Genome sequence of the acid-tolerant Burkholderia sp. strain WSM2230 from Karijini National Park,Australia

Burkholderia sp. strain WSM2230 is an aerobic, motile, Gram-negative, non-spore-forming acid-tolerant rod isolated from acidic soil collected in 2001 from Karijini National Park, Western Australia, using Kennedia coccinea (Coral Vine) as a host. WSM2230 was initially effective in nitrogen-fixation with K. coccinea, but subsequently lost symbiotic competence. Here we describe the features of Burkholderia sp. strain WSM2230, together with genome sequence information and its annotation. The 6,309,801 bp high-quality-draft genome is arranged into 33 scaffolds of 33 contigs containing 5,590 protein-coding genes and 63 RNA-only encoding genes. The genome sequence of WSM2230 failed to identify nodulation genes and provides an explanation for the observed failure of the laboratory grown strain to nodulate. The genome of this strain is one of 100 sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.     

Supercritical carbon dioxide extraction of hydrocarbons from the microalgaBotryococcus braunii

Samples of the microalgaBotryococcus braunii were submitted to supercritical fluid extraction with carbon dioxide at 40 °C and pressures of 12.5, 20.0 and 30.0 MPa. The extraction yield and the fraction of the hydrocarbons in the extracts both increased with pressure and at 30 MPa these compounds were obtained rapidly. This behaviour is associated with the localization of the hydrocarbons outside the cell wall. In the extracts, which are fluid, golden and limpid, chlorophyll and phospholipids were not detected.Author for correspondence     

Pharmacokinetics of ergosterol in rats using rapid resolution liquid chromatography-atmospheric pressure chemical ionization multi-stage tandem mass spectrometry and rapid resolution liquid chromatography/tandem mass spectrometry

Rapid resolution liquid chromatography/tandem multi-stage mass spectrometry (RRLC-MS(n)) and rapid resolution liquid chromatography/tandem mass spectrometry (RRLC/MS/MS) methods were developed for the identification and quantification of ergosterol and its metabolites from rat plasma, urine and faeces. Two metabolites (ERG1 and ERG2) were identified by RRLC/MS(n). The concentrations of the ergosterol were determined by RRLC/MS/MS. The separation was performed on an Agilent Zorbax SB-C18 with the mobile phase consisting of methanol and water (containing 0.1% formic acid). The detection was carried out by means of atmospheric pressure chemical ionization mass spectrometry in positive ion mode with multiple reaction monitoring (MRM). Linear calibration curves were obtained in the concentration range of 7-2000, 6-2000 and 8-7500 ng/mL for plasma, urine and faecal homogenate, respectively. The intra- and inter-day precision values (RSD) were below 10%. The method was applied to the pharmacokinetic properties and elimination pathway of ergosterol in rats.     

Formulating a fluorogenic assay to evaluate S-adenosyl-L-methionine analogues as protein methyltransferase cofactors

Protein methyltransferases (PMTs) catalyze arginine and lysine methylation of diverse histone and nonhistone targets. These posttranslational modifications play essential roles in regulating multiple cellular events in an epigenetic manner. In the recent process of defining PMT targets, S-adenosyl-L-methionine (SAM) analogues have emerged as powerful small molecule probes to label and profile PMT targets. To examine efficiently the reactivity of PMTs and their variants on SAM analogues, we transformed a fluorogenic PMT assay into a ready high throughput screening (HTS) format. The reformulated fluorogenic assay is featured by its uncoupled but more robust character with the first step of accumulation of the commonly-shared reaction byproduct S-adenosyl-L-homocysteine (SAH), followed by SAH-hydrolase-mediated fluorogenic quantification. The HTS readiness and robustness of the assay were demonstrated by its excellent Z' values of 0.83-0.95 for the so-far-examined 8 human PMTs with SAM as a cofactor (PRMT1, PRMT3, CARM1, SUV39H2, SET7/9, SET8, G9a and GLP1). The fluorogenic assay was further implemented to screen the PMTs against five SAM analogues (allyl-SAM, propargyl-SAM, (E)-pent-2-en-4-ynyl-SAM (EnYn-SAM), (E)-hex-2-en-5-ynyl-SAM (Hey-SAM) and 4-propargyloxy-but-2-enyl-SAM (Pob-SAM)). Among the examined 8 × 5 pairs of PMTs and SAM analogues, native SUV39H2, G9a and GLP1 showed promiscuous activity on allyl-SAM. In contrast, the bulky SAM analogues, such as EnYn-SAM, Hey-SAM and Pob-SAM, are inert toward the panel of human PMTs. These findings therefore provide the useful structure-activity guidance to further evolve PMTs and SAM analogues for substrate labeling. The current assay format is ready to screen methyltransferase variants on structurally-diverse SAM analogues.     

