Preliminary Ground-Based Observation for the Soil Moisture Measurement Validation of ADEOS II-AMSR/AMSR-E

研究概要:AMSR-E(2002年3月打ち上げ予定)とAMSR (2002年11月打ち上げ予定)の土壌水分測定アルゴリズムの検証のために、モンゴル高原で地上検証用試験地の設定と検証のための準備観測(モニタリングと集中土壌水分・植物水分移動観測)を行った。試験地内の降雨分布および土壌水分と植物水分の分布には地域的な差異があることが分かると共に変化幅も検証に値するものであることが分かった

Dicer-2-Dependent Activation of <i>Culex</i> Vago Occurs via the TRAF-Rel2 Signaling Pathway

<div><p>Despite their importance as vectors of human and livestock diseases, relatively little is known about innate antiviral immune pathways in mosquitoes and other insects. Previous work has shown that <i>Culex</i> Vago (<i>Cx</i>Vago), which is induced and secreted from West Nile virus (WNV)-infected mosquito cells, acts as a functional homolog of interferon, by activating Jak-STAT pathway and limiting virus replication in neighbouring cells. Here we describe the Dicer-2-dependent pathway leading to WNV-induced <i>Cx</i>Vago activation. Using a luciferase reporter assay, we show that a NF-κB-like binding site in <i>Cx</i>Vago promoter region is conserved in mosquito species and is responsible for induction of <i>Cx</i>Vago expression following WNV infection. Using dsRNA-based gene knockdown, we show that the NF-κB ortholog, Rel2, plays significant role in the signaling pathway that activates <i>Cx</i>Vago in mosquito cells <i>in vitro</i> and <i>in vivo</i>. Using similar approaches, we also show that TRAF, but not TRAF-3, is involved in activation of Rel2 after viral infection. Overall the study shows that a conserved signaling pathway, which is similar to mammalian interferon activation pathway, is responsible for the induction and antiviral activity of <i>Cx</i>Vago.</p></div

<i>Cx</i>Vago activation occurs via Dicer2-TRAF-Rel2 mediated pathway.

<p>Hsu cells were transfected with CxRel2 plasmid (with C-terminal V5 tag) and dsRNA against TRAF (<b>A</b>) or Dicer-2 (<b>B</b>). At 24 h post-transfection, the cells were infected with WNV and cell lysates was collected 48 hpi. Western blot was performed using anti-V5 and anti-beta actin antibodies. Representative blots shown here. (<b>C</b>) Female <i>Culex</i> mosquitoes were microinjected with dsRNAs against GFP, Rel2 or TRAF. After 48 h, the mosquitoes were infected with WNV (Kunjin strain). Total RNA was collected at 48 hpi and real-time RT-qPCR was performed using <i>Cx</i>Vago, <i>Cx</i>Rel2, <i>Cx</i>TRAF and WNV NS1 primers. Error bars represents standard error from experiment with assays performed in duplicates (Student's t-test *p<0.05 compared to dsRNA GFP).</p

The Rel-binding site is responsible for WNV-induced <i>Cx</i>Vago activation.

<p>(<b>A</b>) Hsu cells were transfected with <i>Renilla</i> luciferase plasmid containing Vago promoter region (∼2 kb region upstream of gene, Vago-Luc) or ∼1 kb promoter fragments (Vago1-989-Luc and Vago990-2007-Luc). Cells were also transfected with firefly luciferase plasmid under the insect promoter (<i>OpIE2</i>) as an internal control. As a positive control, cells were transfected with Renilla luciferase under OpIE2 promoter (OpIE2-Luc). After 24 h, the cells were infected with WNV and luciferase activity was measured at 24 hpi. (<b>B</b>) Hsu cells were transfected with <i>Renilla</i> luciferase plasmid containing Vago promoter (Vago-Luc) or smaller fragments of the promoter along with plasmid containing the firefly luciferase reporter (OpIE2 promoter, internal control). After 24 h, the cells were infected with WNV and luciferase activity was measured at 24 hpi. The schematics of the promoter region used in the assay is provided in the right panel. The solid promoter regions were involved in inducing luciferase expression after WNV infection. (<b>C</b>) Hsu cells were transfected with <i>Renilla</i> luciferase plasmid containing Vago promoter (Vago-Luc) or promoter with a mutated Rel-binding site (VagoRel-Luc) along with firefly luciferase plasmid (OpIE2 promoter, internal control). As a positive control, cells were transfected with Renilla luciferase under OpIE2 promoter (OpIE2-Luc). After 24 h, the cells were infected with WNV and luciferase activity was measured at 24 hpi. The <i>Renilla</i> luciferase activity values were standardised using firefly luciferase activity values (R/F) and resulting values were plotted as bar graphs. Error bars represents standard error from three separate experiments with assays performed in triplicate and values were compared to ‘Vago-Luc’ (Student's t-test *p<0.05).</p

<i>Cx</i>TRAF, but not <i>Cx</i>TRAF3, is required for activation of WNV-induced <i>Cx</i>Vago.

