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Craig Winstanley

Publications and source records attributed to Craig Winstanley.

27 records · Page 2Linked to original sources
PubMed2002

Persistent and aggressive bacteria in the lungs of cystic fibrosis children.

There have been enormous improvements in life expectancy of patients with cystic fibrosis, especially with improved nutrition and better understanding of the basic cellular defects. However, infection in particular with Pseudomonas aeruginosa and Burkholderia cepacia, has the greatest effect in decreasing life expectancy. Although infections can be prevented by rigorous infection control procedures, early aggressive antimicrobial chemotherapy and established infection managed by antibiotics, they are not completely effective. A greater understanding of how the bacteria evade the host defences and produce infection is needed.

Bacterial Infections↗
PubMed2002

Distribution of type III secretion gene clusters in Burkholderia pseudomallei, B. thailandensis and B. mallei.

Burkholderia pseudomallei, the causative agent of melioidosis, carries a cluster of genes closely related in organisation to the type III secretion (TTS) system gene clusters of the plant pathogens Ralstonia solanacearum and Xanthomonas spp. The TTS gene cluster (TTS1) is present only in B. pseudomallei and not in avirulent B. thailandensis. Adjacent to the gene cluster encoding putative secreton structural proteins lie a number of open reading frames (ORFs) encoding putative proteins with little or no homology to known proteins, with the exception of one predicted protein with homology to Pseudomonas syringae HrpK. In both R. solanacearum and Xanthomonas spp., genes in this location encode secreted effector proteins. RT-PCR analysis indicated that TTS genes, including two of these ORFs, are expressed in broth at 37 degrees C. Analysis of genome sequence data identified a second cluster of TTS genes (TTS2) present in both B. pseudomallei and B. mallei (99% identity). However, B. mallei appears to lack the TTS1 gene cluster. PCR assays indicated that TTS2 was also present in B. thailandensis. TTS1 and TTS2 are similar in gene organisation, but nucleotide sequences are sufficiently divergent to suggest that the two TTS systems may have different roles.

Amino Acid Sequence↗
PubMed2002

Spot the difference: applications of subtractive hybridisation to the study of bacterial pathogens.

Comparison of DNA from virulent strains of bacterial pathogens with DNA from less virulent or avirulent close relatives allows the identification of those genomic regions that are present only in virulent strains. Such regions are often associated with pathogenicity islands (PIs) and their characterisation can lead to a greater understanding of the pathogenesis of infectious diseases. There is now a large database of bacterial genomic sequences that provides useful reference information with which to compare the genomes of strains that exhibit variations in virulence or host preferences. Subtractive hybridisation (SH) and its sister method, suppression subtractive hybridisation (SSH), are techniques designed to identify those regions present in one genome but absent from another. The application of these techniques has led to the identification of PIs, mobile genetic elements and variations in virulence gene expression in a range of bacterial pathogens.

Bacteria↗
PubMed2002

Distribution of fim3 and flaA TTGE sequence types amongst isolates of Bordetella bronchiseptica from different host animals.

Isolates of Bordetella bronchiseptica associated with different animal hosts were analysed by fim3 and flaA genotyping by temporal temperature gradient gel electrophoresis (TTGE). All the isolates from cats (n = 138), dogs (n = 42) and pigs (n = 13) could be assigned to one of two fim3 and one of three flaA TTGE sequence types, respectively. Two avian isolates and a marmoset isolate exhibited novel fim3 sequence types. Dominant but different TTGE sequence types were apparent in isolates from dogs and pigs for both fim3 (87.5% and 100%, respectively, of isolates were the dominant type) and flaA (95% and 92%, respectively, of isolates were the dominant type). There was a wider distribution of TTGE sequence types amongst cat isolates. As both fimbriae and flagella have been implicated in adherence of bordetellae to host cells, sequence variations in fimbrial proteins and FlaA may have a role to play in host preferences.

Amino Acid Sequence↗
PubMed2002

Use of subtractive hybridization to identify a diagnostic probe for a cystic fibrosis epidemic strain of Pseudomonas aeruginosa.

