Evolutionary implications of a rRNA-based phylogeny of Harpellales

The Harpellales (Trichomycetes) are endosymbiotic microfungi, mostly unculturable and predominantly associated with larval aquatic insects worldwide. Molecular phylogenies including ‘gut fungi’ have included at most only four axenic isolates of the 38 genera of Harpellales. Cladistic analyses were used to infer the phylogeny of the Harpellales using partial 18S or 28S nu-rRNA sequences generated for 16 genera of Harpellales, with 64 of 72 sequences generated from unculturable samples. Both analyses placed Orphella outside an otherwise monophyletic group of Harpellales, more closely allied to the Kickxellales. The current classification recognizing two families is not corroborated and continued use of the family Legeriomycetaceae may not be supportable. The largest genera of Harpellales, Smittium and Stachylina, were polyphyletic and the 28S rRNA sequences separate Smittium culisetae from the remainder of its genus. The cladograms did not support the consistent mapping of important morphological taxonomic characters, including trichospore shape and zygospore type or appendage numbers for both. This study demonstrates the use of microscopic thalli from host guts for molecular phylogenies and suggests the need for more data from the remaining Harpellales, especially with the future inclusion of protein-coding genes.     

Side chain oxygenated cholesterol regulates cellular cholesterol homeostasis through direct sterol-membrane interactions

Side chain oxysterols exert cholesterol homeostatic effects by suppression of sterol regulatory element-binding protein maturation and promoting degradation of hydroxymethylglutaryl-CoA reductase. To examine whether oxysterol-membrane interactions contribute to the regulation of cellular cholesterol homeostasis, we synthesized the enantiomer of 25-hydroxycholesterol. Using this unique oxysterol probe, we provide evidence that oxysterol regulation of cholesterol homeostatic responses is not mediated by enantiospecific oxysterol-protein interactions. We show that side chain oxysterols, but not steroid ring-modified oxysterols, exhibit membrane expansion behavior in phospholipid monolayers and bilayers in vitro. This behavior is non-enantiospecific and is abrogated by increasing the saturation of phospholipid acyl chain constituents. Moreover, we extend these findings into cultured cells by showing that exposure to saturated fatty acids at concentrations that lead to endoplasmic reticulum membrane phospholipid remodeling inhibits oxysterol activity. These studies implicate oxysterol-membrane interactions in acute regulation of sterol homeostatic responses and provide new insights into the mechanism through which oxysterols regulate cellular cholesterol balance.     

Monitoring the Dissemination of the Broad-Host-Range Plasmid pB10 in Sediment Microcosms by Quantitative PCR

Studying the transfer of specific mobile genetic elements in complex environmental matrices remains difficult because suitable molecular tools are not yet available to back up classical culture-dependent approaches. In this report, we show that quantitative PCR could be used to monitor the dissemination of the broad-host-range plasmid pB10 in sediment microcosms. This approach lies in the differential measurement of the host and plasmid DNAs used to inoculate the microcosms, using a particular design of quantitative PCR primers/probes where we took advantage of the mosaic aspect of the bacterial genomes to achieve a highly specific quantitative PCR detection system.Despite the progress made in our understanding of the basic mechanisms involved in horizontal gene transfer (12) and our awareness of their implications in bacterial evolution (1, 5), the dynamics of the genetic phenomenon involved still need to be evaluated in environmental contexts. As a matter of fact, our current vision of gene transfer is limited to either transfer experiments carried out in vitro, when talking about the basic mechanisms involved, or retrospective analyses, when studying the spread of relevant genetic markers among environmental bacteria. In situ transfer experiments were also of great interest, but they reached some limitations since no convenient tools could avoid selection and detection difficulties (11, 15, 16). Classically, the quantification of gene transfer involves culture-based methods and selection, which present major drawbacks when dealing with environmental samples (11, 15): first, it is believed that less than 1% of the environmental bacteria are cultivable, and second, the transferred genes may present a narrow host range of expression, which restricts their proper selection. As a consequence, culture-based approaches often lead to an underestimation of the bacterial counts, therefore limiting our perception of the extent of gene transfer in complex environments (11). Lately, elegant alternatives making use of plasmids tagged with green fluorescent protein genes for the fluorescent detection of transconjugants in situ have been developed (4, 11). Despite a better evaluation of the transfer frequencies, these approaches are restricted to the detection of transconjugants expressing fluorescent proteins well, and they still require the genetic alteration of the element studied. Molecular techniques such as PCR and quantitative PCR (qPCR) also offer the advantage of being both culture independent and gene expression independent since they are based solely on the specific detection of a given nucleotide sequence (10). Nevertheless, because many DNA markers are shared by various genetic entities/genomes, molecular approaches have mostly been restricted to the quantification of redundant conserved sequences in microbial communities rather than being used to specifically monitor the fate of a given mobile genetic element in complex environmental matrices. In this report, we show that the dissemination of the broad-host-range plasmid pB10 can be monitored in sediment microcosms by using a qPCR-based approach if specific primers are carefully designed.     

