Gamma-phage lysin PlyG sequence-based synthetic peptides coupled with Qdot-nanocrystals are useful for developing detection methods for Bacillus anthracis by using its surrogates, B. anthracis-Sterne and B. cereus-4342

<p>Abstract</p> <p>Background</p> <p>Previous reports of site-directed deletion analysis on gamma (γ)-phage lysin protein (PlyG) have demonstrated that removal of a short amino acid sequence in the C-terminal region encompassing a 10-amino acid motif (190LKMTADFILQ199) abrogates its binding activity specific to the cell wall of <it>Bacillus anthracis</it>. Whether short synthetic peptides representing the10-amino acid PlyG putative binding motif flanked by surrounding N- and C-terminal residues also selectively bind to the bacterial cell wall has not been evaluated. If such peptides do demonstrate selective binding to the cell wall, they could serve as bio-probes towards developing detection technologies for <it>B. anthracis</it>. Furthermore, by using <it>B. anthracis </it>(Sterne, 34F2), an animal vaccine and <it>B. cereus</it>-<it>4342</it>, a γ-phage susceptible rare strain as surrogates of <it>B. anthracis</it>, development of proof-of-concepts for <it>B. anthracis </it>are feasible.</p> <p>Results</p> <p>Using four different methods, we evaluated six synthetic peptides representing the putative binding motif including flanking sequences (PlyG-P1 through P6) for the bacterial cell wall binding capacity. Our analysis identified PlyG-P1, PlyG-P3 and PlyG-P5 to have binding capability to both <it>B. anthracis </it>(Sterne, 34F2) and <it>B. cereus-</it>4342. The peptides however did not bind to <it>B. cereus</it>-11778, <it>B. thuringiensis</it>, and <it>B. cereus</it>-10876 suggesting their specificity for <it>B. anthracis</it>-Sterne and <it>B. cereus</it>-<it>4342</it>. PlyG-P3 in combination with fluorescent light microscopy detected even a single bacterium in plasma spiked with the bacteria.</p> <p>Conclusion</p> <p>Overall, these studies illustrate that the short 10-amino acid sequence 'LKMTADFILQ' in fact is a stand-alone bacterial cell wall-binding motif of PlyG. In principle, synthetic peptides PlyG-P1, PlyG-P3 and PlyG-P5, especially PlyG-P3 coupled with Qdot-nanocrystals are useful as high-sensitivity bio-probes in developing detection technologies for <it>B. anthracis</it>.</p

A tracked approach for automated NMR assignments in proteins (TATAPRO)

A novel automated approach for the sequence specific NMR assignments of 1HN, 13C&#945;, 13C&#946;, 13C'/1H&#945; and 15N spins in proteins, using triple resonance experimental data, is presented. The algorithm, TATAPRO (Tracked AuTomated Assignments in Proteins) utilizes the protein primary sequence and peak lists from a set of triple resonance spectra which correlate 1HN and 15N chemical shifts with those of 13C&#945;, 13C&#946; and 13C'/1H&#945;. The information derived from such correlations is used to create a 'master list' consisting of all possible sets of 1HN i, 15Ni, 13C&#945; i, 13C&#946; i, 13C'i/1H&#945; i, 13C&#945; i-1, 13C&#946; i-1 and 13C'i-1/ 1H&#945; i-1 chemical shifts. On the basis of an extensive statistical analysis of 13C&#945; and 13C&#946; chemical shift data of proteins derived from the BioMagResBank (BMRB), it is shown that the 20 amino acid residues can be grouped into eight distinct categories, each of which is assigned a unique two-digit code. Such a code is used to tag individual sets of chemical shifts in the master list and also to translate the protein primary sequence into an array called pps array. The program then uses the master list to search for neighbouring partners of a given amino acid residue along the polypeptide chain and sequentially assigns a maximum possible stretch of residues on either side. While doing so, each assigned residue is tracked in an array called assig array, with the two-digit code assigned earlier. The assig_array is then mapped onto the pps array for sequence specific resonance assignment. The program has been tested using experimental data on a calcium binding protein from Entamoeba histolytica (Eh-CaBP, 15 kDa) having substantial internal sequence homology and using published data on four other proteins in the molecular weight range of 18-42 kDa. In all the cases, nearly complete sequence specific resonance assignments (&gt; 95%) are obtained. Furthermore, the reliability of the program has been tested by deleting sets of chemical shifts randomly from the master list created for the test proteins

