Showing papers by "John Radcliffe Hospital published in 2003"

Non-invasive mapping of connections between human thalamus and cortex using diffusion imaging

Abstract: Evidence concerning anatomical connectivities in the human brain is sparse and based largely on limited post-mortem observations. Diffusion tensor imaging has previously been used to define large white-matter tracts in the living human brain, but this technique has had limited success in tracing pathways into gray matter. Here we identified specific connections between human thalamus and cortex using a novel probabilistic tractography algorithm with diffusion imaging data. Classification of thalamic gray matter based on cortical connectivity patterns revealed distinct subregions whose locations correspond to nuclei described previously in histological studies. The connections that we found between thalamus and cortex were similar to those reported for non-human primates and were reproducible between individuals. Our results provide the first quantitative demonstration of reliable inference of anatomical connectivity between human gray matter structures using diffusion data and the first connectivity-based segmentation of gray matter.

Regulatory T cells in transplantation tolerance.

Abstract: The identification and characterization of regulatory T (T(Reg)) cells that can control immune responsiveness to alloantigens have opened up exciting opportunities for new therapies in transplantation. After exposure to alloantigens in vivo, alloantigen-specific immunoregulatory activity is enriched in a population of CD4+ T cells that express high levels of CD25. In vivo, common mechanisms seem to underpin the activity of CD4+CD25+ T(Reg) cells in both naive and manipulated hosts. However, the origin, allorecognition properties and molecular basis for the suppressive activity of CD4+CD25+ T(Reg) cells, as well as their relationship to other populations of regulatory cells that exist after transplantation, remain a matter of debate..

The Bloom's syndrome helicase suppresses crossing over during homologous recombination

Abstract: Mutations in BLM, which encodes a RecQ helicase, give rise to Bloom's syndrome, a disorder associated with cancer predisposition and genomic instability. A defining feature of Bloom's syndrome is an elevated frequency of sister chromatid exchanges. These arise from crossing over of chromatid arms during homologous recombination, a ubiquitous process that exists to repair DNA double-stranded breaks and damaged replication forks. Whereas crossing over is required in meiosis, in mitotic cells it can be associated with detrimental loss of heterozygosity. BLM forms an evolutionarily conserved complex with human topoisomerase IIIalpha (hTOPO IIIalpha), which can break and rejoin DNA to alter its topology. Inactivation of homologues of either protein leads to hyper-recombination in unicellular organisms. Here, we show that BLM and hTOPO IIIalpha together effect the resolution of a recombination intermediate containing a double Holliday junction. The mechanism, which we term double-junction dissolution, is distinct from classical Holliday junction resolution and prevents exchange of flanking sequences. Loss of such an activity explains many of the cellular phenotypes of Bloom's syndrome. These results have wider implications for our understanding of the process of homologous recombination and the mechanisms that exist to prevent tumorigenesis.

Randomized study to assess the effectiveness of slow- and moderate-release polymer-based paclitaxel-eluting stents for coronary artery lesions

Abstract: Background— Early clinical studies demonstrated the feasibility of local paclitaxel delivery in reducing restenosis after treatment of de novo coronary lesions in small patient populations. Methods and Results— We conducted a randomized, double-blind trial of 536 patients at 38 medical centers evaluating slow-release (SR) and moderate-release (MR) formulations of a polymer-based paclitaxel-eluting stent (TAXUS) for revascularization of single, primary lesions in native coronary arteries. Cohort I compared TAXUS-SR with control stents, and Cohort II compared TAXUS-MR with a second control group. The primary end point was 6-month percent in-stent net volume obstruction measured by intravascular ultrasound. Secondary end points were 6-month angiographic restenosis and 6- and 12-month incidence of major adverse cardiac events, a composite of cardiac death, myocardial infarction, and repeat revascularization. At 6 months, percent net volume obstruction within the stent was significantly lower for TAXUS stents ...

