Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA-2

Alysis distance. The values above 2.0 represent statistically significant hot-spots while below -2.0 are statistically significant cold-spots. The majority of census block groups fall in the statistically insignificant middle categories.<p><b>Copyright information:</b></p><p>Taken from "Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA"</p><p>http://www.ij-healthgeographics.com/content/7/1/19</p><p>International Journal of Health Geographics 2008;7():19-19.</p><p>Published online 1 May 2008</p><p>PMCID:PMC2396613.</p><p></p

Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA-0

Alysis distance. The values above 2.0 represent statistically significant hot-spots while below -2.0 are statistically significant cold-spots. The majority of census block groups fall in the statistically insignificant middle categories.<p><b>Copyright information:</b></p><p>Taken from "Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA"</p><p>http://www.ij-healthgeographics.com/content/7/1/19</p><p>International Journal of Health Geographics 2008;7():19-19.</p><p>Published online 1 May 2008</p><p>PMCID:PMC2396613.</p><p></p

Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA-1

E while the WNV disease incidence rates were derived by dividing the number of cases in a county by the population and multiplying by one million to get the rate per million people.<p><b>Copyright information:</b></p><p>Taken from "Landscape, demographic, entomological, and climatic associations with human disease incidence of West Nile virus in the state of Iowa, USA"</p><p>http://www.ij-healthgeographics.com/content/7/1/19</p><p>International Journal of Health Geographics 2008;7():19-19.</p><p>Published online 1 May 2008</p><p>PMCID:PMC2396613.</p><p></p

Discovery and profiling of miRNAs in <i>B. malayi</i> exosome-like vesicles.

<p><b>(A) Comparative abundance of miRNAs in L3 ELV and tissue-derived samples</b>. miRNA discovery and abundance estimation was carried out using the mirDeep2 pipeline. The 20 miRNAs with highest expression in each sample were retained for comparison and abundance was normalized with respect to total miRNA-mapping reads within each sample. Normalized read count is plotted on a log scale for ELV and tissue miRNAs to provide a relative ordering of fractional abundance. Bma-let-7 only appears in the highly expressed subset, and a number of miRNAs with perfect mature sequence identity to host homologs are highlighted (outer blue circle). <b>(B) Sequence conservation between <i>B. malayi</i> ELV-origin miRNAs and the host ( <i>H. sapiens</i> ) miRNA complement</b>. Reduced heat map showing one-to-one homology between a given <i>B. malayi</i> miRNA and its nearest matching human counterpart in terms of percent identity. Bma-let-7, bma-miR-1, bma-miR-9, bma-miR-92, and bma-miR-100b (white asterisks) share 100% identity with a host miRNA, while bma-miR-34 shows high identity with a host miRNA (21/23 nucleotides). This <i>B. malayi</i> miRNA subset (shown in blue) contains potential modulators of host gene expression. <b>(C) Sequence conservation between <i>B. malayi</i> ELV-origin miRNA seed sites and host ( <i>H. sapiens</i> ) miRNA seed sites</b>. miRNAs sharing perfectly conserved seed sites, defined here as nucleotides 2–8 of the mature miRNA, are marked (blue circles).</p

Protein content of <i>B. malayi</i> exosome-like vesicles.

<p>GO functional annotation of 32 proteins identified in ELVs isolated from <i>B. malayi</i> L3 stage parasites.</p

<i>Brugia malayi</i> ELV miRNA sequence homology to nematode and mammalian host miRNAs.

<p>miRNAs from <i>B. malayi</i>, <i>A. suum</i>, <i>C. elegans</i>, <i>H. sapiens</i>, and <i>M. musculus</i> were grouped by seed site sequence identity (nucleotides 2–8) for multiple sequence alignments. Alignments are shown for bma-let-7, bma-miR-9 and bma-miR-993. bma-let-7 is shown as an example of a <i>Brugia</i> ELV miRNA that exhibits both seed site and full length sequence conservation extending to other parasitic and free-living nematodes, as well as to mammalian host species. bma-miR-9 and bma-miR-993 are presented as examples where conserved parasite miRNAs have clear host homologs, yet lack one-to-one <i>C. elegans</i> orthologs. The complete set of alignments can be found in <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0004069#pntd.0004069.s003" target="_blank">S1 Fig</a>.</p

Annotation of <i>Brugia</i> ELV proteome.

<p>Homology-based annotation of <i>B. malayi</i> ELV proteins reveals hallmarks of mammalian exosomes, including HSP70 and translation elongation factors. Ribosomal proteins, histones, ras-related proteins, cathepsins, ATP synthase subunits, and other homologs of identified <i>Brugia</i> ELV proteins have also been reported in exosomes derived from various cell types.</p><p>Annotation of <i>Brugia</i> ELV proteome.</p

<i>Brugia</i> exosome-like vesicles (ELVs) elicit a classically activated phenotype in J774A.1 macrophages.

<p>(A) J774A.1 (5 × 10⁵) were treated with approximately 4 × 10⁸ purified L3 stage ELVs, live L3 stage parasites (10 worms) or naïve culture media (control) and supernatents collected after 48 hr. The presence of 32 cytokines/chemokines was simultaneously assayed using the Milliplex MAP Mouse Cytokine/Chemokine kit (EDM Millipore) interfaced with a Bio-Plex System (Bio-Rad) utilizing Luminex xMAP technology (Luminex). The quantification of identified cytokines is presented. The cytokine profile generated by ELV treatment is consistent with a classically activated phenotype. (B) Cytokine response to ELV treatment is compared to LPS (200 ng/mL). The close correlation of responses indicates ELV treatment generates a classically activated phenotype. (C) J774A.1 (5 × 10⁵) were treated with high dose LPS (200 ng/mL), low dose LPS (0.003 ng/mL), ELV or naïve culture media (control) for 24 hr, supernatant collected and assayed for G-CSF using a Mouse G-CSF Quantikine ELISA kit (R&D Systems). The absence of response to low dose LPS suggests the classically activated response is not due to LPS-like contamination.</p

<i>Brugia</i> exosome-like vesicles (ELVs) are internalized by J774A.1 macrophages.

<p>(A and D) J774A.1 macrophages were labeled with PKH26 (red) and counterstained with DAPI (blue) to visualize nuclei. (B and E) <i>B. malayi</i> L3 stage ELVs were purified from a 24 hr parasite culture and labeled with PKH67 (green). 3 × 10⁵ J774A.1 were co-incubated with approximately 3 × 10⁷ labeled ELVs for 6 hrs at 37°C and washed repeatedly to remove unbound ELVs. Vesicles internalized by macrophages appear diffusely throughout cytoplasm and focused in discrete puncta associated with the cell membrane. (C and F) Merged images showing internalization of parasite ELVs. All images were acquired using a using a Leica TCS SP5 X Confocal/multiphoton microscope system with 20X (A-C) or 60X (D-F) objectives. Scale bars: 10 <i>μ</i>m (A-C) and 25 <i>μ</i>m (D-F).</p

Isolation of Small RNAs from larval and adult <i>B. malayi</i> ELV fractions.

<p>Bioanalyzer data are shown for RNAs isolated from L3, adult male, and adult female <i>Brugia</i> preparations. L3 ELVs contain significant amounts of small RNAs in the 25–200 nt range (25 and 200 nt reference peaks labeled), while adult male and female vesicle preparations yield fewer RNAs. Vesicle fractions were prepared from 300 L3 and 30 adults in 24 hr culture incubations. Despite the much higher total tissue amounts used in adult culture, we detect much higher levels of small RNAs in L3-derived ELVs.</p