Schematic of the evolutionary history among HA, NP, NA and M segments of H1N1 IAVs.

<p>The schematic view is based on the phylogenetic trees of avian, swine, and human origin H1N1 IAVs and the most recent common ancestor (MRCA) for (A) HA and NP gene segments and (B) NA and M gene segments. North American avian (light blue) and 2009 pandemic H1N1 viruses (pink) are highlighted.</p

The Genomic Contributions of Avian H1N1 Influenza A Viruses to the Evolution of Mammalian Strains

<div><p>Among the influenza A viruses (IAVs) in wild aquatic birds, only H1, H2, and H3 subtypes have caused epidemics in humans. H1N1 viruses of avian origin have also caused 3 of 5 pandemics. To understand the reappearance of H1N1 in the context of pandemic emergence, we investigated whether avian H1N1 IAVs have contributed to the evolution of human, swine, and 2009 pandemic H1N1 IAVs. On the basis of phylogenetic analysis, we concluded that the polymerase gene segments (especially PB2 and PA) circulating in North American avian H1N1 IAVs have been reintroduced to swine multiple times, resulting in different lineages that led to the emergence of the 2009 pandemic H1N1 IAVs. Moreover, the similar topologies of hemagglutinin and nucleoprotein and neuraminidase and matrix gene segments suggest that each surface glycoprotein coevolved with an internal gene segment within the H1N1 subtype. The genotype of avian H1N1 IAVs of Charadriiformes origin isolated in 2009 differs from that of avian H1N1 IAVs of Anseriformes origin. When the antigenic sites in the hemagglutinin of all 31 North American avian H1N1 IAVs were considered, 60%-80% of the amino acids at the antigenic sites were identical to those in 1918 and/or 2009 pandemic H1N1 viruses. Thus, although the pathogenicity of avian H1N1 IAVs could not be inferred from the phylogeny due to the small dataset, the evolutionary process within the H1N1 IAV subtype suggests that the circulation of H1N1 IAVs in wild birds poses a continuous threat for future influenza pandemics in humans.</p></div

Data_Sheet_1_Pneumococcal LytR Protein Is Required for the Surface Attachment of Both Capsular Polysaccharide and Teichoic Acids: Essential for Pneumococcal Virulence.doc

<p>The LytR-Cps-Psr family proteins are commonly present in Gram-positive bacteria, which have been shown to implicate in anchoring cell wall-related glycopolymers to the peptidoglycan. Here, we report the cellular function of SPD_1741 (LytR) in Streptococcus pneumoniae and its role in virulence of pneumococci. Pneumococcal ΔlytR mutants have been successfully constructed by replacing the lytR gene with erm cassette. The role of LytR in pneumococcal growth was determined by growth experiments, and surface accessibility of the LytR protein was analyzed using flow cytometry. Transmission electron microscopy (TEM) and immunoblotting were used to reveal the changes in capsular polysaccharide (CPS). Dot blot and ELISA were used to quantify the amount of teichoic acids (TAs). The contribution of LytR on bacterial virulence was assessed using in vitro phagocytosis assays and infection experiments. Compared to the wild-type strain, the ΔlytR mutant showed a defect in growth which merely grew to a maximal OD₆₂₀ of 0.2 in the liquid medium. The growth of the ΔlytR mutant could be restored by addition of recombinant ΔTM-LytR protein in culture medium in a dose-dependent manner. TEM results showed that the D39ΔlytR mutant was impaired in the surface attachment of CPS. Deletion of lytR gene also impaired the retention of TAs on the surface of pneumococci. The reduction of CPS and TAs on the pneumocccal cells were confirmed using Dot blot and ELISA assays. Compared to wild-type D39, the ΔlytR mutant was more susceptible to the phagocytosis. Animal studies showed that the ability to colonize the nasophaynx and virulence of pneumococci were affected by impairment of the lytR gene. Collectively, these results suggest that pneumococcal LytR is involved in anchoring both the CPS and TAs to cell wall, which is important for virulence of pneumococci.</p

Evolutionary relatedness of PB2 gene segments of H1N1 IAVs of avian, swine, and human origins isolated from North America and Eurasia.

<p>The taxa are colored based on their host and geography: red-North American avian isolates from the St. Jude repository, light blue-North American avian isolates, purple-Eurasian avian isolates, green- swine isolates from North America, grey- swine isolates from Eurasia (both Eurasian avian-like swine and Eurasian classical swine), dark blue-human isolates from North America; orange- human isolates from Eurasia, and pink- 2009 pandemic isolates. The nodes with spillover avian H1N1 IAVs are denoted with light blue arrows.</p

Expression of MDM4 in retinoblastoma.