Activation of the Human Orbitofrontal Cortex to a Liquid Food Stimulus is Correlated with its Subjective Pleasantness

Abstract: Single-neuron recording studies in non-human primates indicate that orbitofrontal cortex neurons represent the reward value of the sight, smell and taste of food, and even changes in the relative reward value, but provide no direct evidence on brain activity that is correlated with subjective reports of the pleasantness of food. In this fMRI investigation we report a significant correlation between the activation of a region of the human orbitofrontal cortex and the decrease in subjective pleasantness when a liquid food is eaten to satiety. Moreover, a cluster of voxels in the orbitofrontal cortex showed a decrease in its activation that was specific to the particular liquid food consumed in a meal, providing a neural correlate of sensory-specific satiety to a liquid whole food in humans. This sensory-specific reduction in activation of the orbitofrontal cortex correlating with subjective pleasantness is consistent with an important role for the orbitofrontal cortex in human emotion and motivation, and associated subjective states.

Original antigenic sin and apoptosis in the pathogenesis of dengue hemorrhagic fever.

Abstract: Dengue virus presents a growing threat to public health in the developing world. Four major serotypes of dengue virus have been characterized, and epidemiological evidence shows that dengue hemorrhagic fever (DHF), the more serious manifestation of the disease, occurs more frequently upon reinfection with a second serotype. We have studied dengue virus–specific T-cell responses in Thai children. During acute infection, few dengue-responsive CD8+ T cells were recovered; most of those present showed an activated phenotype and were undergoing programmed cell death. Many dengue-specific T cells were of low affinity for the infecting virus and showed higher affinity for other, probably previously encountered strains. Profound T-cell activation and death may contribute to the systemic disturbances leading to DHF, and original antigenic sin in the T-cell responses may suppress or delay viral elimination, leading to higher viral loads and increased immunopathology.

Comprehensive Epitope Analysis of Human Immunodeficiency Virus Type 1 (HIV-1)-Specific T-Cell Responses Directed against the Entire Expressed HIV-1 Genome Demonstrate Broadly Directed Responses, but No Correlation to Viral Load

Abstract: Cellular immune responses play a critical role in the control of human immunodeficiency virus type 1 (HIV-1); however, the breadth of these responses at the single-epitope level has not been comprehensively assessed. We therefore screened peripheral blood mononuclear cells (PBMC) from 57 individuals at different stages of HIV-1 infection for virus-specific T-cell responses using a matrix of 504 overlapping peptides spanning all expressed HIV-1 proteins in a gamma interferon-enzyme-linked immunospot (Elispot) assay. HIV-1-specific T-cell responses were detectable in all study subjects, with a median of 14 individual epitopic regions targeted per person (range, 2 to 42), and all 14 HIV-1 protein subunits were recognized. HIV-1 p24-Gag and Nef contained the highest epitope density and were also the most frequently recognized HIV-1 proteins. The total magnitude of the HIV-1-specific response ranged from 280 to 25,860 spot-forming cells (SFC)/10(6) PBMC (median, 4,245) among all study participants. However, the number of epitopic regions targeted, the protein subunits recognized, and the total magnitude of HIV-1-specific responses varied significantly among the tested individuals, with the strongest and broadest responses detectable in individuals with untreated chronic HIV-1 infection. Neither the breadth nor the magnitude of the total HIV-1-specific CD8+-T-cell responses correlated with plasma viral load. We conclude that a peptide matrix-based Elispot assay allows for rapid, sensitive, specific, and efficient assessment of cellular immune responses directed against the entire expressed HIV-1 genome. These data also suggest that the impact of T-cell responses on control of viral replication cannot be explained by the mere quantification of the magnitude and breadth of the CD8+-T-cell response, even if a comprehensive pan-genome screening approach is applied.

RecQ helicases: caretakers of the genome.

Abstract: RecQ helicases are highly conserved from bacteria to man. Germline mutations in three of the five known family members in humans give rise to debilitating disorders that are characterized by, amongst other things, a predisposition to the development of cancer. One of these disorders--Bloom's syndrome--is uniquely associated with a predisposition to cancers of all types. So how do RecQ helicases protect against cancer? They seem to maintain genomic stability by functioning at the interface between DNA replication and DNA repair.

