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Usher 1 patients are born profoundly deaf and then develop retinal degeneration. Thus they are readily identified before the onset of retinal degeneration, making gene therapy a viable strategy to prevent their blindness. Here, we have investigated the use of adeno-associated viruses (AAVs) for the delivery of the Usher 1B gene, MYO7A, to retinal cells in cell culture and in Myo7a-null mice. MYO7A cDNA, under control of a smCBA promoter, was packaged in single AAV2 and AAV5 vectors and as two overlapping halves in dual AAV2 vectors. The 7.9-kb smCBA-MYO7A exceeds the capacity of an AAV vector; packaging of such oversized constructs into single AAV vectors may involve fragmentation of the gene. Nevertheless, the AAV2 and AAV5 single vector preparations successfully transduced photoreceptor and retinal pigment epithelium cells, resulting in functional, full-length MYO7A protein and correction of mutant phenotypes, suggesting successful homologous recombination of gene fragments. With discrete, conventional-sized dual AAV2 vectors, full-length MYO7A was detected, but the level of protein expression was variable, and only a minority of cells showed phenotype correction. Our results show that MYO7A therapy with AAV2 or AAV5 single vectors is efficacious; however, the dual AAV2 approach proved to be less effective.

This study is focused to optimising a bio-ethanols auto-thermal reforming (ATR) process over 5% Rh/Al2O3 catalyst homemade, to produce syn-gas for fuel cell application. The effect of oxygen addition for a given S/C molar ratio ranging from 2.1 to 6.3 at T=923 K was investigated, varying the O2/EtOH molar ratio between 0.2 and 1.1. The results have identified an optimum range where the ethanol conversion is 100%, CH4 and CO productions go down and hydrogen yield reaches a maximum before decreasing because of the prevalent oxidizing action of oxygen. The obtained data, compared with those from steam reforming (SR) process, gave a very clear indication about the potentiality of ATR process. [Copyright &y& Elsevier]

The performance of MgO supported Pd, Rh, Ni and Co catalysts in the steam reforming of simulated bio-ethanol at MC fuel cell operative conditions (T=650 °C) has been investigated. Rh/MgO shows the best performance both in terms of activity and stability, however, it seems not to be so selective towards H2 production. Ni, Co and Pd catalysts are affected by deactivation mainly due to metal sintering. Ni/MgO displays the best performance in terms of H2 selectivity (>95%). A very low coke formation rate was observed on Rh/MgO catalyst, however, even on Ni/MgO catalyst coke formation occurs with modest rate. Kinetics measurements have shown that a significant difference in terms of metals specific activity exist: Rh sites resulted to be 2.2, 3.7 and 5.8 times more active than Pd, Co and Ni, respectively. [Copyright &y& Elsevier]

The 90-kilobase (kb) virulence plasmid of Salmonella typhimurium is responsible for invasion from the intestines to mesenteric lymph nodes and spleens of orally inoculated mice. We used Tn5 and aminoglycoside phosphotransferase (aph) gene insertion mutagenesis and deletion mutagenesis of a previously identified 14-kb virulence region to reduce this virulence region to 7.8kb. The 7.8-kb virulence region subcloned into a low copy-number vector conferred a wild-type level of splenic infection to virulence plasmid-cured S. typhimurium and conferred essentially a wild-type oral LD50. Insertion mutagenesis identified five loci essential for virulence, and DNA sequence analysis of the virulence region identified six open reading frames. Expected protein products were identified from four of the six genes, with three of the proteins identified as doublet bands in Escherichia coli minicells. Three of the five mutated genes were able to be complemented by clones containing only the corresponding wild-type gene. Only one of the five deduced amino acid sequences, that of the positive regulatory element, SpvR, possessed significant homology to other proteins. The codon usage for the virulence genes showed no codon bias, which is consistent with the low levels of expression observed for the corresponding proteins. Consensus promoters for several different sigma factors were identified upstream of several of the genes, whereas only consensus Rho-dependent termination sequences were observed between certain of the genes. The operon structure of this virulence region therefore appears to be complex. The construction of the cloned 7.8-kb virulence region and the determination of the DNA sequence will aid in the further genetic analysis of the five plasmid-encoded virulence genes of S. typhimurium.