolated from the ISA disease outbreaks in Chile diverged from ISAV isolates from Norway around 1996 and was disseminated around 2005, based on surface glycoprotein gene sequences

Background

All viruses in the family Bunyaviridae possess a tripartite genome, consisting of a small, a medium, and a large RNA segment. Bunyaviruses therefore possess considerable evolutionary potential, attributable to both intramolecular changes and to genome segment reassortment. Hantaviruses (family Bunyaviridae, genus Hantavirus) are known to cause human hemorrhagic fever with renal syndrome or hantavirus pulmonary syndrome. The primary reservoir host of Sin Nombre virus is the deer mouse (Peromyscus maniculatus), which is widely distributed in North America. We investigated the prevalence of intramolecular changes and of genomic reassortment among Sin Nombre viruses detected in deer mice in three western states.

Methods

Portions of the Sin Nombre virus small (S) and medium (M) RNA segments were amplified by RT-PCR from kidney, lung, liver and spleen of seropositive peromyscine rodents, principally deer mice, collected in Colorado, New Mexico and Montana from 1995 to 2007. Both a 142 nucleotide (nt) amplicon of the M segment, encoding a portion of the G2 transmembrane glycoprotein, and a 751 nt amplicon of the S segment, encoding part of the nucleocapsid protein, were cloned and sequenced from 19 deer mice and from one brush mouse (P. boylii), S RNA but not M RNA from one deer mouse, and M RNA but not S RNA from another deer mouse.

Results

Two of 20 viruses were found to be reassortants. Within virus sequences from different rodents, the average rate of synonymous substitutions among all pair-wise comparisons (π_s) was 0.378 in the M segment and 0.312 in the S segment sequences. The replacement substitution rate (π_a) was 7.0 × 10^-4 in the M segment and 17.3 × 10^-4 in the S segment sequences. The low π_a relative to π_s suggests strong purifying selection and this was confirmed by a Fu and Li analysis. The absolute rate of molecular evolution of the M segment was 6.76 × 10^-3 substitutions/site/year. The absolute age of the M segment tree was estimated to be 37 years. In the S segment the rate of molecular evolution was 1.93 × 10^-3 substitutions/site/year and the absolute age of the tree was 106 years. Assuming that mice were infected with a single Sin Nombre virus genotype, phylogenetic analyses revealed that 10% (2/20) of viruses were reassortants, similar to the 14% (6/43) found in a previous report.

Conclusion

Age estimates from both segments suggest that Sin Nombre virus has evolved within the past 37–106 years. The rates of evolutionary changes reported here suggest that Sin Nombre virus M and S segment reassortment occurs frequently in nature.     

White Blood Cell Counts as Risk Markers of Developing Metabolic Syndrome and Its Components in the Predimed Study

Background

The Metabolic Syndrome (MetS) is a cluster of metabolic abnormalities that includes hyperglucemia, hypertension, dyslipidemia and central obesity, conferring an increased risk of cardiovascular disease. The white blood cell (WBC) count has been proposed as a marker for predicting cardiovascular risk. However, few prospective studies have evaluated the relationship between WBC subtypes and risk of MetS.

Methods

Participants were recruited from seven PREDIMED study centers. Both a baseline cross-sectional (n = 4,377) and a prospective assessment (n = 1,637) were performed. Participants with MetS at baseline were excluded from the longitudinal analysis. The median follow-up was 3.9 years. Anthropometric measurements, blood pressure, fasting glucose, lipid profile and WBC counts were assessed at baseline and yearly during the follow-up. Participants were categorized by baseline WBC and its subtype count quartiles. Adjusted logistic regression models were fitted to assess the risk of MetS and its components.

Results

Of the 4,377 participants, 62.6% had MetS at baseline. Compared to the participants in the lowest baseline sex-adjusted quartile of WBC counts, those in the upper quartile showed an increased risk of having MetS (OR, 2.47; 95%CI, 2.03–2.99; P-trend<0.001). This association was also observed for all WBC subtypes, except for basophils. Compared to participants in the lowest quartile, those in the top quartile of leukocyte, neutrophil and lymphocyte count had an increased risk of MetS incidence. Leukocyte and neutrophil count were found to be strongly associated with the MetS components hypertriglyceridemia and low HDL-cholesterol. Likewise, lymphocyte counts were found to be associated with the incidence of the MetS components low HDL-cholesterol and high fasting glucose. An increase in the total WBC during the follow-up was also associated with an increased risk of MetS.

Conclusions

Total WBC counts, and some subtypes, were positively associated with MetS as well as hypertriglyceridemia, low HDL-cholesterol and high fasting glucose, all components of MetS.

Trial registration

Controlled-Trials.comISRCTN35739639.