Urticalean rosids: circumscription, rosid ancestry, and phylogenetics based on rbcL, trnL-F, and ndhF sequences

To address the composition of the urticalean rosids, the relationships of the component families (maximally Cannabaceae, Cecropiaceae, Celtidaceae, Moraceae, Ulmaceae, and Urticaceae) and analyze evolution of morphological characters, we analyzed sequence variation for a large sampling of these families and various rosid outgroups using rbcL, trnL-F, and ndhF plastid regions. Urticalean rosids are derived out of a lineage including Barbeyaceae, Dirachmaceae, Elaeagnaceae, and Rhamnaceae, with Rosaceae less closely related; thus, they are imbedded within Rosales. Ulmaceae are the sister to all remaining families. Cannabaceae are derived out of a subclade of Celtidaceae; this expanded family should be called Cannabaceae. Cecropiaceae are derived within Urticaceae and are polyphyletic with Poikilospermum derived elsewhere within Urticaceae; this expanded family should be called Urticaceae. Monophyletic Moraceae are sister to this expanded Urticaceae. Support for these relationships comes from a number of morphological characters (floral sexuality, presence or absence of hypanthium, stamen type and dehiscence, pollen pore number, ovule position, and embryo alignment) and chromosome numbers. Most fruit types, in terms of ecological dispersal, are derived independently multiple times and are strongly correlated with habitat.     

Understanding mechanisms of novel gene expression in polyploids

Polyploidy has long been recognized as a prominent force shaping the evolution of eukaryotes, especially flowering plants. New phenotypes often arise with polyploid formation and can contribute to the success of polyploids in nature or their selection for use in agriculture. Although the causes of novel variation in polyploids are not well understood, they could involve changes in gene expression through increased variation in dosage-regulated gene expression, altered regulatory interactions, and rapid genetic and epigenetic changes. New research approaches are being used to study these mechanisms and the results should provide a more complete understanding of polyploidy.     

Treatment of experimental allergic encephalomyelitis (EAE) induced by guinea pig myelin basic protein epitope 72-85 with a human MBP(87-99) analogue and effects of cyclic peptides

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory and demyelinating disease of the central nervous system and is an animal model of multiple sclerosis (MS). In the present report, a linear analogue and a series of cyclic semi-mimetic peptides were designed and synthesized based on the human myelin basic protein (MBP(87-99)) epitope (Val87-His-Phe-Phe-Lys-Asn-Ile-Val-Thr-Pro-Arg-Thr-Pro90) and on Copolymer I (a mixture of random polymers of Ala, Gln, Lys and Tyr used to treat MS). These analogues were designed looking for suppressors of EAE induced by guinea pig MBP(72-85) epitope (Gln-Lys-Ser-Gln-Arg-Ser-Gln-Asp-Glu-Asn-Pro-Val) in Lewis rats. The linear analogue [Arg91,Ala96]MBP(87-99), in which Arg substitutes Lys91 and Ala substitutes Pro96, was found to be a strong inhibitor which when administered to Lewis rats together with the encephalitogenic agonist MBP(72-85) completely prevented the induction of EAE. In contrast, three N- and C-termini amide-linked cyclic semi-mimetic peptides, [cyclo-Phe-Arg-Asn-Ile-Val-Thr-Ala-Acp (1), cyclo-Phe-Ala-Arg-Gln-Acp (2), cyclo-Tyr-Ala-Lys-Gln-Acp (3)] as well as a Lys side chain and C-terminous cyclic semi mimetic peptide cyclo(Lys, Acp)-Phe-Lys-Asn-Ile-Val-Thr-Ala-Acp (4) which contain segments of MBP(87-99) or are constituted from immunophoric residues of copolymer 1, were ineffective in inducing or inhibiting EAE in Lewis rats. However co-injection of cyclic analogues with MBP(72-85) delayed the onset of EAE indicating a modulatory effect on the EAE activity of MBP(72-85). These findings suggest that molecule length, size of cyclic moiety and backbone conformation are important elements for immunogenic activity. Moreover blockade of MBP(72-85) induced EAE by the unrelated peptide [Arg91,Ala56]MBP(87-99) could indicate that the mechanism of inhibition is not due to binding competition but rather due to the delivery of a negative signal by the antagonist which overcomes the agonist response possibly through the activation of antigen specific regulatory T cells.     

