West Nile Virus Detection in Urine

We report West Nile virus (WNV) RNA in urine collected from a patient with encephalitis 8 days after symptom onset. Viral RNA was detected by reverse transcriptase–polymerase chain reaction (RT-PCR). Sequence and phylogenetic analysis confirmed the PCR product to have ≥99% similarity to the WNV strain NY 2000-crow3356

Bacillus anthracis Protease InhA Increases Blood-Brain Barrier Permeability and Contributes to Cerebral Hemorrhages

Hemorrhagic meningitis is a fatal complication of anthrax, but its pathogenesis remains poorly understood. The present study examined the role of B. anthracis-secreted metalloprotease InhA on monolayer integrity and permeability of human brain microvasculature endothelial cells (HBMECs) which constitute the blood-brain barrier (BBB). Treatment of HBMECs with purified InhA resulted in a time-dependent decrease in trans-endothelial electrical resistance (TEER) accompanied by zonula occluden-1 (ZO-1) degradation. An InhA-expressing B. subtilis exhibited increased permeability of HBMECs, which did not occur with the isogenic inhA deletion mutant (ΔinhA) of B. anthracis, compared with the corresponding wild-type strain. Mice intravenously administered with purified InhA or nanoparticles-conjugated to InhA demonstrated a time-dependent Evans Blue dye extravasation, leptomeningeal thickening, leukocyte infiltration, and brain parenchymal distribution of InhA indicating BBB leakage and cerebral hemorrhage. Mice challenged with vegetative bacteria of the ΔinhA strain of B. anthracis exhibited a significant decrease in leptomeningeal thickening compared to the wildtype strain. Cumulatively, these findings indicate that InhA contributes to BBB disruption associated with anthrax meningitis through proteolytic attack on the endothelial tight junctional protein zonula occluden (ZO)-1

The 2MASS Redshift Survey - Description and Data Release

We present the results of the 2MASS Redshift Survey (2MRS), a ten-year project to map the full three-dimensional distribution of galaxies in the nearby Universe. The 2 Micron All-Sky Survey (2MASS) was completed in 2003 and its final data products, including an extended source catalog (XSC), are available on-line. The 2MASS XSC contains nearly a million galaxies with Ks <= 13.5 mag and is essentially complete and mostly unaffected by interstellar extinction and stellar confusion down to a galactic latitude of |b|=5 deg for bright galaxies. Near-infrared wavelengths are sensitive to the old stellar populations that dominate galaxy masses, making 2MASS an excellent starting point to study the distribution of matter in the nearby Universe. We selected a sample of 44,599 2MASS galaxies with Ks =5 deg (>= 8 deg towards the Galactic bulge) as the input catalog for our survey. We obtained spectroscopic observations for 11,000 galaxies and used previously-obtained velocities for the remainder of the sample to generate a redshift catalog that is 97.6% complete to well-defined limits and covers 91% of the sky. This provides an unprecedented census of galaxy (baryonic mass) concentrations within 300 Mpc. Earlier versions of our survey have been used in a number of publications that have studied the bulk motion of the Local Group, mapped the density and peculiar velocity fields out to 50 Mpc, detected galaxy groups, and estimated the values of several cosmological parameters. Additionally, we present morphological types for a nearly-complete sub-sample of 20,860 galaxies with Ks = 10 deg.Comment: Accepted for publication in The Astrophysical Journal Supplement Series. The 2MRS catalogs and a version of the paper with higher-resolution figures can be found at http://tdc-www.harvard.edu/2mrs

Bacillus anthracis Interacts with Plasmin(ogen) to Evade C3b-Dependent Innate Immunity

The causative agent of anthrax, Bacillus anthracis, is capable of circumventing the humoral and innate immune defense of the host and modulating the blood chemistry in circulation to initiate a productive infection. It has been shown that the pathogen employs a number of strategies against immune cells using secreted pathogenic factors such as toxins. However, interference of B. anthracis with the innate immune system through specific interaction of the spore surface with host proteins such as the complement system has heretofore attracted little attention. In order to assess the mechanisms by which B. anthracis evades the defense system, we employed a proteomic analysis to identify human serum proteins interacting with B. anthracis spores, and found that plasminogen (PLG) is a major surface-bound protein. PLG efficiently bound to spores in a lysine- and exosporium-dependent manner. We identified a-enolase and elongation factor tu as PLG receptors. PLG-bound spores were capable of exhibiting anti-opsonic properties by cleaving C3b molecules in vitro and in rabbit bronchoalveolar lavage fluid, resulting in a decrease in macrophage phagocytosis. Our findings represent a step forward in understanding the mechanisms involved in the evasion of innate immunity by B. anthracis through recruitmen

Western blot of TJ proteins in HBMECs after treatment with InhA.

