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U Römling

Publications and source records attributed to U Römling.

46 records · Page 3Linked to original sources
PubMed1994

A major Pseudomonas aeruginosa clone common to patients and aquatic habitats.

The genomic relatedness of 573 Pseudomonas aeruginosa strains from environmental and clinical habitats was examined by digesting the genome with the rare-cutting enzyme SpeI. Thirty-nine strains were collected from environmental habitats mainly of aquatic origin, like rivers, lakes, or sanitary facilities. Four hundred fifty strains were collected from 76 patients with cystic fibrosis (CF) treated at four different centers, and 25 additional clinical isolates were collected from patients suffering from other diseases. Twenty-nine P. aeruginosa isolates were collected from the environment of one CF clinic. Thirty strains from culture collections were of environmental and clinic origin. A common macrorestriction fingerprint pattern was found in 13 of 46 CF patients, 5 of 29 environmental isolates from the same hospital, in a single ear infection isolate from another hospital, and 8 of 38 isolates from aquatic habitats about 300 km away from the CF clinic. The data indicate that closely related variants of one major clone (called clone C) persisted in various spatially and temporally separated habitats. Southern analysis of the clonal variants with six gene probes and two probes for genes coding for rRNA revealed almost the same hybridization patterns. With the exception of the phenotypically rapidly evolving CF isolates, the close relatedness of the strains of the clone was also shown by their identical responses in pyocin typing, phage typing, and serotyping. Besides clone C, three other P. aeruginosa clones were isolated from more than one clinical or environmental source.

Cystic Fibrosis↗
PubMed1993

Comparative mapping of the Pseudomonas aeruginosa PAO genome with rare-cutter linking clones or two-dimensional pulsed-field gel electrophoresis protocols.

The Spe1 map of the Pseudomonas aeruginosa PAO (DSM 1707) chromosome was constructed by utilizing two-dimensional pulsed-field gel electrophoresis (PFGE) and rare-cutter linking clones. After end-labeling and fluorescence staining of macrorestriction fragments had been combined, the two-dimensional PFGE analyses of partial-total digests and reciprocal double digests were sufficient for the placement of all fragments on the genomic map. Spe1 linking fragments were isolated from BamH1, Pst1, and EcoR1 genomic libraries of P. aeruginosa PAO. After separation of the heterogeneously sized populations of Spe1-linearized and uncut circular plasmid DNAs by field inversion polyacrylamide gel electrophoresis, the gel-eluted linear DNAs were recircularized and subcloned. The 46 analyzed Spe1 linking clones recognized 16 of the 38 fragment links on the Spe1 genome map of P. aeruginosa PAO. The alignment with linking clones was consistent with that obtained from two-dimensional PFGE mapping protocols.

DNA, Bacterial↗
PubMed1992

Localization of alg, opr, phn, pho, 4.5S RNA, 6S RNA, tox, trp, and xcp genes, rrn operons, and the chromosomal origin on the physical genome map of Pseudomonas aeruginosa PAO.

The genes encoding the rrn operons, the 4.5S and 6S RNAs, elements of protein secretion, and outer membrane proteins F and I, and regulatory as well as structural genes for exotoxin A, alkaline phosphatase, and alginate and tryptophan biosynthesis, were assigned on the SpeI/DpnI macrorestriction map of the Pseudomonas aeruginosa PAO chromosome. The zero point of the map was relocated to the chromosomal origin of replication.

Chromosome Mapping↗
PubMed1991

The impact of two-dimensional pulsed-field gel electrophoresis techniques for the consistent and complete mapping of bacterial genomes: refined physical map of Pseudomonas aeruginosa PAO.

The SpeI/DpnI map of the 5.9 Mb Pseudomonas aeruginosa PAO (DSM 1707) genome was refined by two-dimensional (2D) pulsed-field gel electrophoresis techniques (PFGE) which allow the complete and consistent physical mapping of any bacterial genome of interest. Single restriction digests were repetitively separated by PFGE employing different pulse times and ramps in order to detect all bands with optimum resolution. Fragment order was evaluated from the pattern of 2D PFGE gels: 1. Partial-complete digestion. A partial restriction digest was separated in the first dimension, redigested to completion, and subsequently perpendicularly resolved in the second dimension. 2D-gel comparisons of the ethidium bromide stain of all fragments and of the autoradiogram of end-labeled partial digestion fragments was nearly sufficient for the construction of the macrorestriction map. 2. Reciprocal gels. A complete restriction digest with enzyme A was run in the first dimension, redigested with enzyme B, and separated in the second orthogonal direction. The order of restriction digests was reverse on the second gel. In case of two rare-cutters, fragments were visualized by ethidium bromide staining or hybridization with genomic DNA. If a frequent and a rare cutter were employed, linking fragments were identified by end-labeling of the first digest. 3. A few small fragments were isolated by preparative PFGE and used as a probe for Southern analysis.--38 SpeI and 15 DpnI fragments were positioned on the map. The zero point was relocated to the 'origin of replication'. The anonymous mapping techniques described herein are unbiased by repetitive DNA, unclonable genomic regions, unfavourable location of restriction sites, or cloning artifacts as frequently encountered in other top-down or bottom-up approaches.