Extracellular α-amylase from Thermus filiformis Ork A2: purification and biochemical characterization

An extracellular α-amylase produced by the thermophilic bacterium Thermus filiformis Ork A2 was purified from cell-free culture supernatant by ion exchange chromatography. The molecular mass was estimated to be 60 000 Da by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The enzyme was rich in both basic and hydrophobic amino acids, presenting the following NH₂-terminal amino acid sequence: Thr-Ala-Asp-Leu-Ile-Val-Lys-Ile-Asn-Phe. Amylolytic activity on soluble starch was optimal at pH 5.5–6.0 and 95°C, and the enzyme was stable in the pH range of 4.0–8.0. Calcium enhanced thermostability at temperatures above 80°C, increasing the half-life of activity to more than 8 h at 85°C, 80 min at 90°C, and 19 min at 95°C. Ethylenediaminetetraacetic acid (EDTA) inhibited amylase activity, the inhibition being reversed by the addition of calcium or strontium ions. The α-amylase was also inhibited by copper and mercuric ions, and p-chloromercuribenzoic acid, the latter being reversed in the presence of dithiothreitol. Dithiothreitol and β-mercaptoethanol activated the enzyme. The α-amylase exhibited Michaelis-Menten kinetics for starch, with a K _m of 5.0 mg·ml⁻¹ and k _cat/K _m of 5.2 × 10⁵ ml·mg⁻¹ s⁻¹. Similar values were obtained for amylose, amylopectin, and glycogen. The hydrolysis pattern was similar for maltooligosaccharides and polysaccharides, with maltose being the major hydrolysis product. Glucose and maltotriose were generated as secondary products, although glucose was produced in high levels after a 6-h digestion. To our knowledge this is the first report of the characterization of an α-amylase from a strain of the genus Thermus. Received: June 2, 1997 / Accepted: September 16, 1997     

Uranium(III) poly(pyrazol-1-yl)borate complexes

Reactions of the adduct UCl₃·(THF)_x (THF = tetrahydrofuran) with compounds of the type K[H_nBL_4?n], where L = pyrazole or 3,5-dimethylpyrazole, are presented. Based on the results obtained the two compounds UCl₂H₂BPz₂·THF and UCl₂HBPz₃·THF were isolated.     

Anatomy and photosynthetic parameters of roots and leaves of two shade-adapted orchids, Dichaea cogniauxiana Shltr. and Epidendrum secundum Jacq.

This study compares photosynthetic and structural features of Dichaea cogniauxiana and Epidendrum secundum leaves and roots. The diurnal titratable acidity fluctuations indicated crassulacean acid metabolism (CAM) in E. secundum leaves, associated with anatomical features like thick cuticle, large and vacuolated cells, and reduced stomata size and frequency. Roots of both species had chloroplasts in their cortical parenchyma. However, neither the roots nor D. cogniauxiana leaves did show tissue sap acidity fluctuations. This indicates C₃ metabolism in these organs. This lack of oscillation of organic acids in Epidendrum roots was at odds with a CAM-like ¹³C ratio, suggesting that in spite of active CO₂ fixation in roots during the day, the bulk of carbon is imported from the leaves. Roots of both species showed Fv/Fm, ΔF/Fm′, ETR values similar to reports from other non-foliar photosynthetic organs. Besides reducing root carbon cost, root photosynthesis may also be important by alleviating potential hypoxia, since water-saturated velamen severely impedes the gas exchange between radicular cortex.     

Detection and decay rates of prey and prey symbionts in the gut of a predator through metagenomics

DNA methods are useful to identify ingested prey items from the gut of predators, but reliable detection is hampered by low amounts of degraded DNA. PCR‐based methods can retrieve minute amounts of starting material but suffer from amplification biases and cross‐reactions with the predator and related species genomes. Here, we use PCR‐free direct shotgun sequencing of total DNA isolated from the gut of the harlequin ladybird Harmonia axyridis at five time points after feeding on a single pea aphid Acyrthosiphon pisum. Sequence reads were matched to three reference databases: Insecta mitogenomes of 587 species, including H. axyridis sequenced here; A. pisum nuclear genome scaffolds; and scaffolds and complete genomes of 13 potential bacterial symbionts. Immediately after feeding, multicopy mtDNA of A. pisum was detected in tens of reads, while hundreds of matches to nuclear scaffolds were detected. Aphid nuclear DNA and mtDNA decayed at similar rates (0.281 and 0.11 h^?1 respectively), and the detectability periods were 32.7 and 23.1 h. Metagenomic sequencing also revealed thousands of reads of the obligate Buchnera aphidicola and facultative Regiella insecticola aphid symbionts, which showed exponential decay rates significantly faster than aphid DNA (0.694 and 0.80 h^?1, respectively). However, the facultative aphid symbionts Hamiltonella defensa, Arsenophonus spp. and Serratia symbiotica showed an unexpected temporary increase in population size by 1–2 orders of magnitude in the predator guts before declining. Metagenomics is a powerful tool that can reveal complex relationships and the dynamics of interactions among predators, prey and their symbionts.     

