A patient was identified who had been continuously colonized with Campylobacter organisms for over a decade. Whole-genome sequencing and antimicrobial susceptibility testing suggested that the ...Campylobacter population had adapted to the host by altering motility and becoming resistant to prescribed antibiotics.
Abstract
Background
Campylobacteriosis is inflammation of the gastrointestinal tract as a result of Campylobacter infection. Most campylobacteriosis cases are acute and self-limiting, with Campylobacter excretion ceasing a few weeks after symptoms cease. We identified a patient with fecal specimens positive for Campylobacter jejuni (ST45) intermittently during a 10-year period.
Methods
Sixteen Campylobacter isolates were collected from the patient during 2006–2016. The isolates’ genomes were sequenced to determine their relatedness, and their antimicrobial susceptibility patterns and motility were measured to determine the effects of antibiotic therapy and long-term excretion on the Campylobacter population.
Results
Phylogenetic analyses estimated that the isolates shared a date of common ancestor between 1998 and 2006, coinciding with the onset of symptoms for the patient. Genomic analysis identified selection for changes in motility, and antimicrobial susceptibility testing suggested that the Campylobacter population developed resistance to several antibiotics coinciding with periods of antibiotic therapy.
Conclusions
The patient was consistently colonized with organisms from a Campylobacter population that adapted to the internal environment of the patient. Genomic and phylogenetic analyses can give insight into a patient’s infection history and the effect of antimicrobial treatment on Campylobacter populations in this unusual situation of long-term colonization of an individual.
Integrated surveillance of infectious multi-source diseases using a combination of epidemiology, ecology, genetics and evolution can provide a valuable risk-based approach for the control of ...important human pathogens. This includes a better understanding of transmission routes and the impact of human activities on the emergence of zoonoses. Until recently New Zealand had extraordinarily high and increasing rates of notified human campylobacteriosis, and our limited understanding of the source of these infections was hindering efforts to control this disease. Genetic and epidemiological modeling of a 3-year dataset comprising multilocus sequence typed isolates from human clinical cases, coupled with concurrent data on food and environmental sources, enabled us to estimate the relative importance of different sources of human disease. Our studies provided evidence that poultry was the leading cause of human campylobacteriosis in New Zealand, causing an estimated 58–76% of cases with widely varying contributions by individual poultry suppliers. These findings influenced national policy and, after the implementation of poultry industry-specific interventions, a dramatic decline in human notified cases was observed in 2008. The comparative-modeling and molecular sentinel surveillance approach proposed in this study provides new opportunities for the management of zoonotic diseases.
A community resource project recently launched by the National Human Genome Research Institute to sequence large-insert clones from many individuals, systematically discovering and resolving these ...complex variants at the DNA sequence level, is described. The project includes the discovery of variants through development of clone resources, sequence resolution of variants, and accurate typing of variants in individuals of African, European or Asian ancestry.
Cancers with chromosomal instability (CIN) are held to be aneuploid/polyploid with multiple large-scale gains/deletions, but the processes underlying CIN are unclear and different types of CIN might ...exist. We investigated colorectal cancer cell lines using array-comparative genomic hybridization (CGH) for copy number changes and single-copy number polymorphism (SNP) microarrays for allelic loss (LOH). Many array-based CGH changes were not found by LOH because they did not cause true reduction-to-homozygosity. Conversely, many regions of SNP-LOH occurred in the absence of copy number change, comprising an average per cell line of 2 chromosomes with complete LOH; 1-2 terminal regions of LOH (mitotic recombination); and 1 interstitial region of LOH. SNP-LOH detected many novel changes, representing possible locations of uncharacterized tumor suppressor loci. Microsatellite unstable (MSI+) lines infrequently showed gains/deletions or whole-chromosome LOH, but their near-diploid karyotypes concealed mitotic recombination frequencies similar to those of MSI- lines. We analyzed p53 and chromosome 18q (SMAD4) in detail, including mutation screening. Almost all MSI- lines showed LOH and/or deletion of p53 and 18q; some near-triploid lines had acquired three independent changes at these loci. We found consistent results in primary colorectal cancers. Overall, the distributions of mitotic recombination and whole-chromosome LOH in the MSI- cell lines differed significantly from random, with some lines having much higher than expected levels of these changes. Moreover, lines with more LOH changes had significantly fewer copy number changes. These data suggest that CIN is not synonymous with copy number change and some cancers have a specific tendency to whole-chromosome deletion and regain or to mitotic recombination.