Identification of XMRV Infection-Associated microRNAs in Four Cell Types in Culture

INTRODUCTION: XMRV is a gammaretrovirus that was thought to be associated with prostate cancer (PC) and chronic fatigue syndrome (CFS) in humans until recently. The virus is culturable in various cells of human origin like the lymphocytes, NK cells, neuronal cells, and prostate cell lines. MicroRNAs (miRNA), which regulate gene expression, were so far not identified in cells infected with XMRV in culture. METHODS: Two prostate cell lines (LNCaP and DU145) and two primary cells, Peripheral Blood Lymphocytes [PBL] and Monocyte-derived Macrophages [MDM] were infected with XMRV. Total mRNA was extracted from mock- and virus-infected cells at 6, 24 and 48 hours post infection and evaluated for microRNA profile in a microarray. RESULTS: MicroRNA expression profiles of XMRV-infected continuous prostate cancer cell lines differ from that of virus-infected primary cells (PBL and MDMs). miR-193a-3p and miRPlus-E1245 observed to be specific to XMRV infection in all 4 cell types. While miR-193a-3p levels were down regulated miRPlus-E1245 on the other hand exhibited varied expression profile between the 4 cell types. DISCUSSION: The present study clearly demonstrates that cellular microRNAs are expressed during XMRV infection of human cells and this is the first report demonstrating the regulation of miR193a-3p and miRPlus-E1245 during XMRV infection in four different human cell types

Performance of PCA3 and TMPRSS2:ERG urinary biomarkers in prediction of biopsy outcome in the Canary Prostate Active Surveillance Study (PASS).

BackgroundFor men on active surveillance for prostate cancer, biomarkers may improve prediction of reclassification to higher grade or volume cancer. This study examined the association of urinary PCA3 and TMPRSS2:ERG (T2:ERG) with biopsy-based reclassification.MethodsUrine was collected at baseline, 6, 12, and 24 months in the multi-institutional Canary Prostate Active Surveillance Study (PASS), and PCA3 and T2:ERG levels were quantitated. Reclassification was an increase in Gleason score or ratio of biopsy cores with cancer to ≥34%. The association of biomarker scores, adjusted for common clinical variables, with short- and long-term reclassification was evaluated. Discriminatory capacity of models with clinical variables alone or with biomarkers was assessed using receiver operating characteristic (ROC) curves and decision curve analysis (DCA).ResultsSeven hundred and eighty-two men contributed 2069 urine specimens. After adjusting for PSA, prostate size, and ratio of biopsy cores with cancer, PCA3 but not T2:ERG was associated with short-term reclassification at the first surveillance biopsy (OR = 1.3; 95% CI 1.0-1.7, p = 0.02). The addition of PCA3 to a model with clinical variables improved area under the curve from 0.743 to 0.753 and increased net benefit minimally. After adjusting for clinical variables, neither marker nor marker kinetics was associated with time to reclassification in subsequent biopsies.ConclusionsPCA3 but not T2:ERG was associated with cancer reclassification in the first surveillance biopsy but has negligible improvement over clinical variables alone in ROC or DCA analyses. Neither marker was associated with reclassification in subsequent biopsies

Turbulent Diffusion and Turbulent Thermal Diffusion of Aerosols in Stratified Atmospheric Flows