Magnetic resonance: an introduction to ultrashort TE (UTE) imaging.

Abstract: The background underpinning the clinical use of ultrashort echo-time (UTE) pulse sequences for imaging tissues or tissue components with short T2s is reviewed. Tissues properties are discussed, and tissues are divided into those with a majority of short T2 relaxation components and those with a minority. Features of the basic physics relevant to UTE imaging are described including the fact that when the radiofrequency pulse duration is of the order T2, rotation of tissue magnetization into the transverse plane is incomplete. Consequences of the broad line-width of short T2 components are also discussed including their partial saturation by off-resonance fat suppression pulses as well as multislice and multiecho imaging. The need for rapid data acquisition of the order T2 is explained. The basic UTE pulse sequence with its half excitation pulse and radial imaging from the center of k-space is described together with options that suppress fat and/or long T2 components. Image interpretation is discussed. Clinical features of the imaging of cortical bone, tendons, ligaments, menisci, and periosteum as well as brain, liver, and spine are illustrated. Short T2 components in all of these tissues may show high signals. Possible future developments are outlined as are technical limitations.

How Clonal Is Staphylococcus aureus

Abstract: Staphylococcus aureus is an important human pathogen and represents a growing public health burden owing to the emergence and spread of antibiotic-resistant clones, particularly within the hospital environment. Despite this, basic questions about the evolution and population biology of the species, particularly with regard to the extent and impact of homologous recombination, remain unanswered. We address these issues through an analysis of sequence data obtained from the characterization by multilocus sequence typing (MLST) of 334 isolates of S. aureus, recovered from a well-defined population, over a limited time span. We find no significant differences in the distribution of multilocus genotypes between strains isolated from carriers and those from patients with invasive disease; there is, therefore, no evidence from MLST data, which index variation within the stable “core” genome, for the existence of hypervirulent clones of this pathogen. Examination of the sequence changes at MLST loci during clonal diversification shows that point mutations give rise to new alleles at least 15-fold more frequently than does recombination. This contrasts with the naturally transformable species Neisseria meningitidis and Streptococcus pneumoniae, in which alleles change between 5- and 10-fold more frequently by recombination than by mutation. However, phylogenetic analysis suggests that homologous recombination does contribute toward the evolution of this species over the long term. Finally, we note a striking excess of nonsynonymous substitutions in comparisons between isolates belonging to the same clonal complex compared to isolates belonging to different clonal complexes, suggesting that the removal of deleterious mutations by purifying selection may be relatively slow.

Clonal Relationships between Invasive and Carriage Streptococcus pneumoniae and Serotype- and Clone-Specific Differences in Invasive Disease Potential

Abstract: By use of multilocus sequence typing, Streptococcus pneumoniae isolates causing invasive disease (n=150) were compared with those from nasopharyngeal carriage (n=351) among children in Oxford. The prevalence of individual clones (sequence types) and serotypes among isolates from invasive disease was related to their prevalence in carriage, and an odds ratio (OR) for invasive disease was calculated for the major clones and serotypes. All major carried clones and serotypes caused invasive disease, although their ability to do so varied greatly. Thus, 2 serotype 14 clones were approximately 10-fold overrepresented among disease isolates, compared with carriage isolates, whereas a serotype 3 clone was approximately 10-fold underrepresented. The lack of heterogeneity between the ORs of different clones of the same serotype, and analysis of isolates of the same genotype, but different serotype, suggested that capsular serotype may be more important than genotype in the ability of pneumococci to cause invasive disease.

Enhanced T-cell immunogenicity of plasmid DNA vaccines boosted by recombinant modified vaccinia virus Ankara in humans.

