BackgroundPseudomonas aeruginosa is considered to grow in a biofilm in cystic fibrosis (CF) chronic lung infections. Bacterial cell motility is one of the main factors that have been connected with P. aeruginosa adherence to both biotic and abiotic surfaces. In this investigation, we employed molecular and microscopic methods to determine the presence or absence of motility in P. aeruginosa CF isolates, and statistically correlated this with their biofilm forming ability in vitro.ResultsOur investigations revealed a wide diversity in the production, architecture and control of biofilm formation. Of 96 isolates, 49% possessed swimming motility, 27% twitching and 52% swarming motility, while 47% were non-motile. Microtitre plate assays for biofilm formation showed a range of biofilm formation ability from biofilm deficient phenotypes to those that formed very thick biofilms. A comparison of the motility and adherence properties of individual strains demonstrated that the presence of swimming and twitching motility positively affected biofilm biomass. Crucially, however, motility was not an absolute requirement for biofilm formation, as 30 non-motile isolates actually formed thick biofilms, and three motile isolates that had both flagella and type IV pili attached only weakly. In addition, CLSM analysis showed that biofilm-forming strains of P. aeruginosa were in fact capable of entrapping non-biofilm forming strains, such that these 'non-biofilm forming' cells could be observed as part of the mature biofilm architecture.ConclusionsClinical isolates that do not produce biofilms in the laboratory must have the ability to survive in the patient lung. We propose that a synergy exists between isolates in vivo, which allows "non biofilm-forming" isolates to be incorporated into the biofilm. Therefore, there is the potential for strains that are apparently non-biofilm forming in vitro to participate in biofilm-mediated pathogenesis in the CF lung.

Molecular diagnostic tests, based on the detection and identification of nucleic acids in human biological samples, are increasingly employed in the diagnosis of infectious diseases and may be of future benefit to CF microbiology services. Our growing understanding of the complex polymicrobial nature of CF airway infection has highlighted current and likely future shortcomings in standard diagnostic practices. Failure to detect fastidious or slow growing microbes and misidentification of newly emerging pathogens could potentially be addressed using culture-independent molecular technologies with high target specificity. This review considers existing molecular diagnostic tests in the context of the key requirements for an envisaged CF microbiology focussed assay. The issues of assay speed, throughput, detection of multiple pathogens, data interpretation and antimicrobial susceptibility testing are discussed.

Lung disease is the main cause of morbidity and mortality in cystic fibrosis (CF), and involves chronic infection and perturbed immune responses. Tissue damage is mediated mostly by extracellular proteases, but other cellular proteins may also contribute to damage through their effect on cell activities and/or release into sputum fluid by means of active secretion or cell death.We employed MudPIT (multidimensional protein identification technology) to identify sputum cellular proteins with consistently altered abundance in adults with CF, chronically infected with , compared with healthy controls. Ingenuity Pathway Analysis, Gene Ontology, protein abundance and correlation with lung function were used to infer their potential clinical significance.Differentially abundant proteins relate to Rho family small GTPase activity, immune cell movement/activation, generation of reactive oxygen species, and dysregulation of cell death and proliferation. Compositional breakdown identified high abundance of proteins previously associated with neutrophil extracellular traps. Furthermore, negative correlations with lung function were detected for 17 proteins, many of which have previously been associated with lung injury.These findings expand our current understanding of the mechanisms driving CF lung disease and identify sputum cellular proteins with potential for use as indicators of disease status/prognosis, stratification determinants for treatment prescription or therapeutic targets.

Cystic fibrosis is one of the most common life-limiting inherited disorders. Its clinical impact manifests chiefly in the lung, pancreas, gastrointestinal tract and sweat glands, with lung disease typically being most detrimental to health. The median age for survival has increased dramatically over the past decades, largely thanks to advances in understanding of the mechanisms and consequences of disease, leading to the development of better therapies and treatment regimes. The discovery of dysregulated protein biomarkers linked to cystic fibrosis has contributed considerably to this end. This article outlines clinical trials targeting known protein biomarkers, and the current and future contributions of proteomic techniques to cystic fibrosis research. The treatments described range from those designed to provide functional copies of the mutant protein responsible for cystic fibrosis, to others addressing the associated symptoms of chronic inflammation. Preclinical research has employed proteomics to help elucidate pathways and processes implicated in disease that might present opportunities for therapy or prognosis. Global analyses of cystic fibrosis have detected the differential expression of proteins involved in inflammation, proteolytic activity and oxidative stress, which are recognized symptoms of the cystic fibrosis phenotype. The dysregulation of other processes, such as the complement and mitochondrial systems, has also been implicated. A number of studies have focused specifically on proteins that interact with the cystic fibrosis protein, with the goal of restoring its normal proteostasis. Consequently, proteins involved in synthesis, folding, degradation, translocation and localization of the protein have been identified as potential therapeutic targets. Cystic fibrosis patients are prone to lung infections that are thought to contribute to chronic inflammation, and thus proteomic studies have also searched for microbiological biomarkers to use in early infection diagnosis or as indicators of virulence. The review concludes by proposing a future role for proteomics in the high-throughput validation of protein biomarkers under consideration as outcome measures for use in clinical trials and routine disease monitoring.

BackgroundUrinary Tract Infections (UTIs) are common bacterial infections, second only to respiratory tract infections and particularly prevalent within primary care. Conventional detection of UTIs is culture, however, return of results can take between 24 and 72 hours. The introduction of a point of care (POC) test would allow for more timely identification of UTIs, facilitating improved, targeted treatment. This study aimed to obtain consensus on the criteria required for a POC UTI test, to meet patient need within primary care.MethodsCriteria for consideration were compiled by the research team. These criteria were validated through a two-round Delphi process, utilising an expert panel of healthcare professionals from across Europe and United States of America. Using web-based questionnaires, panellists recorded their level of agreement with each criterion based on a 5-point Likert Scale, with space for comments. Using median response, interquartile range and comments provided, criteria were accepted/rejected/revised depending on pre-agreed cut-off scores.ResultsThe first round questionnaire presented thirty-three criteria to the panel, of which 22 were accepted. Consensus was not achieved for the remaining 11 criteria. Following response review, one criterion was removed, while after revision, the remaining 10 criteria entered the second round. Of these, four were subsequently accepted, resulting in 26 criteria considered appropriate for a POC test to detect urinary infections.ConclusionThis study generated an approved set of criteria for a POC test to detect urinary infections. Criteria acceptance and comments provided by the healthcare professionals also supports the development of a multiplex point of care UTI test.