<p>(A) HBMECs cells were treated with increasing concentrations of InhA (0.1, 0.3, 1, 3 µg/ml from left to right) for 24 h at 37°C. Western blots were probed with antibodies to ZO-1 (225 kDa), occludin (56 kDa), claudin-1 (22 KDa), and JAM-1 (39 kDa). (B) Densitometry of ZO-1 protein bands from (A). All samples analyzed (<i>n</i> = 4) were normalized to the intensity of corresponding β-actin bands. (C) Western blot analysis of time-dependent degradation of ZO-1 in HBMECs. Black arrows indicate two isoforms for ZO-1. <i>C</i>, control; <i>N</i>, Npr599; and <i>I</i>, InhA; α+ and α-, two splice variants of α domain. (D) Immunofluorescence of ZO-1 in HBMECs treated with or without 0.25 µg/ml of cytochalasin D for 1 h prior to incubation with varying concentrations of InhA at 37°C for 12 h. Panels show HBMECs treated as follows: untreated (a); InhA-treated (b); cytochalasin D-treated (c); InhA-treated after cytochalasin D treatment (d). Scale bar: 20 µm. (E) Western blot of His₆-tagged rZO-1. Degradation of purified rZO-1 after treatment with 0.1 µg/ml of InhA for indicated time at 37°C. Putative cleavage sites within the rZO-1fragment after treatment with InhA were deduced from immunoblot using the molecular masses of the cleavage products. SH3, Src homology 3; GK, guanylate kinase homolog; α, 80-amino-acid splice variant; -, acidic domain; PRK, proline-rich domain. β-Actin band intensity was used as a loading control for all Western blots. The results shown in (A)-(D) are representative of 4 independent experiments.</p

EB extravasation in the brains of InhA-challenged mice.

<p>Brain vascular permeability was measured using EB dye extravasation from whole mouse brains after challenge with InhA through retro-orbital injection. (A) Representative pictures of mouse brains at indicated time points. (B) Quantitation of the EB dye extravasation in the brain. The mean ± SD are presented as µg of EB dye per gram of brain (<i>n</i> = 5 in the InhA-injected groups; <i>n</i> = 3 in the control group). *<i>P</i><0.05 <i>vs.</i> control group.</p

InhA contributes to increased permeability of infected HBMEC monolayers.

<p>(A, B) Real-time TEER of HBMECs (about 6×10⁴ cells/well) treated with <i>ΔinhA</i> of <i>B. anthracis</i> Ames 35 (<i>ΔinhA</i>, red line in panel A) and InhA-expressing <i>B. subtilis</i> (<i>inhA+</i>, red line in panel B) at MOI of 10. Signals corresponding to parental strains (WT) and culture medium without bacteria are shown as blue and black lines, respectively. Arrows indicate time points when bacteria were added to the cells. (C) Transwell permeability assays of HBMECs infected with <i>B. anthracis</i> (<i>Ba</i>) and <i>B. subtilis</i> (<i>Bs</i>) strains as indicated above for 4 h. Leakage of FITC-dextran added to the monolayers for 2 h was determined by measuring fluorescence in the bottom chamber at 485/538 nm. *<i>P</i><0.01.</p

InhA contributes to BBB breakdown during <i>B. anthracis</i> infection in mice.

<p>Histopathology of representative H&E-stained brain sections from mice challenge as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0017921#pone-0017921-g006" target="_blank">Figure 6</a>. (A, B) Control mice; (C, D) Moribund mice challenged with <i>B. anthracis</i> Ames 35. Meningeal thickening is shown by double arrows. (E, F) Moribund mice infected with <i>B. anthracis ΔinhA</i>. Boxes in C, E show the areas enlarged in D, F, respectively.</p

InhA contributes to virulence of <i>B. anthracis</i> infection in mice.

<p>Animals were injected intravenously into the tail vein with indicated CFU of the early-log-phase vegetative bacteria of Ames 35 (WT) and <i>ΔinhA</i> strains, correspondingly. Mice were monitored twice daily for mortality and survival. (A) Kaplan–Meier survival curve (<i>n</i> = 5 in both groups). Control mice (<i>n</i> = 3) injected intravenously with PBS did not die (data not shown). Data were analyzed using the log-rank test. (B) Bacterial load in brains (left panel, <i>n</i> = 8) and blood (<i>n</i> = 5, right panel) in mice challenged with 5×10⁵ CFU as in (A) at 48 h post challenge. Bar in (B) represents median bacterial CFU.</p