Interlaboratory comparison of real-time polymerase chain reaction methods to detect Coxiella burnetii, the causative agent of Q fever

The bacterium Coxiella burnetii, which has a wide host range, causes Q fever. Infection with C burnetii can cause abortions, stillbirth, and the delivery of weak offspring in ruminants. Coxiella burnetii infection is zoonotic, and in human beings it can cause chronic, potentially fatal disease. Real-time polymerase chain reaction (PCR) is increasingly being used to detect the organism and to aid in diagnosis both in human and animal cases. Many different real-time PCR methods, which target different genes, have been described. To assess the comparability of the C. burnetii real-time PCR assays in use in different European laboratories, a panel of nucleic acid extracts was dispatched to 7 separate testing centers. The testing centers included laboratories from both human and animal health agencies. Each laboratory tested the samples using their in-house real-time PCR methods. The results of this comparison show that the most common target gene for real-time PCR assays is the IS1111 repeat element that is present in multiple copies in the C. burnetii genome. Many laboratories also use additional real-time PCR tests that target single-copy genes. The results of the current study demonstrate that the assays in use in the different laboratories are comparable, with general agreement of results for the panel of samples.

Investigation of the effect of the preparation method on the activity and stability of Au/CeZrO4 catalysts for the low temperature water gas shift reaction

The impact of the preparation method on the activity and stability of gold supported on ceria-zirconia low temperature water-gas shift (WGS) catalysts have been investigated. The influence of the gold deposition method, nature of the gold precursor, nature of the washing solution, drying method, Ce: Zr ratio of the support and sulfation of the support have been evaluated. The highest activity catalysts were obtained using a support with a Ce: Zr mole ratio 1: 1, HAuCl4 as the gold precursor deposited via deposition precipitation using sodium carbonate as the precipitation agent and the catalyst washed with water or 0.1 M NH4OH solution. In addition, the drying used was found to be critical with drying under vacuum at room temperature found to be most effective.

Rate acceleration of the Baylis-Hillman reaction in polar solvents (water and formamide). Dominant role of hydrogen bonding, not hydrophobic effects, is implicated

A substantial acceleration of the Baylis-Hillman reaction between cyclohexenone and benzaldehyde has been observed when the reaction is conducted in water. Several different amine catalysts were tested, and as with reactions conducted in the absence of solvent, 3-hydroxyquinuclidine was found to be the optimum catalyst in terms of rate. The reaction has been extended to other aldehyde electrophiles including pivaldehyde. Attempts to extend this work to acrylates was only partially successful as rapid hydrolysis of methyl and ethyl acrylates occurred under the base-catalyzed and water-promoted conditions. However, tert-butyl acrylates were sufficiently stable to couple with relatively reactive electrophiles. Further studies on the use of polar solvents revealed that formamide also provided significant acceleration and the use of 5 equiv of formamide (optimum amount) gave faster rates than reactions conducted in water. Using formamide, further acceleration was achieved in the presence of Yb(OTf)(3) (5 mol %). The scope of the new conditions was tested with a range of Michael acceptors and benzaldehyde and with a range of electrophiles and ethyl acrylate. The origin of the rate acceleration is discussed.

Evidence for a hypoxic fixation reaction leading to the induction of ssb and dsb in irradiated DNA

Purpose: To measure hypoxic chemical fixation processes of radiation damage in both isolated plasmid DNA and in GSH-depleted E. coli cells.

Pan-serotypic detection of foot-and-mouth disease virus using a minor groove binder probe reverse transcription polymerase chain reaction assay

A novel assay for the pan-serotypic detection of foot-and-mouth disease virus (FMDV) was designed using a 5' conjugated minor groove binder (MGB) probe real-time RT-PCR system. This assay targets the 3D region of the FMDV genome and is capable of detecting 20 copies of a transcribed RNA standard. The linear range of the test was eight logs from 2 x 10(1) to 2 x 10(8) copies and amplification time was approximately 2 h. Using a panel of 83 RNA samples from representative FMDV isolates, the diagnostic sensitivity of this test was shown to be equivalent to a TaqMan real-time RT-PCR that targets the 5' untranslated region of FMDV. Furthermore, the assay does not detect viruses causing similar clinical diseases in pigs such as swine vesicular disease virus and vesicular stomatitis virus, nor does it detect marine caliciviruses causing vesicular exanthema. The development of this assay provides a useful tool for the differential diagnosis of FMD, potentially for use in statutory or emergency testing programmes, or for detection of FMDV RNA in research applications. (C) 2011 Elsevier B.V. All rights reserved.

Enhancement Factor for Gas Absorption in a Finite Liquid Layer. Part 1: Instantaneous Reaction in a Liquid in Plug Flow

An approximate analysis of gas absorption with instantaneous reaction in a liquid layer of finite thickness in plug flow is presented. An approximate solution to the enhancement factor for the case of unequal diffusivities between the dissolved gas and the liquid reactant has been derived and validated by numerical simulation. Depending on the diffusivity ratio of the liquid reactant to the dissolved gas (?), the enhancement factor tends to be either lower or higher than the prediction of the classical enhancement factor equation based on the penetration theory (Ei,pen) at Fourier numbers typically larger than 0.1. An empirical correlation valid for all Fourier numbers is proposed to allow a quick estimation of the enhancement factor, which describes the prediction of the approximate solution and the simulation data with a relative error below 5?% under the investigated conditions (? = 0.34, Ei,pen = 21000).

Distinct functional domains of the Salmonella enterica WbaP transferase that is involved in the initiation reaction for synthesis of the O antigen subunit

WbaP is a membrane enzyme that initiates O antigen synthesis in Salmonella enterica by catalysing the transfer of galactose 1-phosphate (Gal-1-P) onto undecaprenyl phosphate (Und-P). WbaP possesses at least three predicted structural domains: an N-terminal region containing four transmembrane helices, a large central periplasmic loop, and a C-terminal domain containing the last transmembrane helix and a large cytoplasmic tail. In this work, we investigated the contribution of each region to WbaP function by constructing a series of mutant WbaP proteins and using them to complement O antigen synthesis in DeltawbaP mutants of S. enterica serovars Typhi and Typhimurium. Truncated forms of WbaP lacking the periplasmic loop exhibited altered chain-length distributions in O antigen polymerization, suggesting that this central domain is involved in modulating the chain-length distribution of the O polysaccharide. The N-terminal and periplasmic domains were dispensable for complementation of O antigen synthesis in vivo, suggesting that the C-terminal domain carries the sugar-phosphate transferase activity. However, despite the fact that they complemented the synthesis of O antigen in the DeltawbaP mutant in vivo, membrane extracts containing WbaP derivatives without the N-terminal domain failed to transfer radioactive Gal from UDP-Gal into a lipid-rich fraction. These results suggest that the N-terminal region of WbaP, which contains four transmembrane domains, is essential for the insertion or stability of the protein in the bacterial membrane. We propose that the domain structure of WbaP enables this protein not only to function in the transfer of Gal-1-P to Und-P but also to establish critical interactions with additional proteins required for the correct assembly of O antigen in S. enterica.

Dual template synthesis of a highly mesoporous SSZ-13 zeolite with improved stability in the methanol-to-olefins reaction

The dual template synthesis of zeolite SSZ-13 by use of trimethyl-adamantanammonium hydroxide and a diquaternary-ammonium mesoporogen induces considerable mesoporosity without impeding zeolite microporosity. The strongly improved accessibility of Bronsted sites in mesoporous SSZ-13 increases its stability during application as an acid catalyst in the methanol-to-olefins reaction.