Abstract: In animals, effective immune responses against malignancies and against several infectious pathogens, including malaria, are mediated by T cells. Here we show that a heterologous prime-boost vaccination regime of DNA either intramuscularly or epidermally, followed by intradermal recombinant modified vaccinia virus Ankara (MVA), induces high frequencies of interferon (IFN)-gamma-secreting, antigen-specific T-cell responses in humans to a pre-erythrocytic malaria antigen, thrombospondin-related adhesion protein (TRAP). These responses are five- to tenfold higher than the T-cell responses induced by the DNA vaccine or recombinant MVA vaccine alone, and produce partial protection manifest as delayed parasitemia after sporozoite challenge with a different strain of Plasmodium falciparum. Such heterologous prime-boost immunization approaches may provide a basis for preventative and therapeutic vaccination in humans.

Multilocus Sequence Typing System for Group B Streptococcus

Abstract: A multilocus sequence typing (MLST) system was developed for group B streptococcus (GBS). The system was used to characterize a collection (n = 152) of globally and ecologically diverse human strains of GBS that included representatives of capsular serotypes Ia, Ib, II, III, V, VI, and VIII. Fragments (459 to 519 bp) of seven housekeeping genes were amplified by PCR for each strain and sequenced. The combination of alleles at the seven loci provided an allelic profile or sequence type (ST) for each strain. A subset of the strains were characterized by restriction digest patterning, and these results were highly congruent with those obtained with MLST. There were 29 STs, but 66% of isolates were assigned to four major STs. ST-1 and ST-19 were significantly associated with asymptomatic carriage, whereas ST-23 included both carried and invasive strains. All 44 isolates of ST-17 were serotype III clones, and this ST appeared to define a homogeneous clone that was strongly associated with neonatal invasive infections. The finding that isolates with different capsular serotypes had the same ST suggests that recombination occurs at the capsular locus. A web site for GBS MLST was set up and can be accessed at http://sagalactiae.mlst.net. The GBS MLST system offers investigators a valuable typing tool that will promote further investigation of the population biology of this organism.

Taste-olfactory convergence, and the representation of the pleasantness of flavour, in the human brain

Abstract: The functional architecture of the central taste and olfactory systems in primates provides evidence that the convergence of taste and smell information onto single neurons is realized in the caudal orbitofrontal cortex (and immediately adjacent agranular insula). These higher-order association cortical areas thus support flavour processing. Much less is known, however, about homologous regions in the human cortex, or how taste-odour interactions, and thus flavour perception, are implemented in the human brain. We performed an event-related fMRI study to investigate where in the human brain these interactions between taste and odour stimuli (administered retronasally) may be realized. The brain regions that were activated by both taste and smell included parts of the caudal orbitofrontal cortex, amygdala, insular cortex and adjoining areas, and anterior cingulate cortex. It was shown that a small part of the anterior (putatively agranular) insula responds to unimodal taste and to unimodal olfactory stimuli, and that a part of the anterior frontal operculum is a unimodal taste area (putatively primary taste cortex) not activated by olfactory stimuli. Activations to combined olfactory and taste stimuli where there was little or no activation to either alone (providing positive evidence for interactions between the olfactory and taste inputs) were found in a lateral anterior part of the orbitofrontal cortex. Correlations with consonance ratings for the smell and taste combinations, and for their pleasantness, were found in a medial anterior part of the orbitofrontal cortex. These results provide evidence on the neural substrate for the convergence of taste and olfactory stimuli to produce flavour in humans, and where the pleasantness of flavour is represented in the human brain.

Different representations of pleasant and unpleasant odours in the human brain.