Blotting, Southern↗
PubMed1991

Physical and genetic map of the Neisseria gonorrhoeae strain MS11-N198 chromosome.

A macro-restriction map of the Neisseria gonorrhoeae chromosome was constructed using the enzymes Nhel and Spel. Combinations of one- and two-dimensional electrophoresis of completely or partially digested chromosomal DNA were performed to align the restriction fragments. The chromosome is circular, with an estimated size of 2.33 Mb +/- 35 kb. A genetic map was derived from the physical map; positions of over 60 defined loci were determined by Southern hybridization.

Blotting, Southern↗
PubMed1989

A physical genome map of Pseudomonas aeruginosa PAO.

A complete macrorestriction map of the 5.9 Mb genome of Pseudomonas aeruginosa PAO (DSM 1707) was constructed by the combination of various one- and two-dimensional pulsed field gel electrophoresis techniques. A total of 51 restriction sites (36 SpeI sites, 15 DpnI sites) were placed on the physical map yielding an average resolution of 110 kb. Several genes encoding virulence factors and enzymes of metabolic pathways were located on the anonymous map by Southern hybridization. Distances between the gene loci were similar on the genetic and physical maps, suggesting an even distribution of genome mobility throughout the bacterial chromosome. The four rRNA operons were organized in pairs of inverted repeats. The two-dimensional macro-restriction techniques described herein are generally applicable for the genome mapping of any prokaryote and lower eukaryote which yields resolvable fragment patterns on two-dimensional pulsed field gels.

Base Sequence↗
PubMedDate not supplied

Physical genome analysis of bacteria.

Pulsed-field gel electrophoresis (PFGE) is a general analytical tool to separate large DNA molecules and may therefore be applied to problems from all areas of bacteriology. The genome size of bacteria covers the range of 0.6 to 10 megabase pairs. For genome fingerprinting, the bacterial chromosome is cleaved with a restriction endonuclease that gives a resolvable and informative number of five to one hundred fragments on the PFGE gel. Restriction enzymes are chosen according to GC content, degree of methylation, and codon usage of the respective bacterial genus. Macrorestriction fingerprinting allows the identification of bacterial strains and the distinction between related and unrelated strains. If fragment patterns of several restriction digestions are quantitatively evaluated, strains can be classified according to genetic relatedness at the level of genus, species, and biovar. In particular, members of a clonal lineage can be uncovered. Hence, any problem from applied, environmental, and clinical microbiology may be addressed by PFGE restriction analysis where the spatiotemporal spread of a bacterial clone is of interest. In bacterial genomics, PFGE is employed for the top-down construction of macrorestriction maps of the chromosome which yields data about genome organization, mobile genetic elements, and the arrangement of gene loci and gene families. The genomic diversity of a bacterial species is elucidated by comparative chromosome mapping. Map positions of restriction sites and gene loci of interest serve as landmarks to assess the extent of gross chromosomal modification, namely insertions, deletions and inversions. Intra- and interspecies comparisons of genome organization provide insights into the structure and diversity of bacterial populations and the phylogeny of bacterial taxa.

Bacteria↗
PubMedDate not supplied

Pulsed-field gel electrophoresis analysis of a Pseudomonas aeruginosa pathovar.

This paper presents the genome organization and mobility of Pseudomonas aeruginosa strains that had been isolated in half-year intervals from 30 patients with cystic fibrosis since the onset of colonization over a 2- to 8-year period. The chromosomes were digested with DraI or SpeI, separated by pulsed-field gel electrophoresis, blotted and hybridized with probes encoding housekeeping or virulence genes. Strains were differentiated by relatedness of macrorestriction fingerprints. After some turnover of strains during the first two years of colonization, each patient had acquired a set of strains that diversified during the course of the disease. In the majority of patients, two clonal lineages were found to account for colonization in the air passages but each lung habitat was characterized by some specific signature of bands in the macrorestriction fragment pattern.

Cystic Fibrosis↗
PubMedDate not supplied

A PacI/SwaI map of the Pseudomonas aeruginosa PAO chromosome.

The PacI/SwaI macrorestriction map of the 5.9 Mb Pseudomonas aeruginosa PAO (DSM 1707) genome was constructed by fragment size comparisons of complete single and double digests and hybridization with probes of known genomic localization. P. aeruginosa PAO contains five recognition sites for PacI and six sites for SwaI. An integrated DpnI/PacI/SpeI/SwaI map was obtained by fragment pattern and Southern blot hybridization analyses of all combinations of double digestions and by two-dimensional pulsed-field gel electrophoresis mapping protocols.

Base Sequence↗