The validity and importance of identifying these four subgroups is demonstrated. Previous definitions may over-estimate the prevalence of chronic infection.

Various management strategies were introduced at the Leeds Regional Cystic Fibrosis (CF) Unit in an attempt to reduce the prevalence of chronic Pseudomonas aeruginosa respiratory infection, previously thought to be inevitable in most children with CF. These included neonatal screening (1975), regular microbiological monitoring (1975), early antibiotic treatment of first isolations of P. aeruginosa (1985), intensive intravenous antibiotic treatment where nebulized antibiotics failed to eradicate P. aeruginosa (1988), and separate clinics for patients chronically infected with P. aeruginosa and uninfected patients (1991). The aim of this study was to assess the impact of these interventions. All 232 patients receiving full-time care at the Leeds Paediatric CF Centre during the period January 1990-December 2000 were categorized into four groups: never grown P. aeruginosa; free of P. aeruginosa for at least 1 year; intermittent grower of P. aeruginosa with 50% of months with samples positive for P. aeruginosa over the previous 12 months. The yearly prevalence of patients having chronic P. aeruginosa infection fell significantly during the study, from 24.5% in 1990 to 18.1% in 2000 (P < 0.05), despite an increase in mean age of patients from 7.73 to 9.42 years. The number of patients aged less than 11 years who had chronic P. aeruginosa infection fell from 23.8% in January 1990 to only 4.3% by December 2000. The annual incidence and mean age of first acquisition of P. aeruginosa did not alter significantly. In conclusion, antipseudomonal management strategies were associated with both reduced prevalence, and an increase in the mean age of onset of chronic P. aeruginosa infection.

During adenovirus infection, following capsid dissociation, core protein VII enters the host cell nucleus complexed with adenovirus DNA. In order to determine whether protein VII may have an active role in this nuclear import, regions of the preVII gene were amplified by PCR, and further oligonucleotide mutants were designed with site-directed mutation of codons for the basic amino acids arginine and lysine. Fragments were cloned into a mammalian expression plasmid to express the peptides as N-terminal fusions to enhanced green fluorescent protein. Results demonstrate that preVII protein contains both nuclear and nucleolar targeting sequences. Such signals may be important in the delivery of adenovirus DNA to the host cell nucleus during adenovirus infection. Furthermore, the data suggest that protein VII may bind to human chromosomes by means of two distinct domains, one sharing homology with the N-terminal regulatory tail of histone H3.Enclosed by approximately 12 capsid proteins the human adenovirus genome, comprising approximately 36 kb of linear double-stranded DNA, is non-covalently bound to the viral core proteins Mu, V, and VII (Vayda et al., 1983). Proteins VII and Mu are tightly associated with the viral DNA (Vayda et al., 1983;Chatterjee et al., 1985), whilst protein V may form a link between the viral DNA-core protein complex and the viral capsid (Matthews & Russell, 1998).In human adenoviruses, both protein VII and Mu are encoded by the late transcription unit L2 (Alestrom et al., 1984). Mature protein VII of human adenovirus 2 and 5 contains 174 amino acids and is formed from its precursor by adenovirus-encoded protease-mediated cleavage of a 24 amino acid N-terminal segment (Sung et al., 1983;Alestrom et al., 1984;Webster et al., 1989). The mature protein has four highly basic domains containing arginineand lysine-rich sequences, separated by predicted alphahelices. The interaction of protein VII with DNA appears to be charge-based between these basic regions and the phosphate backbone of 90-150 DNA bp (Vayda & Flint, 1987), resulting in the DNA being considerably condensed as a result of superfolding (Black & Center, 1979;Sato & Hosokawa, 1984). Protein VII associates more efficiently with double-stranded DNA than single-stranded (Sato & Hosokawa, 1984). Binding of both proteins VII and Mu to DNA is not DNA sequence-specific (Russell & Precious, 1982).During adenovirus infection, following capsid dissociation, protein VII remains complexed to adenovirus DNA, and this complex enters the nucleus via the nuclear pore (Greber et al., 1997). Protein VII inhibits adenovirus DNA synthesis (Korn & Horwitz, 1986) and transcription in vitro (Nakanishi et al., 1986) and thus is then likely to dissociate from DNA to allow these processes to occur. Antibodies raised against protein Mu for use in Western blots and ELISA cross-react with protein VII, presumably due to the homologous unusual arginine-rich sequences present in both proteins (Lunt et al., 1988). Protein VII has homologues in every adenovirus studi...

Gene therapy holds promise for the treatment of a range of inherited diseases, such as cystic fibrosis. However, efficient delivery and expression of the therapeutic transgene at levels sufficient to result in phenotypic correction of cystic fibrosis pulmonary disease has proved elusive. There are many reasons for this lack of progress, both macroscopically in terms of airway defence mechanisms and at the molecular level with regard to effective cDNA delivery. This review of approaches to cystic fibrosis gene therapy covers these areas in detail and highlights recent progress in the field. For gene therapy to be effective in patients with cystic fibrosis, the cDNA encoding the cystic fibrosis transmembrane conductance regulator protein must be delivered effectively to the nucleus of the epithelial cells lining the bronchial tree within the lungs. Expression of the transgene must be maintained at adequate levels for the lifetime of the patient, either by repeat dosage of the vector or by targeting airway stem cells. Clinical trials of gene therapy for cystic fibrosis have demonstrated proof of principle, but gene expression has been limited to 30 days at best. Results suggest that viral vectors such as adenovirus and adeno-associated virus are unsuited to repeat dosing, as the immune response reduces the effectiveness of each subsequent dose. Nonviral approaches, such as cationic liposomes, appear more suited to repeat dosing, but have been less effective. Current work regarding non-viral gene delivery is now focused on understanding the mechanisms involved in cell entry, endosomal escape and nuclear import of the transgene. There is now increasing evidence to suggest that additional ligands that facilitate endosomal escape or contain a nuclear localization signal may enhance liposome-mediated gene delivery. Much progress in this area has been informed by advances in our understanding of the mechanisms by which viruses deliver their genomes to the nuclei of host cells.