Abstract: Odours are important in emotional processing; yet relatively little is known about the representations of the affective qualities of odours in the human brain. We found that three pleasant and three unpleasant odours activated dissociable parts of the human brain. Pleasant but not unpleasant odours were found to activate a medial region of the rostral orbitofrontal cortex. Further, there was a correlation between the subjective pleasantness ratings of the six odours given during the investigation with activation of a medial region of the rostral orbitofrontal cortex. In contrast, a correlation between the subjective unpleasantness ratings of the six odours was found in regions of the left and more lateral orbitofrontal cortex. Moreover, a double dissociation was found with the intensity ratings of the odours, which were not correlated with the BOLD signal in the orbitofrontal cortex, but were correlated with the signal in medial olfactory cortical areas including the pyriform and anterior entorhinal cortex. Activation was also found in the anterior cingulate cortex, with a middle part of the anterior cingulate activated by both pleasant and unpleasant odours, and a more anterior part of the anterior cingulate cortex showing a correlation with the subjective pleasantness ratings of the odours. Thus the results suggest that there is a hedonic map of the sense of smell in brain regions such as the orbitofrontal cortex, and these results have implications for understanding the psychiatric and related problems that follow damage to these brain areas.

Transcription of antisense RNA leading to gene silencing and methylation as a novel cause of human genetic disease.

Abstract: Transcription of antisense RNA leading to gene silencing and methylation as a novel cause of human genetic disease

NKT cells enhance CD4+ and CD8+ T cell responses to soluble antigen in vivo through direct interaction with dendritic cells.

Abstract: Modification in the function of dendritic cells (DC), such as that achieved by microbial stimuli or T cell help, plays a critical role in determining the quality and size of adaptive responses to Ag. NKT cells bearing an invariant TCR ( i NKT cells) restricted by nonpolymorphic CD1d molecules may constitute a readily available source of help for DC. We therefore examined T cell responses to i.v. injection of soluble Ag in the presence or the absence of i NKT cell stimulation with the CD1d-binding glycolipid α-galactosylceramide (α-GalCer). Considerably enhanced CD4 + and CD8 + T cell responses were observed when α-GalCer was administered at the same time as or close to OVA injection. This enhancement was dependent on the involvement of i NKT cells and CD1d molecules and required CD40 signaling. Studies in IFN-γR −/− mice indicated that IFN-γ was not required for the adjuvant effect of α-GalCer. Consistent with this result, enhanced T cell responses were observed using OCH, an analog of α-GalCer with a truncated sphingosine chain and a reduced capacity to induce IFN-γ. Splenic DC from α-GalCer-treated animals expressed high levels of costimulatory molecules, suggesting maturation in response to i NKT cell activation. Furthermore, studies with cultured DC indicated that potentiation of T cell responses required presentation of specific peptide and α-GalCer by the same DC, implying conditioning of DC by i NKT cells. The i NKT-enhanced T cell responses resisted challenge with OVA-expressing tumors, whereas responses induced in the absence of i NKT stimulation did not. Thus, i NKT cells exert a significant influence on the efficacy of immune responses to soluble Ag by modulating DC function.

Tumor Lymphangiogenesis : A Novel Prognostic Indicator for Cutaneous Melanoma Metastasis and Survival

Abstract: Malignant melanomas of the skin are distinguished by their propensity for early metastatic spread via lymphatic vessels to regional lymph nodes, and lymph node metastasis is a major determinant for the staging and clinical management of melanoma. However, the importance of tumor-induced lymphangiogenesis for lymphatic melanoma spread has remained unclear. We investigated whether tumor lymphangiogenesis occurs in human malignant melanomas of the skin and whether the extent of tumor lymphangiogenesis may be related to the risk for lymph node metastasis and to patient survival, using double immunostains for the novel lymphatic endothelial marker LYVE-1 and for the panvascular marker CD31. Tumor samples were obtained from clinically and histologically closely matched cases of primary melanomas with early lymph node metastasis (n = 18) and from nonmetastatic melanomas (n = 19). Hot spots of proliferating intratumoral and peritumoral lymphatic vessels were detected in a large number of melanomas. The incidence of intratumoral lymphatics was significantly higher in metastatic melanomas and correlated with poor disease-free survival. Metastatic melanomas had significantly more and larger tumor-associated lymphatic vessels, and a relative lymphatic vessel area of >1.5% was significantly associated with poor disease-free and overall survival. In contrast, no differences in the density of tumor-associated blood vessels were found. Vascular endothelial growth factor and vascular endothelial growth factor-C expression was equally detected in a minority of cases in both groups. Our results reveal tumor lymphangiogenesis as a novel prognostic indicator for the risk of lymph node metastasis in cutaneous melanoma.

