Carbohydrate hydrogels with stabilized phage particles for bacterial biosensing: bacterium diffusion studies

Bacteriophage particles have been reported as potentially useful in the development of diagnosis tools for pathogenic bacteria as they specifically recognize and lyse bacterial isolates thus confirming the presence of viable cells. One of the most representative microorganisms associated with health care services is the bacterium Pseudomonas aeruginosa, which alone is responsible for nearly 15 % of all nosocomial infections. In this context, structural and functional stabilization of phage particles within biopolymeric hydrogels, aiming at producing cheap (chromogenic) bacterial biosensing devices, has been the goal of a previous research effort. For this, a detailed knowledge of the bacterial diffusion profile into the hydrogel core, where the phage particles lie, is of utmost importance. In the present research effort, the bacterial diffusion process into the biopolymeric hydrogel core was mathematically described and the theoretical simulations duly compared with experimental results, allowing determination of the effective diffusion coefficients of P. aeruginosa in the agar and calcium alginate hydrogels tested.Financial support to Victor M. Balcao, via an Invited Research Scientist fellowship (FAPESP Ref. No. 2011/51077-8) by Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP, Sao Paulo, Brazil), is hereby gratefully acknowledged

Genetic vaccination of mice with plasmids encoding the NS1 non-structural protein from tick-borne encephalitis virus and dengue 2 virus.

Although there is a safe, inexpensive and efficacious vaccine against yellow fever, vaccination against other flavivirus diseases is less successful. There is no licensed vaccine against dengue fever and current vaccines against tick-borne encephalitis (TBE) and Japanese encephalitis are expensive and require several injections. Furthermore novel vaccines containing only virus envelope proteins may raise fears over antibody mediated enhancement (ADE) of disease. Here we report the successful use of genetic vaccination against TBE in an experimental animal model using a plasmid containing the coding sequence of a non-structural protein (NS1). Such vaccines would provide inexpensive protection against disease, without raising concerns over inducing ADE on subsequent exposure to heterotypic infectious virus. Attempts to generate chaemeric plasmids to protect against both TBE and dengue fever were less successful. Although these chaemeric plasmids directed the synthesis and secretion of the virus NS1 protein normally, no protection was observed against either TBE or dengue fever

X-ray emission studies of the valence band of nanodiamonds annealed at different temperatures

X-ray emission spectroscopy has been applied to examine the electronic structure of onion-like carbon (OLC) generated by the annealing treatment of nanodiamonds (ND). The C Kα spectra of OLC produced in the temperature range of 1600-1900 K were found to be markedly different from the spectrum of particles formed at 2140 K and to be characterized by better ordering of graphitic shells. The latter spectrum was shown to be very similar to the C Kα of polycrystalline graphite, while the former ones exhibited a significant increase of the high-energy maximum that might be caused by the defect structure of graphitic networks forming at the intermediate temperatures. The experimental spectra were compared with the theoretical spectra from quantum-chemical semiempirical AM1 calculation of several models: a fullerene molecule, C240, having icosahedral structure, a C240 molecule incorporating a greater number of nonhexagonal rings, and a holed structure formed by removing pentagons from the icosahedral molecule. The density of high-energy electronic states in the valence band of the graphitic cage was found to be practically invariant to a change in ring statistics but to significantly increase because of localization of electrons on the zigzag sites of a hole boundary

Topology and electronic structure of onion-like carbon and graphite/diamond nanocomposites

Annealing of nanodiamond at moderate temperature makes it possible to produce structures being intermediate in the carbon transformation from sp3 - to sp2-state (graphite/diamond nanocomposites) and onion-like carbon (OLC). Electron microscopy shows such structures involve cage shells with spacing close to graphite. X-ray emission spectroscopy has been applied to examine the electronic structure of OLC and graphite/diamond nanocomposites. The CKα-spectra of OLC produced in the temperature range of 1600-1900 K were found to be markedly different from the spectrum of particles formed at 2140 K and characterized by better ordering of graphitic shells. The latter spectrum was shown to be very Similar to the CKα- spectrum of polycrystalline graphite, while the former ones exhibited a significant increase of high-energy maximum that might be caused by the holed defect structure of graphitic networks forming at the intermediate annealing temperature. To interpret experimental spectra, the quantum-chemical semiempirical AM1 calculation of icosahedral C%40 cage and that with holed defects was carried out. The lack of at least 22% atoms in an internal carbon cage was found to be essential to provide an increase of density of high-energy electronic states similar to that observed in the spectrum of OLC produced at 1900 K

