Exclusion of deceased donors post-procurement of tissues

The EU Tissues and Cells Directive (2004/23/EC, 2006/17/EC, 2006/86/EC) (EUTCD) provides standards for quality and safety for all aspects of banking of tissues and cells for clinical applications. Commission Directive 2006/17/EC stipulates that the complete donor record with all the medical information is assessed for suitability before releasing tissues for clinical use. The aim of this study was to investigate the medical reasons for post-procurement donor exclusion, to identify the various potential sources for gathering information about donors’ medical and behavioural history and to evaluate their contribution to maximising the safety of donations. Information was collected from the Tissue Services (TS) records of 1000 consecutive deceased donors submitted to National Health Service Blood and Transplant (NHSBT) medical officers for authorisation for release for subsequent tissue processing and then for transplantation. Of the 1000 donors 60 (6%) were excluded because they did not fulfil the donor selection requirements of the EUTCD and NHSBT donor selection guidelines. The main reasons for medical exclusion were the presence of significant local or systemic infection in 32 donors (53% of those excluded for medical reasons) and a history of past or occult malignancy in 9 donors (15% of those excluded for medical reasons) which was not identified prior to procurement. The information leading to post-procurement exclusion was obtained from autopsy reports in 35 of the 60 excluded donors for medical reasons (58%) and from the general practitioner for 10 donors (17% of those excluded for medical reasons). In summary, careful evaluation of complete donor records reduces the potential risk of disease transmission by tissue allografts and ensures compliance with regulations and guidelines. The findings may lead to changes in donor selection policies with the aim of improving efficiency without compromising safety.     

N-Linked Glycosylation of the Hemagglutinin Protein Influences Virulence and Antigenicity of the 1918 Pandemic and Seasonal H1N1 Influenza A Viruses

The hemagglutinin (HA) protein is a major virulence determinant for the 1918 pandemic influenza virus; however, it encodes no known virulence-associated determinants. In comparison to seasonal influenza viruses of lesser virulence, the 1918 H1N1 virus has fewer glycosylation sequons on the HA globular head region. Using site-directed mutagenesis, we found that a 1918 HA recombinant virus, of high virulence, could be significantly attenuated in mice by adding two additional glycosylation sites (asparagine [Asn] 71 and Asn 286) on the side of the HA head. The 1918 HA recombinant virus was further attenuated by introducing two additional glycosylation sites on the top of the HA head at Asn 142 and Asn 172. In a reciprocal experimental approach, deletion of HA glycosylation sites (Asn 142 and Asn 177, but not Asn 71 and Asn 104) from a seasonal influenza H1N1 virus, A/Solomon Islands/2006 (SI/06), led to increased virulence in mice. The addition of glycosylation sites to 1918 HA and removal of glycosylation sites from SI/06 HA imposed constraints on the theoretical structure surrounding the glycan receptor binding sites, which in turn led to distinct glycan receptor binding properties. The modification of glycosylation sites for the 1918 and SI/06 viruses also caused changes in viral antigenicity based on cross-reactive hemagglutinin inhibition antibody titers with antisera from mice infected with wild-type or glycan mutant viruses. These results demonstrate that glycosylation patterns of the 1918 and seasonal H1N1 viruses directly contribute to differences in virulence and are partially responsible for their distinct antigenicity.     

The toxicity and biological effect of Pedalium murex L. extracts on the tobacco cutworm,Spodoptera litura (Fabr.) larvae

Petroleum ether, acetone, water and aqueous extract of Pedalium murex L. (Pedalaceae) leaves, root and fruit were tested against fourth instar larvae of the tobacco cutworm, Spodoptera litura (Fabr.) under laboratory conditions using leaf dip method. Larval mortality; larval, pupal and adult periods; pupal weight, pupation and adult emergence; larval and pupal deformities of S. litura were recorded. All the tested solvent extracts of P. murex were effective against S. litura life stages by causing mortality in a dose dependent manner. However, the efficacy was more significant with respect to acetone (leaf and root) and petroleum ether (root) at higher concentrations (0.8%) which leads to 100% larval mortality. The water and aqueous extracts of root caused 86.6 and 88.0% larval mortality respectively at 4% concentration. Among the plant parts tested, root had more importance followed by leaf and fruit. Plant extracts extended the larval, pupal and adult periods; reduced the pupal weight, pupal and adult emergence and caused larval–pupal deformities. Total life time was highly prolonged in acetone extracts of P. murex root. Though all the extracts were found to have insecticide activity, acetone and petroleum ether extracts of P. murex root can be used as an effective alternative to modern synthetic insecticides. Bioactive principles from these extracts can be isolated, identified and integrated in S. litura management.     

Chitosan-induced defence responses in tomato plants against early blight disease caused by Alternaria solani (Ellis and Martin) Sorauer

The in vitro antifungal properties of chitosan and its role in protection of tomato from early blight disease were evaluated. Chitosan inhibited the radial and submerged growth of Alternaria solani at 1?mg/ml and control tomato plants from blight pathogen. Chitosan was able to induce the level of chitinase activity and new isoforms of chitinase, resulting in the reduction of early blight disease severity in tomato leaves. These results suggested the role of chitosan in activation of defence responses as well as protecting tomato plants from A. solani infection.     