Methodological Framework for World Health Organization Estimates of the Global Burden of Foodborne Disease

Background

The Foodborne Disease Burden Epidemiology Reference Group (FERG) was established in 2007 by the World Health Organization to estimate the global burden of foodborne diseases (FBDs). This paper describes the methodological framework developed by FERG''s Computational Task Force to transform epidemiological information into FBD burden estimates.

Methods and Findings

The global and regional burden of 31 FBDs was quantified, along with limited estimates for 5 other FBDs, using Disability-Adjusted Life Years in a hazard- and incidence-based approach. To accomplish this task, the following workflow was defined: outline of disease models and collection of epidemiological data; design and completion of a database template; development of an imputation model; identification of disability weights; probabilistic burden assessment; and estimating the proportion of the disease burden by each hazard that is attributable to exposure by food (i.e., source attribution). All computations were performed in R and the different functions were compiled in the R package ''FERG''. Traceability and transparency were ensured by sharing results and methods in an interactive way with all FERG members throughout the process.

Conclusions

We developed a comprehensive framework for estimating the global burden of FBDs, in which methodological simplicity and transparency were key elements. All the tools developed have been made available and can be translated into a user-friendly national toolkit for studying and monitoring food safety at the local level.     

Selective Cytotoxicity of 1,3,4-Thiadiazolium Mesoionic Derivatives on Hepatocarcinoma Cells (HepG2)

In this work, we evaluated the cytotoxicity of mesoionic 4-phenyl-5-(2-Y, 4-X or 4-X-cinnamoyl)-1,3,4-thiadiazolium-2-phenylamine chloride derivatives (MI-J: X=OH, Y=H; MI-D: X=NO₂, Y=H; MI-4F: X=F, Y=H; MI-2,4diF: X=Y=F) on human hepatocellular carcinoma (HepG2), and non-tumor cells (rat hepatocytes) for comparison. MI-J, M-4F and MI-2,4diF reduced HepG2 viability by ~ 50% at 25 μM after 24-h treatment, whereas MI-D required a 50 μM concentration, as shown by 3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide assays. The cytotoxicity was confirmed with lactate dehydrogenase assay, of which activity was increased by 55, 24 and 16% for MI-J, MI-4F and MI-2,4diF respectively (at 25 μM after 24 h). To identify the death pathway related to cytotoxicity, the HepG2 cells treated by mesoionic compounds were labeled with both annexin V and PI, and analyzed by flow cytometry. All compounds increased the number of doubly-stained cells at 25 μM after 24 h: by 76% for MI-J, 25% for MI-4F and MI-2,4diF, and 11% for MI-D. It was also verified that increased DNA fragmentation occurred upon MI-J, MI-4F and MI-2,4diF treatments (by 12%, 9% and 8%, respectively, at 25 μM after 24 h). These compounds were only weakly, or not at all, transported by the main multidrug transporters, P-glycoprotein, ABCG2 and MRP1, and were able to slightly inhibit their drug-transport activity. It may be concluded that 1,3,4-thiadiazolium compounds, especially the hydroxy derivative MI-J, constitute promising candidates for future investigations on in-vivo treatment of hepatocellular carcinoma.     

Clinical Profiles and Factors Associated with Death in Adults with Dengue Admitted to Intensive Care Units,Minas Gerais,Brazil

The purpose of our study was to describe the clinical profile of dengue-infected patients admitted to Brazilian intensive care units (ICU) and evaluate factors associated with death. A longitudinal, multicenter case series study was conducted with laboratory-confirmed dengue patients admitted to nine Brazilian ICUs situated in Minas Gerais state, southeastern Brazil from January 1, 2008, to December 31, 2013. Demographic, clinical and laboratory data; disease severity scores; and mortality were evaluated. A total of 97 patients were studied. The in-ICU and in-hospital mortality rates were 18.6% and 19.6%, respectively. Patients classified as having severe dengue according to current World Health Organization classifications showed an increased risk of death in a univariate analysis. Nonsurvivors were older, exhibited lower serum albumin concentrations and higher total leukocyte counts and serum creatinine levels. Other risk factors (vomiting, lethargy/restlessness, dyspnea/respiratory distress) were also associated with death in a univariate analysis. Multivariate analysis indicated that in-hospital mortality was significantly associated with Acute Physiology and Chronic Health Evaluation II and the Sequential Organ Failure Assessment score. The ICU and in-hospital mortality observed in this study were higher than values reported in similar studies. An increased frequency of ICU admission due to severe organ dysfunction, higher severity indices and scarcity of ICU beds may partially explain the higher mortality.