Computational fluid dynamics modeling of mass-transfer behavior in a bioreactor for hairy root culture. II. Analysis of ultrasound-intensified process

Recently, cichoric acid production from hairy roots of Echinacea purpurea was significantly improved by ultrasound stimulation in an airlift bioreactor. In this article, the possible mechanism on ultrasound-intensified hairy root culture of E. purpurea in the bioreactor was elucidated with the help of computational fluid dynamics (CFD) simulation, membrane permeability detection, dissolved oxygen concentration detection, confocal laser-scanning microscopy (LSM) observation, and phenylalanine ammonium lyase (PAL) activity analysis. The CFD model developed in Part I was used to simulate the hydrodynamics and oxygen mass transfer in hairy root bioreactor culture stimulated by ultrasound. A dynamic mesh model combined with a changing Schmidt number method was used for the simulation of the ultrasound field. Simulation results and experimental data illustrated that ultrasound intensified oxygen mass transfer in the hairy root clump, which subsequently stimulated root growth and cichoric acid biosynthesis. Ultrasound increased the hairy root membrane permeability, and a high root membrane permeability of 0.359 h(-1) was observed at the bottom region in the bioreactor. LSM observation showed that the change in the membrane permeability recovered to normal in the further culture after ultrasound stimulation. PAL activity in the hairy roots was stimulated by ultrasound increase and was correlated well to cichoric acid accumulation in the hairy roots of E. purpurea.     

Beyond missing heritability: prediction of complex traits

Despite rapid advances in genomic technology, our ability to account for phenotypic variation using genetic information remains limited for many traits. This has unfortunately resulted in limited application of genetic data towards preventive and personalized medicine, one of the primary impetuses of genome-wide association studies. Recently, a large proportion of the "missing heritability" for human height was statistically explained by modeling thousands of single nucleotide polymorphisms concurrently. However, it is currently unclear how gains in explained genetic variance will translate to the prediction of yet-to-be observed phenotypes. Using data from the Framingham Heart Study, we explore the genomic prediction of human height in training and validation samples while varying the statistical approach used, the number of SNPs included in the model, the validation scheme, and the number of subjects used to train the model. In our training datasets, we are able to explain a large proportion of the variation in height (h(2) up to 0.83, R(2) up to 0.96). However, the proportion of variance accounted for in validation samples is much smaller (ranging from 0.15 to 0.36 depending on the degree of familial information used in the training dataset). While such R(2) values vastly exceed what has been previously reported using a reduced number of pre-selected markers (<0.10), given the heritability of the trait (～ 0.80), substantial room for improvement remains.     

ToRCH系列酶标诊断试剂的研制

报道了用辣根过氧化物酶标记的抗人ＩｇＧ和抗人ＩｇＭ（ｕ链）单克隆抗体作第二抗体,用自己培养、纯化的弓形体（Ｔｏ）、风疹病毒（ＲｕＶ）、巨细胞病毒（ＣＭＶ）和单纯疱疹病毒（ＨＳＶ１２）的虫体和病毒抗原包被酶标板,研制出检测ＴｏＲＣＨ系列的特异性ＩｇＧ和ＩｇＭ的间接ＥＬＩＳＡ试剂。质量检定结果表明,该试剂特异性强、本底低,能有效消除ＲＦ因子等干扰因素的影响;灵敏度达１∶１６０～６４０;精密性好,变异系数（Ｃ．Ｖ）在１．４％～９．０％;试剂稳定,３７℃存放４ｄ,各项指标的变化率不超过１５％。     

Neutralization of BDNF Attenuates the in vitro Protective Effects of Olfactory Ensheathing Cell-Conditioned Medium on Scratch-Insulted Retinal Ganglion Cells

Transplantation of olfactory ensheathing cells (OECs) becomes one of the promising strategies in restoring lost functions of injured central nervous system. Elevated level of expressed brain-derived neurotrophic factor (BDNF) was revealed in the previous studies to be related to the protective effects of OECs on injured cortical and brain stem neurons as well as retinal ganglion cells (RGCs), but no evidence has been obtained to demonstrate whether transplanted OECs protect injured central neurons directly by their secreted BDNF. In the present study, the effects of BDNF neutralization on the neuroprotection of adult OEC-conditioned medium (OEC-CM) on scratch-insulted RGCs were examined. The results showed that OEC-CM protected cultured RGCs from scratch insult, and neutralization of BDNF by BDNF neutralizing antibody attenuated such neuroprotection of the medium. It is thus concluded that neurotrophic factors including BDNF secreted by OECs can protect injured OECs in vitro and BDNF plays a major role in such a protection of OECs.     

Influenza Vaccine Induces Intracellular Immune Memory of Human NK Cells

Influenza vaccines elicit antigen-specific antibodies and immune memory to protect humans from infection with drift variants. However, what supports or limits vaccine efficacy and duration is unclear. Here, we vaccinated healthy volunteers with annual vaccine formulations and investigated the dynamics of T cell, natural killer (NK) cell and antibody responses upon restimulation with heterologous or homologous influenza virus strains. Influenza vaccines induced potential memory NK cells with increased antigen-specific recall IFN-γ responses during the first 6 months. In the absence of significant changes in other NK cell markers (CD45RO, NKp44, CXCR6, CD57, NKG2C, CCR7, CD62L and CD27), influenza vaccines induced memory NK cells with the distinct feature of intracellular NKp46 expression. Indeed, surface NKp46 was internalized, and the dynamic increase in NKp46(intracellular)⁺CD56^dim NK cells positively correlated with increased IFN-γ production to influenza virus restimulation after vaccination. In addition, anti-NKp46 antibodies blocked IFN-γ responses. These findings provide insights into a novel mechanism underlying vaccine-induced immunity and NK-related diseases, which may help to design persisting and universal vaccines in the future.