<p><b>A</b>) Structure of the genomic organization of the 11 exons of human <i>MDM4</i>. The exons are to scale as shown but the introns are not to scale. (<b>B</b>) Schematic of the spliced <i>MDM4</i> mRNA that produces the full-length MDM4 protein. The location and identification of Affymetrix gene expression array probesets are shown in red and the location of the real time RT-PCR probe/primer set is shown in blue. (<b>C</b>) Boxplots of the Log₂ expression of each of the 5 probesets that uniquely mapped to <i>MDM4</i> for fetal retina, cell lines, xenografts and 52 primary human retinoblastomas. (<b>D</b>) Real time RT-PCR for MDM2 using Taqman probes as shown in (B) for fetal retina at gestational stage week 20 (FW20), control cell lines (NB1691 and U20S), retinoblastoma cell lines (Y79, Weri1, RB355), primary tumors (SJ33, SJ43, SJ45), and retinoblastoma xenografts (SJ39-X, SJ41-X, SJ42-X and MSK176). Values were normalized to the positive control (U2OS). (<b>E</b>) The same data as shown in (D) but normalized to fetal retina. Each bar is the mean and standard deviation of duplicate experiments. (<b>F</b>) The SAGE data from developing mouse retina shows <i>Mdm4</i> is expressed at significant levels in the developing mouse retina. The gray shaded box represents the limit of detection by SAGE (<2 normalized tags per sample). <i>Gapdh</i> and <i>Gpi1</i> are plotted as ubiquitiously expressed internal controls. (<b>G</b>) Immunoblot of MDM4 (green) protein in a subset of the samples analyzed by real time RT-PCR. Gapdh (red) was used as an internal reference for gel loading and the normalized values are presented below the black and white presentation of the blot for MDM4. The antibody was verified for specificity using a blocking peptide (not shown). Recombinant full length Flag-MDM4 protein was included as a positive control. Multiple bands were detected for MDM4 and were quantitated (below the black and white picture of the blot) and normalized to GAPDH and relative to U2OS cell line.</p

Effects of Ant-25 and Scr on miR-106b∼25 in the ipsilateral cortex 7 days after surgery.

<p>(<b>A</b>–<b>C</b>) <b>A</b>) miR-93, <b>B</b>) miR-106b and <b>C</b>) miR-25 levels in the ischemic cortex after ICV injection of 1 nmol, 2.5 nmol and 4 nmol of Ant-25 or Scr. (D) qRT-PCR measurement of the miR-106b∼25 cluster in the ischemic cortex after ICV injection of 2.5 nmol of Ant-25 or Scr. ^#<b>P</b><0.001 for 2.5 nmol compared to the rTMS group; ^*<b>P</b><0.001 for 2.5 nmol compared to the Scr group. ^##P<0.001 for 4 nmol compared to the rTMS group.</p

Expression changes of miR-25 in the ipsilateral cortex 7 days after surgery.

<p>(A) Electrophoresis of miR-25 and U6 on gel. (B) Relative expressions of miR-25 in different groups. ^#<b>P</b><0.001 compared to the Sham group; ^*<b>P</b><0.001 compared to the MCAO group.</p

Infarct volume assessed by TTC staining 1 day after the tMCAO.

<p>(A) Position of SVZ in the coronal section of brain. Areas imaged for immunofluorescence studies are indicated by box. (B) Coronal brain section stained by TTC 1 day after tMCAO. The white areas without deep red-staining indicate ischemic areas. SVZ, subventricular zone.</p

Neurobehavioral function was improved by rTMS after cerebral ischemia.

<p>NSSs were improved in MCAO rats treated with rTMS as compared with other groups. Data are presented as mean±SD. ^&P<0.001 versus Sham group. ^*P = 0.005 for MCAO group between day 7 and day 1. ^**P<0.001 for rTMS group between day 7 and day 1. ^#P = 0.019 between rTMS group and MCAO group 7 days after surgery.</p

Expression changes of p57 and PTEN in the ipsilateral cortex 7 days after surgery.

<p>(A) Electrophoresis of p57, PTEN and GAPDH on gel. (B) The ratio of the target genes to GAPDH in different groups. ^#<b>P</b> = 0.005 for p57 compared to the Sham group; ^#<b>P</b> = 0.004 for PTEN compared to the Sham group; ^*<b>P</b> = 0.007 for PTEN compared to MCAO group.</p