The paper analyzes the phenomenon of turbulent thermal diffusion in the Earth atmosphere, its relation to the turbulent diffusion and its potential impact on aerosol distribution. This phenomenon was predicted theoretically more than 10 years ago and detected recently in the laboratory experiments. This effect causes a non-diffusive flux of aerosols in the direction of the heat flux and results in formation of long-living aerosol layers in the vicinity of temperature inversions. We demonstrated that the theory of turbulent thermal diffusion explains the GOMOS aerosol observations near the tropopause (i.e., the observed shape of aerosol vertical profiles with elevated concentrations located almost symmetrically with respect to temperature profile). In combination with the derived expression for the dependence of the turbulent thermal diffusion ratio on the turbulent diffusion, these measurements yield an independent method for determining the coefficient of turbulent diffusion at the tropopause. We evaluated the impact of turbulent thermal diffusion to the lower-troposphere vertical profiles of aerosol concentration by means of numerical dispersion modelling, and found a regular upward forcing of aerosols with coarse particles affected stronger than fine aerosols.Comment: 19 pages, 10 figure

Evidence for Smectite Clays from MSL SAM Analyses of Mudstone at Yellowknife Bay, Gale Crater, Mars

Drilled samples of mudstone from the Sheepbed unit at Yellowknife Bay were analyzed by MSL instruments including the Sample Analysis at Mars (SAM) and Chemistry and Mineralogy (CheMin) instruments in MSL's Analytical Laboratory. CheMin analyses revealed the first in situ X-ray diffraction based evidence of clay minerals on Mars, which are likely trioctahedral smectites (e.g., saponite) and comprise approx 20% of the mudstone sample (e.g., Bristow et al., this meeting). SAM analyses, which heated the mudstone samples to 1000 C and monitored volatiles evolved to perform in situ evolved gas analysis mass spectrometry (EGA-MS), resulted in a H2O trace exhibiting a wide evolution at temperatures < 500 C, and an evolution peak at higher temperatures near approx 750 C. The low temperature H2O evolution has many potential contributors, including adsorbed H2O, smectite interlayer H2O, and structural H2O/OH from bassanite and akaganeite (identified by CheMin) and H2O/OH from amorphous phases in the sample. The high temperature H2O is consistent with the evolution of H2O from the dehydroxylation of the smectite clay mineral. Comparison to EGA-MS data collected under SAM-like conditions on a variety of clay mineral reference materials indicate that a trioctahedral smectite, such as saponite, is most consistent with the high temperature H2O evolution observed. There may also be SAM EGA-MS evidence for a small high temperature H2O evolution from scoop samples from the Yellowknife Bay Rocknest sand shadow bedform. As in the mudstone samples, this evolution may indicate the detection of smectite clays, and the idea that minor clays may be present in Rocknest materials that could be expected to be at least partially derived from local sources is reasonable. But, because smectite clays were not definitively observed in CheMin analyses of Rocknest materials, they must be present at much lower abundances than the approx 20% observed in the mudstone samples. This potential detection underscores the complementary nature of the MSL CheMin and SAM instruments for investigations of martian sample mineralogy. Information on the nature of Yellowknife Bay clay minerals may also be available from the detection of H2 evolved during SAM EGA-MS at high temperature. A likely source of at least some of this H2 is H2O evolved from the smectite clays at high temperature, and it is possible these evolutions can be used in a similar fashion to high temperature H2O releases to provide constraints on the clay minerals in a sample. In addition, the D/H of this high temperature H2, as well as the H2O, can be derived from SAM MS and Tunable Laser Spectrometer (TLS) data, respectively. These D/H values may help to inform the provenance of high and low temperature water evolved from martian sample

Vertical structure and size distributions of Martian aerosols from solar occultation measurements

