Biomedical subjects

Fiona M Roche

Publications and source records attributed to Fiona M Roche.

6 recordsLinked to original sources

PubMed2005

Molecular analyses of disease pathogenesis: application of bovine microarrays.

The molecular analysis of disease pathogenesis in cattle has been limited by the lack of availability of tools to analyze both host and pathogen responses. These limitations are disappearing with the advent of methodologies such as microarrays that facilitate rapid characterization of global gene expression at the level of individual cells and tissues. The present review focuses on the use of microarray technologies to investigate the functional pathogenomics of infectious disease in cattle. We discuss a number of unique issues that must be addressed when designing both in vitro and in vivo model systems to analyze host responses to a specific pathogen. Furthermore, comparative functional genomic strategies are discussed that can be used to address questions regarding host responses that are either common to a variety of pathogens or unique to individual pathogens. These strategies can also be applied to investigations of cell signaling pathways and the analyses of innate immune responses. Microarray analyses of both host and pathogen responses hold substantial promise for the generation of databases that can be used in the future to address a wide variety of questions. A critical component limiting these comparative analyses will be the quality of the databases and the complete functional annotation of the bovine genome. These limitations are discussed with an indication of future developments that will accelerate the validation of data generated when completing a molecular characterization of disease pathogenesis in cattle.

Animals↗

PubMed2004

The N-terminal A domain of fibronectin-binding proteins A and B promotes adhesion of Staphylococcus aureus to elastin.

The ability of Staphylococcus aureus to adhere to components of the extracellular matrix is an important mechanism for colonization of host tissues during infection. We have previously shown that S. aureus binds elastin, a major component of the extracellular matrix. The integral membrane protein, elastin-binding protein (EbpS), binds soluble elastin peptides and tropoelastin via its surface-exposed N-terminal domain. In this study, we demonstrate that some strains of S. aureus adhere strongly to immobilized human elastin and that this interaction is independent of EbpS but instead is mediated by the fibronectin-binding proteins, FnBPA and FnBPB. Our results show that EbpS mutant cells adhere to elastin-coated plates, whereas the cells negative for FnBPA and FnBPB do not adhere to the plates. Furthermore, only wild-type cells from the exponential phase of growth adhered when FnBPs were expressed maximally. We show that adherence to elastin promoted by FnBPA was not affected by soluble fibronectin, suggesting that the elastin binding domain is distinct from the fibronectin binding regions. Recombinant FnBPA(37-544) (rFnBPA(37-544)) protein corresponding to the A region of FnBPA and anti-FnBPA(37-544) antibodies inhibited FnBPA-mediated bacterial adherence to immobilized elastin. Finally, recombinant A domain proteins, rFnBPA(37-544) and rFnBPB(37-540), bound immobilized elastin dose-dependently and saturably. This interaction was inhibited by soluble elastin peptides, suggesting a specific receptor-ligand interaction.

Adhesins, Bacterial↗

PubMed2004

ArrayPipe: a flexible processing pipeline for microarray data.

A number of microarray analysis software packages exist already; however, none combines the user-friendly features of a web-based interface with potential ability to analyse multiple arrays at once using flexible analysis steps. The ArrayPipe web server (freely available at www.pathogenomics.ca/arraypipe) allows the automated application of complex analyses to microarray data which can range from single slides to large data sets including replicates and dye-swaps. It handles output from most commonly used quantification software packages for dual-labelled arrays. Application features range from quality assessment of slides through various data visualizations to multi-step analyses including normalization, detection of differentially expressed genes, andcomparison and highlighting of gene lists. A highly customizable action set-up facilitates unrestricted arrangement of functions, which can be stored as action profiles. A unique combination of web-based and command-line functionality enables comfortable configuration of processes that can be repeatedly applied to large data sets in high throughput. The output consists of reports formatted as standard web pages and tab-delimited lists of calculated values that can be inserted into other analysis programs. Additional features, such as web-based spreadsheet functionality, auto-parallelization and password protection make this a powerful tool in microarray research for individuals and large groups alike.