Multilocus Sequence Typing and Evolutionary Relationships among the Causative Agents of Melioidosis and Glanders, Burkholderia pseudomallei and Burkholderia mallei

Abstract: A collection of 147 isolates of Burkholderia pseudomallei, B. mallei, and B. thailandensis was characterized by multilocus sequence typing (MLST). The 128 isolates of B. pseudomallei, the causative agent of melioidosis, were obtained from diverse geographic locations, from humans and animals with disease, and from the environment and were resolved into 71 sequence types. The utility of the MLST scheme for epidemiological investigations was established by analyzing isolates from captive marine mammals and birds and from humans in Hong Kong with melioidosis. MLST gave a level of resolution similar to that given by pulsed-field gel electrophoresis and identified the same three clones causing disease in animals, each of which was also associated with disease in humans. The average divergence between the alleles of B. thailandensis and B. pseudomallei was 3.2%, and there was no sharing of alleles between these species. Trees constructed from differences in the allelic profiles of the isolates and from the concatenated sequences of the seven loci showed that the B. pseudomallei isolates formed a cluster of closely related lineages that were fully resolved from the cluster of B. thailandensis isolates, confirming their separate species status. However, isolates of B. mallei, the causative agent of glanders, recovered from three continents over a 30-year period had identical allelic profiles, and the B. mallei isolates clustered within the B. pseudomallei group of isolates. Alleles at six of the seven loci in B. mallei were also present within B. pseudomallei isolates, and B. mallei is a clone of B. pseudomallei that, on population genetics grounds, should not be given separate species status.

Regulation of Blood and Lymphatic Vascular Separation by Signaling Proteins SLP-76 and Syk

Abstract: Lymphatic vessels develop from specialized endothelial cells in preexisting blood vessels, but the molecular signals that regulate this separation are unknown. Here we identify a failure to separate emerging lymphatic vessels from blood vessels in mice lacking the hematopoietic signaling protein SLP-76 or Syk. Blood-lymphatic connections lead to embryonic hemorrhage and arteriovenous shunting. Expression of slp-76 could not be detected in endothelial cells, and blood-filled lymphatics also arose in wild-type mice reconstituted with SLP-76-deficient bone marrow. These studies reveal a hematopoietic signaling pathway required for separation of the two major vascular networks in mammals.

Predominant role of hypoxia-inducible transcription factor (Hif)-1alpha versus Hif-2alpha in regulation of the transcriptional response to hypoxia.

Abstract: Tumor hypoxia induces the up-regulation of a gene program associated with angiogenesis, glycolysis, adaptation to pH, and apoptosis via the hypoxia-inducible transcription factors (Hifs) 1 and 2. Disruption of this pathway has been proposed as a cancer therapy. Here, we use short interfering RNAs to compare specific inactivation of Hif-1alpha or Hif-2alpha and show markedly different cell type-specific effects on gene expression and cell migration. Remarkably, among a panel of hypoxia-inducible genes, responses were critically dependent on Hif-1 alpha but not Hif-2 alpha in both endothelial and breast cancer cells but critically dependent on Hif-2 alpha in renal carcinoma cells.

RecQ helicases: suppressors of tumorigenesis and premature aging.

Abstract: The RecQ helicases represent a subfamily of DNA helicases that are highly conserved in evolution. Loss of RecQ helicase function leads to a breakdown in the maintenance of genome integrity, in particular hyper-recombination. Germ-line defects in three of the five known human RecQ helicases give rise to defined genetic disorders associated with cancer predisposition and/or premature aging. These are Bloom's syndrome, Werner's syndrome and Rothmund-Thomson syndrome, which are caused by defects in the genes BLM, WRN and RECQ4 respectively. Here we review the properties of RecQ helicases in organisms from bacteria to humans, with an emphasis on the biochemical functions of these enzymes and the range of protein partners that they operate with. We will discuss models in which RecQ helicases are required to protect against replication fork demise, either through prevention of fork breakdown or restoration of productive DNA synthesis.