<p>Hsu cells were transfected with dsRNA against TRAF-3 (TRAF-3 dsRNA) or TRAF (TRAF dsRNA). GFP dsRNA was used as a silencing control. At 24 h post-transfection, the cells were infected with WNV and total RNA was collected 48 hpi. (<b>A</b>) Real-time RT-qPCR was performed on uninfected cells using TRAF-3- or TRAF-specific primers. RpL32 primers were used as internal control. (<b>B</b>) Real-time RT-qPCR was performed using <i>Cx</i>Vago primers with RpL32 as control. (<b>C</b>) Real-time RT-qPCR was performed using WNV NS1 primers with RpL32 as control. (<b>D</b>) Viral titer estimation by plaque assays was conducted on the supernatant media from these cells at 48 hpi. Error bars represents standard error from three separate experiments with assays performed in triplicate (Student's t-test *p<0.05, compared to GFP dsRNA).</p

Vago is up-regulated by flavivirus infection in mosquito cells.

<p>(<b>A</b>) Hsu (<i>Culex quinquefasciatus</i>) and RML12 (<i>Aedes albopictus</i>) cells were infected with WNV and DENV, respectively, and total RNA was collected at 48 hpi. Real-time RT-qPCR was performed using Vago-specific primers. RpL32 (ribosomal protein L32) primers were used as an internal control. Error bars represents standard error from three separate experiments with assays performed in triplicate (Student's t-test *p<0.05, comparing mRNA level between control and virus-infected cells). (<b>B</b>) Cells treated as in (A) were lysed to harvest total protein samples. Western blotting was performed using anti-Vago and anti-beta actin antibodies.</p

Atelier paludisme: an international malaria training course held in Madagascar-1

. 2003; 13 participants.<p><b>Copyright information:</b></p><p>Taken from "Atelier paludisme: an international malaria training course held in Madagascar"</p><p>http://www.malariajournal.com/content/7/1/80</p><p>Malaria Journal 2008;7():80-80.</p><p>Published online 9 May 2008</p><p>PMCID:PMC2394521.</p><p></p

<i>Cx</i>Rel2 is required for activation of WNV-induced <i>Cx</i>Vago.

<p>(<b>A</b>) Hsu cells were transfected with dsRNA against Rel2 (Rel2-dsRNA) or control dsRNA (GFP-dsRNA) along with plasmid containing the Vago-Luciferase reporter (and firefly luciferase control plasmid). At 24 h post-transfection, the cells were infected with WNV and luciferase activity was measured at 24 hpi. The <i>Renilla</i> luciferase activity values were standardised using firefly luciferase activity values (R/F) and resulting values were plotted as bar graphs. * indicates p<0.05 compared to GFP-dsRNA. (<b>B–D</b>) Hsu cells were transfected with dsRNA against Rel2 (Rel2-dsRNA) or control dsRNA (GFP-dsRNA). At 24 h post-transfection, the cells were infected with WNV and total RNA was collected 48 hpi. (<b>B</b>) Real-time RT-qPCR was conducted using primers for <i>Cx</i>Vago, <i>Cx</i>Rel2 and <i>Cx</i>RpL32 (control). * indicates p<0.05 compared to GFP-dsRNA. (<b>C</b>) Real-time RT-qPCR was conducted using primers for WNV-NS1. * indicates p<0.05 compared with WNV infection. (<b>D</b>) Viral titer estimation by plaque assays conducted on the supernatant media from these cells at 48 hpi. Error bars represents standard error from three separate experiments with assays performed in triplicate (Student's t-test *p<0.05 compared to GFP-dsRNA).</p

Atelier paludisme: an international malaria training course held in Madagascar-0

Ther participants and the panel, composed of tutors. These sessions allow the participants to demonstrate their acquisitions, by defending their approach, and aim to evaluate the work done during the week (Photo: courtesy of Dr Olivier Domarle, 2006).<p><b>Copyright information:</b></p><p>Taken from "Atelier paludisme: an international malaria training course held in Madagascar"</p><p>http://www.malariajournal.com/content/7/1/80</p><p>Malaria Journal 2008;7():80-80.</p><p>Published online 9 May 2008</p><p>PMCID:PMC2394521.</p><p></p

Additional file 1: of Assessment of ICount software, a precise and fast egg counting tool for the mosquito vector Aedes aegypti

ICount limitation, examples of low and high egg densities automatically counted with the software. Figure S1. Illustration with “Micro” pictures. Figure S2. Illustration with “Macro” pictures. (DOCX 5469 kb