Gene Expression Profiling↗

PubMed2004

ProbeLynx: a tool for updating the association of microarray probes to genes.

As genome sequence data and gene prediction improve, probes developed for a given microarray experiment should be continuously re-evaluated for their specificity for given genes. ProbeLynx(www.pathogenomics.ca/probelynx) is a new web service which uses current genomic sequence information to re-examine microarray probe specificity and provide annotation updates relevant to determining which gene(s) and transcript(s) are associated with a given probe. Probe sequences (either oligonucleotide- or cDNA-based) are uploaded in FASTA format and the results returned as a tab-delimited flat file for insertion into a spreadsheet application or database management system for further analysis. ProbeLynx has been initially developed to focus on arrays derived from human, mouse, chicken and bovine genomes, but may be expanded to handle other genomic datasets. ProbeLynx offers microarray users the important ability to continuously assess the potential of a probe to cross-hybridize to paralogous genes and the suitability of a given probe to investigate a transcript of interest. By also including the latest gene function annotation information in the output, ProbeLynx provides the critical first step in updating microarray data annotation.

Animals↗

PubMed2003

Characterization of novel LPXTG-containing proteins of Staphylococcus aureus identified from genome sequences.

Twenty-one genes encoding surface proteins belonging to the LPXTG family have been identified by in silico analysis of six Staphylococcus aureus genome sequences. Eleven genes encode previously described proteins, while 10 have not yet been characterized. Of these, eight contain the cell-wall sorting signal LPXTG responsible for covalently anchoring proteins to the cell-wall peptidoglycan. The remaining two, SasF and SasD, harbour a single residue variation in the fourth position of the LPXTG motif (LPXAG). Western blotting of lysostaphin-solubilized S. aureus cell-wall proteins demonstrated the release of SasF in the cell-wall fraction, indicating that proteins carrying LPXAG are sorted normally. Analysis of primary sequences of the Staphylococcus aureus surface (Sas) proteins indicated that several share a similar structural organization and a common signal sequence with previously characterized LPXTG proteins of S. aureus and other Gram-positive cocci. Protein SasG has 128 residue B repeats that are almost identical at the DNA level. PCR analysis indicated that recombinants with repeat length variations are present in the bacterial population whereas they are not detectable in the B-repeat-encoding region of sdrD. The sasG and sasH genes are significantly associated with invasive disease isolates compared to nasal carriage isolates. Several IgG samples purified from patients recovering from S. aureus infections had higher titres against Sas proteins than control IgG, suggesting that expression occurred during infection in some patients.

Adult↗

PubMed2003

The Staphylococcus aureus surface protein SasG and its homologues promote bacterial adherence to human desquamated nasal epithelial cells.

Staphylococcus aureus binds to human desquamated nasal epithelial cells, a phenomenon likely to be important in nasal colonization. ClfB was identified previously as one staphylococcal adhesin that promoted binding to nasal epithelia. In this study, it is shown that the S. aureus surface protein SasG, identified previously by in silico analysis of genome sequences, and two homologous proteins, Pls of S. aureus and AAP of Staphylococcus epidermidis, also promote bacterial adherence to nasal epithelial cells. Conditions for in vitro expression of SasG by S. aureus were not found. Adherence assays were therefore performed with S. aureus and Lactococcus lactis expressing SasG from an expression plasmid. These studies showed that SasG did not bind several ligands typically bound by S. aureus. Significantly, SasG and Pls did promote bacterial adherence to nasal epithelial cells. Furthermore, pre-incubation of epithelial cells with purified recombinant proteins revealed that the N-terminal A regions of SasG, Pls and AAP, but not the B repeats of SasG, inhibited adherence of L. lactis expressing SasG in a dose-dependent fashion. These results suggest that SasG, Pls and AAP bind to the same as-yet-unidentified receptor on the surface of nasal epithelial cells. In addition, expression of SasG, like Pls, reduced adherence of S. aureus to fibronectin and fibrinogen.

Bacterial Adhesion↗