Topology and electronic structure of onion-like carbon and graphite/diamond nanocomposites

Annealing of nanodiamond at moderate temperature makes it possible to produce structures being intermediate in the carbon transformation from sp3 - to sp2-state (graphite/diamond nanocomposites) and onion-like carbon (OLC). Electron microscopy shows such structures involve cage shells with spacing close to graphite. X-ray emission spectroscopy has been applied to examine the electronic structure of OLC and graphite/diamond nanocomposites. The CKα-spectra of OLC produced in the temperature range of 1600-1900 K were found to be markedly different from the spectrum of particles formed at 2140 K and characterized by better ordering of graphitic shells. The latter spectrum was shown to be very Similar to the CKα- spectrum of polycrystalline graphite, while the former ones exhibited a significant increase of high-energy maximum that might be caused by the holed defect structure of graphitic networks forming at the intermediate annealing temperature. To interpret experimental spectra, the quantum-chemical semiempirical AM1 calculation of icosahedral C%40 cage and that with holed defects was carried out. The lack of at least 22% atoms in an internal carbon cage was found to be essential to provide an increase of density of high-energy electronic states similar to that observed in the spectrum of OLC produced at 1900 K

X-ray emission studies of the valence band of nanodiamonds annealed at different temperatures

X-ray emission spectroscopy has been applied to examine the electronic structure of onion-like carbon (OLC) generated by the annealing treatment of nanodiamonds (ND). The C Kα spectra of OLC produced in the temperature range of 1600-1900 K were found to be markedly different from the spectrum of particles formed at 2140 K and to be characterized by better ordering of graphitic shells. The latter spectrum was shown to be very similar to the C Kα of polycrystalline graphite, while the former ones exhibited a significant increase of the high-energy maximum that might be caused by the defect structure of graphitic networks forming at the intermediate temperatures. The experimental spectra were compared with the theoretical spectra from quantum-chemical semiempirical AM1 calculation of several models: a fullerene molecule, C240, having icosahedral structure, a C240 molecule incorporating a greater number of nonhexagonal rings, and a holed structure formed by removing pentagons from the icosahedral molecule. The density of high-energy electronic states in the valence band of the graphitic cage was found to be practically invariant to a change in ring statistics but to significantly increase because of localization of electrons on the zigzag sites of a hole boundary

Comparative structural and functional analysis of orthomyxovirus polymerase cap-snatching domains.

International audienceOrthomyxovirus Influenza A virus (IAV) heterotrimeric polymerase performs transcription of viral mRNAs by cap-snatching, which involves generation of capped primers by host pre-mRNA binding via the PB2 subunit cap-binding site and cleavage 10-13 nucleotides from the 5' cap by the PA subunit endonuclease. Thogotoviruses, tick-borne orthomyxoviruses that includes Thogoto (THOV), Dhori (DHOV) and Jos (JOSV) viruses, are thought to perform cap-snatching by cleaving directly after the cap and thus have no heterogeneous, host-derived sequences at the 5' extremity of their mRNAs. Based on recent work identifying the cap-binding and endonuclease domains in IAV polymerase, we determined the crystal structures of two THOV PB2 domains, the putative cap-binding and the so-called '627-domain', and the structures of the putative endonuclease domains (PA-Nter) of THOV and DHOV. Despite low sequence similarity, corresponding domains have the same fold confirming the overall architectural similarity of orthomyxovirus polymerases. However the putative Thogotovirus cap-snatching domains in PA and PB2 have non-conservative substitutions of key active site residues. Biochemical analysis confirms that, unlike the IAV domains, the THOV and DHOV PA-Nter domains do not bind divalent cations and have no endonuclease activity and the THOV central PB2 domain does not bind cap analogues. On the other hand, sequence analysis suggests that other, non-influenza, orthomyxoviruses, such as salmon anemia virus (isavirus) and Quaranfil virus likely conserve active cap-snatching domains correlating with the reported occurrence of heterogeneous, host-derived sequences at the 5' end of the mRNAs of these viruses. These results highlight the unusual nature of transcription initiation by Thogotoviruses