A multiresistant strain of Pseudomonas aeruginosa is widespread among cystic fibrosis (CF) patients attending clinics in Liverpool, United Kingdom. Suppression subtractive hybridization was used to identify sequences present in the Liverpool CF epidemic strain but absent from strain PAO1. Using dot blot and PCR amplification assays, the prevalence of such sequences among a panel of CF isolates was determined. Several sequences were found only in the Liverpool epidemic strain. Some sequences were present in the Liverpool epidemic strain and in a minority of other isolates, including sequences with homology to genes implicated in O6 serotype and siderophore production. The Liverpool epidemic strain and 81% of nonepidemic isolates contained a sequence identified as part of the PAGI-1 genomic island. Other strains implicated in epidemic spread, which were from Manchester, United Kingdom, and Melbourne, Australia, were also screened. None of the sequences identified was present in the Manchester strain. However, one of two Melbourne strains contained some of the sequences found in the Liverpool epidemic strain. All isolates implicated in epidemic spread and 76% of sporadic isolates contained the exoS gene. A sequence present in all isolates of the Liverpool epidemic strain was used to develop a diagnostic PCR test for identification of the strain from colonies or directly from sputum samples.

Cloning, Molecular↗
PubMed2001

Type III secretion systems and pathogenicity islands.

Some bacterial pathogens have evolved by acquiring pathogenicity islands (PIs), which are clusters of genes encoding virulence traits. PIs encoding the secretion of effector molecules via type III secretion (TTS) systems have been discovered in several gram-negative pathogens. TTS systems are involved in contact-dependent secretion of virulence factors and can facilitate delivery of toxins directly into target cells. The expanding list of bacteria found to contain clusters of TTS genes includes members of the genera Yersinia, Salmonella, Shigella, Escherichia, Pseudomonas, Bordetella, Burkholderia, Chlamydia and a number of plant pathogens or symbionts. This review discusses the current knowledge of the role of TTS PIs in pathogenicity, the genetic organisation and evolution of such systems,and the potential for using TTS systems as targets for novel treatments.

Bacterial Proteins↗
PubMed2001

Flagellin gene PCR-RFLP analysis of a panel of strains from the Burkholderia cepacia complex.

Burkholderia cepacia, an important opportunist pathogen, is genetically heterogeneous. The B. cepacia complex has been subdivided into a number of genospecies or genomovars. A flagellin gene PCR-RFLP method was applied to a representative panel of strains of known genomovar. The technique was able to distinguish strains of B. multivorans from other members of the B. cepacia complex on the basis of amplicon size (typical of type I rather than type II flagellins) with the exception of one genomovar I strain. There was considerable variation in RFLP patterns amongst the panel of strains; only two pairs of strains were indistinguishable with both HaeIII and MspI digestion. Where RFLP patterns matched with both enzymes or a single enzyme, matching strains were always in the same genomovar. It was possible to distinguish the UK cystic fibrosis epidemic strain from all other members of the panel, including nine other genomovar III strains. The level of variation suggests that flagellin genotyping is a useful method for discriminating between B. cepacia strains.

Bacterial Typing Techniques↗
PubMed1999

Analysis of fliC variation among clinical isolates of Burkholderia cepacia.

PCR and restriction fragment length polymorphism (RFLP) typing of flagellin genes (fliC) from 57 clinical isolates of Burkholderia cepacia indicated that only type 11 flagellins were present. Twenty-two isolates previously identified as the epidemic UK cystic fibrosis strain were indistinguishable by this method, as were 11 isolates from a pseudo-outbreak in Senegal. Other clinical isolates, including 19 from disparate sources in Malaysia, were separated into nine fliC RFLP groups, exhibiting a large degree of divergence. When isolates were indistinguishable by fliC genotyping, their similarity was confirmed by whole genome macro-restriction analysis with pulsed-field gel electrophoresis following XbaI digestion. The variation in fliC sequences of B. cepacia was far greater than that with B. pseudomallei, supporting the view that 'B. cepacia', as currently defined, may comprise several different genomic species.

Burkholderia Infections↗