Solar occultations performed with a spectrometer on board the Soviet spacecraft Phobos 2 (Blamont et al. 1991) provided data on the vertical structure of the Martian aerosols in the equatorial region (0[deg]-20[deg] N latitude) near the northern spring equinox (LS = 0[deg]-20[deg]). All measurements were made close to the evening terminator. Five clouds were detected above 45 km altitude and their vertical structure recorded at six wavelengths between 0.28 and 3.7 [mu]m. They have a small vertical extent (3-6 km) and a vertical optical depth less than 0.03. The thermal structure, as derived from saturated profiles of water vapor observed by our instrument in the infrared, does not allow the CO2 frost point to be reached at cloud altitude, strongly suggesting that cloud particles are formed of H2O ice. Under the assumption of spherical particles, a precise determination of their effective radius, which varies from cloud to cloud and with altitude, is obtained and ranges from 0.15 to 0.85 [mu]m; an estimate of the effective variance of the particle size distribution is ~ 0.2. The number density of cloud particles at the peak extinction level is ~1 cm-3. Dust was also observed and monitored at two wavelengths, 1.9 and 3.7 [mu]m, on nine different occasions. The top of the dust opaque layer, defined as the level above which the atmosphere becomes nearly transparent at the wavelengths of observation, is located near 25 km altitude, with variations smaller than +/-3 km from place to place. The scale height of dust at this altitude is 3-4 km. The effective radius of dust particles near the top of the opaque layer is 0.95 +/- 0.25 [mu]m and increases below with a vertical gradient of ~0.05 [mu]m km-1. Assuming that particles are levitated by eddy mixing, the eddy diffusion coefficient, K, is found to be ~106 cm2 sec-1 at 25 km and 105-106 cm2 sec-1 at 50 km using, respectively, dust and cloud observations. An effective variance of 0.25 (+/-50%) for the dust size distribution is obtained on the basis of a simple theoretical model for the observed vertical gradient of the effective radius of dust particles. Three clouds observed by Viking at midlatitude during the northern summer are reanalyzed. The analysis gives K [approximate] 106 cm2 sec-1 below 50 km altitude and at least 107 cm2 sec-1 above. Since the clouds seen from Phobos 2 are observed at twilight, which coincides with the diurnal maximum of the ambient temperature, they can be assumed to be in a steady state. If their thermodynamic state were to vary quickly during the day, our optical thickness at twilight would correspond to unrealistic values in earlier hours when the temperature is lower. Clouds are well fitted by theoretical profiles obtained assuming the steady state. An atmospheric temperature of 165-170 K at ~50 km is inferred. The negative temperature gradient above the cloud is large (1.5-2 K km-1). A parallel is established between these thin clouds and the polar mesospheric clouds observed on Earth. It is shown that upwelling in equatorial regions at equinox could be a significant factor in levitating cloud particles.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/30061/1/0000431.pd

The Atmospheric Chemistry Suite (ACS) of Three Spectrometers for the ExoMars 2016 Trace Gas Orbiter

The Atmospheric Chemistry Suite (ACS) package is an element of the Russian contribution to the ESA-Roscosmos ExoMars 2016 Trace Gas Orbiter (TGO) mission. ACS consists of three separate infrared spectrometers, sharing common mechanical, electrical, and thermal interfaces. This ensemble of spectrometers has been designed and developed in response to the Trace Gas Orbiter mission objectives that specifically address the requirement of high sensitivity instruments to enable the unambiguous detection of trace gases of potential geophysical or biological interest. For this reason, ACS embarks a set of instruments achieving simultaneously very high accuracy (ppt level), very high resolving power (>10,000) and large spectral coverage (0.7 to 17 μm—the visible to thermal infrared range). The near-infrared (NIR) channel is a versatile spectrometer covering the 0.7–1.6 μm spectral range with a resolving power of ∼20,000. NIR employs the combination of an echelle grating with an AOTF (Acousto-Optical Tunable Filter) as diffraction order selector. This channel will be mainly operated in solar occultation and nadir, and can also perform limb observations. The scientific goals of NIR are the measurements of water vapor, aerosols, and dayside or night side airglows. The mid-infrared (MIR) channel is a cross-dispersion echelle instrument dedicated to solar occultation measurements in the 2.2–4.4 μm range. MIR achieves a resolving power of >50,000. It has been designed to accomplish the most sensitive measurements ever of the trace gases present in the Martian atmosphere. The thermal-infrared channel (TIRVIM) is a 2-inch double pendulum Fourier-transform spectrometer encompassing the spectral range of 1.7–17 μm with apodized resolution varying from 0.2 to 1.3 cm−1. TIRVIM is primarily dedicated to profiling temperature from the surface up to ∼60 km and to monitor aerosol abundance in nadir. TIRVIM also has a limb and solar occultation capability. The technical concept of the instrument, its accommodation on the spacecraft, the optical designs as well as some of the calibrations, and the expected performances for its three channels are described