Effects of Dietary Chromium (III) Picolinate on Growth Performance,Respiratory Rate,Plasma Variables,and Carcass Traits of Pigs Fed High-Fat Diets

We investigated the effects of supplemental chromium (Cr) as Cr (III) picolinate on pigs fed high-fat diets (HFD) in a 56-day experiment. Thirty-two crossbred pigs (9.6 kg) were allotted to four treatments with four blocks and two pigs/pen. Treatments included: (1) low-fat diet (fat < 3.5%; LFD) with no Cr, (2) HFD (fat > 30%) with no Cr, (3) HFD with 1,000 ppb Cr, and (4) HFD with 2,000 ppb Cr. Pigs fed HFD gained weight faster, consumed less, and had lower feed:gain (p < 0.05). Pigs fed HFD had higher respiration rates than pigs fed LFD on d 41 (p < 0.05). Plasma insulin on d 14 linearly decreased with Cr (p = 0.05). Plasma cholesterol concentrations were higher in the pigs fed HFD than those fed LFD, but were largely unaffected by supplemental Cr. Consumption of HFD resulted in greater carcass weight, perirenal fat, and backfat measures (p < 0.01) compared with the LFD group. Cr resulted in linear reductions of hot carcass weight (p = 0.08) and average backfat (p < 0.05). The effects of Cr on carcass fat measures were more pronounced in castrated males than in females. These results indicate that Cr attenuates some effects of a HFD, mainly body fat accretion of pigs, and especially in castrated pigs.     

Partitioning of Minimotifs Based on Function with Improved Prediction Accuracy

Background

Minimotifs are short contiguous peptide sequences in proteins that are known to have a function in at least one other protein. One of the principal limitations in minimotif prediction is that false positives limit the usefulness of this approach. As a step toward resolving this problem we have built, implemented, and tested a new data-driven algorithm that reduces false-positive predictions.

Methodology/Principal Findings

Certain domains and minimotifs are known to be strongly associated with a known cellular process or molecular function. Therefore, we hypothesized that by restricting minimotif predictions to those where the minimotif containing protein and target protein have a related cellular or molecular function, the prediction is more likely to be accurate. This filter was implemented in Minimotif Miner using function annotations from the Gene Ontology. We have also combined two filters that are based on entirely different principles and this combined filter has a better predictability than the individual components.

Conclusions/Significance

Testing these functional filters on known and random minimotifs has revealed that they are capable of separating true motifs from false positives. In particular, for the cellular function filter, the percentage of known minimotifs that are not removed by the filter is ∼4.6 times that of random minimotifs. For the molecular function filter this ratio is ∼2.9. These results, together with the comparison with the published frequency score filter, strongly suggest that the new filters differentiate true motifs from random background with good confidence. A combination of the function filters and the frequency score filter performs better than these two individual filters.     

Phase I study of safety and immunogenicity of an Escherichia coli-derived recombinant protective antigen (rPA) vaccine to prevent anthrax in adults

Background

The fatal disease caused by Bacillus anthracis is preventable with a prophylactic vaccine. The currently available anthrax vaccine requires a lengthy immunization schedule, and simpler and more immunogenic options for protection against anthrax are a priority for development. In this report we describe a phase I clinical trial testing the safety and immunogenicity of an anthrax vaccine using recombinant Escherichia coli-derived, B. anthracis protective antigen (rPA).

Methodology/Principal Findings

A total of 73 healthy adults ages 18–40 were enrolled and 67 received 2 injections separated by 4 weeks of either buffered saline placebo, or rPA formulated with or without 704 µg/ml Alhydrogel® adjuvant in increasing doses (5, 25, 50, 100 µg) of rPA. Participants were followed for one year and safety and immunologic data were assessed. Tenderness and warmth were the most common post-injection site reactions. No serious adverse events related to the vaccine were observed. The most robust humoral immune responses were observed in subjects receiving 50 µg of rPA formulated with Alhydrogel® with a geometric mean concentration of anti-rPA IgG antibodies of 283 µg/ml and a toxin neutralizing geometric 50% reciprocal geometric mean titer of 1061. The highest lymphoproliferative peak cellular response (median Lymphocyte Stimulation Index of 29) was observed in the group receiving 25 µg Alhydrogel®-formulated rPA.

Conclusions/Significance

The vaccine was safe, well tolerated and stimulated a robust humoral and cellular response after two doses.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00057525","term_id":"NCT00057525"}}NCT00057525     

Psorantin,a unique methylene linked dimer of vismin and kenganthranol E,two anthranoid derivatives from the fruits of Psorospermum aurantiacum (Hypericaceae)

Two prenylated anthranoids, psorantin and kenganthranol E, were isolated from the fruit of Psorospermum aurantiacum together with the known compounds ferruginin B, vismin, vismion D, haronginanthrone, kenganthranol B, kenganthraquinone, 1,7-dihydroxyxanthone, paradisiol, fridelin, fridelinol and betulinic acid. Their structures were determined on the basis of spectral data and by comparison with data reported in the literature as well as with authentic specimens for the known compounds. The structures of the new compounds were determined as bis-[3,8,9-trihydroxy-6-methyl-4,4-bis-(3,3-dimethylallyl)anthracenyl]methane (1) and 1,8,10-trihydroxy-6-methyl-4,5-bis-(3,3-dimethylallyl)-2,3-(2,2-dimethylpyrano)anthrone (2). Psorantin (1) is a dimer of vismin formed through a methylene linkage. The two new compounds when tested against the microbial strains Bacillus megaterium, Escherichia coli, Chlorella fusca and Microbotryum violaceum showed no activity.     

Dissecting the transcriptional phenotype of ribosomal protein deficiency: implications for Diamond-Blackfan Anemia

Defects in genes encoding ribosomal proteins cause Diamond Blackfan Anemia (DBA), a red cell aplasia often associated with physical abnormalities. Other bone marrow failure syndromes have been attributed to defects in ribosomal components but the link between erythropoiesis and the ribosome remains to be fully defined. Several lines of evidence suggest that defects in ribosome synthesis lead to “ribosomal stress” with p53 activation and either cell cycle arrest or induction of apoptosis. Pathways independent of p53 have also been proposed to play a role in DBA pathogenesis.     

Inheritance as Evolved and Evolving Physiological Processes

Acta Biotheoretica - In this paper, we adopt a physiological perspective in order to produce an intelligible overview of biological transmission in all its diversity. This allows us to put forward...