A novel cold-tolerant Clostridium strain PXYL1 isolated from a psychrophilic cattle manure digester that secretes thermolabile xylanase and cellulase

A Clostridium strain PXYL1 was isolated from a cold-adapted cattle manure biogas digester at 15 degrees C. It could grow at temperatures as low as 5 degrees C up to 50 degrees C with highest specific growth rate at 20 degrees C and is a psychrotroph. It produced extracellular hydrolytic enzymes namely xylanase, endoglucanase, beta-xylosidase, beta-glucosidase and filter paper cellulase, all of which had maximal activity at 20 degrees C. The induction of xylanase was highest on birch wood xylan (37 IU(mg protein)(-1)) compared with xylose (1.11 IU(mg protein)(-1)), cellobiose (1.43 IU(mg protein)(-1)) and glucose (no activity). The xylanase was thermolabile with a half-life of 30 min at 40 degrees C and 8 min at 50 degrees C but stable for over 2 h at 20 degrees C. The crude enzyme released reducing sugars (1.25 g l(-1)) from finger millet flour at 20 degrees C, while commercial food-grade xylanases showed no hydrolysis at this temperature. This is the first report of a Clostridium strain growing at 20 degrees C and producing an array of xylanolytic and cellulolytic enzymes, possessing low temperature optima of 20 degrees C, which may facilitate degradation of plant fibre under low-temperature conditions.     

Performance of halotolerant bacteria associated with Sahara-inhabiting halophytes Atriplex halimus L. and Lygeum spartum L. ameliorate tomato plant growth and tolerance to saline stress: from selective isolation to genomic analysis of potential determinants

World Journal of Microbiology and Biotechnology - Beauvericin and bassiatin are two valuable compounds with various bioactivities biosynthesized by the supposedly same nonribosomal peptide...     

Activation of pre-mRNA splicing by human RNPS1 is regulated by CK2 phosphorylation

Human RNPS1 was originally characterized as a pre-mRNA splicing activator in vitro and was shown to regulate alternative splicing in vivo. RNPS1 was also identified as a protein component of the splicing-dependent mRNP complex, or exon-exon junction complex (EJC), and a role for RNPS1 in postsplicing processes has been proposed. Here we demonstrate that RNPS1 incorporates into active spliceosomes, enhances the formation of the ATP-dependent A complex, and promotes the generation of both intermediate and final spliced products. RNPS1 is phosphorylated in vivo and interacts with the CK2 (casein kinase II) protein kinase. Serine 53 (Ser-53) of RNPS1 was identified as the major phosphorylation site for CK2 in vitro, and the same site is also phosphorylated in vivo. The phosphorylation status of Ser-53 significantly affects splicing activation in vitro, but it does not perturb the nuclear localization of RNPS1. In vivo experiments indicated that the phosphorylation of RNPS1 at Ser-53 influences the efficiencies of both splicing and translation. We propose that RNPS1 is a splicing regulator whose activator function is controlled in part by CK2 phosphorylation.     

Exonic splicing enhancer-dependent selection of the bovine papillomavirus type 1 nucleotide 3225 3' splice site can be rescued in a cell lacking splicing factor ASF/SF2 through activation of the phosphatidylinositol 3-kinase/Akt pathway

Bovine papillomavirus type 1 (BPV-1) late pre-mRNAs are spliced in keratinocytes in a differentiation-specific manner: the late leader 5' splice site alternatively splices to a proximal 3' splice site (at nucleotide 3225) to express L2 or to a distal 3' splice site (at nucleotide 3605) to express L1. Two exonic splicing enhancers, each containing two ASF/SF2 (alternative splicing factor/splicing factor 2) binding sites, are located between the two 3' splice sites and have been identified as regulating alternative 3' splice site usage. The present report demonstrates for the first time that ASF/SF2 is required under physiological conditions for the expression of BPV-1 late RNAs and for selection of the proximal 3' splice site for BPV-1 RNA splicing in DT40-ASF cells, a genetically engineered chicken B-cell line that expresses only human ASF/SF2 controlled by a tetracycline-repressible promoter. Depletion of ASF/SF2 from the cells by tetracycline greatly decreased viral RNA expression and RNA splicing at the proximal 3' splice site while increasing use of the distal 3' splice site in the remaining viral RNAs. Activation of cells lacking ASF/SF2 through anti-immunoglobulin M-B-cell receptor cross-linking rescued viral RNA expression and splicing at the proximal 3' splice site and enhanced Akt phosphorylation and expression of the phosphorylated serine/arginine-rich (SR) proteins SRp30s (especially SC35) and SRp40. Treatment with wortmannin, a specific phosphatidylinositol 3-kinase/Akt kinase inhibitor, completely blocked the activation-induced activities. ASF/SF2 thus plays an important role in viral RNA expression and splicing at the proximal 3' splice site, but activation-rescued viral RNA expression and splicing in ASF/SF2-depleted cells is mediated through the phosphatidylinositol 3-kinase/Akt pathway and is associated with the enhanced expression of other SR proteins.