The recently described DNA replication-based mechanisms of fork stalling and template switching (FoSTeS) and microhomology-mediated break-induced replication (MMBIR) were previously shown to catalyze ...complex exonic, genic and genomic rearrangements. By analyzing a large number of isochromosomes of the long arm of chromosome X (i(Xq)), using whole-genome tiling path array comparative genomic hybridization (aCGH), ultra-high resolution targeted aCGH and sequencing, we provide evidence that the FoSTeS and MMBIR mechanisms can generate large-scale gross chromosomal rearrangements leading to the deletion and duplication of entire chromosome arms, thus suggesting an important role for DNA replication-based mechanisms in both the development of genomic disorders and cancer. Furthermore, we elucidate the mechanisms of dicentric i(Xq) (idic(Xq)) formation and show that most idic(Xq) chromosomes result from non-allelic homologous recombination between palindromic low copy repeats and highly homologous palindromic LINE elements. We also show that non-recurrent-breakpoint idic(Xq) chromosomes have microhomology-associated breakpoint junctions and are likely catalyzed by microhomology-mediated replication-dependent recombination mechanisms such as FoSTeS and MMBIR. Finally, we stress the role of the proximal Xp region as a chromosomal rearrangement hotspot.
G- and R-bands of metaphase chromosomes are characterized by profound differences in gene density, CG content, replication timing, and chromatin compaction. The preferential localization of ...gene-dense, transcriptionally active, and early replicating chromatin in the nuclear interior and of gene-poor, later replicating chromatin at the nuclear envelope has been demonstrated to be evolutionary-conserved in various cell types. Yet, the impact of different local chromatin features on the radial nuclear arrangement of chromatin is still not well understood. In particular, it is not known whether radial chromatin positioning is preferentially shaped by local gene density per se or by other related parameters such as replication timing or transcriptional activity. The interdependence of these distinct chromatin features on the linear deoxyribonucleic acid (DNA) sequence precludes a simple dissection of these parameters with respect to their importance for the reorganization of the linear DNA organization into the distinct radial chromatin arrangements observed in the nuclear space. To analyze this problem, we generated probe sets of pooled bacterial artificial chromosome (BAC) clones from HSA 11, 12, 18, and 19 representing R/G-band-assigned chromatin, segments with different gene density and gene loci with different expression levels. Using multicolor 3D flourescent in situ hybridization (FISH) and 3D image analysis, we determined their localization in the nucleus and their positions within or outside the corresponding chromosome territory (CT). For each BAC data on local gene density within 2- and 10-Mb windows, as well as GC (guanine and cytosine) content, replication timing and expression levels were determined. A correlation analysis of these parameters with nuclear positioning revealed regional gene density as the decisive parameter determining the radial positioning of chromatin in the nucleus in contrast to band assignment, replication timing, and transcriptional activity. We demonstrate a polarized distribution of gene-dense vs gene-poor chromatin within CTs with respect to the nuclear border. Whereas we confirm previous reports that a particular gene-dense and transcriptionally highly active region of about 2 Mb on 11p15.5 often loops out from the territory surface, gene-dense and highly expressed sequences were not generally found preferentially at the CT surface as previously suggested.