Enhanced Immunogenicity and Protective Efficacy Against Mycobacterium tuberculosis of Bacille Calmette-Guérin Vaccine Using Mucosal Administration and Boosting with a Recombinant Modified Vaccinia Virus Ankara

Abstract: Heterologous prime-boost immunization strategies can evoke powerful T cell immune responses and may be of value in developing an improved tuberculosis vaccine. We show that recombinant modified vaccinia virus Ankara, expressing Mycobacterium tuberculosis Ag 85A (M.85A), strongly boosts bacille Calmette-Guerin (BCG)-induced Ag 85A specific CD4(+) and CD8(+) T cell responses in mice. A comparison of intranasal (i.n.) and parenteral immunization of BCG showed that while both routes elicited comparable T cell responses in the spleen, only i.n. delivery elicited specific T cell responses in the lung lymph nodes, and these responses were further boosted by i.n. delivery of M.85A. Following aerosol challenge with M. tuberculosis, i.n. boosting of BCG with either BCG or M.85A afforded unprecedented levels of protection in both the lungs (2.5 log) and spleens (1.5 log) compared with naive controls. Protection in the lung correlated with the induction of Ag 85A-specific, IFN-gamma-secreting T cells in lung lymph nodes. These findings support further evaluation of mucosally targeted prime-boost vaccination approaches for tuberculosis.

The Phenotypic Consequences of CFTR Mutations

Abstract: Cystic fibrosis is a common autosomal recessive disorder that primarily affects the epithelial cells in the intestine, respiratory system, pancreas, gall bladder and sweat glands. Over one thousand mutations have currently been identified in the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene that are associated with CF disease. There have been many studies on the correlation of the CFTR genotype and CF disease phenotype; however, this relationship is still not well understood. A connection between CFTR genotype and disease manifested in the pancreas has been well described, but pulmonary disease appears to be highly variable even between individuals with the same genotype. This review describes the current classification of CFTR mutation classes and resulting CF disease phenotypes. Complex disease alleles and modifier genes are discussed along with alternative disorders, such as disseminated bronchiectasis and pancreatitis, which are also thought to result from CFTR mutations.

Characterization of encapsulated and noncapsulated Haemophilus influenzae and determination of phylogenetic relationships by multilocus sequence typing.

Abstract: A multilocus sequence typing (MLST) scheme has been developed for the unambiguous characterization of encapsulated and noncapsulated Haemophilus influenzae isolates The sequences of internal fragments of seven housekeeping genes were determined for 131 isolates, comprising a diverse set of 104 serotype a, b, c, d, e, and f isolates and 27 noncapsulated isolates Many of the encapsulated isolates had previously been characterized by multilocus enzyme electrophoresis (MLEE), and the validity of the MLST scheme was established by the very similar clustering of isolates obtained by these methods Isolates of serotypes c, d, e, and f formed monophyletic groups on a dendrogram constructed from the differences in the allelic profiles of the isolates, whereas there were highly divergent lineages of both serotype a and b isolates Noncapsulated isolates were distinct from encapsulated isolates and, with one exception, were within two highly divergent clusters The relationships between the major lineages of encapsulated H influenzae inferred from MLEE data could not be discerned on a dendrogram constructed from differences in the allelic profiles, but were apparent on a tree reconstructed from the concatenated nucleotide sequences Recombination has not therefore completely eliminated phylogenetic signal, and in support of this, for encapsulated isolates, there was significant congruence between many of the trees reconstructed from the sequences of the seven individual loci Congruence was less apparent for noncapsulated isolates, suggesting that the impact of recombination is greater among noncapsulated than encapsulated isolates The H influenzae MLST scheme is available at wwwmlstnet, it allows any isolate to be compared with those in the MLST database, and (for encapsulated isolates) it assigns isolates to their phylogenetic lineage, via the Internet