Using published high-resolution data on S-phase replication timing, we determined the three-dimensional (3D) nuclear arrangement of 33 very-early-replicating and 31 very-late-replicating loci. We ...analyzed diploid human, non-human primate and rearranged tumor cells by 3D fluorescence in situ hybridization with the aim of investigating the impact of chromosomal structural changes on the nuclear organization of these loci. Overall, their topology was found to be largely conserved between cell types, species and in tumor cells. Early-replicating loci were localized in the nuclear interior, whereas late-replicating loci showed a broader distribution with a higher preference for the periphery than for late-BrdU-incorporation foci. However, differences in the spatial arrangement of early and late loci of chromosome 2, as compared with those from chromosome 5, 7 and 17, argue against replication timing as a major driving force for the 3D radial genome organization in human lymphoblastoid cell nuclei. Instead, genomic properties, and local gene density in particular, were identified as the decisive parameters. Further detailed comparisons of chromosome 7 loci in primate and tumor cells suggest that the inversions analyzed influence nuclear topology to a greater extent than the translocations, thus pointing to geometrical constraints in the 3D conformation of a chromosome territory.
Array painting is a technique that uses microarray technology to rapidly map chromosome translocation breakpoints. Previous methods to map translocation breakpoints have used fluorescence in situ ...hybridization (FISH) and have consequently been labor-intensive, time-consuming and restricted to the low breakpoint resolution imposed by the use of metaphase chromosomes. Array painting combines the isolation of derivative chromosomes (chromosomes with translocations) and high-resolution microarray analysis to refine the genomic location of translocation breakpoints in a single experiment. In this protocol, we describe array painting by isolation of derivative chromosomes using a MoFlo flow sorter, amplification of these derivatives using whole-genome amplification and hybridization onto commercially available oligonucleotide microarrays. Although the sorting of derivative chromosomes is a specialized procedure requiring sophisticated equipment, the amplification, labeling and hybridization of DNA is straightforward, robust and can be completed within 1 week. The protocol described produces good quality data; however, array painting is equally achievable using any combination of the available alternative methodologies for chromosome isolation, amplification and hybridization.
Mutations in the transcription factor encoding TFAP2A gene underlie branchio-oculo-facial syndrome (BOFS), a rare dominant disorder characterized by distinctive craniofacial, ocular, ectodermal and ...renal anomalies. To elucidate the range of ocular phenotypes caused by mutations in TFAP2A, we took three approaches. First, we screened a cohort of 37 highly selected individuals with severe ocular anomalies plus variable defects associated with BOFS for mutations or deletions in TFAP2A. We identified one individual with a de novo TFAP2A four amino acid deletion, a second individual with two non-synonymous variations in an alternative splice isoform TFAP2A2, and a sibling-pair with a paternally inherited whole gene deletion with variable phenotypic expression. Second, we determined that TFAP2A is expressed in the lens, neural retina, nasal process, and epithelial lining of the oral cavity and palatal shelves of human and mouse embryos—sites consistent with the phenotype observed in patients with BOFS. Third, we used zebrafish to examine how partial abrogation of the fish ortholog of TFAP2A affects the penetrance and expressivity of ocular phenotypes due to mutations in genes encoding bmp4 or tcf7l1a. In both cases, we observed synthetic, enhanced ocular phenotypes including coloboma and anophthalmia when tfap2a is knocked down in embryos with bmp4 or tcf7l1a mutations. These results reveal that mutations in TFAP2A are associated with a wide range of eye phenotypes and that hypomorphic tfap2a mutations can increase the risk of developmental defects arising from mutations at other loci.
As normal as normal can be? Carter, Nigel P
Nature genetics,
200409, 2004-Sep, 2004-9-00, 20040901, Letnik:
36, Številka:
9
Journal Article
Recenzirano
Odprti dostop
Two papers report that large-scale copy-number variations, ranging in size from 100 kb to 2 Mb, are distributed widely throughout the human genome, and that a high proportion of them encompass known ...genes. This unexpected level of genome variation has implications for our view of human genetic diversity and phenotypic variation.
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